RESUMO
CD25, also known as the interleukin-2 receptor α chain (IL-2Rα), is highly expressed on regulatory T cells (Tregs), but relatively lower on effector T cells (Teffs). This makes it a potential target for Treg depletion, which can be used in tumor immunotherapy. However, marketed anti-CD25 antibodies (Basiliximab and Daclizumab) were originally developed as immunosuppressive drugs to prevent graft rejection, because these antibodies can block IL-2 binding to CD25 on Teffs, which in turn destroys the function of Teffs. Recent studies have shown that non-IL-2-blocking anti-CD25 antibodies have displayed exciting antitumor effects. Here, we screened out a non-IL-2-blocking anti-CD25 monoclonal antibody (mAb) 7B7 by hybridoma technology, and confirmed its antitumor activity via depleting Tregs in a CD25 humanized mouse model. Subsequently, we verified that the humanized 7B7, named as h7B7-15S, has comparable activities to 7B7, and that its Treg depletion is further increased when combined with anti-CTLA-4, leading to enhanced remodeling of the tumor immune microenvironment. Moreover, our findings reveal that the Fab form of h7B7-15S has the ability to deplete Tregs, independent of the Fc region. Taken together, our studies expand the application of anti-CD25 in tumor immunotherapy and provide insight into the underlying mechanism.
Assuntos
Anticorpos Monoclonais , Neoplasias , Camundongos , Animais , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Imunossupressores , Linfócitos T Reguladores , Microambiente TumoralRESUMO
BACKGROUND: Leukemia inhibitory factor (LIF) is a multifunctional member of the IL-6 cytokine family that activates downstream signaling pathways by binding to the heterodimer consisting of LIFR and gp130 on the cell surface. Previous research has shown that LIF is highly expressed in various tumor tissues (e.g. pancreatic cancer, breast cancer, prostate cancer, and colorectal cancer) and promotes cancer cell proliferation, migration, invasion, and differentiation. Moreover, the overexpression of LIF correlates with poor clinicopathological characteristics. Therefore, we hypothesized that LIF could be a promising target for the treatment of cancer. In this work, we developed the antagonist antibody 1G11 against LIF and investigated its anti-tumor mechanism and its therapeutic efficacy in mouse models. RESULTS: A series of single-chain variable fragments (scFvs) targeting LIF were screened from a naive human scFv phage library. These scFvs were reconstructed in complete IgG form and produced by the mammalian transient expression system. Among the antibodies, 1G11 exhibited the excellent binding activity to human, cynomolgus monkey and mouse LIF. Functional analysis demonstrated 1G11 could block LIF binding to LIFR and inhibit the intracellular STAT3 phosphorylation signal. Interestingly, 1G11 did not block LIF binding to gp130, another LIF receptor that is involved in forming the receptor complex together with LIFR. In vivo, intraperitoneal administration of 1G11 inhibited tumor growth in CT26 and MC38 models of colorectal cancer. IHC analysis demonstrated that p-STAT3 and Ki67 were decreased in tumor tissue, while c-caspase 3 was increased. Furthermore, 1G11 treatment improves CD3+, CD4 + and CD8 + T cell infiltration in tumor tissue. CONCLUSIONS: We developed antagonist antibodies targeting LIF/LIFR signaling pathway from a naive human scFv phage library. Antagonist anti-LIF antibody exerts antitumor effects by specifically reducing p-STAT3. Further studies revealed that anti-LIF antibody 1G11 increased immune cell infiltration in tumor tissues.
