Immuntoxins were synthesized by conjugating a plant toxic ricin with to three different monclonal antibodiesMoAb) directed against markers of human pre- B lymphocyte leukemic cells. It is useful to eliminate residual ...Immuntoxins were synthesized by conjugating a plant toxic ricin with to three different monclonal antibodiesMoAb) directed against markers of human pre- B lymphocyte leukemic cells. It is useful to eliminate residual leukemic cells from bone marrow for preventing leukemia relapse after autologous bone marrow transplantation(ABMT). In the present work, the elimination of human leukemic cell line (Nalm-6) by three immunotoxins (anti-CD9 and anti-CD10) were observed. In addition, the proliferation of hematopoietic progenitors (CFU-GM. BFU E and CFU- mix) were not apparently inhibited by the immunotoxins in the range of effective concentrations. The possibility for utilizating immunotoxins in ABMT was discussed.展开更多
In the past decade,an increased amount of clinicallyloriented research involving immunotoxins has been published. Immunotoxins are a group of artificially-made cytotoxic molecules targeting cancer cells.These molecule...In the past decade,an increased amount of clinicallyloriented research involving immunotoxins has been published. Immunotoxins are a group of artificially-made cytotoxic molecules targeting cancer cells.These molecules composed of a targeting moiety,such as a ligand or an antibody,linked to toxin moiety,which is a toxin with either truncated or deleted cell-binding domain that prevents it from binding to normal cells.Immunotoxins can be divided into two categories:chemically conjugated immunotoxins and recombinant ones.The immunotoxins of the first category have shown limited efficacy in clinical trials in patients with hematologic malignancies and solid tumors.Within the last few years,single-chain immunotoxins provide enhanced therapeutic efficacy over conjugated forms and result in improved antitumor activity.In this review,we briefly illustrate the design of the immunotoxins and their applications in clinical trials.Cellular & Molecular Immunology.2005;2(2):106-112.展开更多
Objective To induce islet grafting tolerance by intravenous injection of anti CD 4, anti CD 8 immunotoxins and donor soluble antigen Methods Fourteen days or 7 days prior to transplantation, the immunotoxin of...Objective To induce islet grafting tolerance by intravenous injection of anti CD 4, anti CD 8 immunotoxins and donor soluble antigen Methods Fourteen days or 7 days prior to transplantation, the immunotoxin of anti CD 4, anti CD 8 200?μg respectively, and donor soluble antigen 500?μg were intravenously injected and then 500 donor islets were transplanted under the left renal subcapsular space of diabetes recipients (Sprague Dawley rats) Results The islet grafting survival time for those recipients pretreated with immunotoxin and donor soluble antigen was >60 days ( P <0 01) The immunotoxins, donor soluble antigen treatment alone might only slightly prolong the grafting survival time Conclusion The anti CD 4, anti CD 8 immunotoxins jointly used with donor soluble antigen can induce donor specific immunotolerance展开更多
A novel ribosome-inactivating protein designated Moschatin from the mature seeds of pumpkin (Cucurbita moschata) has been successively purified to homogeneity, using ammonium sulfate precipitation, CM-cellulose 52 col...A novel ribosome-inactivating protein designated Moschatin from the mature seeds of pumpkin (Cucurbita moschata) has been successively purified to homogeneity, using ammonium sulfate precipitation, CM-cellulose 52 column chromatography, Blue Sepharose CL-6B Affinity column chromatography and FPLC size-exclusion column chromatography. Moschatin is a type 1 RIP with a pI of 9.4 and molecular weight of~29 kD. It is a rRNA N-glycosidase and potently blocked the protein synthesis in the rabbit reticulocyte lysate with a IC_(50) of 0.26 nM. Using the anti-human melanoma McAb Ng76, a novel immunotoxin Moschatin-Ng76 was prepared successfully and it efficiently inhibited the growth of targeted melanoma cells M_(21) with a IC_(50) of 0.04 nM, 1500 times lower than that of free Moschatin. The results implied that Moschatin could be used as a new potential anticancer agent.展开更多
