<QaBoxq="A therapy for which organ would benefit most people that have worse health than you do?"a="Probably the lung. The pancreas is important too, but stomach problems are usually less pressing than difficulty in breathing."/>
<QaBoxq="You mentioned that doing sport is difficult with CF, why?"a="Hygiene. In the lockers and the showers but also with the equipment."/>
<QaBoxq="Do you feel restricted in your free time activities?"a="No, I always had good alternatives. For example, going swimming at an open-air swimming pool instead of a lake. "/>
<QaBoxq="Would you have more freedom when you are better protected from Pseudomonas spcc. and other potential infections? "a="text"/>
<QaBoxq="text"a="Definitely. That is a big increase in the quality of life and that is a win. It also changes the picture people have of the illness. Of course being protected by prevention is good already but effective therapies for infections increase the sense of freedom even more. "/>
<QaBoxq="Would you have more freedom when you are better protected from Pseudomonas spcc. and other potential infections? "a="Definitely. That is a big increase in the quality of life and that is a win. It also changes the picture people have of the illness. Of course being protected by prevention is good already but effective therapies for infections increase the sense of freedom even more. "/>
<QaBoxq="You said you are afraid every time you must go for a swab, why is that? "a="I am afraid of getting an infection. That still could be a death sentence. "/>
<QaBoxq="Are rooms with air conditioning a problem due to the possible germs in the air conditioners? "a="No, there is usually enough movement. But humidifiers are bad because of the pond water. "/>
<QaBoxq="You mentioned going to the hairdresser is problematic. Could you elaborate? "a="There are many possible sources of ponding water and with that, infections. That and the hygiene aspect in general. I am visited by my hairdresser, and he only uses a specific spray bottle to wet my hair that I keep and dry thoroughly between uses."/>
heading:"Interview with a specialized physiotherapist regarding breathing therapy for cystic fibrosis patients",
interviewtabid:"westhoffinv",
cardtext:"",
language:"de",
quote:"The more we know, the more opportunities we have.",
aimofcontact:"The objective of the contact was to gain in-depth insights into the treatment and care of children with cystic fibrosis. The therapist's expertise was intended to help develop a better understanding of the challenges and necessary measures in the treatment of this chronic disease. In addition, the aim was to ascertain how the therapy is implemented in everyday life and which specific approaches and methods are particularly effective.",
insights:"The interview yielded valuable insights into the regular implementation of the therapy, the use of aids and the adaptation of exercises to the individual needs of the patients. It was notable that the therapy has improved considerably thanks to better medication and adapted exercises, with a concomitant increase in life expectancy for children affected by cystic fibrosis. Of particular interest was the emphasis on the importance of sport and exercise, which should not only be therapeutically effective, but also increase quality of life. ",
implementation:"The following statement by Katrin Westhoff had a particularly profound impact on our project: 'The more we know, the more opportunities we have.' We learned from the interview that the current medication is already helping many patients to a huge extent, but that there is still a significant opportunity for improvement. After all, successful gene therapy would markedly enhance the quality of life for those affected. The findings of this project will be disseminated to the relevant researchers in order to facilitate the rapid improvement of the quality of life of all cystic fibrosis patients, regardless of their mutation. ",
summary:"The objective of our discussion with a therapist was to gain a comprehensive understanding of the treatment and care of children with cystic fibrosis. The interview provided invaluable insights into the therapy's implementation, highlighting the significant advancements in medication and tailored exercises that have led to improved patient outcomes and increased life expectancy. A key takeaway was the emphasis on the role of sports and exercise, not just for therapeutic efficacy but also for enhancing overall quality of life. It underscored the potential for further enhancements in care through successful gene therapy, motivating us to share our findings with relevant researchers to help elevate the quality of life for all cystic fibrosis patients, regardless of their genetic mutations.",
months:"May",
interview:<>
<QaBoxq="From what age do the patients come to you? How long do they stay? How many patients do you treat?"a="The patients come to us at a very early age. A definite diagnosis is made after 6 weeks at the latest. Once diagnosed, the whole family is genetically tested, and children are sent for physiotherapy, often starting in the hospital. Currently, we have 8 children with cystic fibrosis in our practice, which is relatively small compared to other diseases. We have slightly more CF patients because we specialize in it."/>
<QaBoxq="What kind of exercises do you do?"a="We do a lot of breathing therapy and have attended special training courses for CF that introduced new techniques. The current gold standard is autogenous drainage according to Chevallier, which effectively removes mucus. We follow a general routine: 1. wet inhalation to bind mucus, 2. drainage to expel mucus, and 3. antibiotics to work optimally on clean lungs. We also use special belts for compressing 'magic points' to enhance lung ventilation."/>
<QaBoxq="When does drainage start?"a="We start drainage in newborns to prevent mucus from settling."/>
<QaBoxq="Are there special exercises that can also be done at home?"a="Yes, parents are instructed on exercises that can also be performed at home."/>
<QaBoxq="How often does the therapy take place?"a="Therapy usually occurs once a week or every two weeks. Thanks to improved medication, children are now better off. The therapy has evolved significantly, making it easier to cough up mucus and improving life expectancy. Exercise should be enjoyable and a part of daily life from the age of 8 or 9."/>
<QaBoxq="What would happen if no physiotherapy was performed?"a="It’s difficult to predict, but without therapy, children often become more mucousy, leading to worsened ventilation. Specific therapy is crucial, especially during infections."/>
<QaBoxq="How do you measure success (in terms of lung function test, exercise, etc.)?"a="Success is measured subjectively by listening to breathing and observing skin color. A well-ventilated lung shows a 'full barrel' appearance, while wheezing indicates poor ventilation. In clinics, lung function tests, CO2 measurements, and 'finger clip' tests are used, though results can be influenced by the child."/>
<QaBoxq="Do the exercises bring relief or are they preventative for further complaints?"a="The exercises serve both to relieve acute infections and to prevent further issues. Fewer lung infections reduce the likelihood of mucus adhesions."/>
<QaBoxq="Are there any tools to perform therapy?"a="Yes, devices like the 'flutter' or 'cornet' help with exhalation. They create vibrations that loosen mucus in the lungs and should be used by all children with lung diseases."/>
<QaBoxq="What complaints do patients bring with them?"a="Patients typically have lung problems, dry lung mucosa, and pancreatic issues leading to poor metabolism, requiring enzyme therapy before meals. Some children experience growth disorders and less commonly, excessive perspiration."/>
<QaBoxq="Are pancreatic complaints also treated by physiotherapists?"a="Pancreatic complaints are rarely treated with physiotherapy, except in cases of inflammation, where patients may be admitted to the hospital. Techniques like massage or kinesiology tape can help with constipation."/>
<QaBoxq="Are there any special hygiene guidelines for you when working with cystic fibrosis patients?"a="Hygiene is crucial when treating CF patients. We separate children with and without infections (e.g., Pseudomonas) and enforce strict disinfection protocols. Only children with similar infection statuses are treated on the same day."/>
<QaBoxq="Are the specific exercises customized? And if so, how do you know which therapy is the right one for which patient?"a="Exercises are tailored to each patient's situation, focusing on mucus removal and lung ventilation. Each therapist may have their own preferred exercises and techniques."/>
<QaBoxq="Do patients always go to the same physiotherapist?"a="Yes, if therapy is effective, patients tend to remain with the same physiotherapist."/>
<QaBoxq="How many physiotherapists offer muco-therapy?"a="The exact number is unknown, but several child therapists in the region provide cystic fibrosis therapy."/>
<QaBoxq="How are the relatives educated?"a="Education often begins in the maternity ward with a sweat test. In Gütersloh, all children are referred to Bethel for immediate CF care. Parents often experience trauma as children can be severely ill despite appearing healthy."/>
<QaBoxq="What are the limitations of individual medicine?"a="Drug effectiveness can vary, and some are only approved from a certain age. Improved medications can significantly enhance quality of life and life expectancy."/>
summary:"We engaged in a valuable discussion with Jan-Phillip Gerhards regarding prime editing and pegRNAs, leveraging his internship experience with these technologies. Key insights included the effectiveness of silent edits, which can enhance the safety of our PrimeGuide by modifying DNA sequences without altering the resultant protein, thereby preventing pegRNA rebinding. We also learned the importance of optimizing the primer binding site (PBS) length to achieve ideal temperatures close to 37°C, recommending lengths of 16-17 nucleotides. Additionally, we discovered the potential benefits of using non-annotated regions with overhangs for cloning, while also recognizing concerns about circRNA steric hindrance by Cas9. These insights directly informed our project, allowing us to refine our pegRNA design and cloning strategies, ultimately enhancing the precision and efficiency of our gene editing approach.",
months:"May"
},
{
vorname:"Katrin",
nachnname:"Westhoff",
job:"Physiotherapist",
affiliation:"Independent",
pictureurl:pics['westhoff'],
tag:"Medical Professional",
heading:"Interview with a specialized physiotherapist regarding breathing therapy for cystic fibrosis patients",
interviewtabid:"westhoffinv",
cardtext:"",
language:"de",
quote:"The more we know, the more opportunities we have.",
aimofcontact:"The objective of the contact was to gain in-depth insights into the treatment and care of children with cystic fibrosis. The therapist's expertise was intended to help develop a better understanding of the challenges and necessary measures in the treatment of this chronic disease. In addition, the aim was to ascertain how the therapy is implemented in everyday life and which specific approaches and methods are particularly effective.",
insights:"The interview yielded valuable insights into the regular implementation of the therapy, the use of aids and the adaptation of exercises to the individual needs of the patients. It was notable that the therapy has improved considerably thanks to better medication and adapted exercises, with a concomitant increase in life expectancy for children affected by cystic fibrosis. Of particular interest was the emphasis on the importance of sport and exercise, which should not only be therapeutically effective, but also increase quality of life. ",
implementation:"The following statement by Katrin Westhoff had a particularly profound impact on our project: 'The more we know, the more opportunities we have.' We learned from the interview that the current medication is already helping many patients to a huge extent, but that there is still a significant opportunity for improvement. After all, successful gene therapy would markedly enhance the quality of life for those affected. The findings of this project will be disseminated to the relevant researchers in order to facilitate the rapid improvement of the quality of life of all cystic fibrosis patients, regardless of their mutation. ",
summary:"The objective of our discussion with a therapist was to gain a comprehensive understanding of the treatment and care of children with cystic fibrosis. The interview provided invaluable insights into the therapy's implementation, highlighting the significant advancements in medication and tailored exercises that have led to improved patient outcomes and increased life expectancy. A key takeaway was the emphasis on the role of sports and exercise, not just for therapeutic efficacy but also for enhancing overall quality of life. It underscored the potential for further enhancements in care through successful gene therapy, motivating us to share our findings with relevant researchers to help elevate the quality of life for all cystic fibrosis patients, regardless of their genetic mutations.",
months:"May",
interview:<>
<QaBoxq="From what age do the patients come to you? How long do they stay? How many patients do you treat?"a="The patients come to us at a very early age. A definite diagnosis is made after 6 weeks at the latest. Once diagnosed, the whole family is genetically tested, and children are sent for physiotherapy, often starting in the hospital. Currently, we have 8 children with cystic fibrosis in our practice, which is relatively small compared to other diseases. We have slightly more CF patients because we specialize in it."/>
<QaBoxq="What kind of exercises do you do?"a="We do a lot of breathing therapy and have attended special training courses for CF that introduced new techniques. The current gold standard is autogenous drainage according to Chevallier, which effectively removes mucus. We follow a general routine: 1. wet inhalation to bind mucus, 2. drainage to expel mucus, and 3. antibiotics to work optimally on clean lungs. We also use special belts for compressing 'magic points' to enhance lung ventilation."/>
<QaBoxq="When does drainage start?"a="We start drainage in newborns to prevent mucus from settling."/>
<QaBoxq="Are there special exercises that can also be done at home?"a="Yes, parents are instructed on exercises that can also be performed at home."/>
<QaBoxq="How often does the therapy take place?"a="Therapy usually occurs once a week or every two weeks. Thanks to improved medication, children are now better off. The therapy has evolved significantly, making it easier to cough up mucus and improving life expectancy. Exercise should be enjoyable and a part of daily life from the age of 8 or 9."/>
<QaBoxq="What would happen if no physiotherapy was performed?"a="It’s difficult to predict, but without therapy, children often become more mucousy, leading to worsened ventilation. Specific therapy is crucial, especially during infections."/>
<QaBoxq="How do you measure success (in terms of lung function test, exercise, etc.)?"a="Success is measured subjectively by listening to breathing and observing skin color. A well-ventilated lung shows a 'full barrel' appearance, while wheezing indicates poor ventilation. In clinics, lung function tests, CO2 measurements, and 'finger clip' tests are used, though results can be influenced by the child."/>
<QaBoxq="Do the exercises bring relief or are they preventative for further complaints?"a="The exercises serve both to relieve acute infections and to prevent further issues. Fewer lung infections reduce the likelihood of mucus adhesions."/>
<QaBoxq="Are there any tools to perform therapy?"a="Yes, devices like the 'flutter' or 'cornet' help with exhalation. They create vibrations that loosen mucus in the lungs and should be used by all children with lung diseases."/>
<QaBoxq="What complaints do patients bring with them?"a="Patients typically have lung problems, dry lung mucosa, and pancreatic issues leading to poor metabolism, requiring enzyme therapy before meals. Some children experience growth disorders and less commonly, excessive perspiration."/>
<QaBoxq="Are pancreatic complaints also treated by physiotherapists?"a="Pancreatic complaints are rarely treated with physiotherapy, except in cases of inflammation, where patients may be admitted to the hospital. Techniques like massage or kinesiology tape can help with constipation."/>