Assuntos
Fator Inibidor de Leucemia , Anticorpos de Cadeia Única , Animais , Humanos , Anticorpos de Cadeia Única/imunologia , Anticorpos de Cadeia Única/farmacologia , Camundongos , Fator Inibidor de Leucemia/imunologia , Fator Inibidor de Leucemia/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Fator de Transcrição STAT3/metabolismo , Fator de Transcrição STAT3/imunologia , Receptor gp130 de Citocina/imunologia , Receptor gp130 de Citocina/metabolismo , Receptor gp130 de Citocina/antagonistas & inibidores , Biblioteca de Peptídeos , Transdução de Sinais , Feminino , Macaca fascicularis , Camundongos Endogâmicos BALB C , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
5T4, a membrane protein, is overexpressed in many tumor tissues but rarely expressed in normal tissues. Here, CHO-5T4+ cells were generated and served as the antigen to immunize mice. Hybridoma techniques were employed to produce monoclonal antibodies (mAbs). The recombinant protein of human IgG Fc-fused extracellular domain of 5T4 (5T4 ECD-Fc) was obtained from transient expression in HEK293F cells. The fusion protein 5T4 ECD-Fc and CHO-5T4+ cells were respectively utilized to screen anti-5T4 antibodies that could bind to the native antigen. In preliminary screening, three hundred and fifty mAbs were obtained. Via surface plasmon resonance and flow cytometry screening, seven anti-5T4 mAbs stood out. Among them, H6 showed a high affinity (KDâ¯=â¯1.6â¯×â¯10-11â¯M) and internalization percentage (36% for 1â¯h and 80% for 4â¯h). The molecular weight and isoelectric point of H6 were determined by LC-MS and iCIEF. Moreover, the specific reactivity of H6 was demonstrated by western blotting, flow cytometry, and immunohistochemistry, respectively. In conclusion, we produced human recombinant protein of 5T4 extracellular domain and developed high-affinity internalizing monoclonal antibodies which may be applied in the 5T4-targeting ADC therapy and basic research.
Assuntos
Anticorpos Monoclonais Murinos , Afinidade de Anticorpos , Antineoplásicos Imunológicos , Fragmentos Fc das Imunoglobulinas , Glicoproteínas de Membrana , Proteínas Recombinantes de Fusão , Animais , Anticorpos Monoclonais Murinos/química , Anticorpos Monoclonais Murinos/imunologia , Antineoplásicos Imunológicos/química , Antineoplásicos Imunológicos/imunologia , Células CHO , Cricetulus , Feminino , Humanos , Fragmentos Fc das Imunoglobulinas/biossíntese , Fragmentos Fc das Imunoglobulinas/química , Fragmentos Fc das Imunoglobulinas/imunologia , Fragmentos Fc das Imunoglobulinas/isolamento & purificação , Glicoproteínas de Membrana/biossíntese , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/isolamento & purificação , Camundongos , Camundongos Endogâmicos BALB C , Domínios Proteicos , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/isolamento & purificaçãoRESUMO
Human cystatin C (HCC), encoded by cystatin 3 gene, is a 13.3kDa endogenous cysteine proteinase inhibitor and an important biomarker of renal function. However, expressing recombinant cystatin C is difficult because of low yield and inclusion bodies in Escherichia coli (E. coli). In this study, we cloned HCC gene into pET-22b vector containing PelB leader signal sequence, which could direct the protein to the bacterial periplasm. Large amounts of soluble HCC could be efficiently expressed in the bacterial periplasm at 16°C with 0.1mM IPTG induction. The recombinant HCC was isolated in high purity by cation exchange chromatography and gel filtration chromatography. Furthermore, the HCC was characterized by circular dichroism (CD) and dynamic light scattering (DLS), and displayed biological activity against papain. Here, we provide a method to produce large amounts of soluble mature HCC in E. coli.