Schistosomiasis is one of the most prevalent parasitic diseases in China, and hepatic fibrosis caused by schistosome infection is the principal cause of death. The aim of this study was to evaluate the efficacy of NPl...Schistosomiasis is one of the most prevalent parasitic diseases in China, and hepatic fibrosis caused by schistosome infection is the principal cause of death. The aim of this study was to evaluate the efficacy of NPll-4- derived immunotoxin scFv-artesunate on Schistosoma japonicum-induced hepatic fibrosis. A single-chain variable fragment （scFv） was generated from the murine anti-Schistosoma japonicum （S. japanicum） monoclonal antibody NP11-4. The scFv was expressed as a soluble protein and purified by Ni-affinity chromatography. After conjuga- tion with artesunate, the binding ability with soluble egg antigens （SEA） was determined by an enzyme-linked immunosorbent assay （ELISA）. The biological activity of purified scFv, scFv-artesunate （immunotoxin）, and artesunate was detected in vivo. Image-Pro Plus software was used to analyze the size of egg granuloma and the extent of liver fibrosis. The recombinant scFv expession vector was constructed and expressed successfully. After purification by a His-trap Ni-affinity column, the scFv yield was approximately 0.8 mg/L of culture medium. ELISA results showed that chemical conjugation did not affect the binding activity of the immunotoxin. Our animal experiments indicated that the immunotoxin could significantly reduce the size of egg granuloma in the liver and inhibit hepatic fibrosis. The immunotoxin could be used as a promising candidate in the targeted therapy of S. .japonicum-induced hepatic fibrosis.展开更多
In the present study,daunomydn （DIM） was chosen to conjugate cowdently with amonoclonal antitibody,MGb<sub>2</sub>,against human gastric cancer cells via a cis-aconitic anhydride linker（directly） or a ...In the present study,daunomydn （DIM） was chosen to conjugate cowdently with amonoclonal antitibody,MGb<sub>2</sub>,against human gastric cancer cells via a cis-aconitic anhydride linker（directly） or a dextmn bridge （indirectly,） The mo-lar ratio of MGb<sub>2</sub> to DM in the conjugates was1:6 （direct method） and 1:54 （indirect method）,respectively.The ELISA results revealed theantibody after conjugation retained antigen-binding capacity.The conjugates showed a highly se-lective cytotoxicity to the target cells.In lh cytotoxidty test,cytotoxidty of the conjugates wasgreater than that of free DM or irrevalent conjugates to human gastric cancer cell lines,SGC-7901and very low as far as non-target cells （HeLa） were concerned.It is sugared that theselective cytotoxidty on target cells of the conjugates is mediated by the monoclonal antibody.展开更多
In the present study, an indirect assay was employed to investigate 5 anti-gastric cancer monoclonal antibodies for their cytotoxic potential as ricin A chain-containing immunotoxins. The tumor cell, were treated with...In the present study, an indirect assay was employed to investigate 5 anti-gastric cancer monoclonal antibodies for their cytotoxic potential as ricin A chain-containing immunotoxins. The tumor cell, were treated with dilutions of tested antibody followed by ricin A chain coupled to goat anti-mouse immunoglobulin. The cytotoxic effect was determined with tetrazolium colorimetric assay. The results showed that among the 5 antibodies chosen, MGb2 and MG7 could be well used for preparation of effective A chain immunotoxins.展开更多
intraventricular injection of the immunotoxin,192-sap,that selectively de stroys neurons expressing the low affinity neurotrophin receptor,p75NGFr,efficiently and selectively destroys the cholinergic neurons of the ha...intraventricular injection of the immunotoxin,192-sap,that selectively de stroys neurons expressing the low affinity neurotrophin receptor,p75NGFr,efficiently and selectively destroys the cholinergic neurons of the hasal forebrain(CBF).In the present study,we sought to determine if there was a correlation between the degree of CBF neuron loss and alteration in passive avoidance behavior.Anesthetized,adult, male Sprague Dawley rats were stereotactically injected with 4μg of either 192-sap or OX7-sap,a control immunotoxin that recognizes the Thy 1 surface antigen and destroys cerebellar Purkinje neu rons. 6￣8 weeks later,immunotoxin and naive control rats were tested on step-through passive avoidance paradigm. After behavior-testing, all rats were sacrificed and brain sections processed for histochemical demonstration of AChE and immunohistochemical demonstration of p75NGFr.The numbers of neurons in specific regions of the CBF were counted from the P75NGFr,staining and the intensity of dorsolateral neocortical staining for AChE were assessed using image analysis. The magnitude of cell loss was similar(67 % ￣70%) for the entire CBF, the Nbm and septum/DBB. The seventy of passive avoidance impairment was significantly correlated to cell loss in the entire CBF (r= 0.748,23 df, P<0.001) and in the Nbm (r = 0.778, 23 df, P< 0.001) but not the septum/DBB (r=0. 419,23 df, P>0. 05).Behavioral impairment also correlated significantly to loss of conical AchE staining intensity ipsilateral to the intraventricular injections (r = 0.796,15df,P< 0.001).These fndings show that loss of Nbm, but not of septum/DBB,cholinergic neurons is proportional to impairment in passive avoidance behavior suggesting a role for Nbm-neocortex cholinergic innervation in this type of learning.(Supported by the Department of Veterans Affairs.)展开更多
