OBJECTIVE: To identify a gene engineering antibody against cystic echinococcosis in liver. METHODS: A single chain of variable fragment of human antibodies (ScFvs) was selected from the library by using affinity selec...OBJECTIVE: To identify a gene engineering antibody against cystic echinococcosis in liver. METHODS: A single chain of variable fragment of human antibodies (ScFvs) was selected from the library by using affinity selection technique with the recombinant antigen on solid surface. The positive clones were demonstrated by ELISA and their DNA sequences were also determined. RESULTS: The DNA sequence data showed that the antibody gene is composed of 768bp. In addition, a specific combination capacity with recombinant Echinococcus granulosus antigen B (r-EgB) was demonstrated by ELISA. CONCLUSION: The obtained gene engineering antibody against r-EgB may have potential implications in immunological treatment and drug targeting delivery.展开更多
With gene engineering EB virus membrane antigen as the diagnostic antigen, indirect immunofluo-rescence (IF) assay was used to detect IgA antibody against EB virus membrane antigen (MA-IgA) in sera from 202 nasopharyn...With gene engineering EB virus membrane antigen as the diagnostic antigen, indirect immunofluo-rescence (IF) assay was used to detect IgA antibody against EB virus membrane antigen (MA-IgA) in sera from 202 nasopharyngeal carcinoma (NPC) patients and 315 controls (normal and patients with other tumors). MA-IgA antibody was positive in 96.8% of the pretreatment NPC patients with a GMT of 1:36.3. MA-IgA detection by this method was more sensitive than EA-IgA detection by IE. In contrast, patients with tumors other than NPC were negative for MA-IgA antibody. 9.1% of VCA-IgA positive persons were MA-IgA positive with a GMT of less than 1:5. No MA-IgA positive was found in VCA-IgA negatives. The results indicated that this method was relatively specific. In the treatment group, the positive rate and GMT of MA-IgA antibody declined with increase in survival time and the decline was faster than VCA-IgA. When recurrence or distant metastasis developed, similar to VCA-IgA and EA-IgA antibodies, the positive rate and GMT of MA-IgA antibody increased to its pretreatment level. Therefore, MA-IgA detection might be valuable in the early diagnosis and monitor of NPC.展开更多
Monoclonal antibodies(mAbs)have become a major class of therapeutic agents providing effective alternatives to treating various human diseases.To date,15 mAbs have been approved by regulatory agencies in the world for...Monoclonal antibodies(mAbs)have become a major class of therapeutic agents providing effective alternatives to treating various human diseases.To date,15 mAbs have been approved by regulatory agencies in the world for clinical use in oncology indications.The selectivity and specificity,the unique pharmacokinetics,and the ability to engage and activate the host immune system differentiate these biologics from traditional small molecule anticancer drugs.mAb-based regimens have brought clinical benefits,including improvements in overall survival,to patients with a variety of cancers.Many challenges still remain,however,to fully realize the potential of these new medicines.With our further understanding of cancer biology,mechanism of antibody action,and advancement of antibody engineering technologies,many novel antibody formats or antibody-derived molecules are emerging as promising new generation therapeutics.Carefully designed and engineered,they retain the advantage of specificity and selectivity of original antibodies,but in the meantime acquire additional special features such as improved pharmacokinetics,increased selectivity,and enhanced anticancer efficacy.Promising clinical results are being generated with these newly improved antibody-based therapeutics.展开更多
High-affinity antibodies are widely used in diagnostics and for the treatment of human diseases.However,most antibodies are isolated from semi-synthetic libraries by phage display and do not possess in vivo affinity m...High-affinity antibodies are widely used in diagnostics and for the treatment of human diseases.However,most antibodies are isolated from semi-synthetic libraries by phage display and do not possess in vivo affinity maturation,which is triggered by antigen immunization.It is therefore necessary to engineer the affinity of these antibodies by way of in vitro assaying.In this study,we optimized the affinity of two human monoclonal antibodies which were isolated by phage display in a previous related study.For the 42A1 antibody,which targets the liver cancer antigen glypican-3,the variant T57H in the second complementarity-determining region of the heavy chain(CDR-H2)exhibited a 2.6-fold improvement in affinity,as well as enhanced cell-binding activity.For the I4A3 antibody to severe acute respiratory syndrome coronavirus 2,beneficial single mutations in CDR-H2 and CDR-H3 were randomly combined to select the best synergistic mutations.Among these,the mutation S53P-S98T improved binding affinity(about 3.7 fold)and the neutralizing activity(about 12 fold)compared to the parent antibody.Taken together,single mutations of key residues in antibody CDRs were enough to increase binding affinity with improved antibody functions.The mutagenic combination of key residues in different CDRs creates additive enhancements.Therefore,this study provides a safe and effective in vitro strategy for optimizing antibody affinity.展开更多