Assuntos
Cistatina C/biossíntese , Cistatina C/química , Cistatina C/isolamento & purificação , Escherichia coli/metabolismo , Expressão Gênica , Periplasma/metabolismo , Cistatina C/genética , Escherichia coli/genética , Humanos , Periplasma/genética , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificaçãoRESUMO
OBJECTIVE: Ovarian cancer is 1 kind of a highly malignant gynecologic tumor, and current treatments have not achieved satisfactory effects. Human epidermal growth factor receptor 2 (HER2)-targeted therapies including trastuzumab and trastuzumab-DM1 (T-DM1) (antibody-cytotoxic drug conjugates) have been applied to treat HER2-overexpressing breast cancers in clinic. In the present study, we explored whether T-DM1 could effectively treat HER2-positive human ovarian carcinoma in vitro and in vivo. METHODS: HER2 expressions of 6 ovarian cancer cell lines and 2 breast carcinoma cell lines were validated, and the binding capacity of T-DM1 to HER2-positive ovarian cancer SKOV3 cells were analyzed by flow cytometry. Nude mice bearing intraperitoneal and subcutaneous SKOV3 xenografts were used to investigate the antitumor effect of T-DM1. RESULTS: High HER2 expressions in SKOV3 cell lines were detected. The binding capacity of T-DM1 to HER2-positive SKOV3 cells was in a similar manner comparing with trastuzumab. In vitro, T-DM1 showed strong growth inhibitory on SKOV3 cells, with IC50 values of 0.15 nmol/L. Nude mice bearing intraperitoneal and subcutaneous SKOV3 xenografts were used to investigate the antitumor effects of T-DM1 in vivo. In subcutaneous xenografts model, T-DM1 (30 mg/kg and 10 mg/kg) indicated significant anticancer effects. It is noteworthy that tumors were completely eradicated in the T-DM1 (30 mg/kg) group, and no regrowth was observed in a long time after the termination of the treatment. In the peritoneal xenograft model, tumor nodules in 3 of 7 mice were hardly observed in the abdominal cavity of mice after intraperitoneal injection of T-DM1 (30 mg/kg). At the same time, tumor nodules from the other 4 mice weighed on the average of only 0.07 g versus 1.77 g in control group. CONCLUSIONS: Our data showed that T-DM1 possessed promising antitumor effects on HER2-overexpressing ovarian cancer in mouse model, which provided valuable references for the future clinical trials.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Proliferação de Células/efeitos dos fármacos , Maitansina/análogos & derivados , Neoplasias Epiteliais e Glandulares/tratamento farmacológico , Neoplasias Ovarianas/tratamento farmacológico , Ado-Trastuzumab Emtansina , Animais , Anticorpos Monoclonais Humanizados/farmacologia , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Carcinoma Epitelial do Ovário , Linhagem Celular Tumoral , Feminino , Genes erbB-2 , Humanos , Células MCF-7 , Maitansina/farmacologia , Maitansina/uso terapêutico , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Epiteliais e Glandulares/genética , Neoplasias Epiteliais e Glandulares/patologia , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/patologia , Trastuzumab , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Testis-expressed sequence 101 (TEX101) was found to be highly expressed in testis and involved in acrosome reaction in previous studies. Recently, the metastasis suppressor function of TEX101 in cancer was disclosed, but the comprehensive investigation of its expression has rarely been reported. In this study, the expression features of TEX101 in normal human organs and seminoma were systematically analyzed. RESULTS: Immunohistochemistry demonstrated intense staining of TEX101 in human testis tissues; however, its expression in 27 other types of normal human organs, including the ovary, was negligible. Higher expression of TEX101 was observed in the spermatocytes and spermatids of the testis, but relatively lower staining was detected in spermatogonia. Western blotting showed a single TEX101 band of 38 kDa in human testis, but it did not correspond to the predicted molecular weight of its mature form at 21 KDa. Furthermore, we examined seminoma tissues by immunohistochemistry and found that none of the 36 samples expressed TEX101. CONCLUSIONS: Our data confirmed TEX101 to be a testis protein that could be related to the maturation process of male germ cells. The lack of TEX101 in seminoma indicated its potential role in tumor progression. This characteristic expression of TEX101 could provide a valuable reference for understanding its biological functions.
Assuntos
Proteínas de Membrana/metabolismo , Epitélio Seminífero/metabolismo , Seminoma/metabolismo , Neoplasias Testiculares/metabolismo , Western Blotting , Diferenciação Celular , Epitélio/metabolismo , Feminino , Trato Gastrointestinal/metabolismo , Humanos , Imuno-Histoquímica , Tecido Linfoide/metabolismo , Masculino , Tecido Nervoso/metabolismo , Especificidade de Órgãos/fisiologia , Ovário/metabolismo , Epitélio Seminífero/patologia , Seminoma/patologia , Maturação do Esperma/fisiologia , Espermatozoides/crescimento & desenvolvimento , Neoplasias Testiculares/patologia , Testículo/metabolismo , Testículo/patologiaRESUMO
Treatment for cancer can induce a series of secreted factors into the tumor microenvironment, which can affect cancer progression. Wingless-type MMTV (mouse mammary tumor virus) integration site 16B (WNT16B) is a new member of the WNT family and has been reported to play growth-related roles in previous studies. In this study, we found WNT16B could be expressed and secreted into the microenvironment by human ovarian fibroblasts after DNA damage-associated treatment, including chemotherapy drugs and radiation. We also demonstrated that fibroblast-derived WNT16B could result in accumulation of ß-catenin in dendritic cells and secretion of interleukin-10 (IL-10) and transforming growth factor beta (TGF-ß), which contributed to the differentiation of regulatory T cells in a co-culture environment. These results shed light on the roles of WNT16B in immune regulation, especially in regard to cancer treatment.