Elevated collagen levels within breast tumors are strongly associated with tumor progression and present a barrier to effective therapeutic agent penetration within the tumor microenvironment(TME),leading to poor clin...Elevated collagen levels within breast tumors are strongly associated with tumor progression and present a barrier to effective therapeutic agent penetration within the tumor microenvironment(TME),leading to poor clinical outcomes.To address this challenge,we engineered a probiotic strain to degrade collagen within the TME by selectively colonizing in tumors and releasing bacterial collagenase in a lysis-dependent manner.Initially,we constructed a therapeutic bacterial strain designed to lyse within the TME and release an encoded immunotoxin comprising a nanobody targeting CD47(CD47nb)and a modified Pseudomonas exotoxin A(PE38KDEL).The introduction of collagenase-expressing bacteria,in conjunction with therapeutic immunotoxin,reduced collagen fiber levels within the TME,resulting in inhibited tumor growth and prolonged survival in a murine model of breast cancer.Furthermore,we investigated the broader applicability of the collagenase-expressing bacterial strain in combination with chemotherapeutic drugs,such as doxorubicin.Remarkably,synergistic antitumor effects were observed in mice treated with this combination therapy.In conclusion,our study demonstrates that probiotic delivery of bacterial collagenase offers a promising adjuvant treatment strategy for selectively degrading intratumoral collagen,thereby improving the efficacy of anticancer therapies in breast cancer.展开更多
TROP-2, a tumor-associated antigen, has been implicated in the progression of various epithelial tumors. Due to its favorable expression profile, TROP-2 has emerged as a promising target for antibody-drug conjugates (...TROP-2, a tumor-associated antigen, has been implicated in the progression of various epithelial tumors. Due to its favorable expression profile, TROP-2 has emerged as a promising target for antibody-drug conjugates (ADCs) based anti-tumor therapies. Although ADCs have shown efficacy in cancer treatment, their application in solid tumors is hindered by their high molecular weight, poor tumor penetration, and release of cytotoxic molecules. Therefore, a recombinant immunotoxin was developed based on a shark-derived variable domain of immunoglobulin new antigen receptor (VNAR) antibody. VNARs are only one-tenth the size of IgG antibodies and possess remarkable tissue penetration capabilities and high stability. In this study, a shark VNAR phage display library was created, leading to the identification of shark VNAR-5G8 that targets TROP-2. VNAR-5G8 exhibited a high affinity and cellular internalization ability towards cells expressing high levels of TROP-2. Epitope analysis revealed that VNAR-5G8 recognizes a hidden epitope consisting of CRD and TY-1 on TROP-2. Subsequently, VNAR-5G8 was fused with a truncated form of Pseudomonas exotoxin (PE38) to create the recombinant immunotoxin (5G8-PE38), which exhibited significant anti-tumor activity in vitro and in vivo. Overall, this study highlights the promise of 5G8-PE38 as a valuable candidate for cancer therapy.展开更多
Objective To study the selective toxicity of immunotoxin (IT) on T cells in cord blood and simultaneously determine its effect on hematopoietic progenitor cells Methods The percentage of CD 5 and CD 8 T c...Objective To study the selective toxicity of immunotoxin (IT) on T cells in cord blood and simultaneously determine its effect on hematopoietic progenitor cells Methods The percentage of CD 5 and CD 8 T cell subsets in cord blood (CB) and bone marrow (BM) as well as peripheral blood (PB) was measured by immunoenzymatic labeling of monoclonal antibodies using immune complexes of alkaline phosphatase and monoclonal anti alkaline phosphatase (APAAP complexes) One way mixed lymphocyte cultures (MLC) were performed to compare the proliferative response of CB with that of PB The proliferative capability of cord blood T cells and T lymphocyte transformation capacity were evaluated in the presence of anti CD 8 or anti CD 5 immunotoxin by one way MLC and colorimetric MTT (tetrazolium) assay, respectively The effect of IT on the growth of hematopoietic progenitor cell of colony forming unit granulocyte and macrophage (CFU GM), burst forming unit erythroid(BFU E), multipotential hemotapoietic progenitors (CFU Mix) from CB were estimated by colony forming assays Results A certain proportion of CD 5 and CD 8 T cells existed in CB The alloproliferative capacity of CB was similar to that of PB CD 5: Ricin at a dosage of 1×10 10 -1×10 8 mmol/L and CD 8: Ricin concentration in the range of 1×10 9 -1×10 8 mmol/L effectively decreased both the proliferative capability of T cells in MLC during CB and T cell transformation Over the dosage of 1×10 10 -1×10 9 mmol/L, both kinds of IT didn't obviously affect the growth of hematopoietic progenitor cells Conclusion CD 5: Ricin and CD 8: Ricin may effectively deplete T cells and may not significantly inhibit the function of hemaptopoietic cells at a specific dosage展开更多
