diff --git a/src/contents/safety.tsx b/src/contents/safety.tsx index 11ab0d86c9edc5ec69c80e781610a69103724d38..d7dae1737cb5073680db8d877765700cd7953ce7 100644 --- a/src/contents/safety.tsx +++ b/src/contents/safety.tsx @@ -144,32 +144,32 @@ export const Safety: React.FC = () =>{ </p> <H4 text="PAM disrupt" ></H4> <p> - A key safety mechanism incorporated in our design of the Prime Editing complex is the disruption of the PAM sequence. For the nickase enzyme to function properly, it must bind directly to the DNA strand, a process that is facilitated by the presence of a specific sequence called the PAM (Protospacer Adjacent Motif). This critical interaction occurs through the recognition of the PAM sequence by the nickase itself. To achieve PAM disruption, the pegRNA (prime editing guide RNA) is specifically designed in a way so that the PAM sequence is situated within the reverse transcription template (RTT) of the pegRNA. By introducing a silent mutation within the RT template into the PAM sequence. Therefore the PAM sequence is effectively eliminated after the gene editing process is successfully completed <SupScrollLink label="desc-1"/>. As a result of that, the PAM sequence is no longer present on the DNA strand, preventing the nickase from binding again at the same location. This reduction in repeated or undesired binding of the nickase enhances the safety of our prime editing complex, minimizing the risk of unintended edits or off-target effects in subsequent steps. Ultimately, this feature contributes very much to the overall safety and reliability of the prime editing process. + A key safety mechanism incorporated in our design of the Prime Editing complex is the disruption of the PAM sequence. For the nickase enzyme to function properly, it must bind directly to the DNA strand, a process that is facilitated by the presence of a specific sequence called the PAM (Protospacer Adjacent Motif). This critical interaction occurs through the recognition of the PAM sequence by the nickase itself. To achieve PAM disruption, the pegRNA (prime editing guide RNA) is specifically designed in a way so that the PAM sequence is situated within the reverse transcription template (RTT) of the pegRNA. By introducing a silent mutation within the RT template into the PAM sequence. Therefore the PAM sequence is effectively eliminated after the gene editing process is successfully completed <SupScrollLink label="1"/>. As a result of that, the PAM sequence is no longer present on the DNA strand, preventing the nickase from binding again at the same location. This reduction in repeated or undesired binding of the nickase enhances the safety of our prime editing complex, minimizing the risk of unintended edits or off-target effects in subsequent steps. Ultimately, this feature contributes very much to the overall safety and reliability of the prime editing process. </p> <H4 text="pegRNA design - Spacer"></H4> <p> - Biosafety is also guaranteed by the careful selection of the spacer, which plays a critical role in guiding the complex to its intended target site <SupScrollLink label="desc-2"/>. To ensure both precision and safety, we meticulously chose and rigorously checked the spacer using the CRISPick software <SupScrollLink label="desc-3"/>. This allowed us to evaluate whether our Spacer would be likely to target other regions than our target site and therefore allowing us to analyse and predict potential off-target effects, ensuring that erroneous edits are minimised. By optimising the spacer selection, we have not only significantly enhanced the overall editing efficiency, striking a balance between precision and performance, but especially ensured the utmost accuracy in directing the Prime Editor, further contributing to the safety of the editing process. [Bild 1] + Biosafety is also guaranteed by the careful selection of the spacer, which plays a critical role in guiding the complex to its intended target site <SupScrollLink label="2"/>. To ensure both precision and safety, we meticulously chose and rigorously checked the spacer using the CRISPick software <SupScrollLink label="3"/>. This allowed us to evaluate whether our Spacer would be likely to target other regions than our target site and therefore allowing us to analyse and predict potential off-target effects, ensuring that erroneous edits are minimised. By optimising the spacer selection, we have not only significantly enhanced the overall editing efficiency, striking a balance between precision and performance, but especially ensured the utmost accuracy in directing the Prime Editor, further contributing to the safety of the editing process. [Bild 1] </p> <H4 text="Riboswitch"></H4> <p> - Riboswitches are segments of an RNA strand that bind to small molecules, causing them to change their secondary structure by forming hairpin structures. This process regulates gene expression at the translation level by preventing ribosomes from binding at the RBS and translating the coding region on the RNA strand. 0For our project we also considered an ion-sensitive riboswitch, specifically dependent on sodium ions (Naâº), as a regulatory mechanism. The secondary structure of this riboswitch prevents the binding of ribosomes to the ribosome binding site (RBS) under normal conditions, thus inhibiting the translation of the subsequent mRNA. When sodium ions bind to the riboswitch, a structural change occurs, exposing the RBS, which allows for the translation of the mRNA and the production of our fusion protein which is the main component of our prime editing system and therefore of enormous importance for it to work <SupScrollLink label="desc-4"/>. In the context of the CFTR mutation and its effects on the cell, the elevated Na⺠levels play a crucial role. Due to the dysfunctional CFTR channel, which fails to properly function as a chloride channel, the ENaC channel (epithelial sodium channel) becomes upregulated. This upregulation results in an increased transport of sodium ions into the cell, leading to a higher intracellular sodium concentration. This elevated Na⺠concentration creates a specific ionic environment that could potentially be utilized to regulate our Prime-Editing complex in a targeted manner. Given these specific ionic changes in the cell, we could have a disease-specific regulation of our Prime-Editing system based on the ionic situation typical of this condition. However, despite the initial promise of this approach, after further research, we concluded that the riboswitch, even considering the ion levels within epithelial cells, is overall too nonspecific and therefore too unreliable as a regulatory mechanism. Although the ion levels in CFTR cells are much lower, there are still low concentrations of sodium ions, which can lead to the riboswitch not being completely switched off. + Riboswitches are segments of an RNA strand that bind to small molecules, causing them to change their secondary structure by forming hairpin structures. This process regulates gene expression at the translation level by preventing ribosomes from binding at the RBS and translating the coding region on the RNA strand. 