Assuntos
Diferenciação Celular , Células Dendríticas/metabolismo , Fibroblastos/metabolismo , Neoplasias Ovarianas/metabolismo , Linfócitos T Reguladores/metabolismo , Proteínas Wnt/metabolismo , beta Catenina/metabolismo , Antígenos de Diferenciação de Linfócitos T/genética , Antígenos de Diferenciação de Linfócitos T/metabolismo , Células Cultivadas , Feminino , Humanos , Interleucina-10/genética , Interleucina-10/metabolismo , Ovário/citologia , Linfócitos T Reguladores/citologia , Fator de Crescimento Transformador beta/metabolismo , Microambiente Tumoral , Proteínas Wnt/genética , beta Catenina/genéticaRESUMO
7-Ethyl-10-hydroxycamptothecin (SN38), the active metabolite of irinotecan, exerts a 100-fold to 1000-fold higher effect than irinotecan itself against several tumor cell lines. However, the water insolubility of SN38 has prevented its direct use as an antitumor drug in the clinic. To improve the water solubility and antitumor efficacy, SN38 was covalently attached to the only free sulfhydryl at cysteine-34 on the BSA site specifically through a thiol-binding linker to form a prodrug BSA-SN38 conjugate (BSA : SN38=1 : 1). The water solubility of this conjugate was similar to albumin using the current method. Also, SN38 loading in this conjugate became controllable. Size-exclusion chromatography purification and UV characterization of the SDS-PAGE electrophoresis product were carried out. Then, an MTT assay was carried out to test the antitumor effect of this conjugate on five colon cancer cell lines in vitro. The 72 h IC50 values of the BSA-SN38 conjugate ranged from 1.5 to 6.1 µmol/l. A colorectal peritoneal carcinomatosis model in mice was established to determine the intraperitoneal chemotherapy effect of the BSA-SN38 conjugate. The BSA-SN38 conjugate at an SN38 equivalent dose of 10 mg/kg/day was administrated every 4 days. Eighteen days after manipulation, the mice were euthanized and the tumors in the abdominal cavity were collected and weighed. Tumors in the BSA-SN38 conjugate treatment group (m=0.21 ± 0.15 g) were found to be significantly (P=5) lighter than those in the NS control group (m=4.74±0.73 g). The results indicated that this water-soluble BSA-SN38 conjugate exerted a strong antitumor effect on colorectal carcinoma.
Assuntos
Antineoplásicos Fitogênicos/síntese química , Antineoplásicos Fitogênicos/farmacologia , Camptotecina/análogos & derivados , Soroalbumina Bovina/química , Animais , Camptotecina/química , Linhagem Celular Tumoral , Neoplasias Colorretais/tratamento farmacológico , Ensaios de Seleção de Medicamentos Antitumorais/métodos , Feminino , Células HCT116 , Humanos , Irinotecano , Camundongos , Camundongos Endogâmicos BALB C , Neoplasias Peritoneais/tratamento farmacológico , Solubilidade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Antibodyâdrug conjugates (ADCs), which combine the advantages of monoclonal antibodies with precise targeting and payloads with efficient killing, show great clinical therapeutic value. The ADCs' payloads play a key role in determining the efficacy of ADC drugs and thus have attracted great attention in the field. An ideal ADC payload should possess sufficient toxicity, low immunogenicity, high stability, and modifiable functional groups. Common ADC payloads include tubulin inhibitors and DNA damaging agents, with tubulin inhibitors accounting for more than half of the ADC drugs in clinical development. However, due to clinical limitations of traditional ADC payloads, such as inadequate efficacy and the development of acquired drug resistance, novel highly efficient payloads with diverse targets and reduced side effects are being developed. This perspective summarizes the recent research advances of traditional and novel ADC payloads with main focuses on the structure-activity relationship studies, co-crystal structures, and designing strategies, and further discusses the future research directions of ADC payloads. This review also aims to provide valuable references and future directions for the development of novel ADC payloads that will have high efficacy, low toxicity, adequate stability, and abilities to overcome drug resistance.