INTRODUCTIONLung cancer is the leading cause for cancer-related deaths in both genders throughout the world. In the United States alone, there were 224,390 estimated new lung cancer cases and 158,080 estimated deaths ...INTRODUCTIONLung cancer is the leading cause for cancer-related deaths in both genders throughout the world. In the United States alone, there were 224,390 estimated new lung cancer cases and 158,080 estimated deaths in 2016.展开更多
Background Previous studies using knockout mice document a key role for the integrin CD103 in promoting organ allograft rejection and graft-versus-host disease. However, a determination of whether blockade of the CD10...Background Previous studies using knockout mice document a key role for the integrin CD103 in promoting organ allograft rejection and graft-versus-host disease. However, a determination of whether blockade of the CD103 pathway represents a viable therapeutic strategy for intervention in these processes has proven problematic due to the lack of reagents that efficiently deplete CD103＋ cells from wild type hosts. To circumvent this problem, in the present study, we invented an anti-CD103 immunotoxin （M290-SAP）. We investigated whether M290-SAP has capacity to eliminate CD103-expressing cells in vivo and protect transplanted islets from destroying by host immune cells.Methods Flow cytometry was used to analyze the efficacy of M290-SAP in depleting CD103-expressing cells in vivo.Then using allogenic islet transplantation models as well as NOD mice with recent onset type 1 diabetes, the therapeutic efficacy of CD103-expressing cell depletion was addressed.Results M290-SAP dramatically reduces the frequency and absolute numbers of CD103-expressing leukocytes in peripheral lymphatic tissues of treated mice. Balb/c islets transplanted into streptozotocin-induced diabetic C57BL/6 mice under single M290-SAP treatment showed an indefinite survival time compared with untreated mice, M290-treated mice and IgG-SAP treated mice （mean survival time, ＞100 days vs. ＜20 days）. C57BL/6 islets transplanted into hyperglycemic NOD mice under single M290-SAP treatment showed a pronounced delay in allograft rejection compared with untreated mice （mean survival time 12-13 days vs. ＜7 days）. Immunological analysis of mice with long-term islet allograft survival revealed an obvious atrophy thymus and severe downregulation of alloimmunity of CD8 subpopulation response to allogenic stimulation.Conclusion Regardless of the underlying mechanisms, these data document that depletion of CD103-expressing cells represents a viable strategy for therapeutic intervention in islet allograft rejection.展开更多
1 Introduction In recent years, significant progress has been made in applying immunotoxin (IT)in the therapy of leukemia and marrow transplantation. By 1990, several ITs havebeen put into clinical trials under the pe...1 Introduction In recent years, significant progress has been made in applying immunotoxin (IT)in the therapy of leukemia and marrow transplantation. By 1990, several ITs havebeen put into clinical trials under the permission of FDA (Foodand Drug Administration, USA).展开更多
The 28 ku membrane protein is usually over-expressl m human bnasl bmast cancer and other tumor cells. licould be a larget for tumor therapy . By using genetie engineermg teehmgues.a 606 immunotoxin (sefv606-PE40) was ...The 28 ku membrane protein is usually over-expressl m human bnasl bmast cancer and other tumor cells. licould be a larget for tumor therapy . By using genetie engineermg teehmgues.a 606 immunotoxin (sefv606-PE40) was construeted by joining the 606 single-chain antibdy (SeFv606) with the truncll Pseudonwnas exotoxin A (PE40), SeFv606 contains both the heavy and light-chnia variable domams of 606 monoelonal antibody. Which speeifieally ree-ognizes the 28 ku protein. The bacterial expression level og 606 imnmmotoxin is 3.3%. about 5.5 mg ml baeterial lysate.lsing singlc-step llisTrap (Nr2 chelating) column chronnetogaphy, the reeombinant peptide was obtained with a purit of 33.2%.This baeterial espressed 606 immunotosin binuls to MDA-231 human breast-tumer ccll surfaee and kill these cells with a median lethal dosage of 92 ngnd.展开更多