<QaBoxq="Are there any special hygiene guidelines for you when working with cystic fibrosis patients?"a="Hygiene is crucial when treating CF patients. We separate children with and without infections (e.g., Pseudomonas) and enforce strict disinfection protocols. Only children with similar infection statuses are treated on the same day."/>
<QaBoxq="Are the specific exercises customized? And if so, how do you know which therapy is the right one for which patient?"a="Exercises are tailored to each patient's situation, focusing on mucus removal and lung ventilation. Each therapist may have their own preferred exercises and techniques."/>
<QaBoxq="Do patients always go to the same physiotherapist?"a="Yes, if therapy is effective, patients tend to remain with the same physiotherapist."/>
<QaBoxq="How many physiotherapists offer muco-therapy?"a="The exact number is unknown, but several child therapists in the region provide cystic fibrosis therapy."/>
<QaBoxq="How are the relatives educated?"a="Education often begins in the maternity ward with a sweat test. In Gütersloh, all children are referred to Bethel for immediate CF care. Parents often experience trauma as children can be severely ill despite appearing healthy."/>
<QaBoxq="What are the limitations of individual medicine?"a="Drug effectiveness can vary, and some are only approved from a certain age. Improved medications can significantly enhance quality of life and life expectancy."/>
summary:"Our discussion with Mattijs Bulcaen from KU Leuven provided critical insights into the complexities of using prime editing for cystic fibrosis therapy. As we began designing our prime editor, Mattijs highlighted challenges specific to targeting the CFTR F508del deletion, including the influence of mismatch repair systems and chromatin organization on editing efficiency. He introduced us to advanced techniques, such as PE3b systems and dsgRNAs, and recommended using the 3’ stem loop motif from his research to enhance our pegRNA design. Additionally, he advised utilizing HEK cell lines for screening due to their ease of handling and reduced mismatch repair activity. These insights directly influenced our design choices and helped refine our approach to developing an effective prime editing strategy.",
months:"june"
},
{
vorname:"'Der Teuto ruft'",
nachnname:"",
pictureurl:pics['teuto'],
tag:"Education",
heading:"Educational city tour for young and old",
summary:"Julia's insights shifted our focus to the support systems surrounding CF patients. She highlighted the societal implications of CF, including rising healthcare costs due to the long-term nature of treatment and the financial burdens faced by families. Additionally, Julia emphasized the emotional strain that accompanies the illness, alongside the complexities of parenting a child with CF. Importantly, she affirmed that most children adapt well to inhalative therapies, reinforcing our planned delivery method for gene therapy. This interview enriched our understanding of the challenges faced by families and enabled us to better compare the experiences of CF patients in Germany to those in other countries.",
months:"june"
},
{
title:"Prof. Dr.",
vorname:"David",
nachnname:"Liu",
job:" Richard Merkin Professor and director of the Merkin Institute of Transformative Technologies in Healthcare",
affiliation:"vice chair of the faculty at the Broad Institute of MIT and Harvard",
pictureurl:pics['david'],
tag:"Academia",
language:"en",
heading:"Influence of research by David Liu on our design decisions ",
interviewtabid:"liu",
cardtext:"",
quote:"X",
aimofcontact:[<p>David Liu is the principal investigator responsible for the development of the prime editing systems and his laboratory is actively working on improving prime editors, also for application in CFTR mutation F508del. </p>],
a="It’s mainly interdisciplinary. A lot of funding comes from industry, like BioNTech, or foundations like Mukoviszidose e.V., which funds research on cystic fibrosis. But in terms of practical research, it’s usually biologists or biotechnologists. Without industry support, research can struggle due to a lack of funding, so having backing is essential."/>
</>,
summary:"Our discussion addressed the complexities of cystic fibrosis (CF) treatments, focusing on gene therapy and health insurance processes. We learned about the regulatory challenges gene therapies face, particularly regarding the European Medicines Agency (EMA) and the AMNOG process for reimbursement assessments. Public insurers impose stricter guidelines than private insurers, emphasizing the importance of early intervention in CF and the need for adaptable policies for atypical cases. We recognized the high costs associated with gene therapies and incorporated cost-benefit analysis into our project planning. Following the interview, we refined our approach to include straightforward delivery methods and attended a GxP course for regulatory compliance. Engaging with start-ups further informed our practical implementation strategies, ensuring our project aligns with both scientific and regulatory needs.",
summary:"The aim of the contact was to address challenges in LNP transfection and improve formulation protocols for enhanced gene delivery. Dr. Radukic highlighted the importance of lipid-to-nucleic acid ratios, recommending adjustments like 10:1 for better transfection efficiency. He also emphasized optimizing pH and buffer composition, as well as strict storage and mixing practices. Additionally, quality control measures such as fluorescence testing and zeta potential analysis were suggested to ensure LNP stability. These insights were implemented into the project, improving transfection efficiency and paving the way for future LNP applications.",
months:"June"
},
{
vorname:"Joshua",
nachnname:"Bauder",
job:"parent and activist",
affiliation:"CF vests worldwide",
pictureurl:pics['joshua'],
tag:"Patient",
heading:"Interview with a CF Parent and Global Advocate on Worldwide Support and Perspectives",
interviewtabid:"joshua",
cardtext:"",
language:"en",
quote:"We’ve had to sit by and watch people die, knowing that better treatment exists but is inaccessible. ",
aimofcontact:[<p>We contacted the organization <ahref="https://www.cfvww.org/">CF vests worldwide</a> with the aim to hear more diverse perspectives beyond Germany.
After the founder Rod connected us with Joshua, Joshua was so kind to conduct an interview with us not only about the perspectives and
stories he heard but also about his personal experiences with his daughter and living in a country where CF care is very hard to get. Joshua
(from the USA) and his family live in Thailand where he and his wife run a children’s home. Their daughter is the only child with CF.</p>,
<p>It is possible to learn more about Joshua and his family though the <ahref="https://thebonnellfoundation.org/cf-vests-worldwide/">
podcast of the Bonnel foundation</a>.</p>],
insights:[<p> Joshua showed us just how dire the situation is for CF patients is in some regions. It was shocking to hear there is only one doctor
knowledgeable about CF in Thailand and that many doctors dismiss the possibility of CF due to racial bias and misinformation. Additionally, we confirmed how much the accessibility
of care depends on the healthcare system, as we already touched on during the interview with <HPLinktoOtherHPTabtab="nicole"text="Nicole Friedlein"/>,. On the parenting level, Joshua brought in many perspectives contrary to what we previously heard. In the interview with <HPLinktoOtherHPTabtab="maxfirst"text="Max"/>,, we learned he vehemently avoids ponding water while Joshua’s daughter is allowed to roam around with no such restrictions. Neither have chronic infections.</p>],
implementation:[<p>The interview with Josh made us realize we too needed to look at the reason why we chose F508del. Did we, too, fall for bias?
Despite a change of target not being feasible anymore, we looked into it and traced back our steps that led to our decision. We did not find as much
information about other mutations when first researching cystic fibrosis, especially in the context of prime editing. Mattijs Bulceans's paper on
targeting the mutations L227R and N1303K <TabScrollLinktab="joshua"scrollId="desc-1"num="1"/> was one of few papers. After explicitly searching for cystic fibrosis records for specific countries and
regions, we uncovered a moderate number of papers examining CF in Asia and other regions we previously did not know much about. The very first article
supported Joshua's hypotheses and painted a sad picture: Among other things, it describes the case of a four-month-old boy who was diagnosed with cystic
fibrosis. Nothing unusual in itself, but the circumstances are depressing. Two of the three siblings born before him died within months of birth and had
previously presented with symptoms of cystic fibrosis. He was the first to be diagnosed. A sweat test aimed at cystic fibrosis was not available at the
hospital, so one was improvised. Later on, a genetic test revealed the presence of 508del. <TabScrollLinktab="joshua"scrollId="desc-2"num="2"/> We found ourselves and our lack of knowledge in good
company as we found papers as new as from 2020 (14 years after the previously mentioned paper) containing statements such as “recent reports suggest
that CF does occur in Asia” <TabScrollLinktab="joshua"scrollId="desc-3"num="3"/>. Fortunately, there is a rising number of cystic fibrosis experts for Asia and other previously overlooked regions
such as Africa. <TabScrollLinktab="joshua"scrollId="desc-4"num="4"/> We chose to not only look at the scientific data but also into anecdotal evidence. To find the latter, we searched official
and private websites and chatrooms for information and experiences of patients. In the end, we found narratives from most ethnic backgrounds
about being dismissed and often misdiagnosed. Of course, this is not an occurrence unique to cystic fibrosis. Our conclusion is that yes,
we did fall for bias. But regardless of ethnicity, 508del occurs and is overall the most prevalent mutation as was confirmed in our interview
with CF expert Sriram .... This experience was uncomfortable as we felt the pressure to be thorough and deliver a perfect project. What would
have been more devastating than realizing we made a wrong choice at the very core? We made the conscious decision to invest our resources into
figuring out if we indeed made a mistake and we want to encourage other teams to do the same. iGem stands for innovation – but also for growth.
Especially in the context of Integrated Human Practices, it is important to examine both the positive and the negative to create a project with a
summary:"Joshua, a CF parent living in Thailand, shared his experiences about the severe challenges of accessing CF care in regions like Southeast Asia. His story highlighted the racial bias and lack of medical knowledge about CF in these areas. This interview prompted the team to reflect on their focus on the F508del mutation, questioning if their research was biased towards more commonly studied mutations. After revisiting their research process, they found that the F508del mutation remains globally relevant, yet the experience reinforced the importance of addressing gaps in healthcare and research for underrepresented regions.",
<QaBoxq="Can you educate us about your academic career?"a="I did my doctorate 30 years ago at Bielefeld University and then worked at the Max Planck Institute in Göttingen a lot with the patch-clamp technique. Today, I’m head of the working group Cellular Neurophysiology of the medicine faculty of Bielefeld University."/>
summary:"In summary, through the interview with Prof. Dr. Wischmeyer and the collaboration with his employee Dr. Oliver Dräger, we gained valuable insights and optimized our approach to effectively investigate and measure the functionality of the CFTR ion channel, thereby determining the efficiency of our prime editing strategy.",
months:"june"
},
{
vorname:"Joshua",
nachnname:"Bauder",
job:"parent and activist",
affiliation:"CF vests worldwide",
pictureurl:pics['joshua'],
tag:"Patient",
heading:"Interview with a CF Parent and Global Advocate on Worldwide Support and Perspectives",
interviewtabid:"joshua",
cardtext:"",
language:"en",
quote:"We’ve had to sit by and watch people die, knowing that better treatment exists but is inaccessible. ",
aimofcontact:[<p>We contacted the organization <ahref="https://www.cfvww.org/">CF vests worldwide</a> with the aim to hear more diverse perspectives beyond Germany.
After the founder Rod connected us with Joshua, Joshua was so kind to conduct an interview with us not only about the perspectives and
stories he heard but also about his personal experiences with his daughter and living in a country where CF care is very hard to get. Joshua
(from the USA) and his family live in Thailand where he and his wife run a children’s home. Their daughter is the only child with CF.</p>,
<p>It is possible to learn more about Joshua and his family though the <ahref="https://thebonnellfoundation.org/cf-vests-worldwide/">
podcast of the Bonnel foundation</a>.</p>],
insights:[<p> Joshua showed us just how dire the situation is for CF patients is in some regions. It was shocking to hear there is only one doctor
knowledgeable about CF in Thailand and that many doctors dismiss the possibility of CF due to racial bias and misinformation. Additionally, we confirmed how much the accessibility
of care depends on the healthcare system, as we already touched on during the interview with <HPLinktoOtherHPTabtab="nicole"text="Nicole Friedlein"/>,. On the parenting level, Joshua brought in many perspectives contrary to what we previously heard. In the interview with <HPLinktoOtherHPTabtab="maxfirst"text="Max"/>,, we learned he vehemently avoids ponding water while Joshua’s daughter is allowed to roam around with no such restrictions. Neither have chronic infections.</p>],
implementation:[<p>The interview with Josh made us realize we too needed to look at the reason why we chose F508del. Did we, too, fall for bias?
Despite a change of target not being feasible anymore, we looked into it and traced back our steps that led to our decision. We did not find as much
information about other mutations when first researching cystic fibrosis, especially in the context of prime editing. Mattijs Bulceans's paper on
targeting the mutations L227R and N1303K <TabScrollLinktab="joshua"scrollId="desc-1"num="1"/> was one of few papers. After explicitly searching for cystic fibrosis records for specific countries and
regions, we uncovered a moderate number of papers examining CF in Asia and other regions we previously did not know much about. The very first article
supported Joshua's hypotheses and painted a sad picture: Among other things, it describes the case of a four-month-old boy who was diagnosed with cystic
fibrosis. Nothing unusual in itself, but the circumstances are depressing. Two of the three siblings born before him died within months of birth and had
previously presented with symptoms of cystic fibrosis. He was the first to be diagnosed. A sweat test aimed at cystic fibrosis was not available at the
hospital, so one was improvised. Later on, a genetic test revealed the presence of 508del. <TabScrollLinktab="joshua"scrollId="desc-2"num="2"/> We found ourselves and our lack of knowledge in good
company as we found papers as new as from 2020 (14 years after the previously mentioned paper) containing statements such as “recent reports suggest
that CF does occur in Asia” <TabScrollLinktab="joshua"scrollId="desc-3"num="3"/>. Fortunately, there is a rising number of cystic fibrosis experts for Asia and other previously overlooked regions
such as Africa. <TabScrollLinktab="joshua"scrollId="desc-4"num="4"/> We chose to not only look at the scientific data but also into anecdotal evidence. To find the latter, we searched official
and private websites and chatrooms for information and experiences of patients. In the end, we found narratives from most ethnic backgrounds
about being dismissed and often misdiagnosed. Of course, this is not an occurrence unique to cystic fibrosis. Our conclusion is that yes,
we did fall for bias. But regardless of ethnicity, 508del occurs and is overall the most prevalent mutation as was confirmed in our interview
with CF expert Sriram .... This experience was uncomfortable as we felt the pressure to be thorough and deliver a perfect project. What would
have been more devastating than realizing we made a wrong choice at the very core? We made the conscious decision to invest our resources into
figuring out if we indeed made a mistake and we want to encourage other teams to do the same. iGem stands for innovation – but also for growth.
