AIM： To evaluate whether intratumoral expression of measles virus fusogenic membrane glycoproteins H and F （MV-FMG）, encoded by an adenovirus vector Ad.MV-HI F, alone or in combination with local coexpression of cy...AIM： To evaluate whether intratumoral expression of measles virus fusogenic membrane glycoproteins H and F （MV-FMG）, encoded by an adenovirus vector Ad.MV-HI F, alone or in combination with local coexpression of cytokines （IL-2, IL-12, IL-18, IL-21 or GM-CSF）, can serve as a platform for inducing tumor-specific immune responses in colon cancer.METHODS： We used confocal laser scanning microscopy and flow cytometry to analyze cell-cell fusion after expression of MV-FMG by dye colocalization. In a syngeneic bilateral subcutaneous MC38 and Colon26 colon cancer model in C57BL/6 and BALB/c mice, we assessed the effect on both the directly vector-treated tumor as well as the contralateral, not directly vector- treated tumor. We assessed the induction of a tumorspecific cytotoxic T lymphocyte （CTL） response with a lactate dehydrogenase （LDH） release assay.RESULTS： We demonstrated in vitro that transduction of MC38 and Colon26 cells with Ad.MV-H/F resulted in dye colocalization, indicative of cell-cell fusion, in addition, in the syngeneic bilateral tumor model we demonstrated a significant regression of the directly vector-inoculated tumor upon intratumoral expression of MV-FMG alone or in combination with the tested cytokines. We observed the highest anti-neoplastic efficacy with MV-FMG and lL-21 coexpression. The degree of tumor regression of the not directly vector-treated tumor correlated with the anti-neoplastic response of the directly vector-treated tumor. This regression was mediated by a tumor-specific CTL response.CONCLUSION： Our data indicate that intratumoral expression of measles virus fusogenic membrane glycoproteins is a promising tool both for direct tumor treatment as well as for tumor vaccination approaches that can be further enhanced by cytokine coexpression.展开更多
Background:Chimeric antigen receptor(CAR)-based immune cell therapies attack neighboring cancer cells after receptor recognition but are unable to directly affect distant tumor cells.This limitation may contribute to ...Background:Chimeric antigen receptor(CAR)-based immune cell therapies attack neighboring cancer cells after receptor recognition but are unable to directly affect distant tumor cells.This limitation may contribute to their inefficiency in treating solid tumors,given the restricted intratumoral infiltration and immunosuppressive tumor microenvironment.Therefore,cell–cell fusion as a cell-killing mechanism might develop a novel cytotherapy aimed at improving the efficacy against solid tumors.Methods:We constructed a fusogenic protein,fusion-associated small transmembrane(FAST)p14 of reptilian reovirus,into cancer cells and mesenchymal stem cells(MSCs),which cocultured with various colon cancer cells and melenoma cells to validate its ability to induce cell fusion and syncytia formation.RNA sequencing,quantitative reverse transcription polymerase chain reaction,and Western blot were performed to elucidate the mechanism of syncytia death.Cell viability assay was employed to assess the killing effects of MSCs carrying the p14 protein(MSCs-p14),which was also identified in the subcutaneous tumor models.Subsequently,the Tet-On system was introduced to enhance the controllability and safety of therapy.Results:Cancer cells incorporated with fusogenic protein p14 FAST from reovirus fused together to form syncytia and subsequently died through apoptosis and pyroptosis.MSCs-p14 cocultured with different cancer cells and effienctly induced cancer cell fusion and caused widespread cancer cell death in vitro.In mouse tumor models,mMSCs-p14 treatment markedly suppressed tumor growth and also enhanced the activity of natural killer cells and macrophages.Controllability and safety of MSCs-p14 therapy were further improved by introducing the tetracycline-controlled transcriptional system.Conclusion:MSC-based cytotherapy carrying viral fusogenic protein in this study kills cancer cells by inducing cell–cell fusion.It has demonstrated definite efficacy in treating solid tumors and is worth considering for clinical development.展开更多