Objective To investigate the special effect of donor antigen trichosnthin (TCS) conjugate on depleting the immune reactive cells that respond to xenograft, and thus inducing specific xenograft tolerance. Methods R...Objective To investigate the special effect of donor antigen trichosnthin (TCS) conjugate on depleting the immune reactive cells that respond to xenograft, and thus inducing specific xenograft tolerance. Methods Rat anti mouse antibodies coupled sepharose 4B beads were used to purify mouse H 2 antigens by affinity chromatography. Those purified H 2 antigens were conjugated with TCS by heterobifunctional reagents SPDP [3 (2 pyridyldithio) propionic acid hydroxysuccinimide ester] and 2 IT (2 iminothiolane). The specific inhibition function of Ag TCS to recipient's immune cells was measured in vitro. Ag TCS conjugate (200 μg/kg, iv) was administered to the recipients (rats) 4 days prior to cardiac xenotransplantation of mouse to rat. Results In vitro, the proliferation of immune reactive cells in the recipients pretreated with Ag TCS was inhibited. The Ag TCS conjugate significantly prolonged the cardiac survival time (6.88±1.36 days) compared with cyclosporine A (CsA) group (2.83±0.75 days) (P<0.01). Conclusion Donor Ag TCS conjugate can induce specific tolerance to xenograft.展开更多
文摘Immuntoxins were synthesized by conjugating a plant toxic ricin with to three different monclonal antibodiesMoAb) directed against markers of human pre- B lymphocyte leukemic cells. It is useful to eliminate residual leukemic cells from bone marrow for preventing leukemia relapse after autologous bone marrow transplantation(ABMT). In the present work, the elimination of human leukemic cell line (Nalm-6) by three immunotoxins (anti-CD9 and anti-CD10) were observed. In addition, the proliferation of hematopoietic progenitors (CFU-GM. BFU E and CFU- mix) were not apparently inhibited by the immunotoxins in the range of effective concentrations. The possibility for utilizating immunotoxins in ABMT was discussed.
文摘In the past decade,an increased amount of clinicallyloriented research involving immunotoxins has been published. Immunotoxins are a group of artificially-made cytotoxic molecules targeting cancer cells.These molecules composed of a targeting moiety,such as a ligand or an antibody,linked to toxin moiety,which is a toxin with either truncated or deleted cell-binding domain that prevents it from binding to normal cells.Immunotoxins can be divided into two categories:chemically conjugated immunotoxins and recombinant ones.The immunotoxins of the first category have shown limited efficacy in clinical trials in patients with hematologic malignancies and solid tumors.Within the last few years,single-chain immunotoxins provide enhanced therapeutic efficacy over conjugated forms and result in improved antitumor activity.In this review,we briefly illustrate the design of the immunotoxins and their applications in clinical trials.Cellular & Molecular Immunology.2005;2(2):106-112.
文摘Objective To induce islet grafting tolerance by intravenous injection of anti CD 4, anti CD 8 immunotoxins and donor soluble antigen Methods Fourteen days or 7 days prior to transplantation, the immunotoxin of anti CD 4, anti CD 8 200?μg respectively, and donor soluble antigen 500?μg were intravenously injected and then 500 donor islets were transplanted under the left renal subcapsular space of diabetes recipients (Sprague Dawley rats) Results The islet grafting survival time for those recipients pretreated with immunotoxin and donor soluble antigen was >60 days ( P <0 01) The immunotoxins, donor soluble antigen treatment alone might only slightly prolong the grafting survival time Conclusion The anti CD 4, anti CD 8 immunotoxins jointly used with donor soluble antigen can induce donor specific immunotolerance
文摘A novel ribosome-inactivating protein designated Moschatin from the mature seeds of pumpkin (Cucurbita moschata) has been successively purified to homogeneity, using ammonium sulfate precipitation, CM-cellulose 52 column chromatography, Blue Sepharose CL-6B Affinity column chromatography and FPLC size-exclusion column chromatography. Moschatin is a type 1 RIP with a pI of 9.4 and molecular weight of~29 kD. It is a rRNA N-glycosidase and potently blocked the protein synthesis in the rabbit reticulocyte lysate with a IC_(50) of 0.26 nM. Using the anti-human melanoma McAb Ng76, a novel immunotoxin Moschatin-Ng76 was prepared successfully and it efficiently inhibited the growth of targeted melanoma cells M_(21) with a IC_(50) of 0.04 nM, 1500 times lower than that of free Moschatin. The results implied that Moschatin could be used as a new potential anticancer agent.