RESUMO
The CD40 receptor is a member of the tumour necrosis factor receptor family and is widely expressed on various cell types. The antitumour activity of CD40 agonist antibody has been observed in B-cell-derived malignancies, but its activity on ovarian cancer remains unclear. However, in this paper, we first confirmed that the anti-CD40 agonist antibody could inhibit the growth of ovarian cancer cells and induce apoptosis. This study investigated the expression of CD40 by ovarian carcinoma tissues and cell lines, at the same time, we evaluated the effect of a recombinant soluble human CD40L (rshCD40L) and an anti-CD40 agonist antibody on cell growth and apoptosis. Flow cytometry and immunohistochemistry assay demonstrated that CD40 was expressed on ovarian carcinoma cell lines and primary ovarian carcinoma cells derived from ascites, as well as on ovarian carcinoma tissues. The growth inhibition of rshCD40L and the anti-CD40 agonist antibody on ovarian carcinoma cells was examined by MTT assay, and the proportion of apoptotic tumour cells was analysed by flow cytometry and Hoechst staining. Our study showed that CD40 was expressed on all ovarian carcinoma cell lines and was examined in 86.2% (162/188) of ovarian cancer tissue samples, but not in normal ovarian tissues (n = 20). Treatment with rshCD40L or anti-CD40 agonist antibody significantly inhibited ovarian carcinoma cell growth and induced apoptosis. Theses results suggest that CD40 is expressed on ovarian carcinoma cells, moreover, that rshCD40L and anti-CD40 agonist antibody have therapeutic potential to inhibit human ovarian cancer growth.
Assuntos
Anticorpos Monoclonais Humanizados/uso terapêutico , Antígenos CD40/agonistas , Antígenos CD40/biossíntese , Carcinoma/tratamento farmacológico , Neoplasias Ovarianas/tratamento farmacológico , Adulto , Idoso , Anticorpos Monoclonais Humanizados/imunologia , Apoptose/efeitos dos fármacos , Apoptose/imunologia , Antígenos CD40/imunologia , Carcinoma/imunologia , Linhagem Celular Tumoral , Feminino , Humanos , Pessoa de Meia-Idade , Neoplasias Ovarianas/imunologia , Adulto JovemRESUMO
Penicillium marneffei is a dimorphic fungus that responds to changes in temperature. We performed a comparative analysis of gene expression in mycelial- and yeast-phase P. marneffei cells using high-throughput DNA microarrays. A total of 1,884 differentially expressed genes with annotations in the gene ontology (GO) database were identified between P. marneffei mycelial and yeast cells. These differentially expressed genes mainly belong to 18 categories in the organism's ontology, including reproduction, immunity, metabolism, signaling, etc. Bioinformatics suggests that these differentially expressed genes may help explain the resistance to adverse environments and the virulence of P. marneffei. Nine genes from the results of the DNA microarray experiment were selected for further analysis with real-time quantitative PCR to validate the differential expression. Our data provide a global description of the transcriptional response accompanying adaptation to high temperature in yeast.
Assuntos
Perfilação da Expressão Gênica , Micélio/genética , Penicillium/genética , Biologia Computacional , Análise em Microsséries , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase em Tempo RealRESUMO
The use of a bi-specific antibody (BsAb) is an attractive and specific approach to cancer therapy. We have constructed a fully human recombinant single chain Fv BsAb against CD19 and CD3 that was an effective treatment in an animal model of non-Hodgkin's lymphoma (NHL). The CD19/CD3 BsAb was expressed in CHO cells and purified by Ni-column chromatography. Flow cytometry revealed that the CD19/CD3 BsAb specifically bound to both CD19 and CD3-positive cells. In vitro, the CD19/CD3 BsAb could stimulate T cell proliferation and induce the lysis of cultured Raji cells in the presence of unstimulated T lymphocytes. In vivo, the CD19/CD3 BsAb efficiently inhibited tumour growth in SCID mice of NHL, and the survival time of the mice was significantly prolonged. Therefore, our CD19/CD3 BsAb is a useful tool that could be a suitable candidate for treatment of NHL.