Especially in the context of Integrated Human Practices, it is important to examine both the positive and the negative to create a project with a
summary:"Joshua, a CF parent living in Thailand, shared his experiences about the severe challenges of accessing CF care in regions like Southeast Asia. His story highlighted the racial bias and lack of medical knowledge about CF in these areas. This interview prompted the team to reflect on their focus on the F508del mutation, questioning if their research was biased towards more commonly studied mutations. After revisiting their research process, they found that the F508del mutation remains globally relevant, yet the experience reinforced the importance of addressing gaps in healthcare and research for underrepresented regions.",
heading:"Lipid Nanoparticles in Gene Therapy: perspectives from Corden Pharma ",
interviewtabid:"corden",
cardtext:"",
language:"de",
quote:"Wow, you’re already further along than I was! That’s a really good approach, especially since dry powder formulations can help with stability.",
aimofcontact:[<p>Our goal in reaching out to Dr. Katharina Kolonko, who earned her PhD working on chitosan-based nanoparticles for delivering nucleic acids to human respiratory cells in the context of cystic fibrosis, was to seek her advice on the design, stability, and application of nanoparticles. We wanted to learn from her experience with chitosan-capsaicin nanoparticles, especially the challenges she encountered, and apply her insights to improve our own project. Specifically, we aimed to better understand nanoparticle stability, transfection methods, and how to effectively design our experiments.
</p>],
insights:[<p>Dr. Kolonko provided us with valuable insights into working with nanoparticles, particularly emphasizing the advantages of nanocapsules. She highlighted that nanocapsules are more stable than nano-complexes, which is crucial for experiments involving complex environments and high shear forces. Additionally, her use of capsaicin wasn’t aimed at improving transfection efficiency but was part of a broader strategy targeting specific channels. She also shared practical methods for measuring particle stability and cytotoxicity, giving us clear guidance on tools and techniques that we can apply to our project.
Furthermore, Dr. Kolonko discussed the use of chitosan as a component in nanoparticle formulations. Chitosan, with its positive charge, can interact with mRNA, potentially enhancing the stability of the cargo. As an outlook, we plan to explore modifications using chitosan to improve the stability and performance of our mRNA delivery system. This approach may provide a more robust solution for optimizing nanoparticle formulations in future experiments. </p>],
implementation:[<p>We directly applied Katharina’s insights to improve our nanoparticle design and testing methods. Her recommendations on using nano-capsules and OptiMEM as the transfection medium helped refine our experimental approach. She emphasized using a medium with fewer additives, like OptiMEM, and suggested removing antibiotics 24 hours before transfection to avoid interference, ensuring more controlled and effective conditions. We also explored new stability testing ideas, including nasal spray solutions and dry powder formulations.
We incorporated her insights as follows: </p>,
language:"en",
quote:"The stability of LNPs depends on the specific lipid and RNA components used, but ensuring the overall stability of a new formulation requires rigorous empirical testing under various conditions.",
aimofcontact:[<p>The primary aim of the communication with Steffen Bira and Serra Gürcan from Corden Pharma was to explore the technical aspects and practical applications of Lipid Nanoparticles (LNPs) in advanced medical therapies, including gene therapy and inhalation treatments. The conversation focused on the possibility of using Corden Pharma’s LNP starter kits, understanding the factors affecting the stability of LNPs, and exploring options for incorporating antibodies into LNPs to target specific cells. </p>],
insights:[<p>The discussion with <ahref="https://cordenpharma.com/">Corden Pharma</a>, led by Steffen Bira and Serra Gürcan, offered key insights into LNPs and their applications. While Corden Pharma hasn't extensively explored spray drying for LNPs, they recommended consulting specialists to evaluate its feasibility, especially concerning lipid stability during the process. Stability was highlighted as crucial for inhalation therapies, requiring thorough testing of entire LNP formulations, possibly aided by cryoprotectants and controlled temperatures.
Corden Pharma's LNP starter kits are based on well-researched lipid combinations designed for stability and encapsulation efficiency, making them suitable for multiple experiments. They suggested that modifying lipid components, such as incorporating cholesterol derivatives, could enhance cellular uptake and overall efficacy. Additionally, they confirmed the possibility of incorporating antibodies into LNPs and emphasized the importance of considering intellectual property when selecting lipids for commercial use. They also showed openness to collaboration,
including offering discounts in exchange for recognition in publications.
The interaction with Corden Pharma provided several key insights:</p>,
<ul>
<li><strong>Nano-Capsules Focus:</strong> Based on Katharina’s advice, we prioritized nano-capsules for their enhanced stability over nano-complexes.</li>
<li><strong>Chitosan for Stability:</strong> We're exploring chitosan to improve mRNA delivery system stability due to its positive charge, which binds mRNA to the nanoparticle surface.</li>
<li><strong>OptiMEM for Transfection:</strong> OptiMEM is now our chosen transfection medium, with the suggestion to remove antibiotics 24 hours prior.</li>
<li><strong>MTT Test for Cytotoxicity:</strong> We adopted the MTT test for cytotoxicity due to its simplicity and reliability.</li>
<li><strong>Nasal Spray and Dry Powder Testing:</strong> We are considering testing nanoparticle stability using nasal spray solutions and exploring dry powder formulations.</li>
<li><strong>Capsaicin Exclusion:</strong> As capsaicin did not significantly impact transfection efficiency in Katharina’s research, we decided not to include it in our project.</li>
<li><strong>Spray Drying Feasibility:</strong> Corden Pharma hasn’t explored spray drying extensively; consultation with specialists is recommended for assessing feasibility and lipid stability.</li>
<li><strong>LNP Stability:</strong> Stability of LNPs, particularly for inhalation therapies, needs empirical testing, considering shear forces and the potential use of cryoprotectants or temperature control.</li>
<li><strong>Lipid Selection in Kits:</strong> Starter kits use well-researched lipid combinations, tested for stability, encapsulation efficiency, and potency. They provide materials for multiple experimental batches.</li>
<li><strong>Lipid Modifications:</strong> Exploring alternative lipids (e.g., cholesterol derivatives) could enhance stability and cellular uptake, tailored to project needs.</li>
<li><strong>Antibody Incorporation:</strong> Antibodies can be incorporated into LNPs during preparation or afterward, depending on targeting requirements.</li>
<li><strong>Intellectual Property:</strong> IP considerations are crucial when selecting lipids for LNP formulations, as many lipids are patented.</li>
<li><strong>Collaboration Opportunities:</strong> Corden Pharma is open to offering discounts or forming partnerships, with recognition in publications or acknowledgments.</li>
</ul>
],
interview:<>
<QaBoxq="How did you approach the design of Lipid Nanoparticles (LNPs)? What were the first steps you took at that time? Were you already familiar with LNPs, or was that a completely new experience for you?"a="I started working with nanoparticles during my bachelor’s thesis. I continued with nanoparticles into my master’s thesis, working on a project related to cystic fibrosis. Initially, I worked with nano-complexes, but later switched to nano-capsules due to their stability, especially in cell culture media."/>
<QaBoxq="Since you’re focused on stability and applying high shear forces, could you explain why nano-capsules are more stable than complexes in this context?"a="Nano-capsules are generally more stable in cell culture media compared to nano-complexes, which often react with additives and proteins. However, I didn’t explore shear forces much further. My main goal was to stabilize the particles in cell culture media for testing on cells."/>
<QaBoxq="What kind of cell culture medium did you use for these experiments?"a="For transfection, we used Optimem as the medium, after removing antibiotics from the culture medium 24 hours prior to transfection."/>
<QaBoxq="How long did it take you to get to the point where you used nano-capsules?"a="I only started working with nano-capsules towards the end of my PhD. I spent much of my time with nano-complexes, but when I visited a lab in Leeds, I shifted to nano-capsules. This transition happened quite late, just months before I finished my thesis."/>
<QaBoxq="You mentioned capsaicin in your recent paper. Does it significantly affect transfection efficiency, and is it worth including in our experiments?"a="No, capsaicin didn’t affect transfection efficiency in our experiments. It was included to inhibit the Ina-channel as part of a dual strategy targeting both CFTR and Ina-channels, but it might not be necessary for your project."/>
<QaBoxq="Were there any critical components in the formulation of your nanoparticles that you couldn’t do without?"a="No, the main comparison was between nano-complexes and nano-capsules. Nano-complexes were inconsistent in size and stability, while nano-capsules were stable and smaller, which I believed would work better in later experiments."/>
<QaBoxq="Is there a way to check if the mRNA sticks to the outside of the nanoparticle or ends up inside?"a="I believe the mRNA sticks to the outside. The process involved forming nano-capsules using lecithin and oil, and after the ethanol was evaporated, mRNA was added last. The mRNA likely adhered to the positively charged chitosan on the outside of the capsule."/>
<QaBoxq="Do you remember the ratio of mRNA to nano-capsule?"a="I don’t remember the exact ratio offhand, but it’s documented in my dissertation. I optimized the amount of mRNA that needed to enter the cell for effective results, but didn’t do extensive testing with nano-capsules."/>
<QaBoxq="Was determining the optimal amount of nanoparticles trial and error?"a="Yes, definitely. It involved a lot of optimization."/>
<QaBoxq="You used the MTT test for cytotoxicity. Would you recommend it for us, or are there better alternatives?"a="Yes, the MTT test is simple and reliable. You just need to pipette accurately. We used it frequently, and it worked well."/>
<QaBoxq="How did you assess the stability of the nano-capsules? Did you use microscopy or another method?"a="We used a device called a Zetasizer, which measures size, zeta potential, and polydispersity index (PDI). We used it to assess stability in cell culture medium over time, from half an hour to 24 hours."/>
<QaBoxq="Do you have any advice for our project or anything we might have missed?"a="If you’re planning to use a diffuser for nasal administration, you might want to test the stability of the nanoparticles in a saline solution or standard nasal spray solution. It could be useful to see how they behave in such a medium. Otherwise, it seems like you’re well ahead of where I was!"/>
<QaBoxq="Thank you so much for your time and insights!"a="You’re welcome! I’m glad I could help."/>
],
implementation:[<p>The insights from Corden Pharma had a major impact on our project, especially in selecting lipids critical for LNP stability and optimizing gene therapy applications. Initially, we used the Cayman kit, but it was suboptimal for delivering our Primeguide. After receiving feedback, we switched to Corden Pharma’s kit #2, which includes advanced lipid components like cationic lipids that improve cellular uptake and enhance LNP stability. This shift has significantly boosted the efficiency and robustness of our formulations.