The efficacy and specificity of conventional monoclonal antibody(mAb)drugs in the clinic require further improvement.Currently,the development and application of novel antibody formats for improving cancer immunothera...The efficacy and specificity of conventional monoclonal antibody(mAb)drugs in the clinic require further improvement.Currently,the development and application of novel antibody formats for improving cancer immunotherapy have attracted much attention.Variable region-retaining antibody fragments,such as antigen-binding fragment(Fab),single-chain variable fragment(scFv),bispecific antibody,and bi/trispecific cell engagers,are engineered with humanization,multivalent antibody construction,affinity optimization and antibody masking for targeting tumor cells and killer cells to improve antibody-based therapy potency,efficacy and specificity.In this review,we summarize the application of antibody variable region engineering and discuss the future direction of antibody engineering for improving cancer therapies.展开更多
Objective To study the passive immunization with human monoclonal antibodies as for prophylaxis of human cytomegalovirus （HCMV） infection. Methods Fab monoclonal antibodies to HCMV were recovered by repertoire cloni...Objective To study the passive immunization with human monoclonal antibodies as for prophylaxis of human cytomegalovirus （HCMV） infection. Methods Fab monoclonal antibodies to HCMV were recovered by repertoire cloning of mRNA from a HCMV infected individual. Antigen binding specificity, CDR sequence of VH and VL and neutralizing activity on HCMV AD169 stain were analyzed in vitro. The light and heavy chain Fd fragment genes of Fab antibodies were further cloned into a recombinant baculovirus expression vector pAC-K-Fc to express intact IgG. Secreted products were purified with affinity chromatography using protein G. Results SDS-PAGE and Western blot confirmed the expression of the intact IgG. Immuno-blotting and -precipitation were used to identify HCMV proteins. One Fab monoclonal antibody recognized a conformational HCMV protein. Conclusion IgG antibodies can neutralize the HCMV AD169 strain efficiently at a titer of 2.5 μg/mL and may prove valuable for passive immunoprophylaxis against HCMV infection in humans.展开更多
Monoclonal antibodies(mAbs)are widely utilized as therapeutic drugs for various diseases,such as cancer,autoimmune diseases,and infectious diseases.Using the avian-derived B cell line DT40,we previously developed an a...Monoclonal antibodies(mAbs)are widely utilized as therapeutic drugs for various diseases,such as cancer,autoimmune diseases,and infectious diseases.Using the avian-derived B cell line DT40,we previously developed an antibody display technology,namely,the ADLib system,which rapidly generates antigen-specific mAbs.Here,we report the development of a human version of the ADLib system and showcase the streamlined generation and optimization of functional human mAbs.Tailored libraries were first constructed by replacing endogenous immunoglobulin genes with designed human counterparts.From these libraries,clones producing full-length human IgGs against distinct antigens can be isolated,as exemplified by the selection of antagonistic mAbs.Taking advantage of avian biology,effective affinity maturation was achieved in a straightforward manner by seamless diversification of the parental clones into secondary libraries followed by single-cell sorting,quickly affording mAbs with improved affinities and functionalities.Collectively,we demonstrate that the human ADLib system could serve as an integrative platform with unique diversity for rapid de novo generation and optimization of therapeutic or diagnostic antibody leads.Furthermore,our results suggest that libraries can be constructed by introducing exogenous genes into DT40 cells,indicating that the ADLib system has the potential to be applied for the rapid and effective directed evolution and optimization of proteins in various fields beyond biomedicine.展开更多
