Anti-OX40 Antibody Combined with HBc VLPs Delays Tumor Growth in a Mouse Colon Cancer Model

Objective Combination immunotherapy strategies targeting OX40,a co-stimulatory molecule that can enhance antitumor immunity by modulating the proliferation,differentiation,and effector function of tumor-infiltrating T cells,have attracted much attention for their excellent therapeutic effects.In this study,we aimed to evaluate the antitumor efficacy of combined anti-OX40 and hepatitis B core viruslike particles(HBc VLPs)therapy using a mouse colon cancer model.Methods Humanized B-h OX40 mice were injected subcutaneously with MC38 colon tumor cells and treated with HBc VLPs+anti-h OX40 antibody.Tumor growth was monitored.Flow cytometric analysis was performed to evaluate the populations of T cell subsets in the tumors.Results The combination of anti-OX40 with HBc VLPs resulted in a significant delay in tumor growth,suggesting that a potent antitumor immunity was induced by the combination therapy.Further studies revealed that HBc VLPs+anti-OX40 treatment induced a significant increase in effector T cells(Teffs)and a significant decrease in regulatory T cells(Tregs)in the tumor microenvironment(TME),which accounted for the synergistic antitumor effect of anti-OX40 in combination with HBc VLPs.Conclusion Combination therapy of anti-h OX40 and HBc VLPs provides synergistic antitumor activity in colon cancer-bearing mice,which may represent a potential design strategy for cancer immunotherapy.

Phase I study of chimeric anti-CD20 monoclonal antibody in Chinese patients with CD20-positive non-Hodgkin＇s lymphoma

Objective： This study was designed to determine the safety, pharmacokinetics and biologic effects of a humanmouse chimeric anti-CD20 monoclonal antibody （SCT400） in Chinese padents with CD20-positive B-cell non- Hodgkin＇s lymphoma （CD20 B-cell NHL）. SCT400 has an identical amino acid sequence as rituximab, with the exception of one amino acid in the CH1 domain of the heavy chain, which is common in Asians. Methods： Fifteen patients with CD20＋ B-cell NHL received dose-escalating SCT400 infusions （250 mg/m2： n=3; 375 mg/m2： n=9; 500 mg/m2： n=3） once weekly for 4 consecutive weeks with a 24-week follow-up period. The data of all patients were collected for pharmacoklnetics and pharmacodynamics analyses. Results： No dose-limiting toxicities were observed. Most drug-related adverse events were grade 1 or 2. Two patients had grade 3 or 4 ncutropenia. Under premedication, the drug-related infusion reaction was mild. A rapid, profound and durable depletion of circulating B cells was observed in all dose groups without significant effects on T cell count, natural killer （NK） cell count or immunoglobulin levels. No patient developed anti- SCT400 antibodies during the course of the study. SCT400 serum half-life （Tin）, maximum concentration （Cmax and area under the curve （AUC） generally increased between the first and fourth infusions （P〈0.05）. At the 375 mg/m2 dose, the T1/2 was 122.5±46.7 h vs. 197.0,75.0 11, respectively, and the Cmax was 200.6±20.2 pg/mL vs. 339.1±71.0 ng/mL, respectively. From 250 mg/m2 to 500 mg/m2, the Cmax and AUC increased significantly in a dose-dependent manner （P〈0.05）. Patients with a high tumor burden had markedly lower serum SCT400 concenmations compared with those without or with a low tumor burden. Of the 9 assessable patients, 1 achieved complete response and 2 achieved partial responses. Conclusions; SCT400 is well-tolerated and has encouraging preliminary efficacy in Chinese patients with CD20＋ B-cell NHL.

