Research on the safety of viral vectors
1、Introduction
Viral vectors are the most popular delivery method for cell therapy and gene therapy. In cell and gene therapy, viral vectors expressing anti-tumor, cytotoxic and immunostimulatory genes, such as CAR, cytokines and chemokines, have been approved for the treatment of cancer, metabolic, neurological and ophthalmic diseases. Compared with non-viral vectors, viral vectors have superior gene delivery capabilities, such as high delivery efficiency and minimal damage to cells, and therefore play a central role in cell and gene therapy [1].
2、Retroviruses
(1) Integration sites and safety accidents
Retroviruses (RVs), due to their reverse transcription activity, have RNA genomes that are capable of producing dsDNA copies and integrating into the host genome. Chromosomal integration is beneficial to long-term transgene expression. Integration of gamma retroviral vectors (γ-RVV) tends to be inserted near the transcription start site of the gene. Once γ-RVV was used in gene therapy, it once led to treatment leukemia-induced adverse events in SCID-X1 patients [2]. Therefore, researchers designed a self-inactivating γ-RV vector (SIN-γRVV), which has been proven to be safe in clinical trials [3]. In adenosine deaminase deficiency severe combined immunodeficiency (ADA-SCID), the occurrence of insertional tumors after γ-RVV treatment is extremely rare [4]. The retroviral vectors underlying the cell products currently on the market are all self-losing. Live carrier, high safety. Lentiviral vectors (LVVs) preferentially integrate into active transcription units, and there are also reports of adverse events and random insertion leading to tumorigenesis. In June 2021, Bluebird Biotech suspended 2 clinical trials of LentiGlobin (BB305 LV) for the treatment of sickle cell disease because 2 patients receiving LentiGlobin were diagnosed with acute myeloid leukemia and myelodysplasia respectively. syndrome[5]. Therefore, both lentiviral vectors and retroviral vectors have the risk of causing cancer due to random insertion, but the probability is extremely low (nearly one in 10 million). As of now, among the cell products that have been marketed, whether they use retroviral vectors Whether it is a lentiviral vector or a lentiviral vector, there are no safety incidents caused by random integration.
(2)Risk of replicating viruses
Retroviral vectors or lentiviral vectors are also commonly used vectors to efficiently introduce CAR genes into T cells in CAR-T cell therapy, but they may produce replication-competent retroviruses (RCR) or replication-competent lentiviruses. (Replication Competent Lentivirus, RCL) potential risk of contamination. RCR/RCL can also be integrated into the cell genome, thereby creating the risk of insertional tumors due to integration that activates proto-oncogenes, destroys tumor suppressor genes, or increases the expression of factors that promote cell growth; on the other hand, because it can The production of replication-competent viruses also increases the risk of integration and insertional tumors [6]. In a trial of in vitro infusion of bone marrow progenitor cells, some researchers found that 3 of 10 severely immunodeficient rhesus monkeys treated with γ-RV developed lymphoma [7]. Analysis of the reasons found that RCR was detected in lymphoma tissues, and animals with lymphoma developed gamma-retroviremia, which was also caused by RCR [8]. Therefore, the real culprit of lymphoma in animals is RCR rather than RCR. Gamma-retroviral vectors. Similarly, since the envelope of lentiviral vectors is usually replaced with membrane proteins from other viruses, such as the commonly used VSV-G membrane protein, this may increase the potential risk caused by RCL contamination [6]. Therefore, the contamination risk of RCR/RCL is always a safety issue that we should focus on.
3、DNA virus vector
DNA virus vectors mainly include adenovirus (AV) and adeno-associated virus (AVV). Adenovirus is a non-enveloped double-stranded DNA virus. The gene of the adenovirus vector will not be integrated into the genome after introduction, so there is no risk of random insertion. High immunogenicity, cytotoxicity, and transient transgene expression still make adenoviral vector research challenging. In 1999, an 18-year-old patient with an ornithine transcarbamylase deficiency died after systemic injection of a high-dose adenoviral vector due to an innate immune response to the capsid protein that triggered cytokine release syndrome. syndrome, CRS, also known as cytokine storm). Currently, gene therapy based on adenovirus vectors is mainly used in gene vaccines and anti-tumor treatments [5]. AVV is a non-enveloped parvovirus with single-stranded DNA. Its vector has the characteristics of broad tropism, low immunogenicity and easy production, which is beneficial to clinical application. However, research results show that AAV-delivered RNA interference (RNAi) therapy can cause neurotoxicity in the brain of non-human primates [9] .