0For our project we also considered an ion-sensitive riboswitch, specifically dependent on sodium ions (Naâº), as a regulatory mechanism. The secondary structure of this riboswitch prevents the binding of ribosomes to the ribosome binding site (RBS) under normal conditions, thus inhibiting the translation of the subsequent mRNA. When sodium ions bind to the riboswitch, a structural change occurs, exposing the RBS, which allows for the translation of the mRNA and the production of our fusion protein which is the main component of our prime editing system and therefore of enormous importance for it to work <SupScrollLink label="4"/>. In the context of the CFTR mutation and its effects on the cell, the elevated Na⺠levels play a crucial role. Due to the dysfunctional CFTR channel, which fails to properly function as a chloride channel, the ENaC channel (epithelial sodium channel) becomes upregulated. This upregulation results in an increased transport of sodium ions into the cell, leading to a higher intracellular sodium concentration. This elevated Na⺠concentration creates a specific ionic environment that could potentially be utilized to regulate our Prime-Editing complex in a targeted manner. Given these specific ionic changes in the cell, we could have a disease-specific regulation of our Prime-Editing system based on the ionic situation typical of this condition. However, despite the initial promise of this approach, after further research, we concluded that the riboswitch, even considering the ion levels within epithelial cells, is overall too nonspecific and therefore too unreliable as a regulatory mechanism. Although the ion levels in CFTR cells are much lower, there are still low concentrations of sodium ions, which can lead to the riboswitch not being completely switched off. [Bild 2] As a further approach to developing alternative riboswitch variants, we considered the possibility of an RNA-regulated riboswitch targeting the defective mRNA sequence of the genetically defective CFTR gene. The basic idea behind this concept was that the riboswitch specifically binds to a region on the CFTR mRNA containing the F508Δ mutation. This binding should induce a structural change in the riboswitch on our prime editing complex’s mRNA that ultimately leads to exposure of the RBS to allow translation of the downstream sequence. This mechanism would be designed to react specifically to the defective CFTR mRNA and only cause a change in the secondary structure in the presence of the specific mutation. The riboswitch could thus ensure selective and disease-specific activation of our prime editing complex, which would be of particular interest in the context of genetic diseases such as cystic fibrosis. However, we did not pursue this approach any further. A major reason for this was the lack of sufficient literature providing a sound scientific basis for this specific application of a riboswitch. In addition, our research steered us in a different direction, particularly with regard to the alternative mechanism involving the XBP1 intron to regulate the prime editing system. This alternative seemed more promising and was based on an established regulatory mechanism that is triggered by cellular stress and specifically responds to misfolding processes. </p> <H4 text="XBP1 Intron"></H4> <p> - After extensive research, we discovered a regulatory system in eukaryotic cells, the XBP1 mechanism. The activation of XBP1 is an important mechanism that occurs as part of the Unfolded Protein Response (UPR), a cellular stress response triggered by the accumulation of misfolded proteins in the endoplasmic reticulum (ER). The ER is a key cellular component responsible for protein folding and transport. When many misfolded proteins accumulate in the ER, a specific regulatory mechanism is activated to reduce the stress on the ER. XBP1 activation is controlled by a protein called IRE1α, which is embedded in the ER membrane. IRE1α acts as a sensor for protein misfolding stress in the ER. Once IRE1α detects misfolded proteins, it dimerizes and becomes activated through autophosphorylation. This activation switches on the endoribonuclease activity of IRE1α, which is a crucial step in the activation of XBP1. The mRNA for XBP1 is continuously transcribed in the nucleus and transported to the cytoplasm, where it contains an intron that is not normally spliced out. This intron contains a stop codon, preventing the translation of a functional XBP1 protein. However, when ER stress activates IRE1α, the endoribonuclease domain of IRE1α splices this intron out of the XBP1 mRNA. This is an unconventional splicing event, as it occurs in the cytoplasm rather than in the nucleus. Once the intron is removed, the spliced XBP1 mRNA can be translated into a functional XBP1 protein. This activated XBP1 acts as a transcription factor, turning on genes that increase the protein-folding capacity of the ER and promote the degradation of misfolded proteins. In this way, XBP1 helps the cell cope with ER stress and restore balance in the protein-folding process. Thus, this mechanism originally functions within the cell in the context of ER stress to maintain ER function when protein folding is disrupted. <SupScrollLink label="desc-5"/><SupScrollLink label="desc-6"/> Our idea was therefore to integrate this intron into the mRNA encoding our prime-editing complex and thus use this mechanism to ensure that a functional prime editor is only synthesized when there is a high accumulation of misfolded proteins in the cell (similar to F508del). This would therefore represent an optimal safety aspect, as our fusion protein, which is essential for prime editing, cannot be fully synthesised as long as the genetic defect is not present in the cell. Accordingly, this provides the security that no healthy cells, as well as correctly edited cells, cannot be edited, which is an enormous contribution to biosafety. However, there was too much uncertainty about the extent to which other factors, such as misfolded proteins that are not associated with the CFTR protein, play a role in this mechanism. And since we could not and did not want to take the risk of such factors initiating the system, we decided against using it. To clarify this unknown correlation, we have considered a future experiment in which we want to switch this intron in front of a fluorescent marker and express it in cells with defective CFTR in order to confirm/investigate the dependence of intron splicing and the presence of CFTR F508del. + After extensive research, we discovered a regulatory system in eukaryotic cells, the XBP1 mechanism. The activation of XBP1 is an important mechanism that occurs as part of the Unfolded Protein Response (UPR), a cellular stress response triggered by the accumulation of misfolded proteins in the endoplasmic reticulum (ER). The ER is a key cellular component responsible for protein folding and transport. When many misfolded proteins accumulate in the ER, a specific regulatory mechanism is activated to reduce the stress on the ER. XBP1 activation is controlled by a protein called IRE1α, which is embedded in the ER membrane. IRE1α acts as a sensor for protein misfolding stress in the ER. Once IRE1α detects misfolded proteins, it dimerizes and becomes activated through autophosphorylation. This activation switches on the endoribonuclease activity of IRE1α, which is a crucial step in the activation of XBP1. The mRNA for XBP1 is continuously transcribed in the nucleus and transported to the cytoplasm, where it contains an intron that is not normally spliced out. This intron contains a stop codon, preventing the translation of a functional XBP1 protein. However, when ER stress activates IRE1α, the endoribonuclease domain of IRE1α splices this intron out of the XBP1 mRNA. This is an unconventional splicing event, as it occurs in the cytoplasm rather than in the nucleus. Once the intron is removed, the spliced XBP1 mRNA can be translated into a functional XBP1 protein. This activated XBP1 acts as a transcription factor, turning on genes that increase the protein-folding capacity of the ER and promote the degradation of misfolded proteins. In this way, XBP1 helps the cell cope with ER stress and restore balance in the protein-folding process. Thus, this mechanism originally functions within the cell in the context of ER stress to maintain ER function when protein folding is disrupted. <SupScrollLink label="5"/><SupScrollLink label="6"/> Our idea was therefore