Time-Programmed Delivery of Sorafenib and Anti-CD47 Antibody via a Double-Layer-Gel Matrix for Postsurgical Treatment of Breast Cancer

The highly immunosuppressive microenvironment after surgery has a crucial impact on the recurrence and metastasis in breast cancer patients.Programmable delivery of immunotherapy-involving combinations through a single drug delivery system is highly promising,yet greatly challenging,to reverse postoperative immunosuppression.Here,an injectable hierarchical gel matrix,composed of dual lipid gel(DLG)layers with different soybean phosphatidylcholine/glycerol dioleate mass ratios,was developed to achieve the time-programmed sequential delivery of combined cancer immunotherapy.The outer layer of the DLG matrix was thermally responsive and loaded with sorafenib-adsorbed graphene oxide(GO)nanoparticles.GO under manually controlled near-infrared irradiation generated mild heat and provoked the release of sorafenib first to reeducate tumor-associated macrophages(TAMs)and promote an immunogenic tumor microenvironment.The inner layer,loaded with anti-CD47 antibody(aCD47),could maintain the gel state for a much longer time,enabling the sustained release of aCD47 afterward to block the CD47-signal regulatory proteinα(SIRPα)pathway for a long-term antitumor effect.In vivo studies on 4T1 tumor-bearing mouse model demonstrated that the DLG-based strategy efficiently prevented tumor recurrence and metastasis by locally reversing the immunosuppression and synergistically blocking the CD47-dependent immune escape,thereby boosting the systemic immune responses.

Veterans health administration hepatitis B testing and treatment with anti-CD20 antibody administration

AIM: To evaluate pretreatment hepatitis B virus (HBV) testing, vaccination, and antiviral treatment rates in Veterans Affairs patients receiving anti-CD20 Ab for quality improvement.METHODS: We performed a retrospective cohort study using a national repository of Veterans Health Administration (VHA) electronic health record data. We identified all patients receiving anti-CD20 Ab treatment (2002-2014). We ascertained patient demographics, laboratory results, HBV vaccination status (from vaccination records), pharmacy data, and vital status. The high risk period for HBV reactivation is during anti-CD20 Ab treatment and 12 mo follow up. Therefore, we analyzed those who were followed to death or for at least 12 mo after completing anti-CD20 Ab. Pretreatment serologic tests were used to categorize chronic HBV (hepatitis B surface antigen positive or HBsAg+), past HBV (HBsAg-, hepatitis B core antibody positive or HBcAb+), resolved HBV (HBsAg-, HBcAb+, hepatitis B surface antibody positive or HBsAb+), likely prior vaccination (isolated HBsAb+), HBV negative (HBsAg-, HBcAb-), or unknown. Acute hepatitis B was defined by the appearance of HBsAg+ in the high risk period in patients who were pretreatment HBV negative. We assessed HBV antiviral treatment and the incidence of hepatitis, liver failure, and death during the high risk period. Cumulative hepatitis, liver failure, and death after anti-CD20 Ab initiation were compared by HBV disease categories and differences compared using the χ2 test. Mean time to hepatitis peak alanine aminotransferase, liver failure, and death relative to anti-CD20 Ab administration and follow-up were also compared by HBV disease group.RESULTS: Among 19304 VHA patients who received anti-CD20 Ab, 10224 (53%) had pretreatment HBsAg testing during the study period, with 49% and 43% tested for HBsAg and HBcAb, respectively within 6 mo pretreatment in 2014. Of those tested, 2% (167/10224) had chronic HBV, 4% (326/7903) past HBV, 5% (427/8110) resolved HBV, 8% (628/8110) likely prior HBV vaccination, and 76% (6022/7903) were HBV negative. In those with chronic HBV infection, ≤ 37% received HBV antiviral treatment during the high risk period while 21% to 23% of those with past or resolved HBV, respectively, received HBV antiviral treatment. During and 12 mo after anti-CD20 Ab, the rate of hepatitis was significantly greater in those HBV positive vs negative (P = 0.001). The mortality rate was 35%-40% in chronic or past hepatitis B and 26%-31% in hepatitis B negative. In those pretreatment HBV negative, 16 (0.3%) developed acute hepatitis B of 4947 tested during anti-CD20Ab treatment and follow-up.CONCLUSION: While HBV testing of Veterans has increased prior to anti-CD20 Ab, few HBV+ patients received HBV antivirals, suggesting electronic health record algorithms may enhance health outcomes.