4、Summary
In summary, the application of viral vectors in cell therapy is very safe. Viral vectors are still a reliable, safe and efficient delivery method in the development of cell drugs. However, high attention should be paid to their viruses in related gene therapy and stem cell-related treatments. Carrier safety issues. This requires us to continuously optimize the design and production of viral vector processes and control the quality control system to ensure the clinical safety and effectiveness of cell and gene therapy products, so that cell and gene therapy based on viral vectors can benefit more with minimal safety issues. Many patients.
Reference：
[1] Lundstrom K. Viral Vectors in Gene Therapy: Where Do We Stand in 2023? Viruses. 2023 Mar 7; 15(3):698. doi: 10.3390/v15030698. PMID: 36992407; PMCID: PMC10059137.
[2]Hacein-Bey-Abina S, Garrigue A, Wang GP, Soulier J, Lim A, Morillon E, Clappier E, Caccavelli L, Delabesse E, Beldjord K, Asnafi V, MacIntyre E, Dal Cortivo L, Radford I, Brousse N, Sigaux F, Moshous D, Hauer J, Borkhardt A, Belohradsky BH, Wintergerst U, Velez MC, Leiva L, Sorensen R, Wulffraat N, Blanche S, Bushman FD, Fischer A, Cavazzana-Calvo M. Insertional oncogenesis in 4 patients after retrovirus-mediated gene therapy of SCID-X1. J Clin Invest. 2008 Sep; 118(9):3132-42. doi: 10.1172/JCI35700. PMID: 18688285; PMCID: PMC2496963.
[3]Hacein-Bey-Abina S, Pai SY, Gaspar HB, Armant M, Berry CC, Blanche S, Bleesing J, Blondeau J, de Boer H, Buckland KF, Caccavelli L, Cros G, De Oliveira S, Fernandez KS, Guo D, Harris CE, Hopkins G, Lehmann LE, Lim A, London WB, van der Loo JC, Malani N, Male F, Malik P, Marinovic MA, McNicol AM, Moshous D, Neven B, Oleastro M, Picard C, Ritz J, Rivat C, Schambach A, Shaw KL, Sherman EA, Silberstein LE, Six E, Touzot F, Tsytsykova A, Xu-Bayford J, Baum C, Bushman FD, Fischer A, Kohn DB, Filipovich AH, Notarangelo LD, Cavazzana M, Williams DA, Thrasher AJ. A modified gamma -retrovirus vector for X-linked severe combined immunodeficiency. N Engl J Med. 2014 Oct 9; 371(15):1407-17. doi: 10.1056/NEJMoa1404588. PMID: 25295500; PMCID: PMC4274995.
[4] Pai SY. Built to last: gene therapy for ADA SCID. Blood. 2021 Oct 14; 138 (15) : 1287-1288. The doi: 10.1182 / blood. 2021012300. PMID: 34647983; PMCID: PMC8525332.
[5] Li Manqi, Wei Liping, Tao Qiaoyu et al. Gene therapy carrier, research progress and safety applications [J]. China's pharmaceutical industry magazines, does 2022 (12) : 1671-1682. The DOI: 10.16522 / j.carol carroll nki CJPH. 2022.12.001.
[6] Wu Xueling, Zhao Xiang, Meng Shufang. Risk analysis and control of replicative viruses in CAR T cell therapy products [J]. China Pharmaceutical Journal,2018,32(07):879-885.DOI:10.16153/ J.1002-7777.2018.07.006.