Additionally, Corden Pharma's guidance on lipid modifications and antibody incorporation opened new possibilities for targeted therapies. These insights not only improved our technical approach but also paved the way for potential collaborations, offering cost benefits and increased scientific recognition. The feedback will continue to shape our testing process and improve therapeutic delivery. </p>],
interview:<>
<QaBoxq="Is it possible to dry the LNPs designed by Corden Pharma, such as through spray drying?"a="It has not been confirmed whether LNPs have been successfully dried using spray drying. Further investigation or consultation with a specialized company would be required to determine feasibility."/>
<QaBoxq="How is stability ensured in LNPs, particularly for use in inhalation therapy?"a="The stability largely depends on the specific lipid and RNA components used in the formulation. While the stability of individual lipids can be assessed, the overall stability of a new LNP formulation requires empirical testing under various conditions."/>
<QaBoxq="How are lipid combinations selected for inclusion in the LNP starter kits, and what testing is conducted?"a="Lipid combinations in the LNP starter kits are selected based on known interactions, particularly in formulations containing RNA. Testing is conducted to assess physical-chemical properties, encapsulation efficiency, and overall potency. The kits are designed to provide sufficient material for multiple experimental batches."/>
<QaBoxq="Is it advisable to modify the lipid components in an existing LNP formulation?"a="It is generally advisable to consider alternative lipid components, as different lipids may offer improved stability or efficacy. However, the specific needs of the project will dictate whether changes are necessary."/>
<QaBoxq="Is it possible to incorporate antibodies into LNPs, and what is the recommended approach?"a="Yes. Incorporation of antibodies into LNPs is possible. This can be achieved either during the initial preparation phase or by incubating antibodies with LNPs after their formation, depending on whether surface or internal localization of antibodies is desired. Reference to specific studies may provide further guidance."/>
<QaBoxq="What is Corden Pharma's position on projects involving gene therapy?"a="Corden Pharma operates as a service provider, focusing on the manufacturing of active pharmaceutical ingredients (APIs) and excipients rather than developing therapeutic products. Consideration should be given to the intellectual property status of the lipids used in LNP formulations, particularly for commercial applications."/>
<QaBoxq="Is it possible to obtain a discount on LNP starter kits or establish a collaborative relationship with Corden Pharma?"a="We would need to discuss this internally but generally we would be open to potential collaborations that could involve recognition in publications or other forms of acknowledgment, pending approval from relevant management."/>
</>,
summary:"We identified several crucial insights to guide our project development. Nano-capsules were found to be more stable than nano-complexes, making them our preferred formulation choice. We will utilize chitosan to enhance mRNA stability due to its positive charge, while capsaicin was deemed irrelevant for our purposes. For transfection, we will use OptiMEM as the medium, removing antibiotics 24 hours prior to the procedure. We will assess stability with a Zetasizer and evaluate cytotoxicity using the MTT test. Additionally, we are exploring nasal spray and dry powder formulations to improve nanoparticle delivery. These insights will significantly shape our approach to optimizing mRNA delivery systems.",
months:"August"
summary:"The primary goal of the communication with Steffen Bira and Serra Gürcan from Corden Pharma was to explore the use of Lipid Nanoparticles (LNPs) in gene therapy and inhalation treatments. Corden Pharma recommended consulting specialists for assessing spray drying feasibility, while highlighting the importance of testing LNP stability under various conditions. Their LNP starter kits are optimized for stability and encapsulation efficiency, with potential for lipid modifications to enhance cellular uptake. They also confirmed that antibodies can be incorporated into LNPs and emphasized considering intellectual property when selecting lipids. Based on Corden Pharma’s insights, we switched to their kit #2 for our project, hoping to improve the stability and efficiency of our LNP formulations. Their guidance also opened opportunities for targeted therapies and collaborations. This feedback will continue to enhance our testing and therapeutic approaches.",
summary:"We reached out to Svenja Vinke, a former iGEMer and Postdoc, to gain insights on using phage-assisted continuous evolution (PACE) for optimizing our prime editing. Svenja explained that a PACE approach is not feasible for our nickase candidates due to several reasons: it requires too much time, endonucleases are likely too large for optimization, unspecific cutting can kill bacterial cells, and prime editing is less effective in E. coli than in human cells. Based on Svenja's feedback and other expert opinions, we decided against implementing a PACE system for our project.",
months:"July"
},
{
vorname:"Max",
nachnname:"Beckmann",
job:"Patient and Student",
affiliation:"University Bielefeld",
pictureurl:pics['max'],
tag:"Patient",
heading:"Consultation on University Hygiene Risks and Improvement for Hygiene Concept",
job:"PhD Researcher at Laboratory for Molecular Virology & Gene Therapy",
affiliation:"KU Leuven",
pictureurl:pics['mattijs'],
tag:"Academia",
heading:"Visit Mattijs Bulcaen in Leuven and the Laboratory for Molecular Virology & Gene Therapy",
interviewtabid:"mattijsvisit",
cardtext:"",
language:"en",
quote:"x",
aimofcontact:[<p>After our first interview with Mattijs Bulcaen we stayed in contact via email and eventually visited him in Leuven at his laboratory. Here we wanted to gain further information about the CFTR F508del models and editing confirmation.</p>],
insights:[<p>We spoke about approaches for testing CFTR F508del correction in models and methods of confirmation. In this context we talked about HEK293T cell lines[Link] established in his laboratory that stably overexpress CFTR wild-type and F508del. We also discussed how to handle these cell lines. He explained, that the CFTR is fused with a 3HA tag, that in wild-type CFTR would be exposed to the extracellular space and therefore can be used for immunohistochemical staining of the protein, showing correct protein processing and channelling. Also, this allows for a western blot to be made using 3HA antibodies. Functional recovery of CFTR can also be visualized using halide sensitive eYFP or organoid assay, the ladder Mattijs had established an automated readout. Furthermore, we talked about how to handle Sanger sequencing data to analyse edits and discussed the possibility to avoid the weaknesses of Sanger sequencing by using Nanopore sequencing instead. We asked about the applicability of patch clamp analysis in the context of CFTR and Mattijs said that, to his knowledge, it has not been used to test for successful editing in CFTR.
Lastly Mattijs explained how he plans to deliver the prime editing complex to the patient, and we evaluated the advantages and disadvantages of delivery strategies, including our very own LNP approach.</p>],
implementation:[<p>When planning how to test and confirm editing by our own constructs, we were largely inspired by the information given to us by Mattijs. For example, we tested prime editing in the HEK293 cell lines we spoke about with Mattijs and used halide sensitive eYFP to check for CFTR function. Also, we tried differentiating wild-type and F508del cells using patch clamp. Unfortunately, a lot of the methods mentioned were not usable for us because of time constraints, but are still valuable for future projects and research built upon PreCyse. </p>],
summary:"We visited Mattijs Bulcaen in Leuven to enhance our understanding of CFTR F508del models and editing confirmation. During the visit, we examined HEK293T cell lines that stably overexpress wild-type CFTR and the F508del variant. Mattijs demonstrated how to use a 3HA tag for visualizing protein processing and discussed methods for assessing CFTR functional recovery. We also compared Sanger and Nanopore sequencing techniques and evaluated delivery strategies for our prime editing complex. The insights from this visit guided our project, leading us to test prime editing in HEK293T cells using halide-sensitive eYFP. While some methods were not feasible due to time constraints, they laid the groundwork for future research in the PreCyse project.",
months:"july"
},
{
vorname:"Collaborations",
nachnname:"iGEM Team Linköping ",
type:"meta",
pictureurl:pics['linköping'],
tag:"Milestone",
heading:"Cooperation to create a Lipid Delivery System Handbook",
interviewtabid:"handbook",
cardtext:"",
quoteVorname:"Kaya",
quoteNachname:"Lange",
quote:"We were genuinely excited when Linköping University approached us for collaboration. From the very beginning, their ideas resonated with us, and our shared enthusiasm laid a strong foundation for a productive partnership. We're happy to work together, also with the other teams, and explore new possibilities.",
aimofcontact:[<p>The initial contact for our collaboration came from the iGEM team 2024 of Linköping, Sweden, who approached us with a proposal to create a “Delivery-Based Handbook”[link Handbook]. Their goal was to reduce the steep learning curve associated with these technologies by sharing collective knowledge from multiple teams, including ours. We were excited to contribute and help future teams navigate these challenges more easily. The handbook would serve as a valuable tool. </p>],
insights:[<p>Throughout the collaboration, we gained significant insights, both scientific and collaborative. Initially, our meetings with the Linköping team and other participating teams - Patras, Radboud-University and TERMOSZ-Selye-HUN - were invaluable. These sessions allowed us to exchange ideas and learn how each team planned to use lipid-based delivery systems in their own projects. This mutual sharing of knowledge opened our eyes to new methodologies and potential applications of LNPs and liposomes. We also gained a deeper appreciation for the interdisciplinary nature of these systems. From the challenges of formulating stable particles to optimizing their efficiency in targeting cells, we realized the complexity of the field and how collaboration could help overcome many of these obstacles. By discussing our respective approaches, we were able to pool our expertise, which not only improved our understanding but also ensured that the handbook would be comprehensive and valuable for various iGEM teams, regardless of their specific project focus.
In summary: </p>,
<ul>
<li>Learned different approaches to using LNPs and liposomes in iGEM projects.</li>
<li>Discovered new methods for optimizing LNPs.</li>
<li>Recognized challenges in particle stability and targeted delivery.</li>
<li>Gained appreciation for the interdisciplinary complexity of these systems.</li>
<li>Focused on documenting work to benefit future iGEM teams.</li>
</ul>
],
implementation:[<p>The collaboration expanded our understanding of what's possible, inspiring us to consider new ideas for how we might integrate advanced techniques into our nanoparticle systems in future projects. The collaborative process also encouraged us to document our work more thoroughly, ensuring that future iGEM teams could benefit from both our successes and the challenges we encountered along the way. Beyond the technical improvements, the experience taught us the value of teamwork across borders and disciplines. Each team brought a unique perspective, and by working together, we were able to develop a resource that was far greater than the sum of its parts</p>],
summary:"This collaboration with Linköping and the other iGEM teams was an incredibly enriching experience. Together, we developed a “Delivery-Based Handbook”[link Handbook] that will serve as a valuable resource for future teams working with LNPs and liposomes. The knowledge we gained not only enhanced our project but also strengthened our sense of community within iGEM. We are excited to present the handbook at the Grand Jamboree, where we will finally meet our collaborators in person and celebrate the culmination of our collective efforts. This partnership has shown us the immense power of collaboration, and we are proud to have been part of such a meaningful initiative.",
job:"PhD student Working group: Organic chemistry and biocatalysis ",
affiliation:"University of Bielefeld",
pictureurl:pics['kaihammer'],
tag:"Academia",
heading:"First insights of Enzym Engineering",
interviewtabid:"hammerkai",
language:"de",
cardtext:"",
quote:"x",
aimofcontact:[<p>When we realized that the creation of a nickase from the endonucleases in use was a desired outcome, it became necessary to talk to an expert in the field of enzyme engineering. Our first contact was Kai Schülke, a former iGEMer and PhD student under the guidance of Prof. Dr. Hammer[Link Hammer], who is the leader of the working group organic chemistry and bioanalytics at Bielefeld University.</p>],
insights:[<p>In the process of our interaction with Kai, we learned about the various methods employed in enzyme engineering. He demonstrated the complexity of this field of research and emphasized the importance of choosing the right approach. As a former iGEMer, Kai, inspired by his past experiences, is highly motivated and determined to develop an outstanding project. He pointed out that we cannot rely on classical methods such as directed evolution, but instead should use a rational approach to select mutation candidates. His insights and enthusiasm have encouraged us to think critically and pursue innovative solutions in our work. </p>],
implementation:[<p>We incorporated Kai's insights into our project by shifting our approach to enzyme engineering. By focusing on a more targeted approach, we were able to refine our enzyme optimization process, ensuring that the modifications we made were based on informed, calculated decisions. This not only streamlined our research but also improved the chances of success by reducing the trial-and-error inherent in traditional methods. </p>],
summary:"The team reached out to Kai Schülke, a former iGEM participant and enzyme engineering expert, for guidance on developing a nickase from the endonucleases in use. Kai emphasized the need for a rational, targeted approach rather than traditional methods like directed evolution. His insights helped the team refine their enzyme optimization process, making it more strategic and efficient. This shift reduced trial-and-error efforts and improved the chances of success, driving innovation in their project.",
months:"July"
},
{
title:"M.Sc.",
vorname:"Nils",
nachnname:"Berelsmann",
job:"PhD Working group: Prof. Dr. Gabriele Fischer von Mollard ",
affiliation:"University of Bielefeld",
pictureurl:pics['nilshefe'],
language:"de",
tag:"Academia",
heading:"Adapting expression strategies for Fanzor nickases and exploring the potential of Pichia pastoris for SpuFz1 nickase variants ",
interviewtabid:"nberelsmann",
cardtext:"",
quote:"X",
aimofcontact:[<p>During our interview with Makoto Saito[Linkinterview] about fanzor[link fanzor], it became evident that the expression of our fanzor nickases in yeast is very promising. We then refined our expression strategy for the nickases and approached Nils Berelsmann, who is currently working on his PhD thesis with the yeast strain Pichia pastoris (SMD1163). This particular strain could be ideal for expressing the SpuFz1 nickase variants. Our main aim in contacting Nils was to gain insight and advice on yeast expression and he generously shared his expertise with us. Not only did he give us valuable advice, but he also provided us with the yeast strain itself, along with a corresponding expression vector for possible experiments. He also provided us with detailed protocols and the plasmid map of the vector and gave us practical tips on how to optimize the expression process. His support was invaluable in moving our work forward. </p>],
insights:[<p>Pichia pastoris (SMD1163) is a promising option for expressing SpuFz1 nickase variants. Refining expression strategies based on expert insights is crucil for success. Nils provided practical tips on yeast expression, including optimizing growth conditions and fine-tuning induction protocols.</p>],
implementation:[<p>We adapted our expression strategy for Fanzor nickases in yeast by incorporating the Pichia pastoris strain (SMD1163) and the provided expression vector into our experiments. Following Nils' detailed protocols and plasmid map, we optimized key steps, enhancing expression efficiency and protein yield.</p>],
summary:"The team sought expert advice from Nils to optimize yeast expression for Fanzor nickases. Nils provided invaluable guidance on addressing potential challenges and troubleshooting the process. He supplied the Pichia pastoris (SMD1163) strain along with a suitable expression vector, crucial for expressing SpuFz1 nickase variants. Additionally, he shared detailed protocols for yeast transformation and growth optimization, enabling the team to replicate his methods effectively for their experiments.",
months:"July",
},
{
title:"Dr.",
vorname:"Timm",
nachnname:"Weber",
job:"Staff Scientist, Project- and Quality Manager",
affiliation:"Central Biobank of the University of Bielefeld",
pictureurl:pics['biobank'],
tag:"Academia",
heading:"Discussed the processes involved in the storage, processing, and security of patient samples.",
interviewtabid:"timm",
cardtext:"",
quote:"A biobank is not just a collection of samples; it's a bridge between patient trust and scientific discovery, ensuring that valuable biological data is safeguarded while contributing to future research.",
aimofcontact:"Contact was established with Timm for the purpose of gaining deeper insights into the functioning of the biobank and of deepening our understanding of the processing of patient samples.",
insights:"We were provided with invaluable insights into the quality and project management of the biobank and storage of patient samples. It was of particular interest to note that Biobank OWL occupies a distinctive position in this context, insofar as a trustee is not a mandatory figure within its system and is therefore not provided for as a standard component. However, Biobank OWL has elected to integrate a trustee in order to enhance the security standards for the safeguarding of patient data. This illustrates the biobank's dedication to ensuring the optimal protection and security of sensitive patient data.",
implementation:"The insights gained have facilitated a deeper comprehension of the significance of quality management in the processing of patient samples. This understanding has been integrated into our project processes, thereby enhancing the accuracy and reliability of our procedures. ",
summary:"The interview focused on understanding the operations of the Biobank OWL, particularly in the areas of quality management and sample processing. Provided a detailed overview of biobank activities, including sample collection, storage conditions, and data protection measures",
language:"de",
interview:<>
<QaBoxq="Can you briefly explain to us what exactly a biobank is and what its main tasks are?"