Assuntos
Anticorpos Biespecíficos/imunologia , Antígenos CD19/imunologia , Complexo CD3/imunologia , Linfoma não Hodgkin/imunologia , Linfócitos T Citotóxicos/imunologia , Animais , Anticorpos Biespecíficos/genética , Anticorpos Biespecíficos/uso terapêutico , Células CHO , Proliferação de Células , Cromatografia de Afinidade , Cricetinae , Modelos Animais de Doenças , Humanos , Imunoterapia/métodos , Camundongos , Camundongos SCID , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/uso terapêutico , Resultado do TratamentoRESUMO
A novel series of hybrid molecules combining pyrrolobenzodiazepine (PBD) and anthracenecarboxyimide pharmacophores were designed, synthesized, and tested for in vitro cytotoxicity against various cancer cell lines. The most potent compound from this series, 37b3, exhibited a subnanomolar level of cytotoxicity with an IC50 of 0.17-0.94 nM. 37b3 induced DNA damage and led to tumor cell cycle arrest and apoptosis. We employed 37b3 as a payload to conjugate with trastuzumab to obtain the antibody-drug conjugate (ADC) T-PBA. T-PBA maintained its mode of target and internalization ability of trastuzumab. We demonstrated that T-PBA could be degraded through the lysosomal pathway to release the payload 37b3 after internalization. T-PBA showed a powerful killing effect on Her2-positive cancer cells in vitro. Furthermore, T-PBA significantly inhibited tumor growth in gastric and ovarian cancer xenograft mouse models without overt toxicity. Collectively, these studies suggest that T-PBA represents a promising new ADC that deserves further investigation.
Assuntos
Imunoconjugados , Animais , Benzodiazepinas , Linhagem Celular Tumoral , Humanos , Imunoconjugados/farmacologia , Imunoconjugados/uso terapêutico , Camundongos , Pirróis/farmacologia , Receptor ErbB-2/genética , Trastuzumab , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Anti-CD38 monoclonal antibodies (mAbs), daratumumab, and isatuximab have represented a breakthrough in the treatment of multiple myeloma (MM). Recently, CD38-based mAbs were expected to achieve increasing potential beyond MM, which encouraged us to develop new anti-CD38 mAbs to meet clinical needs. In this study, we developed a novel humanized anti-CD38 antibody, FTL004, which exhibited enhanced pro-apoptotic ability and negligible binding to red blood cells (RBCs). FTL004 presented a better ability to induce direct apoptosis independent of Fc-mediated cross-linking against lymphoma and MM cell lines as well as primary myeloma cells derived from MM patients. For instance, FTL004 induced RPMI 8226 cells with 55% early apoptosis cells compared with 20% in the isatuximab-treated group. Of interest, FTL004 showed ignorable binding to CD38 on human RBCs in contrast to tumor cells, even at concentrations up to 30 µg/mL. Furthermore, with an engineered Fc domain, FTL004 displayed stronger antibody-dependent cellular cytotoxicity (ADCC) against CD38+ malignant cells. In vivo MM and non-Hodgkin lymphoma tumor xenograft models showed that FTL004 possessed an effective anti-tumor effect. Cryo-electron microscopy structure resolved two epitope centers of FTL004 on CD38: one of which was unique while the other partly overlapped with that of isatuximab. Taken together, FTL004 distinguishes it from other CD38 targeting mAbs and represents a potential candidate for the treatment of MM and non-Hodgkin lymphoma.
Assuntos
Antineoplásicos , Linfoma não Hodgkin , Mieloma Múltiplo , Humanos , Mieloma Múltiplo/patologia , Microscopia Crioeletrônica , ADP-Ribosil Ciclase 1 , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linfoma não Hodgkin/tratamento farmacológico , Eritrócitos/patologiaRESUMO
Multiple myelomas (MMs) are etiologically heterogeneous and there are limited treatment options; indeed, current monoclonal antibody therapies have had limited success, so more effective antibodies are urgently needed. Polyclonal antibodies are a possible alternative because they target multiple antigens simultaneously. In this study, we produced polyclonal rabbit anti-murine plasmacytoma cell immunoglobulin (PAb) by immunizing rabbits with the murine plasmacytoma cell line MPC-11. The isolated PAb bound to plasma surface antigens in several MM cell lines, inhibited their proliferation as revealed by MTT assay, and induce apoptosis as indicated by flow cytometry, microscopic observation of apoptotic changes in morphology, and DNA fragmentation on agarose gels. The cytotoxicity of PAb on MPC-11 cell lines was both dose-dependent and time-dependent; PAb exerted a 50% inhibitory effect on MPC-11 cell viability at a concentration of 200 µg/ml in 48 h. Flow cytometry demonstrated that PAb treatment significantly increased the number of apoptotic cells (48.1%) compared with control IgG (8.3%). Apoptosis triggered by PAb was confirmed by activation of caspase-3, -8, and -9. Serial intravenous or intraperitoneal injections of PAb inhibited tumour growth and prolonged survival in mice bearing murine plasmacytoma, while TUNEL assay demonstrated that PAb induced statistically significant apoptosis (P < 0.05) compared to control treatments. We conclude that PAb is an effective agent for in vitro and in vivo induction of apoptosis in multiple myeloma and that exploratory clinical trials may be warranted.