[7]Donahue RE, Kessler SW, Bodine D, McDonagh K, Dunbar C, Goodman S, Agricola B, Byrne E, Raffeld M, Moen R, et al. Helper virus induced T cell lymphoma in nonhuman primates after retroviral mediated gene transfer. J Exp Med. 1992 Oct 1; 176(4):1125-35. doi: 10.1084/ jem.176.4.1125.PMID: 1383375; PMCID: PMC2119385.
[8]Vanin EF, Kaloss M, Broscius C, Nienhuis AW. Characterization of replication-competent retroviruses from nonhuman primates with virus-induced T-cell lymphomas and observations regarding the mechanism of oncogenesis. J Virol. 1994 Jul; 68(7):4241-50. doi: 10.1128/JVI.68.7.4241-4250.1994. PMID: 8207799; PMCID: PMC236347.
[9]Keiser MS, Ranum PT, Yrigollen CM, Carrell EM, Smith GR, Muehlmatt AL, Chen YH, Stein JM, Wolf RL, Radaelli E, Lucas TJ 2nd, Gonzalez-Alegre P, Davidson BL. Toxicity after AAV delivery of RNAi expression constructs into nonhuman primate brain. Nat Med. 2021 Nov; 27(11):1982-1989. doi: 10.1038/s41591-021-01522-3. Epub 2021 Oct 18.PMID: 34663988; PMCID: PMC8605996.
Disclaimer: Shenzhen Cell Valley is committed to the research of cell and gene therapy, in order to promote emerging technologies and let more people understand the new developments in biomedicine.The content of this article is for information exchange only. This platform remains neutral with respect to the content, statements, and opinion judgments in the article, and does not represent the position and opinions of Shenzhen Cell Valley.The relevant information in this article should not be used for diagnosis or treatment, and cannot replace professional medical advice. Our website will not assume any responsibility.The final interpretation of the above statement belongs to our company’s website. This statement will apply to articles shared on our website at all times. Thank you for your cooperation! Copyright statement: The copyright of the article belongs to Shenzhen Cell Valley. Individuals are welcome to forward it to friends, media or Any unauthorized reproduction by the organization to other platforms will be regarded as infringement.If you need to reprint, please contact email: contact@sz-cell.com
Viral vectors are the most popular delivery method for cell therapy and gene therapy. In cell and gene therapy, viral vectors expressing anti-tumor, cytotoxic and immunostimulatory genes, such as CAR, cytokines and chemokines, have been approved for the treatment of cancer, metabolic, neurological and ophthalmic diseases. Compared with non-viral vectors, viral vectors have superior gene delivery capabilities, such as high delivery efficiency and minimal damage to cells, and therefore play a central role in cell and gene therapy [1].
2、Retroviruses
(1) Integration sites and safety accidents
Retroviruses (RVs), due to their reverse transcription activity, have RNA genomes that are capable of producing dsDNA copies and integrating into the host genome. Chromosomal integration is beneficial to long-term transgene expression. Integration of gamma retroviral vectors (γ-RVV) tends to be inserted near the transcription start site of the gene. Once γ-RVV was used in gene therapy, it once led to treatment leukemia-induced adverse events in SCID-X1 patients [2]. Therefore, researchers designed a self-inactivating γ-RV vector (SIN-γRVV), which has been proven to be safe in clinical trials [3]. In adenosine deaminase deficiency severe combined immunodeficiency (ADA-SCID), the occurrence of insertional tumors after γ-RVV treatment is extremely rare [4]. The retroviral vectors underlying the cell products currently on the market are all self-losing. Live carrier, high safety. Lentiviral vectors (LVVs) preferentially integrate into active transcription units, and there are also reports of adverse events and random insertion leading to tumorigenesis. In June 2021, Bluebird Biotech suspended 2 clinical trials of LentiGlobin (BB305 LV) for the treatment of sickle cell disease because 2 patients receiving LentiGlobin were diagnosed with acute myeloid leukemia and myelodysplasia respectively. syndrome[5]. Therefore, both lentiviral vectors and retroviral vectors have the risk of causing cancer due to random insertion, but the probability is extremely low (nearly one in 10 million). As of now, among the cell products that have been marketed, whether they use retroviral vectors Whether it is a lentiviral vector or a lentiviral vector, there are no safety incidents caused by random integration.