to integrate this intron into the mRNA encoding our prime-editing complex and thus use this mechanism to ensure that a functional prime editor is only synthesized when there is a high accumulation of misfolded proteins in the cell (similar to F508del). This would therefore represent an optimal safety aspect, as our fusion protein, which is essential for prime editing, cannot be fully synthesised as long as the genetic defect is not present in the cell. Accordingly, this provides the security that no healthy cells, as well as correctly edited cells, cannot be edited, which is an enormous contribution to biosafety. However, there was too much uncertainty about the extent to which other factors, such as misfolded proteins that are not associated with the CFTR protein, play a role in this mechanism. And since we could not and did not want to take the risk of such factors initiating the system, we decided against using it. To clarify this unknown correlation, we have considered a future experiment in which we want to switch this intron in front of a fluorescent marker and express it in cells with defective CFTR in order to confirm/investigate the dependence of intron splicing and the presence of CFTR F508del. </p> </Subesction> <Subesction title="Safety aspects of our Airbuddy" id="Biosafety2"> <H4 text="SORT LNP and Cytotoxicity"></H4> <p> - We have carefully considered the biosafety aspects of our delivery system, starting with the decision between Adeno-associated viruses (AAV) or LNPs as delivery systems. Our comparison revealed that the biocompatibility and safety of LNPs are paramount for our approach. That is why we chose selective organ-targeting (SORT) lipid nanoparticles (LNPs) <SupScrollLink label="desc-7"/> in the context of targeted pulmonary mRNA delivery. One of our primary concerns with the LNP was the potential cytotoxicity of polyethylene glycol (PEG), a common stabilizing agent in LNP formulations. Aware of the immune responses PEG can trigger, potentially leading to cytotoxicity <SupScrollLink label="desc-8"/>, we aimed at optimizing its concentration in our SORT LNPs to minimize such reactions while maintaining therapeutic efficacy. By the use of low molecular weight PEG, we addressed this problem. To test weather our approach succeeded, we conducted MTT and proliferation assays to ensure that our LNP posed no cytotoxicity risks. + We have carefully considered the biosafety aspects of our delivery system, starting with the decision between Adeno-associated viruses (AAV) or LNPs as delivery systems. Our comparison revealed that the biocompatibility and safety of LNPs are paramount for our approach. That is why we chose selective organ-targeting (SORT) lipid nanoparticles (LNPs) <SupScrollLink label="7"/> in the context of targeted pulmonary mRNA delivery. One of our primary concerns with the LNP was the potential cytotoxicity of polyethylene glycol (PEG), a common stabilizing agent in LNP formulations. Aware of the immune responses PEG can trigger, potentially leading to cytotoxicity <SupScrollLink label="8"/>, we aimed at optimizing its concentration in our SORT LNPs to minimize such reactions while maintaining therapeutic efficacy. By the use of low molecular weight PEG, we addressed this problem. To test weather our approach succeeded, we conducted MTT and proliferation assays to ensure that our LNP posed no cytotoxicity risks. </p> <H4 text="Precision of our SORT LNP"></H4> <p> - To further improve safety, we focused on reducing off-target effects. By incorporating specific SORT molecules, such as permanently cationic lipids like DOTAP, we ensured that the nanoparticles are systematically directed to the lungs. This precise targeting is particularly beneficial for respiratory diseases, as it enhances therapeutic effectiveness while limiting the impact on non-target organs. Our outlook of antibody conjugation as surface modification of our LNP for cell type-specific delivery, more exactly club cells <SupScrollLink label="desc-9"/> and ionocytes <SupScrollLink label="desc-10"/> as CFTR-expressing lung epithelial cells, would round off this aspect. + To further improve safety, we focused on reducing off-target effects. By incorporating specific SORT molecules, such as permanently cationic lipids like DOTAP, we ensured that the nanoparticles are systematically directed to the lungs. This precise targeting is particularly beneficial for respiratory diseases, as it enhances therapeutic effectiveness while limiting the impact on non-target organs. Our outlook of antibody conjugation as surface modification of our LNP for cell type-specific delivery, more exactly club cells <SupScrollLink label="9"/> and ionocytes <SupScrollLink label="10"/> as CFTR-expressing lung epithelial cells, would round off this aspect. </p> <p> In summary, our design strategy emphasizes both safety and efficacy. The careful optimization of components like PEG 2000 and the use of targeted delivery molecules allow SORT LNPs to deliver therapeutic agents directly to the lungs, reducing systemic exposure and minimizing side effects. This targeted approach ensures more effective treatments, especially for conditions requiring localized intervention. @@ -196,19 +196,19 @@ export const Safety: React.FC = () =>{ Given the sensitive nature of genome editing, our project presents specific biosecurity concerns that need to be assessed and mitigated. </p> <p> - <strong>Dual-Use Potential:</strong> One of the main biosecurity risks is the potential for dual-use of the Prime Editing technology. The system we are developing, while intended for therapeutic use, could be misused to target other genes or genomes for malicious purposes. <SupScrollLink label="desc-11"/> This includes the possibility of weaponizing the technology to induce harmful genetic changes in crops, animals, or even humans. The modular design of our plasmid system, although intended to facilitate optimization, could be exploited to exchange components for harmful applications, thereby increasing the risk of misuse. + <strong>Dual-Use Potential:</strong> One of the main biosecurity risks is the potential for dual-use of the Prime Editing technology. The system we are developing, while intended for therapeutic use, could be misused to target other genes or genomes for malicious purposes. <SupScrollLink label="11"/> This includes the possibility of weaponizing the technology to induce harmful genetic changes in crops, animals, or even humans. The modular design of our plasmid system, although intended to facilitate optimization, could be exploited to exchange components for harmful applications, thereby increasing the risk of misuse. </p> <p> - <strong>Unintendend Dissemination:</strong> Since our approach uses mRNA delivered via LNPs, there is a risk of unintended dissemination into the environment. If the LNPs are not adequately contained or disposed of, there is a possibility that they could be absorbed by non-target organisms, potentially leading to off-target genetic modifications.<SupScrollLink label="desc-12"/> In addition, the mRNA itself could theoretically be transferred between cells, especially if taken up by unintended hosts, raising concerns about unintentional spread in the environment. + <strong>Unintendend Dissemination:</strong> Since our approach uses mRNA delivered via LNPs, there is a risk of unintended dissemination into the environment. If the LNPs are not adequately contained or disposed of, there is a possibility that they could be absorbed by non-target organisms, potentially leading to off-target genetic modifications.<SupScrollLink label="12"/> In addition, the mRNA itself could theoretically be transferred between cells, especially if taken up by unintended hosts, raising concerns about unintentional spread in the environment. </p> <p> - <strong>Unauthorized Access:</strong> The genetic constructs and the detailed methodology of our Prime Editing system must be securely stored and protected.