a="A biobank is a specialized facility that collects, stores, and manages biological samples and associated data for research purposes. Each biobank is unique in its operations and functions. In Bielefeld and Lippe, the Biobank BOWL (Biobank OWL) is responsible for the storage of patient samples. The Data Integration Centre (DIZ) stores data pertaining to these samples. A trustee oversees the pseudonymisation of data, acting as an interface between BOWL and DIZ, ensuring that patient data cannot be directly linked to patient samples."/>
<QaBoxq="What types of samples are collected in your biobank and for what research purposes are they used?"
a="The biobank collects a wide variety of samples, including blood, stool, and soil. Samples may be gathered for specific research projects or for establishing a general repository under 'broad consent.' Researchers wishing to use these samples must apply to the 'use access committee,' which evaluates whether the requested samples and data can be released for their research."/>
<QaBoxq="How large is your biobank? How many samples do you currently store and how many new samples are added on average?"
a="The biobank is still in the process of establishing itself and has not yet reached its full sample capacity. However, it is anticipated to accumulate a significant number of samples in the near future, with several thousand samples expected to be analyzed in dedicated sessions."/>
<QaBoxq="What requirements and criteria must be met for a sample to be included in your biobank?"a="Samples must be processed according to highly detailed protocols, and regular audits are conducted to ensure compliance with all standards."/>
<QaBoxq="Which other research institutions or biobanks do you cooperate with and what form does this cooperation take?"
a="Biobank OWL has a second location in Lippe, in addition to Bielefeld. Collaborations exist with the DIZ, the Treuhand, and three university hospitals. It is anticipated that cooperation with other working groups will increase in the future."/>
<QaBoxq="What specific storage conditions (e.g. temperature, humidity) must be observed for different sample types?"
a="Samples are stored under various temperature conditions, including -20°C, -80°C, and -150°C, along with the use of liquid nitrogen."/>
<QaBoxq="How do you ensure that the samples remain stable and usable over longer periods of time?"
a="Samples are stored in nitrogen for long-term stability."/>
<QaBoxq="What encryption techniques or data protection measures are used in your biobank to prevent unauthorized access to patient data? Are there special regulations for the anonymisation of data and how is it ensured that patients cannot be traced?"
a="Pseudonyms are created using specialized software such as CentraXX or REDcap to protect patient data."/>
<QaBoxq="What rights do patients have in relation to their samples, and how are these rights safeguarded in your biobank?"
a="Patients have the right to revoke their consent at any time, which can be done at the clinic or biobank. The trustee, acting as an intermediary, will notify BOWL and DIZ to destroy the corresponding samples or data."/>
</>,
months:"august"
},
{
vorname:"'Schüler*innen Akademie'",
nachnname:"",
pictureurl:pics['schueler'],
tag:"Education",
heading:"Empowering students through synthetic biology",
<QaBoxq="Would you be willing to support us in our project? Would you dry our LNPs?"a="The spray dryer requires 5 mL of a solution with 5% lipid solids and 0.02% RNA. We’ve published recovery rates of 70%. You can send us the mRNA and LNP components to encapsulate and dry."/>
</>,
summary:"The conversation focused on spray-drying LNPs, emphasizing the shelf life of RNA-based formulations, optimal storage conditions, and technical requirements for the drying process. Corden Pharma shared insights on the protocol, highlighting the need for testing each LNP formulation individually for stability. AI technology is used to optimize LNP formulations, and potential collaborations were discussed, including support for drying LNPs.",
months:"juli"
months:"august"
},
{
title:"Prof. Dr",
vorname:"David",
nachnname:"Liu",
job:" Richard Merkin Professor and director of the Merkin Institute of Transformative Technologies in Healthcare",
affiliation:"vice chair of the faculty at the Broad Institute of MIT and Harvard",
pictureurl:pics['placeholder'],
tag:"Academia",
heading:"Influence of research by David Liu on our design decisions ",
interviewtabid:"liu",
vorname:"Max",
nachnname:"Beckmann",
job:"Patient and Student",
affiliation:"University Bielefeld",
pictureurl:pics['max'],
tag:"Patient",
heading:"Consultation on University Hygiene Risks and Improvement for Hygiene Concept",
interviewtabid:"maxhygiene",
cardtext:"",
quote:"X",
aimofcontact:"David Liu is the principal investigator responsible for the development of the prime editing systems and his laboratory is actively working on improving prime editors, also for application in CFTR mutation F508del.",
language:"de",
quote:"x",
aimofcontact:"",
insights:"",
implementation:"",
summary:"",
months:""
months:"August"
},
{
vorname:"Steffen Bira and",
nachnname:"Serra Gürcan from Corden Pharma",
job:"Associate director",
affiliation:"Corden Pharma",
pictureurl:pics['corden'],
tag:"Industry",
heading:"Lipid Nanoparticles in Gene Therapy: perspectives from Corden Pharma ",
interviewtabid:"corden",
title:"Dr.",
vorname:"Katharina",
nachnname:"Kolonko",
job:"Expert for nanocapsules",
affiliation:"Biologist",
pictureurl:pics['kolonko'],
tag:"Academia",
heading:"Optimizing our mRNA Delivery Systems",
interviewtabid:"kolonkofirst",
cardtext:"",
language:"en",
quote:"The stability of LNPs depends on the specific lipid and RNA components used, but ensuring the overall stability of a new formulation requires rigorous empirical testing under various conditions.",
aimofcontact:[<p>The primary aim of the communication with Steffen Bira and Serra Gürcan from Corden Pharma was to explore the technical aspects and practical applications of Lipid Nanoparticles (LNPs) in advanced medical therapies, including gene therapy and inhalation treatments. The conversation focused on the possibility of using Corden Pharma’s LNP starter kits, understanding the factors affecting the stability of LNPs, and exploring options for incorporating antibodies into LNPs to target specific cells. </p>],
insights:[<p>The discussion with <ahref="https://cordenpharma.com/">Corden Pharma</a>, led by Steffen Bira and Serra Gürcan, offered key insights into LNPs and their applications. While Corden Pharma hasn't extensively explored spray drying for LNPs, they recommended consulting specialists to evaluate its feasibility, especially concerning lipid stability during the process. Stability was highlighted as crucial for inhalation therapies, requiring thorough testing of entire LNP formulations, possibly aided by cryoprotectants and controlled temperatures.
Corden Pharma's LNP starter kits are based on well-researched lipid combinations designed for stability and encapsulation efficiency, making them suitable for multiple experiments. They suggested that modifying lipid components, such as incorporating cholesterol derivatives, could enhance cellular uptake and overall efficacy. Additionally, they confirmed the possibility of incorporating antibodies into LNPs and emphasized the importance of considering intellectual property when selecting lipids for commercial use. They also showed openness to collaboration,
including offering discounts in exchange for recognition in publications.
The interaction with Corden Pharma provided several key insights:</p>,
language:"de",
quote:"Wow, you’re already further along than I was! That’s a really good approach, especially since dry powder formulations can help with stability.",
aimofcontact:[<p>Our goal in reaching out to Dr. Katharina Kolonko, who earned her PhD working on chitosan-based nanoparticles for delivering nucleic acids to human respiratory cells in the context of cystic fibrosis, was to seek her advice on the design, stability, and application of nanoparticles. We wanted to learn from her experience with chitosan-capsaicin nanoparticles, especially the challenges she encountered, and apply her insights to improve our own project. Specifically, we aimed to better understand nanoparticle stability, transfection methods, and how to effectively design our experiments.
</p>],
insights:[<p>Dr. Kolonko provided us with valuable insights into working with nanoparticles, particularly emphasizing the advantages of nanocapsules. She highlighted that nanocapsules are more stable than nano-complexes, which is crucial for experiments involving complex environments and high shear forces. Additionally, her use of capsaicin wasn’t aimed at improving transfection efficiency but was part of a broader strategy targeting specific channels. She also shared practical methods for measuring particle stability and cytotoxicity, giving us clear guidance on tools and techniques that we can apply to our project.
Furthermore, Dr. Kolonko discussed the use of chitosan as a component in nanoparticle formulations. Chitosan, with its positive charge, can interact with mRNA, potentially enhancing the stability of the cargo. As an outlook, we plan to explore modifications using chitosan to improve the stability and performance of our mRNA delivery system. This approach may provide a more robust solution for optimizing nanoparticle formulations in future experiments. </p>],
implementation:[<p>We directly applied Katharina’s insights to improve our nanoparticle design and testing methods. Her recommendations on using nano-capsules and OptiMEM as the transfection medium helped refine our experimental approach. She emphasized using a medium with fewer additives, like OptiMEM, and suggested removing antibiotics 24 hours before transfection to avoid interference, ensuring more controlled and effective conditions. We also explored new stability testing ideas, including nasal spray solutions and dry powder formulations.
We incorporated her insights as follows: </p>,
<ul>
<li><strong>Spray Drying Feasibility:</strong> Corden Pharma hasn’t explored spray drying extensively; consultation with specialists is recommended for assessing feasibility and lipid stability.</li>
<li><strong>LNP Stability:</strong> Stability of LNPs, particularly for inhalation therapies, needs empirical testing, considering shear forces and the potential use of cryoprotectants or temperature control.</li>
<li><strong>Lipid Selection in Kits:</strong> Starter kits use well-researched lipid combinations, tested for stability, encapsulation efficiency, and potency. They provide materials for multiple experimental batches.</li>
<li><strong>Lipid Modifications:</strong> Exploring alternative lipids (e.g., cholesterol derivatives) could enhance stability and cellular uptake, tailored to project needs.</li>
<li><strong>Antibody Incorporation:</strong> Antibodies can be incorporated into LNPs during preparation or afterward, depending on targeting requirements.</li>
<li><strong>Intellectual Property:</strong> IP considerations are crucial when selecting lipids for LNP formulations, as many lipids are patented.</li>
<li><strong>Collaboration Opportunities:</strong> Corden Pharma is open to offering discounts or forming partnerships, with recognition in publications or acknowledgments.</li>
<li><strong>Nano-Capsules Focus:</strong> Based on Katharina’s advice, we prioritized nano-capsules for their enhanced stability over nano-complexes.</li>
<li><strong>Chitosan for Stability:</strong> We're exploring chitosan to improve mRNA delivery system stability due to its positive charge, which binds mRNA to the nanoparticle surface.</li>
<li><strong>OptiMEM for Transfection:</strong> OptiMEM is now our chosen transfection medium, with the suggestion to remove antibiotics 24 hours prior.</li>
<li><strong>MTT Test for Cytotoxicity:</strong> We adopted the MTT test for cytotoxicity due to its simplicity and reliability.</li>
<li><strong>Nasal Spray and Dry Powder Testing:</strong> We are considering testing nanoparticle stability using nasal spray solutions and exploring dry powder formulations.</li>
<li><strong>Capsaicin Exclusion:</strong> As capsaicin did not significantly impact transfection efficiency in Katharina’s research, we decided not to include it in our project.</li>
</ul>
],
implementation:[<p>The insights from Corden Pharma had a major impact on our project, especially in selecting lipids critical for LNP stability and optimizing gene therapy applications. Initially, we used the Cayman kit, but it was suboptimal for delivering our Primeguide. After receiving feedback, we switched to Corden Pharma’s kit #2, which includes advanced lipid components like cationic lipids that improve cellular uptake and enhance LNP stability. This shift has significantly boosted the efficiency and robustness of our formulations.