基金supported by a grant from the National High Technology Research and Development Program of China ("863"ProgramNo.2006AA02Z415)
文摘Schistosomiasis is one of the most prevalent parasitic diseases in China, and hepatic fibrosis caused by schistosome infection is the principal cause of death. The aim of this study was to evaluate the efficacy of NPll-4- derived immunotoxin scFv-artesunate on Schistosoma japonicum-induced hepatic fibrosis. A single-chain variable fragment （scFv） was generated from the murine anti-Schistosoma japonicum （S. japanicum） monoclonal antibody NP11-4. The scFv was expressed as a soluble protein and purified by Ni-affinity chromatography. After conjuga- tion with artesunate, the binding ability with soluble egg antigens （SEA） was determined by an enzyme-linked immunosorbent assay （ELISA）. The biological activity of purified scFv, scFv-artesunate （immunotoxin）, and artesunate was detected in vivo. Image-Pro Plus software was used to analyze the size of egg granuloma and the extent of liver fibrosis. The recombinant scFv expession vector was constructed and expressed successfully. After purification by a His-trap Ni-affinity column, the scFv yield was approximately 0.8 mg/L of culture medium. ELISA results showed that chemical conjugation did not affect the binding activity of the immunotoxin. Our animal experiments indicated that the immunotoxin could significantly reduce the size of egg granuloma in the liver and inhibit hepatic fibrosis. The immunotoxin could be used as a promising candidate in the targeted therapy of S. .japonicum-induced hepatic fibrosis.
文摘In the present study,daunomydn （DIM） was chosen to conjugate cowdently with amonoclonal antitibody,MGb<sub>2</sub>,against human gastric cancer cells via a cis-aconitic anhydride linker（directly） or a dextmn bridge （indirectly,） The mo-lar ratio of MGb<sub>2</sub> to DM in the conjugates was1:6 （direct method） and 1:54 （indirect method）,respectively.The ELISA results revealed theantibody after conjugation retained antigen-binding capacity.The conjugates showed a highly se-lective cytotoxicity to the target cells.In lh cytotoxidty test,cytotoxidty of the conjugates wasgreater than that of free DM or irrevalent conjugates to human gastric cancer cell lines,SGC-7901and very low as far as non-target cells （HeLa） were concerned.It is sugared that theselective cytotoxidty on target cells of the conjugates is mediated by the monoclonal antibody.
文摘In the present study, an indirect assay was employed to investigate 5 anti-gastric cancer monoclonal antibodies for their cytotoxic potential as ricin A chain-containing immunotoxins. The tumor cell, were treated with dilutions of tested antibody followed by ricin A chain coupled to goat anti-mouse immunoglobulin. The cytotoxic effect was determined with tetrazolium colorimetric assay. The results showed that among the 5 antibodies chosen, MGb2 and MG7 could be well used for preparation of effective A chain immunotoxins.
文摘intraventricular injection of the immunotoxin,192-sap,that selectively de stroys neurons expressing the low affinity neurotrophin receptor,p75NGFr,efficiently and selectively destroys the cholinergic neurons of the hasal forebrain(CBF).In the present study,we sought to determine if there was a correlation between the degree of CBF neuron loss and alteration in passive avoidance behavior.Anesthetized,adult, male Sprague Dawley rats were stereotactically injected with 4μg of either 192-sap or OX7-sap,a control immunotoxin that recognizes the Thy 1 surface antigen and destroys cerebellar Purkinje neu rons. 6￣8 weeks later,immunotoxin and naive control rats were tested on step-through passive avoidance paradigm. After behavior-testing, all rats were sacrificed and brain sections processed for histochemical demonstration of AChE and immunohistochemical demonstration of p75NGFr.The numbers of neurons in specific regions of the CBF were counted from the P75NGFr,staining and the intensity of dorsolateral neocortical staining for AChE were assessed using image analysis. The magnitude of cell loss was similar(67 % ￣70%) for the entire CBF, the Nbm and septum/DBB. The seventy of passive avoidance impairment was significantly correlated to cell loss in the entire CBF (r= 0.748,23 df, P<0.001) and in the Nbm (r = 0.778, 23 df, P< 0.001) but not the septum/DBB (r=0. 419,23 df, P>0. 05).Behavioral impairment also correlated significantly to loss of conical AchE staining intensity ipsilateral to the intraventricular injections (r = 0.796,15df,P< 0.001).These fndings show that loss of Nbm, but not of septum/DBB,cholinergic neurons is proportional to impairment in passive avoidance behavior suggesting a role for Nbm-neocortex cholinergic innervation in this type of learning.(Supported by the Department of Veterans Affairs.)
基金sponsored by the National Natural Science Founda-tion of China(22134003)the National Key Research and Develop-ment Program of China(2020YFA0908800).