<SupScrollLink label="desc-13"/> If unauthorized individuals were to gain access to the plasmids, LNP formulations, or editing protocols, there is a risk of the technology being replicated or adapted for unintended, potentially harmful uses. This highlights the importance of proper biosecurity protocols in both physical and digital storage of our project materials. + <strong>Unauthorized Access:</strong> The genetic constructs and the detailed methodology of our Prime Editing system must be securely stored and protected.<SupScrollLink label="13"/> If unauthorized individuals were to gain access to the plasmids, LNP formulations, or editing protocols, there is a risk of the technology being replicated or adapted for unintended, potentially harmful uses. This highlights the importance of proper biosecurity protocols in both physical and digital storage of our project materials. </p> <p> - <strong>Synthetic Biology and information Sharing:</strong> The ease of synthesizing genetic material means that our project information could potentially be used to order similar constructs from commercial synthesis providers.<SupScrollLink label="desc-14"/> While these providers follow biosecurity guidelines, the increasing accessibility of synthetic biology raises the concern of our Prime Editing system being reproduced or modified without our knowledge. This includes potential attempts to bypass safety mechanisms or create variants that evade current regulatory frameworks. + <strong>Synthetic Biology and information Sharing:</strong> The ease of synthesizing genetic material means that our project information could potentially be used to order similar constructs from commercial synthesis providers.<SupScrollLink label="14"/> While these providers follow biosecurity guidelines, the increasing accessibility of synthetic biology raises the concern of our Prime Editing system being reproduced or modified without our knowledge. This includes potential attempts to bypass safety mechanisms or create variants that evade current regulatory frameworks. </p> <p> - <strong>Public Perception and Miscommunication:</strong> There is a biosecurity risk in how our project's technology is communicated to the public.<SupScrollLink label="desc-15"/> Miscommunication or misunderstanding of the project’s intent and capabilities could lead to misinformation, fear, or even attempts to replicate the technology outside of controlled and regulated environments. This could undermine public trust in legitimate therapeutic uses of genome-editing technologies and potentially facilitate misuse. + <strong>Public Perception and Miscommunication:</strong> There is a biosecurity risk in how our project's technology is communicated to the public.<SupScrollLink label="15"/> Miscommunication or misunderstanding of the project’s intent and capabilities could lead to misinformation, fear, or even attempts to replicate the technology outside of controlled and regulated environments. This could undermine public trust in legitimate therapeutic uses of genome-editing technologies and potentially facilitate misuse. </p> </Subesction> <Subesction title="Managing Risks" id="Biosecurity3"> @@ -281,13 +281,13 @@ export const Safety: React.FC = () =>{ <Section title="Bioethics" id="Bioethics"> <div> <p> - Bioethics is an interdisciplinary field of research that addresses ethical issues pertaining to the life sciences and medical research. It plays a pivotal role in contemporary research, particularly in projects that employ human samples or data. This is due to the fact that in these cases, the protection of the rights and dignity of the people involved is of the utmost importance <SupScrollLink label="1"/> <SupScrollLink label="desc-16"/>. In order to ascertain the necessity for an ethics application, an interview was conducted with Eva-Maria Berens, the scientific director of the office of the Ethics Committee at Bielefeld University, as part of the current research project. Following a comprehensive review, it was concluded that an ethics application was not necessary for the specific research project. Nevertheless, a comprehensive patient consent form was developed in conjunction with Eva-Maria Berens to guarantee that the donors of their samples are adequately informed and provide their consent of their own volition. The document guarantees that all pertinent information regarding sample collection, utilisation and storage is provided in an intelligible format. Furthermore, an interview was conducted with Dr. Timm Weber, a representative of the biobank, to discuss the topic of bioethics in greater depth. During the course of the interviews, the ethical aspects of sample storage and utilisation within the biobank were discussed in detail. Particular attention was paid to the responsible handling and protection of the rights of the test subjects. The discussion of bioethics in both interviews emphasises the relevance of ethical principles for research and ensures that it is conducted in accordance with the highest ethical standards. + Bioethics is an interdisciplinary field of research that addresses ethical issues pertaining to the life sciences and medical research. It plays a pivotal role in contemporary research, particularly in projects that employ human samples or data. This is due to the fact that in these cases, the protection of the rights and dignity of the people involved is of the utmost importance <SupScrollLink label="16"/>. In order to ascertain the necessity for an ethics application, an interview was conducted with Eva-Maria Berens, the scientific director of the office of the Ethics Committee at Bielefeld University, as part of the current research project. Following a comprehensive review, it was concluded that an ethics application was not necessary for the specific research project. Nevertheless, a comprehensive patient consent form was developed in conjunction with Eva-Maria Berens to guarantee that the donors of their samples are adequately informed and provide their consent of their own volition. The document guarantees that all pertinent information regarding sample collection, utilisation and storage is provided in an intelligible format. Furthermore, an interview was conducted with Dr. Timm Weber, a representative of the biobank, to discuss the topic of bioethics in greater depth. During the course of the interviews, the ethical aspects of sample storage and utilisation within the biobank were discussed in detail. Particular attention was paid to the responsible handling and protection of the rights of the test subjects. The discussion of bioethics in both interviews emphasises the relevance of ethical principles for research and ensures that it is conducted in accordance with the highest ethical standards. </p> </div> <Subesction title="Gene Therapy" id="Bioethics1"> <div> <p> - The potential of gene therapy to treat genetic diseases is promising, but it is also associated with significant ethical issues. One of the principal challenges is ensuring the safety of the procedure and the potential for unforeseen long-term consequences. Such consequences may only become apparent years after the genetic intervention has taken place. The modification of the germline, which affects not only the individual but also future generations, is a particularly sensitive issue. This gives rise to the question of the extent to which the decisions made today will influence future generations without their consent, thereby jeopardising intergenerational justice <SupScrollLink label="desc-17"/>. Another ethical issue is the potential for misuse for eugenic purposes. While the current focus is on combating disease, future