Additionally, Corden Pharma's guidance on lipid modifications and antibody incorporation opened new possibilities for targeted therapies. These insights not only improved our technical approach but also paved the way for potential collaborations, offering cost benefits and increased scientific recognition. The feedback will continue to shape our testing process and improve therapeutic delivery. </p>],
interview:<>
<QaBoxq="Is it possible to dry the LNPs designed by Corden Pharma, such as through spray drying?"a="It has not been confirmed whether LNPs have been successfully dried using spray drying. Further investigation or consultation with a specialized company would be required to determine feasibility."/>
<QaBoxq="How is stability ensured in LNPs, particularly for use in inhalation therapy?"a="The stability largely depends on the specific lipid and RNA components used in the formulation. While the stability of individual lipids can be assessed, the overall stability of a new LNP formulation requires empirical testing under various conditions."/>
<QaBoxq="How are lipid combinations selected for inclusion in the LNP starter kits, and what testing is conducted?"a="Lipid combinations in the LNP starter kits are selected based on known interactions, particularly in formulations containing RNA. Testing is conducted to assess physical-chemical properties, encapsulation efficiency, and overall potency. The kits are designed to provide sufficient material for multiple experimental batches."/>
<QaBoxq="Is it advisable to modify the lipid components in an existing LNP formulation?"a="It is generally advisable to consider alternative lipid components, as different lipids may offer improved stability or efficacy. However, the specific needs of the project will dictate whether changes are necessary."/>
<QaBoxq="Is it possible to incorporate antibodies into LNPs, and what is the recommended approach?"a="Yes. Incorporation of antibodies into LNPs is possible. This can be achieved either during the initial preparation phase or by incubating antibodies with LNPs after their formation, depending on whether surface or internal localization of antibodies is desired. Reference to specific studies may provide further guidance."/>
<QaBoxq="What is Corden Pharma's position on projects involving gene therapy?"a="Corden Pharma operates as a service provider, focusing on the manufacturing of active pharmaceutical ingredients (APIs) and excipients rather than developing therapeutic products. Consideration should be given to the intellectual property status of the lipids used in LNP formulations, particularly for commercial applications."/>
<QaBoxq="Is it possible to obtain a discount on LNP starter kits or establish a collaborative relationship with Corden Pharma?"a="We would need to discuss this internally but generally we would be open to potential collaborations that could involve recognition in publications or other forms of acknowledgment, pending approval from relevant management."/>
],
interview:<>
<QaBoxq="How did you approach the design of Lipid Nanoparticles (LNPs)? What were the first steps you took at that time? Were you already familiar with LNPs, or was that a completely new experience for you?"a="I started working with nanoparticles during my bachelor’s thesis. I continued with nanoparticles into my master’s thesis, working on a project related to cystic fibrosis. Initially, I worked with nano-complexes, but later switched to nano-capsules due to their stability, especially in cell culture media."/>
<QaBoxq="Since you’re focused on stability and applying high shear forces, could you explain why nano-capsules are more stable than complexes in this context?"a="Nano-capsules are generally more stable in cell culture media compared to nano-complexes, which often react with additives and proteins. However, I didn’t explore shear forces much further. My main goal was to stabilize the particles in cell culture media for testing on cells."/>
<QaBoxq="What kind of cell culture medium did you use for these experiments?"a="For transfection, we used Optimem as the medium, after removing antibiotics from the culture medium 24 hours prior to transfection."/>
<QaBoxq="How long did it take you to get to the point where you used nano-capsules?"a="I only started working with nano-capsules towards the end of my PhD. I spent much of my time with nano-complexes, but when I visited a lab in Leeds, I shifted to nano-capsules. This transition happened quite late, just months before I finished my thesis."/>
<QaBoxq="You mentioned capsaicin in your recent paper. Does it significantly affect transfection efficiency, and is it worth including in our experiments?"a="No, capsaicin didn’t affect transfection efficiency in our experiments. It was included to inhibit the Ina-channel as part of a dual strategy targeting both CFTR and Ina-channels, but it might not be necessary for your project."/>
<QaBoxq="Were there any critical components in the formulation of your nanoparticles that you couldn’t do without?"a="No, the main comparison was between nano-complexes and nano-capsules. Nano-complexes were inconsistent in size and stability, while nano-capsules were stable and smaller, which I believed would work better in later experiments."/>
<QaBoxq="Is there a way to check if the mRNA sticks to the outside of the nanoparticle or ends up inside?"a="I believe the mRNA sticks to the outside. The process involved forming nano-capsules using lecithin and oil, and after the ethanol was evaporated, mRNA was added last. The mRNA likely adhered to the positively charged chitosan on the outside of the capsule."/>
<QaBoxq="Do you remember the ratio of mRNA to nano-capsule?"a="I don’t remember the exact ratio offhand, but it’s documented in my dissertation. I optimized the amount of mRNA that needed to enter the cell for effective results, but didn’t do extensive testing with nano-capsules."/>
<QaBoxq="Was determining the optimal amount of nanoparticles trial and error?"a="Yes, definitely. It involved a lot of optimization."/>
<QaBoxq="You used the MTT test for cytotoxicity. Would you recommend it for us, or are there better alternatives?"a="Yes, the MTT test is simple and reliable. You just need to pipette accurately. We used it frequently, and it worked well."/>
<QaBoxq="How did you assess the stability of the nano-capsules? Did you use microscopy or another method?"a="We used a device called a Zetasizer, which measures size, zeta potential, and polydispersity index (PDI). We used it to assess stability in cell culture medium over time, from half an hour to 24 hours."/>
<QaBoxq="Do you have any advice for our project or anything we might have missed?"a="If you’re planning to use a diffuser for nasal administration, you might want to test the stability of the nanoparticles in a saline solution or standard nasal spray solution. It could be useful to see how they behave in such a medium. Otherwise, it seems like you’re well ahead of where I was!"/>
<QaBoxq="Thank you so much for your time and insights!"a="You’re welcome! I’m glad I could help."/>
</>,
summary:"The primary goal of the communication with Steffen Bira and Serra Gürcan from Corden Pharma was to explore the use of Lipid Nanoparticles (LNPs) in gene therapy and inhalation treatments. Corden Pharma recommended consulting specialists for assessing spray drying feasibility, while highlighting the importance of testing LNP stability under various conditions. Their LNP starter kits are optimized for stability and encapsulation efficiency, with potential for lipid modifications to enhance cellular uptake. They also confirmed that antibodies can be incorporated into LNPs and emphasized considering intellectual property when selecting lipids. Based on Corden Pharma’s insights, we switched to their kit #2 for our project, hoping to improve the stability and efficiency of our LNP formulations. Their guidance also opened opportunities for targeted therapies and collaborations. This feedback will continue to enhance our testing and therapeutic approaches.",
job:"PhD Researcher at Laboratory for Molecular Virology & Gene Therapy",
affiliation:"KU Leuven",
pictureurl:pics['mattijs'],
tag:"Academia",
heading:"Visit Mattijs Bulcaen in Leuven and the Laboratory for Molecular Virology & Gene Therapy",
interviewtabid:"mattijsvisit",
cardtext:"",
language:"en",
quote:"x",
aimofcontact:[<p>After our first interview with Mattijs Bulcaen we stayed in contact via email and eventually visited him in Leuven at his laboratory. Here we wanted to gain further information about the CFTR F508del models and editing confirmation.</p>],
insights:[<p>We spoke about approaches for testing CFTR F508del correction in models and methods of confirmation. In this context we talked about HEK293T cell lines[Link] established in his laboratory that stably overexpress CFTR wild-type and F508del. We also discussed how to handle these cell lines. He explained, that the CFTR is fused with a 3HA tag, that in wild-type CFTR would be exposed to the extracellular space and therefore can be used for immunohistochemical staining of the protein, showing correct protein processing and channelling. Also, this allows for a western blot to be made using 3HA antibodies. Functional recovery of CFTR can also be visualized using halide sensitive eYFP or organoid assay, the ladder Mattijs had established an automated readout. Furthermore, we talked about how to handle Sanger sequencing data to analyse edits and discussed the possibility to avoid the weaknesses of Sanger sequencing by using Nanopore sequencing instead. We asked about the applicability of patch clamp analysis in the context of CFTR and Mattijs said that, to his knowledge, it has not been used to test for successful editing in CFTR.
Lastly Mattijs explained how he plans to deliver the prime editing complex to the patient, and we evaluated the advantages and disadvantages of delivery strategies, including our very own LNP approach.</p>],
implementation:[<p>When planning how to test and confirm editing by our own constructs, we were largely inspired by the information given to us by Mattijs. For example, we tested prime editing in the HEK293 cell lines we spoke about with Mattijs and used halide sensitive eYFP to check for CFTR function. Also, we tried differentiating wild-type and F508del cells using patch clamp. Unfortunately, a lot of the methods mentioned were not usable for us because of time constraints, but are still valuable for future projects and research built upon PreCyse. </p>],
summary:"We visited Mattijs Bulcaen in Leuven to enhance our understanding of CFTR F508del models and editing confirmation. During the visit, we examined HEK293T cell lines that stably overexpress wild-type CFTR and the F508del variant. Mattijs demonstrated how to use a 3HA tag for visualizing protein processing and discussed methods for assessing CFTR functional recovery. We also compared Sanger and Nanopore sequencing techniques and evaluated delivery strategies for our prime editing complex. The insights from this visit guided our project, leading us to test prime editing in HEK293T cells using halide-sensitive eYFP. While some methods were not feasible due to time constraints, they laid the groundwork for future research in the PreCyse project.",
months:"july"
summary:"We identified several crucial insights to guide our project development. Nano-capsules were found to be more stable than nano-complexes, making them our preferred formulation choice. We will utilize chitosan to enhance mRNA stability due to its positive charge, while capsaicin was deemed irrelevant for our purposes. For transfection, we will use OptiMEM as the medium, removing antibiotics 24 hours prior to the procedure. We will assess stability with a Zetasizer and evaluate cytotoxicity using the MTT test. Additionally, we are exploring nasal spray and dry powder formulations to improve nanoparticle delivery. These insights will significantly shape our approach to optimizing mRNA delivery systems.",
heading:"Successful participation of a team member in a 5 day GxP course",
interviewtabid:"gxpcourse",
cardtext:"",
quote:"The GXP course was extremely useful as it provided us with important knowledge that supports our entire team in complying with quality standards. This knowledge will help us to organise our processes efficiently and in accordance with regulations in the future.",
quoteVorname:"Kaya",
quoteNachname:"Lange",
text:[<p>I, Kaya, Team Member of iGEM Bielefeld 2024, recently participated in an intensive one-week GXP (Good Practice) training course, which was pivotal experience for both me and our project. The course covered essential regulatory frameworks, including</p>,
<ul>
<li>Good Laboratory Practice (GLP)</li>
<li>Good Clinical Practice (GCP)</li>
<li>Good Manufacturing Practice (GMP)</li>
</ul>,
<p>
which are all designed to ensure quality, safety, and compliance across every phase of scientific research and development.
As the head of Integrated Human Practices, I found this training particularly valuable. It provided me with a deeper understanding of the rigorous standards that need to be maintained in research, especially concerning ethics, data integrity, and patient safety. I learned how to properly document research processes, ensure the reproducibility of results, and assess and mitigate risks, all while keeping the ethical considerations of our project at the forefront.
I have acquired the ability to create standard operating procedures (SOPs) that guarantee the transparent and traceable documentation of each stage of the research process. This not only facilitates internal organisation but is also crucial for subsequent approvals and audits by regulatory authorities.
It is of paramount importance to ensure the reproducibility of our experiments by maintaining accurate protocols and meticulously documenting all variables. This is of particular importance should the intention be to pursue clinical research at a later stage, as the reproducibility of experiments is a crucial factor in the validity of the results.
I acquired knowledge of techniques for risk assessment, including Failure Mode and Effects Analysis (FMEA). This process enables the identification of potential risks in a project at an early stage, thus facilitating the development of strategies to minimise them. This approach allows us to identify and address potential sources of error before they lead to significant issues.
This knowledge is crucial as we think about the future of our project, particularly if we aim to move our gene therapy approach for cystic fibrosis closer to clinical trials and real-world applications. My participation in the GXP training has equipped me with the necessary tools to potentially guide our team through the complex regulatory landscape, ensuring our work remains aligned with industry standards and ready for the next steps in development.
One of the key speakers during the GXP course was Dr. Marcus Berger [LINK INtreview Beerger], whose expertise was invaluable to me and the entire team. After the course, I had the opportunity to ask Dr. Berger some questions, further deepening my understanding of the practical applications of GXP in research. The connection with Dr. Berger has been highly beneficial, as his insights helped shape key aspects of our project’s development and compliance with industry standards. His guidance will continue to be a valuable resource for our team moving forward.
Through this training, I feel better positioned to contribute to the team’s efforts, ensuring our project adheres to global safety and ethical guidelines. This experience has strengthened our approach and set a solid foundation for future progress, ensuring that our research, public engagement, and potential clinical applications continue to meet the highest regulatory standards. </p>],
type:"meta",
summary:"Kaya, a member of the iGEM Bielefeld 2024 team, completed an intensive one-week GXP (Good Practice) training, which covered Good Laboratory Practice (GLP), Good Clinical Practice (GCP), and Good Manufacturing Practice (GMP). The training provided valuable insights into maintaining high standards of quality, safety, and ethics throughout the research process. Kaya learned crucial skills, such as documenting research processes for reproducibility, creating standard operating procedures (SOPs), and conducting risk assessments using techniques like Failure Mode and Effects Analysis (FMEA). This knowledge is essential for advancing their cystic fibrosis gene therapy project toward clinical trials and ensuring compliance with regulatory standards. Dr. Marcus Berger, a key speaker in the course, provided additional guidance, offering valuable insights that will continue to benefit the team.",
heading:"urgent requirement for a hygiene concept for students with disabilities and immunocompromised employees",
heading:"Hygiene concept for students with disabilities and immunocompromised employees",
interviewtabid:"johannfunke",
cardtext:"",
quote:"The implementation of the hygiene concept is proving more difficult than expected due to the bureaucracy at the university. Nevertheless, the interview gave us a good insight into this labyrinth of regulations and we got started the prozess of implementation.",
heading:"F508del mutation confirmed as the most common CFTR mutation worldwide, including Asia, supporting the efficacy of existing therapies for the majority of patients. ",
heading:"F508del mutation confirmed as the most common CFTR mutation worldwide, including Asia",
summary:"The contact aimed to leverage Philipp’s expertise in ALI cultures to improve our experimental protocols Gained insights into optimizing conditions for primary cell cultures and managing common challenges like fungal contamination",
months:"several times"
},
{
title:"Dr.",
vorname:"Timm",
nachnname:"Weber",
job:"Staff Scientist, Project- and Quality Manager",
affiliation:"Central Biobank of the University of Bielefeld",
pictureurl:pics['biobank'],
tag:"Academia",
heading:"Discussed the processes involved in the storage, processing, and security of patient samples.",
interviewtabid:"timm",
cardtext:"",
quote:"A biobank is not just a collection of samples; it's a bridge between patient trust and scientific discovery, ensuring that valuable biological data is safeguarded while contributing to future research.",
aimofcontact:"Contact was established with Timm for the purpose of gaining deeper insights into the functioning of the biobank and of deepening our understanding of the processing of patient samples.",
insights:"We were provided with invaluable insights into the quality and project management of the biobank and storage of patient samples. It was of particular interest to note that Biobank OWL occupies a distinctive position in this context, insofar as a trustee is not a mandatory figure within its system and is therefore not provided for as a standard component. However, Biobank OWL has elected to integrate a trustee in order to enhance the security standards for the safeguarding of patient data. This illustrates the biobank's dedication to ensuring the optimal protection and security of sensitive patient data.",
implementation:"The insights gained have facilitated a deeper comprehension of the significance of quality management in the processing of patient samples. This understanding has been integrated into our project processes, thereby enhancing the accuracy and reliability of our procedures. ",
summary:"The interview focused on understanding the operations of the Biobank OWL, particularly in the areas of quality management and sample processing. Provided a detailed overview of biobank activities, including sample collection, storage conditions, and data protection measures",
language:"de",
interview:<>
<QaBoxq="Can you briefly explain to us what exactly a biobank is and what its main tasks are?"