In this dispensation of the fourth industrial revolution,protein engineering has become a popular approach for increasing enzymatic activity,stability,and titer in the biosynthesis of natural products.This is attribut...In this dispensation of the fourth industrial revolution,protein engineering has become a popular approach for increasing enzymatic activity,stability,and titer in the biosynthesis of natural products.This is attributed to its numerous advantages(over direct isolation from plants or via chemical synthesis),including decreasing or eliminating reaction byproducts,high precision,moderate handling of intricate and chemically unstable chemicals,overall reusability,and cost efficiency.Recently,protein engineering tools have advanced to redesign and enhance natural product biosynthesis.These methods include direct evolution,substrate engineering,medium engineering,enzyme engineering and immobilization,structure-assisted protein engineering,and advanced computational.Recent successes in implementing these emerging protein engineering technologies were critically discussed in this article.Also,the advantages,limitations,and applications in industrial and medical biotechnology were discussed.Last,future research directions and potential were also highlighted.展开更多
Ion channels play instrumental roles in regulating membrane potential and cross-membrane signal transduction,thus making them attractive targets for understanding various physiological processes and associated disease...Ion channels play instrumental roles in regulating membrane potential and cross-membrane signal transduction,thus making them attractive targets for understanding various physiological processes and associated diseases.Gaining a deeper understanding of their structural and functional properties has significant implications for developing therapeutic interventions.In recent years,nanobodies,single-domain antibody fragments derived from camelids,have emerged as powerful tools in ion channel and synthetic biology research.Their small size,high specificity,and ability to recognize difficult-to-reach epitopes offer advantages over conventional antibodies and biologics.Furthermore,their resemblance to the variable region of human IgG family III reduces immunogenicity concerns.Nanobodies have introduced new opportunities for exploring ion channel structure-function relationships and offer a promising alternative to conventional drugs,which often face challenges such as off-target effects and toxicity.This review highlights recent progress in applying nanobodies to interrogate and modulate ion channel activity,with an emphasis on their potential to overcome current technical and therapeutic limitations.展开更多
Advancements in protein engineering have driven the continuous optimization of T-cell engagers(TCEs),resulting in remarkable clinical outcomes in the treatment of B-cell malignancies.Moreover,developing tri-or multisp...Advancements in protein engineering have driven the continuous optimization of T-cell engagers(TCEs),resulting in remarkable clinical outcomes in the treatment of B-cell malignancies.Moreover,developing tri-or multispecific TCEs has emerged as a promising strategy to address the challenges of tumor heterogeneity and antigen escape.However,considerable obstacles remain,primarily in format design.In this study,we engineered BAFF-based TCEs with various formats that incorporate anti-CD3 Fab or IgG domains fused with BAFF ligands to target BAFF recep tors(BAFFR,BCMA,and TACI).These constructs varied in valency and the presence or absence of long-acting ele ments such as Fc domains or the albumin binding domain consensus sequence(ABDCon).Although the inclusion of an Fc domain did not enhance sustained tumor eradication,variations in valency and spatial configuration profoundly influenced cytotoxicity.We identified TriBAFF/CD3/ABDCon as the optimal trifunctional construct,fea turing an anti-CD3 Fab backbone with BAFF and ABDCon fused to the C-termini of the heavy and light chains.This design facilitates optimal immune synapse formation between the target cells and T cells and effectively controls tumor burdens in various B-cell malignancy models with good tolerability.Notably,TriBAFF/CD3/ABDCon outper formed conventional therapies,including blinatumomab and BAFF-based CAR-T cells,in models of heterogeneous leukemia and aggressive lymphoma.These findings underscore the potential of using natural ligands as antibody-targeting modules and provide valuable insights into the design of the next generation of multispecific TCEs,which hold promise for improving treatment outcomes in a wide range of malignancies and beyond.展开更多