applications could be aimed at 'optimising' human traits, which could result in a worsening of social inequalities. Access to gene therapy is also a significant issue. High costs could limit access to wealthy population groups, which would reinforce existing inequalities <SupScrollLink label="desc-18"/>. The issue of informed consent is also a key aspect. Many patients do not have the necessary knowledge to fully understand the complex risks, which raises ethical questions about their decision-making capacity. Overall, the debate around gene therapy highlights that ethical considerations such as safety, justice and patient rights need to be considered alongside scientific progress <SupScrollLink label="desc-19"/>. + The potential of gene therapy to treat genetic diseases is promising, but it is also associated with significant ethical issues. One of the principal challenges is ensuring the safety of the procedure and the potential for unforeseen long-term consequences. Such consequences may only become apparent years after the genetic intervention has taken place. The modification of the germline, which affects not only the individual but also future generations, is a particularly sensitive issue. This gives rise to the question of the extent to which the decisions made today will influence future generations without their consent, thereby jeopardising intergenerational justice <SupScrollLink label="17"/>. Another ethical issue is the potential for misuse for eugenic purposes. While the current focus is on combating disease, future applications could be aimed at 'optimising' human traits, which could result in a worsening of social inequalities. Access to gene therapy is also a significant issue. High costs could limit access to wealthy population groups, which would reinforce existing inequalities <SupScrollLink label="18"/>. The issue of informed consent is also a key aspect. Many patients do not have the necessary knowledge to fully understand the complex risks, which raises ethical questions about their decision-making capacity. Overall, the debate around gene therapy highlights that ethical considerations such as safety, justice and patient rights need to be considered alongside scientific progress <SupScrollLink label="19"/>. </p> </div> </Subesction> @@ -295,11 +295,11 @@ export const Safety: React.FC = () =>{ <div> <H4 text="Introduction of primary cultures"></H4> <p> - A primary culture is defined as a cell culture that is isolated directly from the tissue of an organism. In our case, the organism is human. The cells are then cultivated in a controlled environment, namely an S2 laboratory <SupScrollLink label="desc-20"/>. Primary cultures are a fundamental biomedical research tool, widely regarded as indispensable due to their capacity for realistic modelling of complex cell interactions. Primary cells are derived directly from the tissue of an organism and, as a consequence, they essentially retain their original properties. Consequently, they mirror the authentic conditions of the target tissue, which is vital for accurately assessing the impact of a therapeutic agent. In contrast, HEK cells represent transformed cell lines that exhibit physiological properties distinct from those of target cells in the human body. The effect of a therapeutic agent is typically limited to a specific cell type. The investigation of cell-specific effects and reactions of an active substance is feasible with the use of primary cells, as these possess the functional characteristics inherent to the cell type under consideration. Although HEK cells are relatively straightforward to cultivate, they are less representative of a number of tissue types and may activate other signalling pathways. The authenticity of the receptors and signalling pathways is guaranteed, as primary cells show the natural expression of receptors, ion channels and other cellular mechanisms. HEK cells are often genetically modified to express specific receptors, which can be useful for simple test systems. However, this does not reflect the complex environment of a real tissue. Given the sensitivity of primary cultures to environmental influences, thus resulting in higher risk of a contamination, it is imperative that researchers employ special safety measures to ensure the safety of themselves and the integrity of the cells. Primary cultures are employed extensively in the development of vaccines, cancer research and the investigation of basic cell processes. + A primary culture is defined as a cell culture that is isolated directly from the tissue of an organism. In our case, the organism is human. The cells are then cultivated in a controlled environment, namely an S2 laboratory <SupScrollLink label="20"/>. Primary cultures are a fundamental biomedical research tool, widely regarded as indispensable due to their capacity for realistic modelling of complex cell interactions. Primary cells are derived directly from the tissue of an organism and, as a consequence, they essentially retain their original properties. Consequently, they mirror the authentic conditions of the target tissue, which is vital for accurately assessing the impact of a therapeutic agent. In contrast, HEK cells represent transformed cell lines that exhibit physiological properties distinct from those of target cells in the human body. The effect of a therapeutic agent is typically limited to a specific cell type. The investigation of cell-specific effects and reactions of an active substance is feasible with the use of primary cells, as these possess the functional characteristics inherent to the cell type under consideration. Although HEK cells are relatively straightforward to cultivate, they are less representative of a number of tissue types and may activate other signalling pathways. The authenticity of the receptors and signalling pathways is guaranteed, as primary cells show the natural expression of receptors, ion channels and other cellular mechanisms. HEK cells are often genetically modified to express specific receptors, which can be useful for simple test systems. However, this does not reflect the complex environment of a real tissue. Given the sensitivity of primary cultures to environmental influences, thus resulting in higher risk of a contamination, it is imperative that researchers employ special safety measures to ensure the safety of themselves and the integrity of the cells. Primary cultures are employed extensively in the development of vaccines, cancer research and the investigation of basic cell processes. </p> <H4 text="Ethics in work with primary cultures"></H4> <p> - The term 'ethics' is used to describe the examination of moral principles that determine the behaviour of individuals or groups <SupScrollLink label="desc-21"/>. In a scientific context, the term 'ethics' encompasses the examination of the moral justifiability of actions and decisions, particularly with regard to the welfare of living beings and the responsible use of resources <SupScrollLink label="desc-22"/>. The isolation of primary cells from living organisms raises ethical questions, particularly in the case of human or animal tissue. In the context of research with animal primary cells, careful consideration must be given to the need for animal suffering and the potential benefits of the research <SupScrollLink label="desc-23"/>. An ethical dilemma frequently arises from the fact that primary cells offer the most meaningful data from a biological standpoint, yet their production is associated with challenges. In this context, the necessity of primary cell cultures is called into question, and the promotion of alternative methods, such as artificially produced tissues or organoids, is advocated where feasible. It is of crucial importance to emphasize the necessity of ethical