a="A biobank is a specialized facility that collects, stores, and manages biological samples and associated data for research purposes. Each biobank is unique in its operations and functions. In Bielefeld and Lippe, the Biobank BOWL (Biobank OWL) is responsible for the storage of patient samples. The Data Integration Centre (DIZ) stores data pertaining to these samples. A trustee oversees the pseudonymisation of data, acting as an interface between BOWL and DIZ, ensuring that patient data cannot be directly linked to patient samples."/>
<QaBoxq="What types of samples are collected in your biobank and for what research purposes are they used?"
a="The biobank collects a wide variety of samples, including blood, stool, and soil. Samples may be gathered for specific research projects or for establishing a general repository under 'broad consent.' Researchers wishing to use these samples must apply to the 'use access committee,' which evaluates whether the requested samples and data can be released for their research."/>
<QaBoxq="How large is your biobank? How many samples do you currently store and how many new samples are added on average?"
a="The biobank is still in the process of establishing itself and has not yet reached its full sample capacity. However, it is anticipated to accumulate a significant number of samples in the near future, with several thousand samples expected to be analyzed in dedicated sessions."/>
<QaBoxq="What requirements and criteria must be met for a sample to be included in your biobank?"a="Samples must be processed according to highly detailed protocols, and regular audits are conducted to ensure compliance with all standards."/>
<QaBoxq="Which other research institutions or biobanks do you cooperate with and what form does this cooperation take?"
a="Biobank OWL has a second location in Lippe, in addition to Bielefeld. Collaborations exist with the DIZ, the Treuhand, and three university hospitals. It is anticipated that cooperation with other working groups will increase in the future."/>
<QaBoxq="What specific storage conditions (e.g. temperature, humidity) must be observed for different sample types?"
a="Samples are stored under various temperature conditions, including -20°C, -80°C, and -150°C, along with the use of liquid nitrogen."/>
<QaBoxq="How do you ensure that the samples remain stable and usable over longer periods of time?"
a="Samples are stored in nitrogen for long-term stability."/>
<QaBoxq="What encryption techniques or data protection measures are used in your biobank to prevent unauthorized access to patient data? Are there special regulations for the anonymisation of data and how is it ensured that patients cannot be traced?"
a="Pseudonyms are created using specialized software such as CentraXX or REDcap to protect patient data."/>
<QaBoxq="What rights do patients have in relation to their samples, and how are these rights safeguarded in your biobank?"
a="Patients have the right to revoke their consent at any time, which can be done at the clinic or biobank. The trustee, acting as an intermediary, will notify BOWL and DIZ to destroy the corresponding samples or data."/>
</>,
months:"august"
},
{
title:"M.Sc.",
vorname:"Nils",
nachnname:"Berelsmann",
job:"PhD Working group: Prof. Dr. Gabriele Fischer von Mollard ",
affiliation:"University of Bielefeld",
pictureurl:pics['nilshefe'],
tag:"Academia",
heading:"Focus on adapting expression strategies for Fanzor nickases and exploring the potential of Pichia pastoris (SMD1163) for SpuFz1 nickase variants ",
interviewtabid:"nberelsmann",
cardtext:"",
quote:"X",
aimofcontact:[<p>During our interview with Makoto Saito[Linkinterview] about fanzor[link fanzor], it became evident that the expression of our fanzor nickases in yeast is very promising. We then refined our expression strategy for the nickases and approached Nils Berelsmann, who is currently working on his PhD thesis with the yeast strain Pichia pastoris (SMD1163). This particular strain could be ideal for expressing the SpuFz1 nickase variants. Our main aim in contacting Nils was to gain insight and advice on yeast expression and he generously shared his expertise with us. Not only did he give us valuable advice, but he also provided us with the yeast strain itself, along with a corresponding expression vector for possible experiments. He also provided us with detailed protocols and the plasmid map of the vector and gave us practical tips on how to optimize the expression process. His support was invaluable in moving our work forward. </p>],
insights:[<p>Pichia pastoris (SMD1163) is a promising option for expressing SpuFz1 nickase variants. Refining expression strategies based on expert insights is crucil for success. Nils provided practical tips on yeast expression, including optimizing growth conditions and fine-tuning induction protocols.</p>],
implementation:[<p>We adapted our expression strategy for Fanzor nickases in yeast by incorporating the Pichia pastoris strain (SMD1163) and the provided expression vector into our experiments. Following Nils' detailed protocols and plasmid map, we optimized key steps, enhancing expression efficiency and protein yield.</p>],
summary:"The team sought expert advice from Nils to optimize yeast expression for Fanzor nickases. Nils provided invaluable guidance on addressing potential challenges and troubleshooting the process. He supplied the Pichia pastoris (SMD1163) strain along with a suitable expression vector, crucial for expressing SpuFz1 nickase variants. Additionally, he shared detailed protocols for yeast transformation and growth optimization, enabling the team to replicate his methods effectively for their experiments.",
months:"July",
},
{
title:"M.Sc.",
vorname:"Kai",
nachnname:"Schülke",
job:"PhD student Working group: Organic chemistry and biocatalysis ",
affiliation:"University of Bielefeld",
pictureurl:pics['kaihammer'],
tag:"Academia",
heading:"First insights of Enzym Engineering",
interviewtabid:"hammerkai",
cardtext:"",
quote:"x",
aimofcontact:[<p>When we realized that the creation of a nickase from the endonucleases in use was a desired outcome, it became necessary to talk to an expert in the field of enzyme engineering. Our first contact was Kai Schülke, a former iGEMer and PhD student under the guidance of Prof. Dr. Hammer[Link Hammer], who is the leader of the working group organic chemistry and bioanalytics at Bielefeld University.</p>],
insights:[<p>In the process of our interaction with Kai, we learned about the various methods employed in enzyme engineering. He demonstrated the complexity of this field of research and emphasized the importance of choosing the right approach. As a former iGEMer, Kai, inspired by his past experiences, is highly motivated and determined to develop an outstanding project. He pointed out that we cannot rely on classical methods such as directed evolution, but instead should use a rational approach to select mutation candidates. His insights and enthusiasm have encouraged us to think critically and pursue innovative solutions in our work. </p>],
implementation:[<p>We incorporated Kai's insights into our project by shifting our approach to enzyme engineering. By focusing on a more targeted approach, we were able to refine our enzyme optimization process, ensuring that the modifications we made were based on informed, calculated decisions. This not only streamlined our research but also improved the chances of success by reducing the trial-and-error inherent in traditional methods. </p>],
summary:"The team reached out to Kai Schülke, a former iGEM participant and enzyme engineering expert, for guidance on developing a nickase from the endonucleases in use. Kai emphasized the need for a rational, targeted approach rather than traditional methods like directed evolution. His insights helped the team refine their enzyme optimization process, making it more strategic and efficient. This shift reduced trial-and-error efforts and improved the chances of success, driving innovation in their project.",
<QaBoxq="How do you assess the differences in the treatment and support of cystic fibrosis patients in Germany compared to other countries?"a="In Europe, especially in Germany, we live at a very high medical level, even if there are challenges such as the shortage of specialists. The difference in prosperity in the USA is greater, but at the same time it is also a driver of innovation. Compared to countries as some third-world countries, where cystic fibrosis patients have to wait in hospitals for appointments between infectious patients, we are complaining at a high level here. There is always room for improvement, but overall we have a brilliant healthcare system."/>
<QaBoxq="Do you feel sufficiently informed by the available sources of information?"a="Yes, the information situation is good. You can find sufficient information from organisations such as the CF Foundation, the CF Trust and Mukoviszidose e.V.. However, you have to get involved yourself and actively seek out the information."/>
<QaBoxq="Is there anything else you would like to tell us?"a="It is important to understand that CF patients are very different, including in terms of their cultural and family background. In addition, surveys and questions should be reviewed in advance by patients or parents to ensure that they are understandable and do not contain unfortunate wording. If you need any help or anything, I would love to help you. Send me the link as soon as the website is ready and I'll give you feedback. I can also circulate the survey in the community. I'm always available if you have any further questions or need support."/>
job:"PhD student Working Group Patel 'Fermentation and Formulation of Biologicals and Chemicals'",
affiliation:"at FH Bielefeld",
pictureurl:pics['moorlach'],
tag:"Academia",
heading:"Gathering information about Chitosan coating for RNA protection",
interviewtabid:"moorlach",
cardtext:"",
language:"de",
quote:"x",
aimofcontact:[<p>The aim of the contact with Benjamin Willem Moorlach, M.Sc., from the Department of Engineering and Mathematics, was to gain a deeper understanding of how Chitosan could be applied in lipid-based nanoparticles (LNPs) and to explore its potential role in our project. We had several questions focusing on the properties of Chitosan, its advantages and disadvantages, and how it could be integrated into LNPs. Benjamin Moorlach provided extensive insights into Chitosan’s interactions with RNA, its behavior, and how we might leverage it for our formulations. </p>],
insights:[<p>From our discussion, we gained valuable insights into the unique properties of Chitosan, a cationic polymer with significant potential to stabilize RNA. Notably, Chitosan offers strong protection against RNases, making it highly beneficial for formulations like lipid-based nanoparticles (LNPs). Another key feature is its heat stability, withstanding temperatures up to 121°C, which makes it suitable for processing methods such as spray drying. However, at higher concentrations (0.5% or more), Chitosan can become toxic, suffocating cells and displaying antimicrobial properties. While it differs from PEG and cannot serve as a direct alternative, Chitosan can be a valuable complement, especially in stabilizing RNA within LNPs.
A critical point Benjamin emphasized is that Chitosan must be in an acidic environment, typically with a pH range of 4 to 6, to remain positively charged. This positive charge is essential for its effective interaction with RNA and successful integration into the LNP system.
One of the most important attributes of Chitosan is its ability to form complexes with RNA, offering a high degree of protection, which is crucial for the stability of LNP formulations. This characteristic makes Chitosan particularly advantageous in enhancing RNA stability during processes like spray drying. However, incorporating Chitosan directly into the lipid shell of LNPs poses challenges due to its hydrophilic nature and incompatible charge ratios, which prevent its use as an external coating on LNPs. Instead, it is more suitable for forming stable RNA-Chitosan complexes that can be encapsulated within the LNP structure, ensuring improved stability and protection.</p>],
implementation:[<p>We have integrated the information by primarily using Chitosan as an RNA stabilizer, rather than embedding it directly into the LNP lipid shell. Benjamin suggested forming Chitosan-RNA complexes first and then encapsulating them within LNPs to ensure the RNA remains stable and functional. For this, Chitosan with a low molecular weight (around 5 kDa) is ideal, as it helps produce smaller particles that can be efficiently encapsulated.
Additionally, Benjamin recommended starting with small-scale tests (about 100 µL) before moving to larger formulations. The ratio of RNA to Chitosan is key to creating negatively charged particles, and a 2:1 ratio should be maintained. We will verify successful encapsulation using microscopic analysis and gel electrophoresis.
This knowledge has directly shaped our approach to using Chitosan. Our focus is now on forming stable RNA-Chitosan complexes, which can be encapsulated in LNPs. We’ve also learned the importance of optimizing concentrations to prevent aggregation or toxicity while ensuring the particles stay within the desired nanometer range. Microscopy and electrophoresis will now be key methods in our protocol to confirm complete RNA encapsulation within the LNPs. </p>],
summary:"In summary, the insights from Benjamin’s expertise were crucial in shaping our understanding of how to integrate Chitosan into our LNP formulations. Chitosan’s protective abilities for RNA, along with its heat stability, make it a valuable component in our project. However, its hydrophilic and cationic nature presents challenges for direct integration into LNP lipid shells, so we are focusing on its use as an encapsulation for the RNA. Benjamin’s advice on concentrations, molecular weight, and complex formation gave us a clear path forward, which will be validated through experimental testing. ",
months:"september"
},
{
vorname:"",
nachnname:"",
job:"Physical and Biophysical Chemistry working group of ",
affiliation:"Bielefeld University",
pictureurl:pics['physik'],
tag:"Academia",
heading:"",
interviewtabid:"biophysik",
cardtext:"",
quote:"",
aimofcontact:"",
insights:"",
implementation:"",
summary:"",
months:""
},
{
vorname:"Collaborations",
nachnname:"iGEM Team Linköping ",
pictureurl:pics['linköping'],
tag:"Other",
heading:"Cooperation to create a Lipid Delivery System Handbook",
interviewtabid:"handbook",
cardtext:"",
quoteVorname:"Kaya",
quoteNachname:"Lange",
quote:"We were genuinely excited when Linköping University approached us for collaboration. From the very beginning, their ideas resonated with us, and our shared enthusiasm laid a strong foundation for a productive partnership. We're happy to work together, also with the other teams, and explore new possibilities.",
aimofcontact:[<p>The initial contact for our collaboration came from the iGEM team 2024 of Linköping, Sweden, who approached us with a proposal to create a “Delivery-Based Handbook”[link Handbook]. Their goal was to reduce the steep learning curve associated with these technologies by sharing collective knowledge from multiple teams, including ours. We were excited to contribute and help future teams navigate these challenges more easily. The handbook would serve as a valuable tool. </p>],
insights:[<p>Throughout the collaboration, we gained significant insights, both scientific and collaborative. Initially, our meetings with the Linköping team and other participating teams - Patras, Radboud-University and TERMOSZ-Selye-HUN - were invaluable. These sessions allowed us to exchange ideas and learn how each team planned to use lipid-based delivery systems in their own projects. This mutual sharing of knowledge opened our eyes to new methodologies and potential applications of LNPs and liposomes. We also gained a deeper appreciation for the interdisciplinary nature of these systems. From the challenges of formulating stable particles to optimizing their efficiency in targeting cells, we realized the complexity of the field and how collaboration could help overcome many of these obstacles. By discussing our respective approaches, we were able to pool our expertise, which not only improved our understanding but also ensured that the handbook would be comprehensive and valuable for various iGEM teams, regardless of their specific project focus.