As of May 1, 2017, 74 antibody-based molecules have been approved by a regulatory authority in a major market. Additionally, there are 70 and 575 antibody- based molecules in phase III and phase 1/11 clinical trials, ...As of May 1, 2017, 74 antibody-based molecules have been approved by a regulatory authority in a major market. Additionally, there are 70 and 575 antibody- based molecules in phase III and phase 1/11 clinical trials, respectively. These total 719 antibody-based clinical stage molecules include 493 naked IgGs, 87 antibody- drug conjugates, 61 bispecific antibodies, 37 total Fc fusion proteins, 17 radioimmunoglobulins, 13 antibody fragments, and 11 immunocytokines. New uses for these antibodies are being discovered each year. For oncol- ogy, many of the exciting new approaches involve anti- body modulation of T-cells. There are over 80 antibodies in clinical trials targeting T cell checkpoints, 26 T-cell- redirected bispecific antibodies, and 145 chimeric anti- gen receptor （CAR） cell-based candidates （all currently in phase I or II clinical trials）, totaling more than 250 T cell interacting clinical stage antibody-based candi- dates. Finally, significant progress has been made recently on routes of delivery, including delivery of proteins across the blood-brain barrier, oral delivery to the gut, delivery to the cellular cytosol, and gene- and viral-based delivery of antibodies. Thus, there are cur- rently at least 864 antibody-based clinical stage mole- cules or cells, with incredible diversity in how they are constructed and what activities they impart. These are followed by a next wave of novel molecules, approa- ches, and new methods and routes of delivery, demon- strating that the field of antibody-based biologics is very innovative and diverse in its approaches to fulfill their promise to treat unmet medical needs.展开更多
As a result of recent breakthroughs in cancer immunotherapies, unprecedented and durable remission, and even cure, has been reported in some patients. Importantly, this progress has been achieved, not by the induction...As a result of recent breakthroughs in cancer immunotherapies, unprecedented and durable remission, and even cure, has been reported in some patients. Importantly, this progress has been achieved, not by the induction of immunity, but by the delivery of immunity in the form of engineered antibodies （cAbs） or effector T cells. However, these single-target technologies have failed to result in a therapeutic effect in some patients, and evidence suggests that further advances depend on an effective strategy for coping with cancer heterogeneity and dynamics. A synthetic immunity （SI） strategy is proposed to achieve this goal. The fundamental basis of SI involves the generation of a panel of cAbs and antibody-retargeted CTLs designed to destroy all cell lineages of a cancer with high specificity. This goal can be achieved only when the composition of the cAbs is determined using a systematic approach, i.e., selecting the antigens targeted by the cAbs based on an epitope-tree illustrating the clonal antigen architecture of the cancer. Integration of technologies that increase the epitope breadth, cAb affinity and T cell activity will further enhance the efficacy of SI. Using DNA vectors to express the eAbs will be a safe, effective and affordable solution.展开更多
In this study, we discussed the necessity of human IgG1 Cγ1 domain for recombinant antibody using computeraided homology modeling method and experimental studies. The heavy （VH） and light （VL） chain variable regi...In this study, we discussed the necessity of human IgG1 Cγ1 domain for recombinant antibody using computeraided homology modeling method and experimental studies. The heavy （VH） and light （VL） chain variable regions of 1-28, a murine IgM-type anti-CD20 mAb, were ligated by linker peptide （Gly4Ser）3 to form the single-chain Fv fragment （scFv）. Then, the engineered antibody （LH1-3） was generated by fusing scFv with the entire IgG1 heavy constant regions. The 3-D structure of LH1-3 was modeled using computer-aided homology modeling method and the binding activity of LH1-3 was evaluated theoretically. Compared to the 3-D structure of the Fv fragment of the parent antibody, the conformation of the active pocket of LH1-3 was remained because of the rigid support of Cγ1. Further experimental results of flow cytometry showed that the engineered anti-CD20 antibody possessed specifically binding activity to CD20-expressing target cells. The anti-CD20 antibody fragments could also mediate complement-dependent cytotoxicity （CDC） of human B-lymphoid cell lines. Our study highlights some interests and advantages of a methodology based on the homology modeling and analysis of molecular structural properties.展开更多
基金The project was supported by a grant from the National Natural Science Fundation of China (No. 39860078) and Xinjiang Natural Science Fundation China (No. 200221101).