Gene therapy is known highly effective for treatment of many diseases;however,its wide use has been severely bottlenecked by lack of safe and effective delivery vectors[1].Cationic polymers are safe nonviral gene vect...Gene therapy is known highly effective for treatment of many diseases;however,its wide use has been severely bottlenecked by lack of safe and effective delivery vectors[1].Cationic polymers are safe nonviral gene vectors with great potential for large-scale applications[2],and widely used to condense the large macromolecules into cationic polymer/DNA complexes(polyplexes)nanoparticles,protect-展开更多
The integrity of the neuronal membrane is crucial for its function and cellular survival; thus, ineffective repair of damaged membranes may be one of the key elements underlying the neuronal degeneration and overall f...The integrity of the neuronal membrane is crucial for its function and cellular survival; thus, ineffective repair of damaged membranes may be one of the key elements underlying the neuronal degeneration and overall functional loss that occurs after spinal cord injury （SCI）. It has been shown that polyethylene glycol （PEG） can reseal axonal membranes following various injuries in multiple in vitro and in vivo injury models. In addition, PEG may also directly prevent the effects of mitochondria-derived oxidative stress on intracellular components. Thus, PEG repairs mechanically injured cells by at least two distinct pathways： resealing of the disrupted plasma membrane and direct protection of mitochondria. Besides repairing primary membrane damage, PEG treatment also results in significant attenuation of oxidative stress, likely due to its capacity to reseal the membrane, thereby breaking the cycle of cellular damage and free-radical production. Based on this, in addition to the practicality of its application, we expect that PEG may be established as an effective treatment for SCI where membrane disruption and mitochondriai damage are implicated.展开更多
Although tumor cell membranes with broad-spectrum antigens have been explored for cancer vaccines for decades,their relatively poor capacity to stimulate immune responses,especially cellular immune responses,has limit...Although tumor cell membranes with broad-spectrum antigens have been explored for cancer vaccines for decades,their relatively poor capacity to stimulate immune responses,especially cellular immune responses,has limited their application.Here,we presented a novel bacterial and cancerous cell membrane fusogenic liposome for co-delivering cell membrane-derived antigens and adjuvants.Meanwhile,a programmed death-ligand 1(PD-L1)inhibitor,JQ-1,was incorporated into the formulation to tackle the up-regulated PD-L1 expression of antigen-presenting cells(APCs)upon vaccination,thereby augmenting its anti-tumor efficacy.The fusogenic liposomes demonstrated significantly improved cellular uptake by APCs and effectively suppressed PD-L1 expression in bone marrow-derived dendritic cells(BMDCs)in vitro.Following subcutaneous vaccination,the nano-vaccines efficiently drained to the tumor-draining lymph nodes(TDLNs),and significantly inhibited PD-L1 expression of both dendritic cells(DCs)and macrophages within the TDLNs and tumors.As a result,the liposomal vaccine induced robust innate and cellular immune responses and inhibited tumor growth in a colorectal carcinoma-burden mouse model.In summary,the fabricated cell membrane-based fusogenic liposomes offer a safe,effective,and easily applicable strategy for tumor immunotherapy and hold potential for personalized cancer immunotherapy.展开更多
Engineered T cells expressing chimeric antigen receptor(CAR)exhibit high response rates in B-cell malignancy treatments and possess therapeutic potentials against various diseases.However,the complicated ex vivo produ...Engineered T cells expressing chimeric antigen receptor(CAR)exhibit high response rates in B-cell malignancy treatments and possess therapeutic potentials against various diseases.However,the complicated ex vivo production process of CAR-T cells limits their application.Herein,we use virus-mimetic fusogenic nanovesicles(FuNVs)to produce CAR-T cells in vivo via membrane fusion-mediated CAR protein delivery.Briefly,the FuNVs are modified using T-cell fusogen,adapted from measles virus or reovirus fusogens via displaying anti-CD3 single-chain variable fragment.The FuNVs can efficiently fuse with the T-cell membrane in vivo,thereby delivering the loaded anti-CD19(aCD19)CAR protein onto T-cells to produce aCD19 CAR-T cells.These aCD19 CAR-T cells alone or in combination with anti-OX40 antibodies can treat B-cell lymphoma without inducing cytokine release syndrome.Thus,our strategy provides a novel method for engineering T cells into CAR-T cells in vivo and can further be employed to deliver other therapeutic membrane proteins.展开更多