(2)Risk of replicating viruses
Retroviral vectors or lentiviral vectors are also commonly used vectors to efficiently introduce CAR genes into T cells in CAR-T cell therapy, but they may produce replication-competent retroviruses (RCR) or replication-competent lentiviruses. (Replication Competent Lentivirus, RCL) potential risk of contamination. RCR/RCL can also be integrated into the cell genome, thereby creating the risk of insertional tumors due to integration that activates proto-oncogenes, destroys tumor suppressor genes, or increases the expression of factors that promote cell growth; on the other hand, because it can The production of replication-competent viruses also increases the risk of integration and insertional tumors [6]. In a trial of in vitro infusion of bone marrow progenitor cells, some researchers found that 3 of 10 severely immunodeficient rhesus monkeys treated with γ-RV developed lymphoma [7]. Analysis of the reasons found that RCR was detected in lymphoma tissues, and animals with lymphoma developed gamma-retroviremia, which was also caused by RCR [8]. Therefore, the real culprit of lymphoma in animals is RCR rather than RCR. Gamma-retroviral vectors. Similarly, since the envelope of lentiviral vectors is usually replaced with membrane proteins from other viruses, such as the commonly used VSV-G membrane protein, this may increase the potential risk caused by RCL contamination [6]. Therefore, the contamination risk of RCR/RCL is always a safety issue that we should focus on.
3、DNA virus vector
DNA virus vectors mainly include adenovirus (AV) and adeno-associated virus (AVV). Adenovirus is a non-enveloped double-stranded DNA virus. The gene of the adenovirus vector will not be integrated into the genome after introduction, so there is no risk of random insertion. High immunogenicity, cytotoxicity, and transient transgene expression still make adenoviral vector research challenging. In 1999, an 18-year-old patient with an ornithine transcarbamylase deficiency died after systemic injection of a high-dose adenoviral vector due to an innate immune response to the capsid protein that triggered cytokine release syndrome. syndrome, CRS, also known as cytokine storm). Currently, gene therapy based on adenovirus vectors is mainly used in gene vaccines and anti-tumor treatments [5]. AVV is a non-enveloped parvovirus with single-stranded DNA. Its vector has the characteristics of broad tropism, low immunogenicity and easy production, which is beneficial to clinical application. However, research results show that AAV-delivered RNA interference (RNAi) therapy can cause neurotoxicity in the brain of non-human primates [9] .
4、Summary
In summary, the application of viral vectors in cell therapy is very safe. Viral vectors are still a reliable, safe and efficient delivery method in the development of cell drugs. However, high attention should be paid to their viruses in related gene therapy and stem cell-related treatments. Carrier safety issues. This requires us to continuously optimize the design and production of viral vector processes and control the quality control system to ensure the clinical safety and effectiveness of cell and gene therapy products, so that cell and gene therapy based on viral vectors can benefit more with minimal safety issues. Many patients.
Reference：
[1] Lundstrom K. Viral Vectors in Gene Therapy: Where Do We Stand in 2023? Viruses. 2023 Mar 7; 15(3):698. doi: 10.3390/v15030698. PMID: 36992407; PMCID: PMC10059137.
[2]Hacein-Bey-Abina S, Garrigue A, Wang GP, Soulier J, Lim A, Morillon E, Clappier E, Caccavelli L, Delabesse E, Beldjord K, Asnafi V, MacIntyre E, Dal Cortivo L, Radford I, Brousse N, Sigaux F, Moshous D, Hauer J, Borkhardt A, Belohradsky BH, Wintergerst U, Velez MC, Leiva L, Sorensen R, Wulffraat N, Blanche S, Bushman FD, Fischer A, Cavazzana-Calvo M. Insertional oncogenesis in 4 patients after retrovirus-mediated gene therapy of SCID-X1. J Clin Invest. 2008 Sep; 118(9):3132-42. doi: 10.1172/JCI35700. PMID: 18688285; PMCID: PMC2496963.