文摘OBJECTIVE: To identify a gene engineering antibody against cystic echinococcosis in liver. METHODS: A single chain of variable fragment of human antibodies (ScFvs) was selected from the library by using affinity selection technique with the recombinant antigen on solid surface. The positive clones were demonstrated by ELISA and their DNA sequences were also determined. RESULTS: The DNA sequence data showed that the antibody gene is composed of 768bp. In addition, a specific combination capacity with recombinant Echinococcus granulosus antigen B (r-EgB) was demonstrated by ELISA. CONCLUSION: The obtained gene engineering antibody against r-EgB may have potential implications in immunological treatment and drug targeting delivery.
文摘With gene engineering EB virus membrane antigen as the diagnostic antigen, indirect immunofluo-rescence (IF) assay was used to detect IgA antibody against EB virus membrane antigen (MA-IgA) in sera from 202 nasopharyngeal carcinoma (NPC) patients and 315 controls (normal and patients with other tumors). MA-IgA antibody was positive in 96.8% of the pretreatment NPC patients with a GMT of 1:36.3. MA-IgA detection by this method was more sensitive than EA-IgA detection by IE. In contrast, patients with tumors other than NPC were negative for MA-IgA antibody. 9.1% of VCA-IgA positive persons were MA-IgA positive with a GMT of less than 1:5. No MA-IgA positive was found in VCA-IgA negatives. The results indicated that this method was relatively specific. In the treatment group, the positive rate and GMT of MA-IgA antibody declined with increase in survival time and the decline was faster than VCA-IgA. When recurrence or distant metastasis developed, similar to VCA-IgA and EA-IgA antibodies, the positive rate and GMT of MA-IgA antibody increased to its pretreatment level. Therefore, MA-IgA detection might be valuable in the early diagnosis and monitor of NPC.
文摘Monoclonal antibodies(mAbs)have become a major class of therapeutic agents providing effective alternatives to treating various human diseases.To date,15 mAbs have been approved by regulatory agencies in the world for clinical use in oncology indications.The selectivity and specificity,the unique pharmacokinetics,and the ability to engage and activate the host immune system differentiate these biologics from traditional small molecule anticancer drugs.mAb-based regimens have brought clinical benefits,including improvements in overall survival,to patients with a variety of cancers.Many challenges still remain,however,to fully realize the potential of these new medicines.With our further understanding of cancer biology,mechanism of antibody action,and advancement of antibody engineering technologies,many novel antibody formats or antibody-derived molecules are emerging as promising new generation therapeutics.Carefully designed and engineered,they retain the advantage of specificity and selectivity of original antibodies,but in the meantime acquire additional special features such as improved pharmacokinetics,increased selectivity,and enhanced anticancer efficacy.Promising clinical results are being generated with these newly improved antibody-based therapeutics.