基金grants from Deutsche Forschungsgemeinschaft, Wilhelm Sander-Stiftung, and Forschungsfrderung Ruhr-Universitt Bochum Medizinische Fakultt to OW
文摘AIM： To evaluate whether intratumoral expression of measles virus fusogenic membrane glycoproteins H and F （MV-FMG）, encoded by an adenovirus vector Ad.MV-HI F, alone or in combination with local coexpression of cytokines （IL-2, IL-12, IL-18, IL-21 or GM-CSF）, can serve as a platform for inducing tumor-specific immune responses in colon cancer.METHODS： We used confocal laser scanning microscopy and flow cytometry to analyze cell-cell fusion after expression of MV-FMG by dye colocalization. In a syngeneic bilateral subcutaneous MC38 and Colon26 colon cancer model in C57BL/6 and BALB/c mice, we assessed the effect on both the directly vector-treated tumor as well as the contralateral, not directly vector- treated tumor. We assessed the induction of a tumorspecific cytotoxic T lymphocyte （CTL） response with a lactate dehydrogenase （LDH） release assay.RESULTS： We demonstrated in vitro that transduction of MC38 and Colon26 cells with Ad.MV-H/F resulted in dye colocalization, indicative of cell-cell fusion, in addition, in the syngeneic bilateral tumor model we demonstrated a significant regression of the directly vector-inoculated tumor upon intratumoral expression of MV-FMG alone or in combination with the tested cytokines. We observed the highest anti-neoplastic efficacy with MV-FMG and lL-21 coexpression. The degree of tumor regression of the not directly vector-treated tumor correlated with the anti-neoplastic response of the directly vector-treated tumor. This regression was mediated by a tumor-specific CTL response.CONCLUSION： Our data indicate that intratumoral expression of measles virus fusogenic membrane glycoproteins is a promising tool both for direct tumor treatment as well as for tumor vaccination approaches that can be further enhanced by cytokine coexpression.
基金supported by grants 82272813,82373027,and 82303488 from the National Natural Science Foundation of Chinagrant BK20220668 from the Natural Science Foundation of Jiangsu Province,China+1 种基金grant CXZX202234 from Jiangsu Province Capability Improvement Project through Science,Technology and Educationgrant KC22233 from the Key Development Project of Science and Technology Bureau in Xuzhou.
文摘Background:Chimeric antigen receptor(CAR)-based immune cell therapies attack neighboring cancer cells after receptor recognition but are unable to directly affect distant tumor cells.This limitation may contribute to their inefficiency in treating solid tumors,given the restricted intratumoral infiltration and immunosuppressive tumor microenvironment.Therefore,cell–cell fusion as a cell-killing mechanism might develop a novel cytotherapy aimed at improving the efficacy against solid tumors.Methods:We constructed a fusogenic protein,fusion-associated small transmembrane(FAST)p14 of reptilian reovirus,into cancer cells and mesenchymal stem cells(MSCs),which cocultured with various colon cancer cells and melenoma cells to validate its ability to induce cell fusion and syncytia formation.RNA sequencing,quantitative reverse transcription polymerase chain reaction,and Western blot were performed to elucidate the mechanism of syncytia death.Cell viability assay was employed to assess the killing effects of MSCs carrying the p14 protein(MSCs-p14),which was also identified in the subcutaneous tumor models.Subsequently,the Tet-On system was introduced to enhance the controllability and safety of therapy.Results:Cancer cells incorporated with fusogenic protein p14 FAST from reovirus fused together to form syncytia and subsequently died through apoptosis and pyroptosis.MSCs-p14 cocultured with different cancer cells and effienctly induced cancer cell fusion and caused widespread cancer cell death in vitro.In mouse tumor models,mMSCs-p14 treatment markedly suppressed tumor growth and also enhanced the activity of natural killer cells and macrophages.Controllability and safety of MSCs-p14 therapy were further improved by introducing the tetracycline-controlled transcriptional system.Conclusion:MSC-based cytotherapy carrying viral fusogenic protein in this study kills cancer cells by inducing cell–cell fusion.It has demonstrated definite efficacy in treating solid tumors and is worth considering for clinical development.