Expression of Anti-CD4 Human/Murine Chimeric Antibody and Their Killer Tumor Activity

From the mouse hybridoma cell line secreting an anti-CD4 monoclonal antibody (McAb), total RNA was prepared. The VH and VL genes were amplified by RT-PCR with family specific primer pairs. The PCR products were cloned into pGEM-T vectors, then tranfected into JM109. The VH and VL genes were snalyzed by automatic DNA sequencer. According to Kabat classification, the VH and VL genes belong to the mouse ig heavy subgroup Ⅱ(A) and x chain subgroupⅢ, respectively. The VH and VL genes were subcloned into pr1-Expr and Pk Expr respectively, then transfected into XL2-Blue. The VH- Pr1 and VL- pk were trans feeted by electroporation into mouse myeloma cell X63Ag8. 653. The transfectoma cells were selected by G418 screening, and then supernatant of cultured transfectoma were analyzed by ELISA and immunofluorescence techniques.We have acquired transfectoma cells secreting anti-CD4 chimeric antibodies.These chimeric antibodies are able to kill tumor cells specifically in vitro.

Tumor neoangiogenesis detection by confocal laser endomicroscopy and anti-CD105 antibody:Pilot study

AIM: To evaluate neoangiogenesis in patients with colon cancer by two fluorescently labeled antibodies on fresh biopsy samples imaged with confocal laser endomicroscopy(CLE).METHODS: CLE is an imaging technique for gastrointestinal endoscopy providing in vivo microscopy at subcellular resolution.An important question in validating tumor angiogenesis is what proportion of the tumor vascular network is represented by preexisting parent tissue vessels and newly formed vessels.CD105(endoglin) represents a proliferation-associated endothelial cell adhesion molecule.In contrast to panendothelial markers,such as CD31,CD105 is preferentially expressed in activated endothelial cells that participate in neovascularization.Thus,we evaluated CD105 and CD31 expression from samples of ten patients with primary rectal adenocarcinoma,using a dedicated endomicroscopy system.A imaging software was used to obtain the Z projection of the confocal serial images from each biopsy sample previously combined into stacks.Vascular density and vessel diameters were measured within two 50 μm x 475 μm rectangular regions of interest centered in the middle of each image in the horizontal and vertical direction.The results were averaged over all the patients and were expressed as the mean ± SE.RESULTS: The use of an anti-CD105 antibody was found to be suitable for the detection of blood vessels in colon cancer.Whereas anti-CD31 antibodies stained blood vessels in both normal and pathologic colon equally,CD105 expression was observed primarily in malignant lesions,with little or no expression in the vessels of the normal mucosa(244.21 ± 130.7 vessels/mm3 in only four patients).The average diameter of antiCD105 stained vessels was 10.97 ± 0.6 μm in tumor tissue,and the vessel density was 2787.40 ± 134.8 vessels/mm3.When using the anti-CD31 antibody,the average diameter of vessels in the normal colon tissue was 7.67 ± 0.5 μm and the vessel density was 3191.60 ± 387.8 vessels/mm3,while in the tumors we obtained an average diameter of 10.88 ± 0.8 μm and a vessel density of 4707.30 ± 448.85 vessels/mm3.Thus,there were more vessels stained with CD31 than CD105(P < 0.05).The average vessel diameter was similar for both CD31 and CD105 staining.A qualitative comparison between CLE vs immunohistochemistry lead to similar results.CONCLUSION: Specific imaging and quantification of tumor microvessels are feasible in human rectal cancer using CLE examination and CD105 immunostaining of fresh tissue samples.