基金supported by the National Natural Science Foundation of China(Grant No.81972284)
文摘High-affinity antibodies are widely used in diagnostics and for the treatment of human diseases.However,most antibodies are isolated from semi-synthetic libraries by phage display and do not possess in vivo affinity maturation,which is triggered by antigen immunization.It is therefore necessary to engineer the affinity of these antibodies by way of in vitro assaying.In this study,we optimized the affinity of two human monoclonal antibodies which were isolated by phage display in a previous related study.For the 42A1 antibody,which targets the liver cancer antigen glypican-3,the variant T57H in the second complementarity-determining region of the heavy chain(CDR-H2)exhibited a 2.6-fold improvement in affinity,as well as enhanced cell-binding activity.For the I4A3 antibody to severe acute respiratory syndrome coronavirus 2,beneficial single mutations in CDR-H2 and CDR-H3 were randomly combined to select the best synergistic mutations.Among these,the mutation S53P-S98T improved binding affinity(about 3.7 fold)and the neutralizing activity(about 12 fold)compared to the parent antibody.Taken together,single mutations of key residues in antibody CDRs were enough to increase binding affinity with improved antibody functions.The mutagenic combination of key residues in different CDRs creates additive enhancements.Therefore,this study provides a safe and effective in vitro strategy for optimizing antibody affinity.
基金CAMS Innovation Fund for Medical Sciences,Grant/Award Number:2021-I2M-1-017。
文摘The efficacy and specificity of conventional monoclonal antibody(mAb)drugs in the clinic require further improvement.Currently,the development and application of novel antibody formats for improving cancer immunotherapy have attracted much attention.Variable region-retaining antibody fragments,such as antigen-binding fragment(Fab),single-chain variable fragment(scFv),bispecific antibody,and bi/trispecific cell engagers,are engineered with humanization,multivalent antibody construction,affinity optimization and antibody masking for targeting tumor cells and killer cells to improve antibody-based therapy potency,efficacy and specificity.In this review,we summarize the application of antibody variable region engineering and discuss the future direction of antibody engineering for improving cancer therapies.
文摘Objective To study the passive immunization with human monoclonal antibodies as for prophylaxis of human cytomegalovirus （HCMV） infection. Methods Fab monoclonal antibodies to HCMV were recovered by repertoire cloning of mRNA from a HCMV infected individual. Antigen binding specificity, CDR sequence of VH and VL and neutralizing activity on HCMV AD169 stain were analyzed in vitro. The light and heavy chain Fd fragment genes of Fab antibodies were further cloned into a recombinant baculovirus expression vector pAC-K-Fc to express intact IgG. Secreted products were purified with affinity chromatography using protein G. Results SDS-PAGE and Western blot confirmed the expression of the intact IgG. Immuno-blotting and -precipitation were used to identify HCMV proteins. One Fab monoclonal antibody recognized a conformational HCMV protein. Conclusion IgG antibodies can neutralize the HCMV AD169 strain efficiently at a titer of 2.5 μg/mL and may prove valuable for passive immunoprophylaxis against HCMV infection in humans.
基金Financial supports were provided in part by the New Energy and Industrial Technology Development Organization(NEDO)and by a Core Research for Evolutional Science and Technology(CREST)grant from the Japan Science and Technology Corporation(JPMJCR18S3).
文摘Monoclonal antibodies(mAbs)are widely utilized as therapeutic drugs for various diseases,such as cancer,autoimmune diseases,and infectious diseases.Using the avian-derived B cell line DT40,we previously developed an antibody display technology,namely,the ADLib system,which rapidly generates antigen-specific mAbs.Here,we report the development of a human version of the ADLib system and showcase the streamlined generation and optimization of functional human mAbs.Tailored libraries were first constructed by replacing endogenous immunoglobulin genes with designed human counterparts.From these libraries,clones producing full-length human IgGs against distinct antigens can be isolated,as exemplified by the selection of antagonistic mAbs.Taking advantage of avian biology,effective affinity maturation was achieved in a straightforward manner by seamless diversification of the parental clones into secondary libraries followed by single-cell sorting,quickly affording mAbs with improved affinities and functionalities.Collectively,we demonstrate that the human ADLib system could serve as an integrative platform with unique diversity for rapid de novo generation and optimization of therapeutic or diagnostic antibody leads.Furthermore,our results suggest that libraries can be constructed by introducing exogenous genes into DT40 cells,indicating that the ADLib system has the potential to be applied for the rapid and effective directed evolution and optimization of proteins in various fields beyond biomedicine.