Assuntos
Apoptose/efeitos dos fármacos , Imunoglobulinas/farmacologia , Mieloma Múltiplo/patologia , Plasmocitoma/metabolismo , Animais , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Marcação In Situ das Extremidades Cortadas , Camundongos , Camundongos Endogâmicos BALB C , Ligação Proteica/efeitos dos fármacos , CoelhosRESUMO
Hepatitis B virus (HBV) infection is the leading cause of liver cirrhosis and hepatocellular carcinoma worldwide. In recent decades, significant progress toward understanding the molecular virology and pathogenesis of HBV infection has been made. In addition, multiple treatment modalities have been developed for persons with HBV infection. In the present study, we demonstrated that IL-4 inhibits the expression of hepatitis B surface antigen and hepatitis B e antigen in a HBV stably transfected hepatocellular carcinoma cell line (HepG2.2.15). To reveal the anti-HBV mechanism of IL-4 by proteomics, 2-DE and MS technology were utilized to profile global changes in protein expression in HepG2.2.15 cells after IL-4 treatment. A total of 56 differentially expressed proteins were identified in IL-4-treated HepG2.2.15 cells. To find out the interaction of these changed proteins by bioinformatics, signaling network analysis with the STRING tool showed that the identified proteins are primarily involved in transcription and proteolysis. Taken together, these results offer valuable clues for understanding the molecular mechanisms of the IL-4-mediated anti-HBV response.
Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/virologia , Vírus da Hepatite B/imunologia , Interleucina-4/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/virologia , Western Blotting , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Análise por Conglomerados , Eletroforese em Gel Bidimensional , Hepatite B/tratamento farmacológico , Hepatite B/imunologia , Antígenos de Superfície da Hepatite B/biossíntese , Antígenos E da Hepatite B/biossíntese , Interações Hospedeiro-Patógeno , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Proteômica/métodos , Reprodutibilidade dos Testes , Transdução de Sinais , Espectrometria de Massas em TandemRESUMO
Small cell lung cancer (SCLC) is an aggressive malignant tumour which accounts for approximately 13-15% of all newly diagnosed lung cancer cases. To date, platinum-based chemotherapy are still the first-line treatments for SCLC. However, chemotherapy resistance and systemic toxicity limit the long-term clinical outcome of first-line treatment in SCLC. Recent years, targeted therapy and immunotherapy have made great breakthrough in cancer therapy, and researchers aim to exploit both as a single agent or in combination with chemotherapy to improve the survival of SCLC patients, but limited effectiveness and the adverse events remain the major obstacles in the treatment of SCLC. To overcome these challenges for SCLC therapies, prevention and early diagnosis for this refractory disease is very important. At the same time, we should reveal more information about the pathogenesis of SCLC and the mechanism of drug resistance. Finally, new treatment strategies should also be taken into considerations, such as repurposing drug, optimising of targets, combination therapy strategies or prognostic biomarkers to enhance therapeutic effects and decrease the adverse events rates in SCLC patients. This article will review the molecular biology characteristics of SCLC and discuss the opportunities and obstacles of the current therapy for SCLC patients.