Inhibitory Effect of Anti-HER-2 Anti-CD3 Bi-specific Antibody on the Growth of Gastric Carcinoma

To evaluate the effect of anti-HER-2 × anti-CD3 bi-specific antibody（BsAb） on the growth of HER-2/neu-expressing human gastric carcinoma in vitro and in vivo, an MTT assay was carried out to test the inhibitive rates of herceptin, anti-CD3 and BsAb antibodies on SGC-7901 gastric carcinoma cells. Immunocytochemistry methods were used to test the HER-2 level of SGC-7901. Nude mice models were employed to test the effect of HER-2 CD3 BsAh combined with effector ceils（ peripheral blood lymphatic cells of healthy human beings） on the growth of tumors in animals. Compared with that of the untreated control group, the tumor cell growth rates in vitro and in vivo will both be significantly inhibited when treated with effector cells combined with anti-CD3 McAb, herceptin or HER2 CD3 BsAb （p 〈0. 05）, and the growth inhibition is the most remarkable in the group treated with HER2 CD3 BsAb combined with effector cells. The growth of tumor xenografts will also be significantly inhibited in the group treated with HER2 CD3 BsAb combined with effector cells when compared with that in the group treated with anti-CD3 McAb or the group treated with herceptin combined with effector cells（p 〈0. 05）. We can conclude that HER-2/neu is possibly a useful target for immunotherapy of gastric carcinoma, and anti-HER2 × anti-CD3 BsAb has evident anti-tumor efficacy both, in vitro and in vivo.

The expression of CD40 and CD40L on the surface of peripheral blood mononuclear cells in asthmatic rats and the effect of anti-CD40L McAb on Th1 and Th2 cytokines

Objective： To investigate the expression of CD40 and CD40 ligand （CD4OL） on the surface of peripheral blood mononuclear cells（PBMCs） in asthmatic rats and the effect of anti-CD40L McAb on cytokines of PBMCs. Methods： Flow cytometry and RT-PCR were used to detect the expression of CD40 and CD40L of PBMCs in asthmatic rats. After the PBMCs was treated with anti-CD40L McAb, ELISA was used to detect the IL-4 and IFN-γ levels of culture supernatants. Results： Compared with the normal control group, the expression of CD40 and CD40L of PBMCs in asthmatic rats increased （P 〈 0.05）. Compared with the untreated group, the level of IL-γ and the ratio of IL-4/IFN-γ decreased after the PBMCs was treated with anti-CD40L McAb（P 〈 0.05）. Conclusion： The expression of CD40 and CD40L on the surface of PBMCs in asthmatic rats was up-regulated. Anti-CD40L McAb can rectify the imbalance of Th1 and Th2 cytokines.

阿尔茨海默病β淀粉样肽40人单链抗体的生物活性

目的测定重组表达的抗β淀粉样肽(Aβ)40的人单链抗体的生物活性。方法通过基因重组表达方式获得抗Aβ40人单链抗体,应用常规ELISA和竞争性ELISA鉴定抗Aβ40人单链抗体的抗原结合活性,以人成神经细胞瘤SH-SY5Y细胞为模型检测抗Aβ40人单链抗体的神经元保护作用。结果重组表达的抗Aβ40的人单链抗体主要位于细菌不溶性内涵物中,经过尿素溶解和金属亲和层析纯化后具有结合Aβ40或Aβ42的活性。与Aβ42组相比,Aβ42加等摩尔抗体组可显著提高SH-SY5Y细胞的存活率(P<0.05)。与Aβ40组相比,Aβ40加等摩尔抗体组亦显著提高SH-SY5Y细胞的存活率(P<0.01)。结论从大肠杆菌系统中重组表达的抗Aβ40人单链抗体可以部分对抗β淀粉样肽的神经毒性。

Evaluation of teplizumab's efficacy and safety in treatment of type 1 diabetes mellitus:A systematic review and meta-analysis