基金funded by the University of Witwatersrand postdoctoral research fellowship obtained by O.Ssupported by the South African Research Chairs Initiative(SARChI)of the Department of Science and Technologythe National Research Foundation(grant 64788 to I.A.).
文摘In this dispensation of the fourth industrial revolution,protein engineering has become a popular approach for increasing enzymatic activity,stability,and titer in the biosynthesis of natural products.This is attributed to its numerous advantages(over direct isolation from plants or via chemical synthesis),including decreasing or eliminating reaction byproducts,high precision,moderate handling of intricate and chemically unstable chemicals,overall reusability,and cost efficiency.Recently,protein engineering tools have advanced to redesign and enhance natural product biosynthesis.These methods include direct evolution,substrate engineering,medium engineering,enzyme engineering and immobilization,structure-assisted protein engineering,and advanced computational.Recent successes in implementing these emerging protein engineering technologies were critically discussed in this article.Also,the advantages,limitations,and applications in industrial and medical biotechnology were discussed.Last,future research directions and potential were also highlighted.
基金support from the National Institutes of Health(R01GM144986,R01CA232017,and R21AI174606 to Y.Z.,as well as R35HL166557,R01DK132286,and R01CA240258 to Y.H.)the Leukemia&Lymphoma Society(to Y.Z.)the Welch Foundation(BE-1913-20220331 to Y.Z.).
文摘Ion channels play instrumental roles in regulating membrane potential and cross-membrane signal transduction,thus making them attractive targets for understanding various physiological processes and associated diseases.Gaining a deeper understanding of their structural and functional properties has significant implications for developing therapeutic interventions.In recent years,nanobodies,single-domain antibody fragments derived from camelids,have emerged as powerful tools in ion channel and synthetic biology research.Their small size,high specificity,and ability to recognize difficult-to-reach epitopes offer advantages over conventional antibodies and biologics.Furthermore,their resemblance to the variable region of human IgG family III reduces immunogenicity concerns.Nanobodies have introduced new opportunities for exploring ion channel structure-function relationships and offer a promising alternative to conventional drugs,which often face challenges such as off-target effects and toxicity.This review highlights recent progress in applying nanobodies to interrogate and modulate ion channel activity,with an emphasis on their potential to overcome current technical and therapeutic limitations.
基金supported by Shenzhen Medical Research Fund(B2402027)the National Natural Science Foundation of China(32171464)+3 种基金Shenzhen Fundamental Research Program(Natural Science Foundation)-Key Basic Research Project(JCYJ20241202125200001)the National Key R&D Program of China(2019YFA0904200 and 2019YFA0906100)Shenzhen Nanshan District Health System Science and Technology Major Project(NSZD2023018)This study is a Nanshan District medical key discipline construction financial support project.
文摘Advancements in protein engineering have driven the continuous optimization of T-cell engagers(TCEs),resulting in remarkable clinical outcomes in the treatment of B-cell malignancies.Moreover,developing tri-or multispecific TCEs has emerged as a promising strategy to address the challenges of tumor heterogeneity and antigen escape.However,considerable obstacles remain,primarily in format design.In this study,we engineered BAFF-based TCEs with various formats that incorporate anti-CD3 Fab or IgG domains fused with BAFF ligands to target BAFF recep tors(BAFFR,BCMA,and TACI).These constructs varied in valency and the presence or absence of long-acting ele ments such as Fc domains or the albumin binding domain consensus sequence(ABDCon).Although the inclusion of an Fc domain did not enhance sustained tumor eradication,variations in valency and spatial configuration profoundly influenced cytotoxicity.We identified TriBAFF/CD3/ABDCon as the optimal trifunctional construct,fea turing an anti-CD3 Fab backbone with BAFF and ABDCon fused to the C-termini of the heavy and light chains.This design facilitates optimal immune synapse formation between the target cells and T cells and effectively controls tumor burdens in various B-cell malignancy models with good tolerability.Notably,TriBAFF/CD3/ABDCon outper formed conventional therapies,including blinatumomab and BAFF-based CAR-T cells,in models of heterogeneous leukemia and aggressive lymphoma.These findings underscore the potential of using natural ligands as antibody-targeting modules and provide valuable insights into the design of the next generation of multispecific TCEs,which hold promise for improving treatment outcomes in a wide range of malignancies and beyond.