responsibility in the collection of primary cultures. It is of the utmost importance that the procedure is carried out with consideration for the rights, and particularly the well-being of the donor. The removal of cells or tissue must be medically justifiable and, moreover, ethically justifiable in every case. To this end, the potential for research use and the possible risks and burdens for the donor must be weighed against each other to ensure careful consideration. However, it is also particularly important to ensure that the donor is involved in the entire process and is able to make an informed decision. The purpose of the research, the use of the cells and possible consequences must also be made transparent at all times. + The term 'ethics' is used to describe the examination of moral principles that determine the behaviour of individuals or groups <SupScrollLink label="21"/>. In a scientific context, the term 'ethics' encompasses the examination of the moral justifiability of actions and decisions, particularly with regard to the welfare of living beings and the responsible use of resources <SupScrollLink label="22"/>. The isolation of primary cells from living organisms raises ethical questions, particularly in the case of human or animal tissue. In the context of research with animal primary cells, careful consideration must be given to the need for animal suffering and the potential benefits of the research <SupScrollLink label="23"/>. An ethical dilemma frequently arises from the fact that primary cells offer the most meaningful data from a biological standpoint, yet their production is associated with challenges. In this context, the necessity of primary cell cultures is called into question, and the promotion of alternative methods, such as artificially produced tissues or organoids, is advocated where feasible. It is of crucial importance to emphasize the necessity of ethical responsibility in the collection of primary cultures. It is of the utmost importance that the procedure is carried out with consideration for the rights, and particularly the well-being of the donor. The removal of cells or tissue must be medically justifiable and, moreover, ethically justifiable in every case. To this end, the potential for research use and the possible risks and burdens for the donor must be weighed against each other to ensure careful consideration. However, it is also particularly important to ensure that the donor is involved in the entire process and is able to make an informed decision. The purpose of the research, the use of the cells and possible consequences must also be made transparent at all times. The obtaining of informed consent represents a fundamental aspect of ethical practice in the collection of primary cells. This process must encompass not only a formal consent procedure, but also the provision of comprehensive information to donors regarding the collection, utilisation and prospective future applications of the cells. The act of consent must be given freely and without undue influence, and donors must be fully aware of the consequences of their participation. Furthermore, donors must be granted the right to revoke their consent at any time without consequence. Prior to the collection of cells, a comprehensive discussion is held with the donor, during which all pertinent details are elucidated and any queries or concerns they may have, are addressed. This guarantees that the donor is adequately informed and is thus able to make an autonomous decision based on a comprehensive understanding of the procedure. The protection of privacy and confidentiality is of paramount importance when working with primary cultures. Given that primary cultures are predominantly human tissue, they contain genetic information and other personal data that is sensitive and deserving of protection. It is therefore of great importance that the data is anonymized and kept strictly confidential in order to protect the identity of the donor. Every person who has access to the data or samples must be obliged to comply with confidentiality standards. It must be ensured that all legal requirements for data protection are met, including compliance with data protection laws such as the GDPR in the EU. diff --git a/src/data/hptimelinedata.tsx b/src/data/hptimelinedata.tsx index 26e6882e7e28703e96b67ac721cea7981c500073..7f3e0b578870830714baf557aae466929aa149d3 100644 --- a/src/data/hptimelinedata.tsx +++ b/src/data/hptimelinedata.tsx @@ -52,6 +52,9 @@ const pics: { [key: string]: string } = { winkeljann: "https://static.igem.wiki/teams/5247/photos/hp/rnhale-winkeljann.jpg", kuehnel: "https://static.igem.wiki/teams/5247/photos/hp/hp-philippk-hnel.jpeg ", wischmeyer: "https://static.igem.wiki/teams/5247/photos/hp/wischmeyer-erhard.webp", + nicole: "https://static.igem.wiki/teams/5247/photos/hp/hp-friedlein-nicole.jpg", + joshua: "https://static.igem.wiki/teams/5247/photos/hp/joshua.jpg", + hammer: "https://static.igem.wiki/teams/5247/photos/hp/hp-hammer.webp", }; /* { @@ -221,20 +224,32 @@ export const timelinedata: Array<TimelineDatenpunkt> = [ }, { title: "Prof. Dr.", - vorname: "Christoph", + vorname: "Wolf-Michael", nachnname: "Weber", job: "", pictureurl: pics['placeholder'], tag: "Academia", - heading: "Feedback Session with Expert", + heading: "Feedback Session with Experts", interviewtabid: "weber", cardtext: "", language: "en", quote: "", - aimofcontact: "", - - insights: "", - implementation: "", + aimofcontact: [<p>The aim of the interview was to gain expert insights on optimizing the delivery of CFTR-mRNA via lung-targeted lipid nanoparticles (LNPs) for cystic fibrosis (CF) treatment. + Specifically, the goal was to explore potential cell targets, challenges in delivery mechanisms, and technical tools for assessing the effectiveness of mRNA therapies like the Ussing chamber system. </p>], + insights: [<p>The experts highlighted the potential of targeting ionocytes, given their key role in CFTR expression, but emphasized the difficulty in accessing them due to their basal position in the respiratory epithelium. + While Prof. Weber found ionocytes to be an intriguing target, Dr. Große-Onnebrink pointed out that there is still limited understanding of their exact role in CF pathology. Both stressed the challenge of penetrating the + mucus barrier in vivo, particularly when using air-liquid interface cultures, and underscored the importance of optimizing particle size to ensure effective delivery to the deeper regions of the lungs. + Prof. Weber also emphasized the need to test whether the system can still transfect cells in the presence of mucus. </p>, + <p>It was suggested to use the Ussing chamber to assess the effectiveness of the delivery system and therapeutic mRNA, though they noted certain challenges with this technique. We also discussed alternatives like organoids, + which offer only indirect measurements of CFTR function, and patch clamping, which, though more precise, is a more complex and expensive method. Additionally, Prof. Weber recommended exploring chitosan-based delivery + systems due to their success in his previous work, suggesting they could be a safer alternative to PEG-lipid systems, which had shown issues with cytotoxicity. </p>], + implementation: [<p>These insights helped refine our iGEM project in several key ways: + <ul> + <li>Cell Targeting: We decided to continue exploring ionocytes as a target but acknowledged the technical hurdles involved. We also expanded our focus to include multiple cell types to test different delivery systems. </li> + <li>Delivery Systems: We began investigating chitosan-based nanoparticles as a safer alternative to PEG-lipid systems. The suggestion to optimize particle size and delivery for inhalation was also integrated into our design. </li> + <li>Experimental