In summary: </p>,
<ul>
<li>Learned different approaches to using LNPs and liposomes in iGEM projects.</li>
<li>Discovered new methods for optimizing LNPs.</li>
<li>Recognized challenges in particle stability and targeted delivery.</li>
<li>Gained appreciation for the interdisciplinary complexity of these systems.</li>
<li>Focused on documenting work to benefit future iGEM teams.</li>
</ul>
],
implementation:[<p>The collaboration expanded our understanding of what's possible, inspiring us to consider new ideas for how we might integrate advanced techniques into our nanoparticle systems in future projects. The collaborative process also encouraged us to document our work more thoroughly, ensuring that future iGEM teams could benefit from both our successes and the challenges we encountered along the way. Beyond the technical improvements, the experience taught us the value of teamwork across borders and disciplines. Each team brought a unique perspective, and by working together, we were able to develop a resource that was far greater than the sum of its parts</p>],
type:"meta",
summary:"This collaboration with Linköping and the other iGEM teams was an incredibly enriching experience. Together, we developed a “Delivery-Based Handbook”[link Handbook] that will serve as a valuable resource for future teams working with LNPs and liposomes. The knowledge we gained not only enhanced our project but also strengthened our sense of community within iGEM. We are excited to present the handbook at the Grand Jamboree, where we will finally meet our collaborators in person and celebrate the culmination of our collective efforts. This partnership has shown us the immense power of collaboration, and we are proud to have been part of such a meaningful initiative.",
heading:"Successful participation of a team member in a 5 day GxP course",
interviewtabid:"gxpcourse",
cardtext:"",
quote:"The GXP course was extremely useful as it provided us with important knowledge that supports our entire team in complying with quality standards. This knowledge will help us to organise our processes efficiently and in accordance with regulations in the future.",
quoteVorname:"Kaya",
quoteNachname:"Lange",
text:[<p>I, Kaya, Team Member of iGEM Bielefeld 2024, recently participated in an intensive one-week GXP (Good Practice) training course, which was pivotal experience for both me and our project. The course covered essential regulatory frameworks, including</p>,
<ul>
<li>Good Laboratory Practice (GLP)</li>
<li>Good Clinical Practice (GCP)</li>
<li>Good Manufacturing Practice (GMP)</li>
</ul>,
<p>
which are all designed to ensure quality, safety, and compliance across every phase of scientific research and development.
As the head of Integrated Human Practices, I found this training particularly valuable. It provided me with a deeper understanding of the rigorous standards that need to be maintained in research, especially concerning ethics, data integrity, and patient safety. I learned how to properly document research processes, ensure the reproducibility of results, and assess and mitigate risks, all while keeping the ethical considerations of our project at the forefront.
I have acquired the ability to create standard operating procedures (SOPs) that guarantee the transparent and traceable documentation of each stage of the research process. This not only facilitates internal organisation but is also crucial for subsequent approvals and audits by regulatory authorities.
It is of paramount importance to ensure the reproducibility of our experiments by maintaining accurate protocols and meticulously documenting all variables. This is of particular importance should the intention be to pursue clinical research at a later stage, as the reproducibility of experiments is a crucial factor in the validity of the results.
I acquired knowledge of techniques for risk assessment, including Failure Mode and Effects Analysis (FMEA). This process enables the identification of potential risks in a project at an early stage, thus facilitating the development of strategies to minimise them. This approach allows us to identify and address potential sources of error before they lead to significant issues.
This knowledge is crucial as we think about the future of our project, particularly if we aim to move our gene therapy approach for cystic fibrosis closer to clinical trials and real-world applications. My participation in the GXP training has equipped me with the necessary tools to potentially guide our team through the complex regulatory landscape, ensuring our work remains aligned with industry standards and ready for the next steps in development.
One of the key speakers during the GXP course was Dr. Marcus Berger [LINK INtreview Beerger], whose expertise was invaluable to me and the entire team. After the course, I had the opportunity to ask Dr. Berger some questions, further deepening my understanding of the practical applications of GXP in research. The connection with Dr. Berger has been highly beneficial, as his insights helped shape key aspects of our project’s development and compliance with industry standards. His guidance will continue to be a valuable resource for our team moving forward.
Through this training, I feel better positioned to contribute to the team’s efforts, ensuring our project adheres to global safety and ethical guidelines. This experience has strengthened our approach and set a solid foundation for future progress, ensuring that our research, public engagement, and potential clinical applications continue to meet the highest regulatory standards. </p>],
type:"meta",
summary:"Kaya, a member of the iGEM Bielefeld 2024 team, completed an intensive one-week GXP (Good Practice) training, which covered Good Laboratory Practice (GLP), Good Clinical Practice (GCP), and Good Manufacturing Practice (GMP). The training provided valuable insights into maintaining high standards of quality, safety, and ethics throughout the research process. Kaya learned crucial skills, such as documenting research processes for reproducibility, creating standard operating procedures (SOPs), and conducting risk assessments using techniques like Failure Mode and Effects Analysis (FMEA). This knowledge is essential for advancing their cystic fibrosis gene therapy project toward clinical trials and ensuring compliance with regulatory standards. Dr. Marcus Berger, a key speaker in the course, provided additional guidance, offering valuable insights that will continue to benefit the team.",
<QaBoxq="What initial steps do we need to take to analyse the market and prepare for market access?"a="A stakeholder analysis of the market participants and a comparison of the new therapy with the standard therapy are required. Physician networks should be identified and the pricing strategy defined, taking into account the GBA and the health insurance funds."/>
<QaBoxq="How do we develop a strategy for the protection of intellectual property and patents?"a="The strategy should include patent applications in the following order: First for the active ingredient and the formulation (product patent), then the manufacturing route as process patent and followed by indication as use patent. Finally, a utility model may also be useful."/>
</>
}
},
{
vorname:"Physical ",
nachnname:"and Biophysical Chemistry ",
job:"Working group ",
affiliation:"University Bielefeld ",
language:"en",
pictureurl:pics['physik'],
tag:"Academia",
heading:"Performance of Experiments for LNP characterization ",
interviewtabid:"biophysik",
cardtext:"",
quote:"x",
aimofcontact:[<p>For our project, we collaborated closely with the Physical Chemistry workgroup to properly categorize our lipid nanoparticles (LNPs). We reached out to them to leverage their expertise and ensure that our characterization was thorough and precise. Marco, Uwe, and Yvonne were instrumental in this effort, not only advising us on appropriate characterization methods but also actively assisting us during the experimental process and data analysis. </p>],
insights:[<p>We employed several analytical techniques, including Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Dynamic Light Scattering (DLS), and zeta potential analysis. TEM and SEM allowed us to visualize the structural morphology of the LNPs, providing detailed images to understand their size and shape on the nanometer scale. DLS was used to measure the size distribution of the particles in solution, while the zeta potential analysis gave us insight into the surface charge, which is crucial for understanding stability in suspension. </p>],
implementation:[<p>Thanks to the guidance and hands-on support of the Physical Chemistry team, we successfully completed these tests, gaining detailed insights into our LNPs that will be crucial for our project's further development. Their expertise not only streamlined the process but also ensured the reliability and accuracy of our results. Here a sneak peak of the results – take a look at the image of our SORT LNP taken via TEM. </p>],
summary:"We collaborated with the Physical Chemistry workgroup to accurately characterize our lipid nanoparticles (LNPs). Their expertise, particularly from Marco, Uwe, and Yvonne, was invaluable in selecting and applying various analytical techniques, including Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Dynamic Light Scattering (DLS), and zeta potential analysis. This collaboration not only enhanced our understanding of the LNPs' size, shape, and stability but also ensured the reliability of our results. With their guidance, we successfully completed our tests, providing crucial insights for the project's advancement.",
job:"PhD Student Faculty of Biology / Working Group",
affiliation:"University Bielefeld ",
language:"en",
pictureurl:pics['hakan'],
tag:"Academia",
heading:"Performance of Experiments for LNP characterization ",
interviewtabid:"hakan",
cardtext:"",
quote:"Just hand me over the strain and the vector, I will try to take care of the rest.",
aimofcontact:[<p>After our interview with Makoto Saito[link] we learned, that he was not able to express the SpuFz1 protein in E. coli and recommended we used yeast to produce it. We were provided with a yeast expression strain and a suitable vector to clone the coding sequence into, but we lacked the necessary know-how and the facilities to transform yeast, select for positive transformants and cultivate the yeast. </p>],
insights:[<p>Hakan generously agreed to carry out the transformation and prepare potential positive transformants for cultivation for us, leaving only the purification of the proteins from the supernatant for us to do. </p>],
implementation:[<p>Hakan performed the transformation of a pPIC9K-n3SpuFz1 construct we created into Yeast. Unfortunately, the first attempt of transformation did not yield any positive clones. However, we value his spontaneous and extensive support as a great contribution to our project. </p>],
summary:"We collaborated with the Physical Chemistry workgroup to accurately characterize our lipid nanoparticles (LNPs). Their expertise, particularly from Marco, Uwe, and Yvonne, was invaluable in selecting and applying various analytical techniques, including Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Dynamic Light Scattering (DLS), and zeta potential analysis. This collaboration not only enhanced our understanding of the LNPs' size, shape, and stability but also ensured the reliability of our results. With their guidance, we successfully completed our tests, providing crucial insights for the project's advancement.",
months:"September",
},
{
title:"M.Sc.",
vorname:"Benjamin",
nachnname:"Moorlach",
job:"PhD student Working Group Patel 'Fermentation and Formulation of Biologicals and Chemicals'",
affiliation:"at FH Bielefeld",
pictureurl:pics['moorlach'],
tag:"Academia",
heading:"Gathering information about Chitosan coating for RNA protection",
interviewtabid:"moorlach",
cardtext:"",
language:"de",
quote:"x",
aimofcontact:[<p>The aim of the contact with Benjamin Willem Moorlach, M.Sc., from the Department of Engineering and Mathematics, was to gain a deeper understanding of how Chitosan could be applied in lipid-based nanoparticles (LNPs) and to explore its potential role in our project. We had several questions focusing on the properties of Chitosan, its advantages and disadvantages, and how it could be integrated into LNPs. Benjamin Moorlach provided extensive insights into Chitosan’s interactions with RNA, its behavior, and how we might leverage it for our formulations. </p>],
insights:[<p>From our discussion, we gained valuable insights into the unique properties of Chitosan, a cationic polymer with significant potential to stabilize RNA. Notably, Chitosan offers strong protection against RNases, making it highly beneficial for formulations like lipid-based nanoparticles (LNPs). Another key feature is its heat stability, withstanding temperatures up to 121°C, which makes it suitable for processing methods such as spray drying. However, at higher concentrations (0.5% or more), Chitosan can become toxic, suffocating cells and displaying antimicrobial properties. While it differs from PEG and cannot serve as a direct alternative, Chitosan can be a valuable complement, especially in stabilizing RNA within LNPs.
A critical point Benjamin emphasized is that Chitosan must be in an acidic environment, typically with a pH range of 4 to 6, to remain positively charged. This positive charge is essential for its effective interaction with RNA and successful integration into the LNP system.
One of the most important attributes of Chitosan is its ability to form complexes with RNA, offering a high degree of protection, which is crucial for the stability of LNP formulations. This characteristic makes Chitosan particularly advantageous in enhancing RNA stability during processes like spray drying. However, incorporating Chitosan directly into the lipid shell of LNPs poses challenges due to its hydrophilic nature and incompatible charge ratios, which prevent its use as an external coating on LNPs. Instead, it is more suitable for forming stable RNA-Chitosan complexes that can be encapsulated within the LNP structure, ensuring improved stability and protection.</p>],
implementation:[<p>We have integrated the information by primarily using Chitosan as an RNA stabilizer, rather than embedding it directly into the LNP lipid shell. Benjamin suggested forming Chitosan-RNA complexes first and then encapsulating them within LNPs to ensure the RNA remains stable and functional. For this, Chitosan with a low molecular weight (around 5 kDa) is ideal, as it helps produce smaller particles that can be efficiently encapsulated.
Additionally, Benjamin recommended starting with small-scale tests (about 100 µL) before moving to larger formulations. The ratio of RNA to Chitosan is key to creating negatively charged particles, and a 2:1 ratio should be maintained. We will verify successful encapsulation using microscopic analysis and gel electrophoresis.
This knowledge has directly shaped our approach to using Chitosan. Our focus is now on forming stable RNA-Chitosan complexes, which can be encapsulated in LNPs. We’ve also learned the importance of optimizing concentrations to prevent aggregation or toxicity while ensuring the particles stay within the desired nanometer range. Microscopy and electrophoresis will now be key methods in our protocol to confirm complete RNA encapsulation within the LNPs. </p>],
summary:"In summary, the insights from Benjamin’s expertise were crucial in shaping our understanding of how to integrate Chitosan into our LNP formulations. Chitosan’s protective abilities for RNA, along with its heat stability, make it a valuable component in our project. However, its hydrophilic and cationic nature presents challenges for direct integration into LNP lipid shells, so we are focusing on its use as an encapsulation for the RNA. Benjamin’s advice on concentrations, molecular weight, and complex formation gave us a clear path forward, which will be validated through experimental testing. ",