文摘Elevated collagen levels within breast tumors are strongly associated with tumor progression and present a barrier to effective therapeutic agent penetration within the tumor microenvironment(TME),leading to poor clinical outcomes.To address this challenge,we engineered a probiotic strain to degrade collagen within the TME by selectively colonizing in tumors and releasing bacterial collagenase in a lysis-dependent manner.Initially,we constructed a therapeutic bacterial strain designed to lyse within the TME and release an encoded immunotoxin comprising a nanobody targeting CD47(CD47nb)and a modified Pseudomonas exotoxin A(PE38KDEL).The introduction of collagenase-expressing bacteria,in conjunction with therapeutic immunotoxin,reduced collagen fiber levels within the TME,resulting in inhibited tumor growth and prolonged survival in a murine model of breast cancer.Furthermore,we investigated the broader applicability of the collagenase-expressing bacterial strain in combination with chemotherapeutic drugs,such as doxorubicin.Remarkably,synergistic antitumor effects were observed in mice treated with this combination therapy.In conclusion,our study demonstrates that probiotic delivery of bacterial collagenase offers a promising adjuvant treatment strategy for selectively degrading intratumoral collagen,thereby improving the efficacy of anticancer therapies in breast cancer.
基金Qingdao Marine Science and Technology Center(No.2022QNLM030003-4 and 8-01,China)Program of National Natural Science Foundation of China(No.82273846)Taishan Scholars Program(No.tsqn202211058,China).
文摘TROP-2, a tumor-associated antigen, has been implicated in the progression of various epithelial tumors. Due to its favorable expression profile, TROP-2 has emerged as a promising target for antibody-drug conjugates (ADCs) based anti-tumor therapies. Although ADCs have shown efficacy in cancer treatment, their application in solid tumors is hindered by their high molecular weight, poor tumor penetration, and release of cytotoxic molecules. Therefore, a recombinant immunotoxin was developed based on a shark-derived variable domain of immunoglobulin new antigen receptor (VNAR) antibody. VNARs are only one-tenth the size of IgG antibodies and possess remarkable tissue penetration capabilities and high stability. In this study, a shark VNAR phage display library was created, leading to the identification of shark VNAR-5G8 that targets TROP-2. VNAR-5G8 exhibited a high affinity and cellular internalization ability towards cells expressing high levels of TROP-2. Epitope analysis revealed that VNAR-5G8 recognizes a hidden epitope consisting of CRD and TY-1 on TROP-2. Subsequently, VNAR-5G8 was fused with a truncated form of Pseudomonas exotoxin (PE38) to create the recombinant immunotoxin (5G8-PE38), which exhibited significant anti-tumor activity in vitro and in vivo. Overall, this study highlights the promise of 5G8-PE38 as a valuable candidate for cancer therapy.
文摘Objective To study the selective toxicity of immunotoxin (IT) on T cells in cord blood and simultaneously determine its effect on hematopoietic progenitor cells Methods The percentage of CD 5 and CD 8 T cell subsets in cord blood (CB) and bone marrow (BM) as well as peripheral blood (PB) was measured by immunoenzymatic labeling of monoclonal antibodies using immune complexes of alkaline phosphatase and monoclonal anti alkaline phosphatase (APAAP complexes) One way mixed lymphocyte cultures (MLC) were performed to compare the proliferative response of CB with that of PB The proliferative capability of cord blood T cells and T lymphocyte transformation capacity were evaluated in the presence of anti CD 8 or anti CD 5 immunotoxin by one way MLC and colorimetric MTT (tetrazolium) assay, respectively The effect of IT on the growth of hematopoietic progenitor cell of colony forming unit granulocyte and macrophage (CFU GM), burst forming unit erythroid(BFU E), multipotential hemotapoietic progenitors (CFU Mix) from CB were estimated by colony forming assays Results A certain proportion of CD 5 and CD 8 T cells existed in CB The alloproliferative capacity of CB was similar to that of PB CD 5: Ricin at a dosage of 1×10 10 -1×10 8 mmol/L and CD 8: Ricin concentration in the range of 1×10 9 -1×10 8 mmol/L effectively decreased both the proliferative capability of T cells in MLC during CB and T cell transformation Over the dosage of 1×10 10 -1×10 9 mmol/L, both kinds of IT didn't obviously affect the growth of hematopoietic progenitor cells Conclusion CD 5: Ricin and CD 8: Ricin may effectively deplete T cells and may not significantly inhibit the function of hemaptopoietic cells at a specific dosage
基金The work was supported by grants from the Young Scientists Fund from the National Natural Science Foundation of China （No. 81401896）, and the Pujiang Talent Program from Shanghai Municipal Human Resource Bureau and Shanghai Science and Technology Committee （No. 14PJ1402000）.