Tools: Based on the discussion, we plan to use an Ussing chamber to measure overall CFTR function in different cell types but will also explore organoid-based approaches for preliminary testing. Additionally, we consulted the medical faculty on the possibility of using patch clamping for more detailed measurements of successful transfection and restored CFTR function. </li> + </ul>, + </p>], }, { vorname: "Exploring new ideas", @@ -506,16 +521,58 @@ export const timelinedata: Array<TimelineDatenpunkt> = [ nachnname: "Friedlein", job: "Research group on fundamental rights", affiliation: "Universität Potsdam", - pictureurl: pics['placeholder'], + pictureurl: pics['nicole'], tag: "Academia", heading: "Discussion on how health insurance companies manage cystic fibrosis patients and gene therapy treatments", interviewtabid: "nicole", cardtext: "", language: "de", - quote: "", + quote: "Public health insurance operates under an economic efficiency principle, meaning the most cost-effective treatments are preferred. But if gene therapies become the only treatment option for certain conditions, they will likely have to be included in the coverage, which could be a challenge for the system.", aimofcontact: "The main objective of the contact was to learn from the discussion on issues related to cystic fibrosis (CF), gene therapy, health insurance processes and regulatory pathways. In particular, we wanted to understand the real-world challenges and technical aspects of gene editing, especially prime editing, as well as the complexities of approval and reimbursement of gene therapies for CF patients.", insights: "The regulatory approval process, particularly by the European Medicines Agency (EMA) for advanced medical devices, has highlighted the bureaucratic hurdles that gene therapies must overcome. We learned that such therapies for cystic fibrosis have to navigate complex European and German regulatory systems. The discussion on the AMNOG process was crucial. We learnt that the additional benefit of a therapy is assessed for reimbursement by the statutory health insurance funds. We implemented this insight in our project by considering the long-term regulatory and economic effects as important milestones for therapy development. We also gained insight into how public and private health insurers may differ in their reimbursement of such therapies. Public insurers have stricter guidelines, while private insurers can be more flexible, but both require strict justification, especially for rare diseases such as cystic fibrosis. Information on newborn screening and genetic counselling covered by public health insurance was crucial to understanding how preventive measures for CF are managed. This underlines the importance of early intervention and diagnosis in our project. Atypical forms of CF, where health insurance companies do not cover treatment due to non-standardised test results, were identified as a key problem. This helped us to recognise the need for more adaptable insurance policies and clearer pathways for the treatment of atypical cases in our project plans. The debate about whether healthcare systems can afford the high costs of gene therapies highlighted an important issue in the current medical landscape. We have incorporated this insight into our project by discussing possible cost-effective alternatives and the need for thorough cost-benefit analysis in the development of treatments.", implementation: "After the interview, we further tailored our project to focus on a simple delivery method. To gain an overview of the regulatory requirements and to better deliver the project, one of our team members attended a GxP course to ensure we met all the necessary standards. To deepen our knowledge of entrepreneurship, we conducted further interviews with start-ups and industrial companies, which gave us important insights into practical implementation. These steps ensure that our project is not only based on scientific research, but also takes into account the practical, regulatory and social aspects that are crucial to bringing new CF therapies to the market. We are currently developing strategies to successfully implement our ideas and the project in the future.", + interview: <> + <QaBox q="To start with this interview. Do you have any questions about this project?" + a="Are you writing a paper on this, or are you conducting actual laboratory research? Or is it primarily literature review? How does your work look?" /> + <QaBox q="It’s not just literature review, though we do start with that. We have a lot of lab work ahead of us. Ideally, we would have a finished construct to present at the end, maybe even a functional gene therapy, though that’s quite ambitious and probably not possible in the short time frame. We’re working on various gene-editing approaches and testing plasmids in HEK cells. We are also exploring Prime Editing and trying to improve its efficiency using different reverse transcriptase enzymes. So, it’s a mix of lab work, literature research, and preparing for a presentation at a competition." + a="Are you writing a formal paper?" /> + <QaBox q="We’re not writing a formal text-based paper, but everything will be available on a website. We will document most of our work on the website, with sub-pages detailing lab work, interviews, and research." + a="What exactly is Prime Editing, and how does it differ from altering the germline? Where in the genome does this therapy act?" /> + <QaBox q="Our current plan is to deliver the therapy via a lipid nanoparticle system, which will be inhaled and go into the lungs. While cystic fibrosis (CF) affects all mucus membranes, the lungs are the most critical area, so we’re focusing on that. The therapy will only target surface cells in the lungs, not the basal cells responsible for producing new lung cells." + a="Thank you for giving me insights into your project." /> + <QaBox q="Do you know how cystic fibrosis (CF) approval works in terms of health insurance and regulatory processes?" + a="The approval process for gene therapies is primarily done through the EMA (European Medicines Agency) under specific EU regulations for Advanced Medical Products, including gene therapies. There is also a national approval process in Germany for individualized treatments, but large-scale therapies must go through the EU process." /> + <QaBox q="Can you share more about the approval and reimbursement processes for CF treatment?" + a="The approval process is separate from reimbursement by public health insurance. CF is considered a rare disease if it affects fewer than five out of 10,000 people, and treatments for rare diseases often face special reimbursement challenges. If more than five out of 10,000 people are affected, the disease is relatively common, and approval and reimbursement go through a different procedure known as the AMNOG process. For more common diseases, an additional benefit (Zusatznutzen) must be demonstrated during the approval process." /> + <QaBox q="Have you heard about issues with reimbursement from private insurance companies?" + a="We’ve heard that private insurance companies can make it difficult to get treatments reimbursed, especially experimental ones. One of our colleagues almost had to go to court to get his treatment reimbursed by his private insurer, which was quite expensive. Eventually, he switched to public insurance, but the situation was difficult." /> + <QaBox q="Why did your colleague have issues with private insurance?" + a="He was privately insured, but the treatment was very expensive, around €16,000 per month, and the insurance company was reluctant to cover it." /> + <QaBox q="Do you need legal information for your project?" + a="Both. We want to be well-informed to identify potential obstacles early on, such as legal restrictions