[3]Hacein-Bey-Abina S, Pai SY, Gaspar HB, Armant M, Berry CC, Blanche S, Bleesing J, Blondeau J, de Boer H, Buckland KF, Caccavelli L, Cros G, De Oliveira S, Fernandez KS, Guo D, Harris CE, Hopkins G, Lehmann LE, Lim A, London WB, van der Loo JC, Malani N, Male F, Malik P, Marinovic MA, McNicol AM, Moshous D, Neven B, Oleastro M, Picard C, Ritz J, Rivat C, Schambach A, Shaw KL, Sherman EA, Silberstein LE, Six E, Touzot F, Tsytsykova A, Xu-Bayford J, Baum C, Bushman FD, Fischer A, Kohn DB, Filipovich AH, Notarangelo LD, Cavazzana M, Williams DA, Thrasher AJ. A modified gamma -retrovirus vector for X-linked severe combined immunodeficiency. N Engl J Med. 2014 Oct 9; 371(15):1407-17. doi: 10.1056/NEJMoa1404588. PMID: 25295500; PMCID: PMC4274995.
[4] Pai SY. Built to last: gene therapy for ADA SCID. Blood. 2021 Oct 14; 138 (15) : 1287-1288. The doi: 10.1182 / blood. 2021012300. PMID: 34647983; PMCID: PMC8525332.
[5] Li Manqi, Wei Liping, Tao Qiaoyu et al. Gene therapy carrier, research progress and safety applications [J]. China's pharmaceutical industry magazines, does 2022 (12) : 1671-1682. The DOI: 10.16522 / j.carol carroll nki CJPH. 2022.12.001.
[6] Wu Xueling, Zhao Xiang, Meng Shufang. Risk analysis and control of replicative viruses in CAR T cell therapy products [J]. China Pharmaceutical Journal,2018,32(07):879-885.DOI:10.16153/ J.1002-7777.2018.07.006.
[7]Donahue RE, Kessler SW, Bodine D, McDonagh K, Dunbar C, Goodman S, Agricola B, Byrne E, Raffeld M, Moen R, et al. Helper virus induced T cell lymphoma in nonhuman primates after retroviral mediated gene transfer. J Exp Med. 1992 Oct 1; 176(4):1125-35. doi: 10.1084/ jem.176.4.1125.PMID: 1383375; PMCID: PMC2119385.
[8]Vanin EF, Kaloss M, Broscius C, Nienhuis AW. Characterization of replication-competent retroviruses from nonhuman primates with virus-induced T-cell lymphomas and observations regarding the mechanism of oncogenesis. J Virol. 1994 Jul; 68(7):4241-50. doi: 10.1128/JVI.68.7.4241-4250.1994. PMID: 8207799; PMCID: PMC236347.
[9]Keiser MS, Ranum PT, Yrigollen CM, Carrell EM, Smith GR, Muehlmatt AL, Chen YH, Stein JM, Wolf RL, Radaelli E, Lucas TJ 2nd, Gonzalez-Alegre P, Davidson BL. Toxicity after AAV delivery of RNAi expression constructs into nonhuman primate brain. Nat Med. 2021 Nov; 27(11):1982-1989. doi: 10.1038/s41591-021-01522-3. Epub 2021 Oct 18.PMID: 34663988; PMCID: PMC8605996.
Disclaimer: Shenzhen Cell Valley is committed to the research of cell and gene therapy, in order to promote emerging technologies and let more people understand the new developments in biomedicine.The content of this article is for information exchange only. This platform remains neutral with respect to the content, statements, and opinion judgments in the article, and does not represent the position and opinions of Shenzhen Cell Valley.The relevant information in this article should not be used for diagnosis or treatment, and cannot replace professional medical advice. Our website will not assume any responsibility.The final interpretation of the above statement belongs to our company’s website. This statement will apply to articles shared on our website at all times. Thank you for your cooperation! Copyright statement: The copyright of the article belongs to Shenzhen Cell Valley. Individuals are welcome to forward it to friends, media or Any unauthorized reproduction by the organization to other platforms will be regarded as infringement.If you need to reprint, please contact email: contact@sz-cell.com