文摘As of May 1, 2017, 74 antibody-based molecules have been approved by a regulatory authority in a major market. Additionally, there are 70 and 575 antibody- based molecules in phase III and phase 1/11 clinical trials, respectively. These total 719 antibody-based clinical stage molecules include 493 naked IgGs, 87 antibody- drug conjugates, 61 bispecific antibodies, 37 total Fc fusion proteins, 17 radioimmunoglobulins, 13 antibody fragments, and 11 immunocytokines. New uses for these antibodies are being discovered each year. For oncol- ogy, many of the exciting new approaches involve anti- body modulation of T-cells. There are over 80 antibodies in clinical trials targeting T cell checkpoints, 26 T-cell- redirected bispecific antibodies, and 145 chimeric anti- gen receptor （CAR） cell-based candidates （all currently in phase I or II clinical trials）, totaling more than 250 T cell interacting clinical stage antibody-based candi- dates. Finally, significant progress has been made recently on routes of delivery, including delivery of proteins across the blood-brain barrier, oral delivery to the gut, delivery to the cellular cytosol, and gene- and viral-based delivery of antibodies. Thus, there are cur- rently at least 864 antibody-based clinical stage mole- cules or cells, with incredible diversity in how they are constructed and what activities they impart. These are followed by a next wave of novel molecules, approa- ches, and new methods and routes of delivery, demon- strating that the field of antibody-based biologics is very innovative and diverse in its approaches to fulfill their promise to treat unmet medical needs.
基金supported by the government funds of Shenzhen,China(SFG 2012.566 and SKC 2012.237)
文摘As a result of recent breakthroughs in cancer immunotherapies, unprecedented and durable remission, and even cure, has been reported in some patients. Importantly, this progress has been achieved, not by the induction of immunity, but by the delivery of immunity in the form of engineered antibodies （cAbs） or effector T cells. However, these single-target technologies have failed to result in a therapeutic effect in some patients, and evidence suggests that further advances depend on an effective strategy for coping with cancer heterogeneity and dynamics. A synthetic immunity （SI） strategy is proposed to achieve this goal. The fundamental basis of SI involves the generation of a panel of cAbs and antibody-retargeted CTLs designed to destroy all cell lineages of a cancer with high specificity. This goal can be achieved only when the composition of the cAbs is determined using a systematic approach, i.e., selecting the antigens targeted by the cAbs based on an epitope-tree illustrating the clonal antigen architecture of the cancer. Integration of technologies that increase the epitope breadth, cAb affinity and T cell activity will further enhance the efficacy of SI. Using DNA vectors to express the eAbs will be a safe, effective and affordable solution.
文摘In this study, we discussed the necessity of human IgG1 Cγ1 domain for recombinant antibody using computeraided homology modeling method and experimental studies. The heavy （VH） and light （VL） chain variable regions of 1-28, a murine IgM-type anti-CD20 mAb, were ligated by linker peptide （Gly4Ser）3 to form the single-chain Fv fragment （scFv）. Then, the engineered antibody （LH1-3） was generated by fusing scFv with the entire IgG1 heavy constant regions. The 3-D structure of LH1-3 was modeled using computer-aided homology modeling method and the binding activity of LH1-3 was evaluated theoretically. Compared to the 3-D structure of the Fv fragment of the parent antibody, the conformation of the active pocket of LH1-3 was remained because of the rigid support of Cγ1. Further experimental results of flow cytometry showed that the engineered anti-CD20 antibody possessed specifically binding activity to CD20-expressing target cells. The anti-CD20 antibody fragments could also mediate complement-dependent cytotoxicity （CDC） of human B-lymphoid cell lines. Our study highlights some interests and advantages of a methodology based on the homology modeling and analysis of molecular structural properties.