Assuntos
Imunoterapia/métodos , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/terapia , Terapia de Alvo Molecular/métodos , Carcinoma de Pequenas Células do Pulmão/imunologia , Carcinoma de Pequenas Células do Pulmão/terapia , Ensaios Clínicos como Assunto/métodos , Humanos , Inibidores de Checkpoint Imunológico/uso terapêuticoRESUMO
Immune checkpoint inhibitors have achieved unprecedented success in cancer immunotherapy. However, the overall response rate to immune checkpoint inhibitor therapy for many cancers is only between 20 and 40%, and even less for colorectal cancer (CRC) patients. Thus, there is an urgent need to develop an efficient immunotherapeutic strategy for CRC. Here, we developed a novel CRC combination therapy consisting of a multiple receptor tyrosine kinase inhibitor (Foretinib) and anti-PD-1 antibody. The combination therapy significantly inhibited tumor growth in mice, led to improved tumor regression without relapse (83% for CT26 tumors and 50% for MC38 tumors) and prolonged overall survival. Mechanistically, Foretinib caused increased levels of PD-L1 via activating the JAK2-STAT1 pathway, which could improve the effectiveness of the immune checkpoint inhibitor. Moreover, the combination therapy remodeled the tumor microenvironment and enhanced anti-tumor immunity by further increasing the infiltration and improving the function of T cells, decreasing the percentage of tumor-associated macrophages (TAMs) and inhibiting their polarization toward the M2 phenotype. Furthermore, the combination therapy inhibited the metastasis of CT26-Luc tumors to the lung in BALB/c mouse by reducing proportions of regulatory T-cells, TAMs and M2 phenotype TAMs in their lungs. This study suggests that a novel combination therapy utilizing both Foretinib and anti-PD-1 antibody could be an effective combination strategy for CRC immunotherapy.
RESUMO
Hematopoiesis in bone marrow declines during aging owing to alteration of the hematopoietic niche. However, due to difficult accessibility and other complexities, senescence-related alteration of the hematopoietic niche is largely unknown. The interstitial fluid of bone marrow (IFBM), a pivotal component of the hematopoietic niche, includes soluble secretory factors that are present between bone marrow cells. To characterize the proteomic profile changes of IFBM during aging, we analyzed the IFBMs of young, adult, and senescent rats using 2-DE combined with ESI/MALDI-Q-TOF MS. Finally, 31 differentially expressed proteins involved in multiple biological functions were identified. Peroxiredoxin 2 (Prx2), down-regulated during aging, was further analyzed and demonstrated that it is produced by bone marrow stromal cells. Interestingly, higher levels of hydrogen peroxide (H(2)O(2)) were detected in the bone marrow with lower Prx2 expression. Moreover, exogenous Prx2 reduced the intracellular H(2)O(2) level in bone marrow stromal cells in vitro. Therefore, Prx2 is implied in the regulation of H(2)O(2) production in the bone marrow during aging. Our data characterized the dynamic protein profiles of the bone marrow microenvironment during aging and we provided clues to elucidate the mechanism of creating a low ROS level in the hematopoietic niche.
Assuntos
Envelhecimento/metabolismo , Medula Óssea/metabolismo , Líquido Extracelular/metabolismo , Regulação da Expressão Gênica/fisiologia , Peróxido de Hidrogênio/metabolismo , Peroxirredoxinas/metabolismo , Proteômica/métodos , Envelhecimento/fisiologia , Animais , Western Blotting , Eletroforese em Gel Bidimensional , Espectrometria de Massas , Ratos , Ratos Sprague-Dawley , Células Estromais/metabolismoRESUMO
Apoptosis plays an important role in embryonic development. PNAS-4 has been demonstrated to induce apoptosis in several cancer cells. In this study, we cloned Xenopus laevis PNAS-4 (xPNAS-4), which is homologous to the human PNAS-4 gene. Bioinformatics analysis for PNAS-4 indicated that xPNAS-4 shared 87.6% identity with human PNAS-4 and 85.5% with mouse PNAS-4. The phylogenetic tree of PNAS-4 protein was also summarized. An analysis of cellular localization using an EGFP-fused protein demonstrated that xPNAS-4 was localized in the perinuclear region of the cytoplasm. RT-PCR analysis revealed that xPNAS-4, as a maternally expressed gene, was present in all stages of early embryo development. Whole-mount in situ hybridization showed that xPNAS-4 was mainly expressed in ectoderm and mesoderm. Furthermore, microinjection of xPNAS-4 mRNA in vivo caused developmental defects manifesting as a small eye phenotype in the Xenopous embryos, and as a small eye or one-eye phenotype in developing zebrafish embryos. In addition, embryos microinjected with xPNAS-4 antisense morpholino oligonucleotides (MOs) exhibited a failure of head development and shortened axis.