or bans on altering certain chromosomes. Although we won’t be running clinical trials, understanding the regulatory landscape is crucial for our future planning." /> + <QaBox q="How does genetic counseling and testing work for CF?" + a="Genetic counseling and testing are usually covered by health insurance if there’s a medical reason, such as a family history or suspicion that the parents might be carriers. However, if both parents are healthy and there’s no family history of CF, insurance might not cover the tests." /> + <QaBox q="Are there differences between public and private insurers for genetic tests?" + a="Public insurance has different regulations than private insurance, but I’m not entirely sure if that leads to different decisions regarding genetic testing. I can look into the public insurance regulations if that would be helpful." /> + <QaBox q="Is newborn screening for CF covered by health insurance?" + a="Yes, newborn screening is part of a set of health examinations for children and adolescents, regulated under §26 SGB V (Social Security Code). Since it’s part of the regular screening process, it’s covered by health insurance without additional requirements." /> + <QaBox q="How does public insurance handle CF treatment when a test comes back negative?" + a="Public health insurance works with standardized guidelines, and if a test comes back negative, it may no longer meet the criteria for coverage. However, if a doctor reconfirms the diagnosis, the treatment should continue to be covered." /> + <QaBox q="Is there no rule that says genetic diseases, once diagnosed, should remain covered since genetics don’t change?" + a="In theory, yes. But the guidelines are usually based on medical evidence at the time, and re-testing can sometimes lead to complications in terms of coverage if the result differs. However, with proper medical documentation, it should be possible to maintain coverage." /> + <QaBox q="Have recent changes in gene therapy costs impacted public health insurance?" + a="Not much has changed. It’s a political and societal question—how willing are we to finance these expensive therapies? Right now, public health insurance operates under an economic efficiency principle, meaning the most cost-effective treatments are preferred. But if gene therapies become the only treatment option for certain conditions, they will likely have to be included in the coverage, and it could be a challenge for the system. There are also ongoing price negotiations between insurers and manufacturers." /> + <QaBox q="Do patents play a significant role in keeping gene therapy costs high?" + a="Yes, patents certainly influence the price, but the production of gene therapies is inherently expensive due to the complex research and manufacturing process." /> + <QaBox q="Would private supplemental insurance be an option for covering expensive gene therapies?" + a="It’s possible that private supplemental insurance could cover these therapies if public health insurance doesn’t. However, this raises concerns about equity and accessibility. If public insurance doesn’t cover it, the burden might fall on private insurance, which could create disparities in access to treatment." /> + <QaBox q="Is gene therapy research driven more by biology or medicine?" + a="It’s definitely interdisciplinary. Both biologists and medical professionals contribute. For example, at our university, the medical and biology faculties collaborate closely. Biologists usually handle the research, while medical professionals focus more on clinical applications." /> + <QaBox q="Do biologists or medical professionals develop gene therapies?" + a="In terms of development, it’s mainly biologists and biotechnologists. Medical professionals get involved primarily in clinical trials. Some doctors do research, but they’re often needed in hospitals, so hands-on development is mostly handled by molecular biologists or biotechnologists." /> + <QaBox q="Does research in genome medicine and gene therapies come from biology, medicine, or both?" + 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." /> + </> }, { vorname: "Katrin", @@ -600,7 +657,7 @@ export const timelinedata: Array<TimelineDatenpunkt> = [ nachnname: "Bauder", job: "parent and activist", affiliation: "CF vests worldwide", - pictureurl: pics['placeholder'], + pictureurl: pics['joshua'], tag: "Patient", heading: "Interview with a CF Parent and Global Advocate on Worldwide Support and Perspectives", interviewtabid: "joshua", @@ -837,7 +894,7 @@ export const timelinedata: Array<TimelineDatenpunkt> = [ nachnname: "Hammer", job: "Junior Professor of Organic Chemistry and Biocatalysis", affiliation: "Universität Bielefeld", - pictureurl: pics['placeholder'], + pictureurl: pics['hammer'], tag: "Academia", heading: "Safety Briefing and Laboratory Practices Advice", interviewtabid: "hammer", @@ -879,10 +936,17 @@ export const timelinedata: Array<TimelineDatenpunkt> = [ cardtext: "", language: "de", quote: "", - aimofcontact: "", - - insights: "", - implementation: "", + aimofcontact: [<p>We contacted Svenja Vinke, a former iGEMer from Bielefeld, to get her insight and her opinion regarding the use of phage assisted continuous evolution + (PACE, see engineering cycle 1[Link]) for our prime editing optimizations. Svenja works at the iGEM Safety and Security Committee. Additionally, she was part of the Biosafety and Security Award Team of Bielefeld University in 2016.</p>], + insights: [<p>Svenja explained, that a PACE approach is not feasible to use for optimization of our nickase candidates because of several reasons:</p>, + <ul> + <li>Implementing a PACE system takes way too much time to use for our project.</li> + <li>Endonucleases might be too big to optimize using PACE.</li> + <li>Unspecific cutting or nicking in the bacterial genome kills the cells, which makes optimization of endonucleases very challenging.</li> + <li>Prime editing in E. coli cells is less effective compared to human cells, which might impair the evolution process.</li> + </ul> + ] , + implementation: [<p>On the basis of Svenja’s and other opinions on the topic, we decided not to try implementing a PACE system. </p>], }, { vorname: "Max", @@ -911,7 +975,7 @@ export const timelinedata: Array<TimelineDatenpunkt> = [ interviewtabid: "berens", cardtext: "", language: "de", - quote: "", + quote: "The most important thing is a detailed letter of consent and a detailed privacy policy. This must explain to the patient as precisely as possible what happens to their cells and data, as well as the time span, which people are involved or have access to the cells and how.", aimofcontact: "The aim of the interview was to get an answer to the question of whether we need an ethics vote for our project or not and to obtain guidelines for dealing with patient cells regarding ethical issues and data protection. ", insights: "The discussion was very informative in terms of how we should approach this topic and focused primarily on the important factors that need to be considered when planning the handling of patient cells. These include which legal principles need to be observed, data protection, ethical considerations and, above all, detailed and specific information for the donor. It also made us look at the situation from many different angles and consider the risks of worst-case scenarios. Overall, this interview was very useful to us, and we were able to use the information we gained to develop a kind of guideline that allowed us to approach this sensitive topic, which was new to us, with a certain degree of confidence. ", implementation: "Based on the knowledge we have gained, we have drawn up guidelines for our handling of the cells. We used this guide when handling the patient cells, to ensure they were handled in an ethically correct manner.",