文摘INTRODUCTIONLung cancer is the leading cause for cancer-related deaths in both genders throughout the world. In the United States alone, there were 224,390 estimated new lung cancer cases and 158,080 estimated deaths in 2016.
文摘Background Previous studies using knockout mice document a key role for the integrin CD103 in promoting organ allograft rejection and graft-versus-host disease. However, a determination of whether blockade of the CD103 pathway represents a viable therapeutic strategy for intervention in these processes has proven problematic due to the lack of reagents that efficiently deplete CD103＋ cells from wild type hosts. To circumvent this problem, in the present study, we invented an anti-CD103 immunotoxin （M290-SAP）. We investigated whether M290-SAP has capacity to eliminate CD103-expressing cells in vivo and protect transplanted islets from destroying by host immune cells.Methods Flow cytometry was used to analyze the efficacy of M290-SAP in depleting CD103-expressing cells in vivo.Then using allogenic islet transplantation models as well as NOD mice with recent onset type 1 diabetes, the therapeutic efficacy of CD103-expressing cell depletion was addressed.Results M290-SAP dramatically reduces the frequency and absolute numbers of CD103-expressing leukocytes in peripheral lymphatic tissues of treated mice. Balb/c islets transplanted into streptozotocin-induced diabetic C57BL/6 mice under single M290-SAP treatment showed an indefinite survival time compared with untreated mice, M290-treated mice and IgG-SAP treated mice （mean survival time, ＞100 days vs. ＜20 days）. C57BL/6 islets transplanted into hyperglycemic NOD mice under single M290-SAP treatment showed a pronounced delay in allograft rejection compared with untreated mice （mean survival time 12-13 days vs. ＜7 days）. Immunological analysis of mice with long-term islet allograft survival revealed an obvious atrophy thymus and severe downregulation of alloimmunity of CD8 subpopulation response to allogenic stimulation.Conclusion Regardless of the underlying mechanisms, these data document that depletion of CD103-expressing cells represents a viable strategy for therapeutic intervention in islet allograft rejection.
基金Bio-High Technology Program from the State Commission of Science and Technology, PRC.
文摘1 Introduction In recent years, significant progress has been made in applying immunotoxin (IT)in the therapy of leukemia and marrow transplantation. By 1990, several ITs havebeen put into clinical trials under the permission of FDA (Foodand Drug Administration, USA).
基金Project Supported by the Chinese Natiotal High-Tech Project
文摘The 28 ku membrane protein is usually over-expressl m human bnasl bmast cancer and other tumor cells. licould be a larget for tumor therapy . By using genetie engineermg teehmgues.a 606 immunotoxin (sefv606-PE40) was construeted by joining the 606 single-chain antibdy (SeFv606) with the truncll Pseudonwnas exotoxin A (PE40), SeFv606 contains both the heavy and light-chnia variable domams of 606 monoelonal antibody. Which speeifieally ree-ognizes the 28 ku protein. The bacterial expression level og 606 imnmmotoxin is 3.3%. about 5.5 mg ml baeterial lysate.lsing singlc-step llisTrap (Nr2 chelating) column chronnetogaphy, the reeombinant peptide was obtained with a purit of 33.2%.This baeterial espressed 606 immunotosin binuls to MDA-231 human breast-tumer ccll surfaee and kill these cells with a median lethal dosage of 92 ngnd.
文摘Objective To investigate the special effect of donor antigen trichosnthin (TCS) conjugate on depleting the immune reactive cells that respond to xenograft, and thus inducing specific xenograft tolerance. Methods Rat anti mouse antibodies coupled sepharose 4B beads were used to purify mouse H 2 antigens by affinity chromatography. Those purified H 2 antigens were conjugated with TCS by heterobifunctional reagents SPDP [3 (2 pyridyldithio) propionic acid hydroxysuccinimide ester] and 2 IT (2 iminothiolane). The specific inhibition function of Ag TCS to recipient's immune cells was measured in vitro. Ag TCS conjugate (200 μg/kg, iv) was administered to the recipients (rats) 4 days prior to cardiac xenotransplantation of mouse to rat. Results In vitro, the proliferation of immune reactive cells in the recipients pretreated with Ag TCS was inhibited. The Ag TCS conjugate significantly prolonged the cardiac survival time (6.88±1.36 days) compared with cyclosporine A (CsA) group (2.83±0.75 days) (P<0.01). Conclusion Donor Ag TCS conjugate can induce specific tolerance to xenograft.