文摘Gene therapy is known highly effective for treatment of many diseases;however,its wide use has been severely bottlenecked by lack of safe and effective delivery vectors[1].Cationic polymers are safe nonviral gene vectors with great potential for large-scale applications[2],and widely used to condense the large macromolecules into cationic polymer/DNA complexes(polyplexes)nanoparticles,protect-
文摘The integrity of the neuronal membrane is crucial for its function and cellular survival; thus, ineffective repair of damaged membranes may be one of the key elements underlying the neuronal degeneration and overall functional loss that occurs after spinal cord injury （SCI）. It has been shown that polyethylene glycol （PEG） can reseal axonal membranes following various injuries in multiple in vitro and in vivo injury models. In addition, PEG may also directly prevent the effects of mitochondria-derived oxidative stress on intracellular components. Thus, PEG repairs mechanically injured cells by at least two distinct pathways： resealing of the disrupted plasma membrane and direct protection of mitochondria. Besides repairing primary membrane damage, PEG treatment also results in significant attenuation of oxidative stress, likely due to its capacity to reseal the membrane, thereby breaking the cycle of cellular damage and free-radical production. Based on this, in addition to the practicality of its application, we expect that PEG may be established as an effective treatment for SCI where membrane disruption and mitochondriai damage are implicated.
基金supported by the National Natural Science Foundation of China(No.82341038)Natural Science Foundation of Sichuan Province(No.2022NSFSC1491)+2 种基金China Postdoctoral Science Foundation Grant(No.2019M663534,China)Sichuan Veterinary Medicine and Drug Innovation Group of China Agricultural Research System(CARS-SVIDIP)the Fundamental Research Funds for the Central Universities and Sichuan University Postdoctoral Interdisciplinary Innovation Fund.
文摘Although tumor cell membranes with broad-spectrum antigens have been explored for cancer vaccines for decades,their relatively poor capacity to stimulate immune responses,especially cellular immune responses,has limited their application.Here,we presented a novel bacterial and cancerous cell membrane fusogenic liposome for co-delivering cell membrane-derived antigens and adjuvants.Meanwhile,a programmed death-ligand 1(PD-L1)inhibitor,JQ-1,was incorporated into the formulation to tackle the up-regulated PD-L1 expression of antigen-presenting cells(APCs)upon vaccination,thereby augmenting its anti-tumor efficacy.The fusogenic liposomes demonstrated significantly improved cellular uptake by APCs and effectively suppressed PD-L1 expression in bone marrow-derived dendritic cells(BMDCs)in vitro.Following subcutaneous vaccination,the nano-vaccines efficiently drained to the tumor-draining lymph nodes(TDLNs),and significantly inhibited PD-L1 expression of both dendritic cells(DCs)and macrophages within the TDLNs and tumors.As a result,the liposomal vaccine induced robust innate and cellular immune responses and inhibited tumor growth in a colorectal carcinoma-burden mouse model.In summary,the fabricated cell membrane-based fusogenic liposomes offer a safe,effective,and easily applicable strategy for tumor immunotherapy and hold potential for personalized cancer immunotherapy.
基金This work was supported by the National Key R&D Program of China(2022YFB3808100)the National Natural Science Foundation of China(32271442,52130301,and 82072048)+2 种基金the Guangdong Basic and Applied Basic Research Foundation(2022B1515020025)the Science and Technology Program of Guangzhou,China(202103030004)the Fundamental Research Funds for the Central Universities(2022ZYGXZR102).
文摘Engineered T cells expressing chimeric antigen receptor(CAR)exhibit high response rates in B-cell malignancy treatments and possess therapeutic potentials against various diseases.However,the complicated ex vivo production process of CAR-T cells limits their application.Herein,we use virus-mimetic fusogenic nanovesicles(FuNVs)to produce CAR-T cells in vivo via membrane fusion-mediated CAR protein delivery.Briefly,the FuNVs are modified using T-cell fusogen,adapted from measles virus or reovirus fusogens via displaying anti-CD3 single-chain variable fragment.The FuNVs can efficiently fuse with the T-cell membrane in vivo,thereby delivering the loaded anti-CD19(aCD19)CAR protein onto T-cells to produce aCD19 CAR-T cells.These aCD19 CAR-T cells alone or in combination with anti-OX40 antibodies can treat B-cell lymphoma without inducing cytokine release syndrome.Thus,our strategy provides a novel method for engineering T cells into CAR-T cells in vivo and can further be employed to deliver other therapeutic membrane proteins.