BACKGROUND Islets of Langerhans beta cells diminish in autoimmune type 1 diabetes mellitus(T1DM).Teplizumab,a humanized anti-CD3 monoclonal antibody,may help T1DM.Its long-term implications on clinical T1DM development,safety,and efficacy are unknown.AIM To assess the effectiveness and safety of teplizumab as a therapeutic intervention for individuals with T1DM.METHODS A systematic search was conducted using four electronic databases(PubMed,Embase,Scopus,and Cochrane Library)to select publications published in peerreviewed journals written in English.The odds ratio(OR)and risk ratio(RR)were calculated,along with their 95%CI.We assessed heterogeneity using Cochrane Q and I2 statistics and the appropriate P value.RESULTS There were 8 randomized controlled trials(RCTs)in the current meta-analysis with a total of 1908 T1DM patients from diverse age cohorts,with 1361 patients receiving Teplizumab and 547 patients receiving a placebo.Teplizumab was found to have a substantial link with a decrease in insulin consumption,with an OR of 4.13(95%CI:1.72 to 9.90).Teplizumab is associated with an improved Cpeptide response(OR 2.49;95%CI:1.62 to 3.81)and a significant change in Glycated haemoglobin A1c(HbA1c)levels in people with type 1 diabetes[OR 1.75(95%CI:1.03 to 2.98)],and it has a RR of 0.71(95%CI:0.53 to 0.95).CONCLUSION In type 1 diabetics,teplizumab decreased insulin consumption,improved C-peptide response,and significantly changed HbA1c levels with negligible side effects.Teplizumab appears to improve glycaemic control and diabetes management with good safety and efficacy.

D2-40标记的微淋巴管密度与直肠癌淋巴转移的关系

目的:研究直肠癌组织中D2-40标记的微淋巴管特点,探讨其与肿瘤淋巴结转移的关系。方法:收集直肠癌和癌旁组织标本90份及正常直肠组织标本30份,用免疫组化SP法检测D2-40表达,计算D2-40标记的微淋巴管密度(LMVD),并结合直肠癌临床病理特征进行相关性分析。结果:肿瘤边缘区LMVD(11.3±4.6)明显高于肿瘤中心区(5.1±2.1)(P<0.01),两者均明显高于正常直肠组织(3.8±2.5)(P<0.01)。直肠癌边缘区高LMVD与肿瘤浸润深度(P<0.05)、淋巴结转移、远处转移和Dukes分期(P<0.01)有关,与患者的性别、年龄、肿瘤大小、部位及分化程度均无关。结论:直肠癌边缘区高LMVD与肿瘤组织的淋巴结转移密切相关。肿瘤边缘区LMVD是判断肿瘤淋巴管生成和淋巴结转移的重要指标。

SV40灭活疫苗免疫恒河猴后的抗体反应

目的 评价SV40灭活疫苗免疫恒河猴后的抗体反应.方法 用SV40灭活疫苗经肌肉注射免疫0.5～1周岁的恒河猴,分别于第1次和第2次免疫后2周,检测血清中和抗体效价,免疫18个月后检测SV40基因组成分.结果 免疫的12只恒河猴在第1次免疫后2周,血清抗体全部阳转,平均抗体滴度为1：16,第2次免疫2周后血清抗体滴度明显增长,平均达到1：64～1：128.用PCR方法未检测到SV40基因组成分.结论 SV40灭活疫苗可以有效诱导恒河猴产生特异性抗体反应.

血清YKL-40在诊断抗黑色素瘤分化相关基因5阳性皮肌炎合并严重肺损伤中的价值

目的:研究血清及支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)中YKL-40(chitinase-3-like-1 protein)在抗黑色素瘤分化相关基因5(anti-melanoma differentiation-associated gene 5,MDA5)阳性皮肌炎(dermatomyositis,DM)合并严重肺损伤中的价值,严重肺损伤包括快速进展间质性肺病(rapidly progressive interstitial lung disease,RP-ILD)和肺部感染。方法:选择2013—2018年中日友好医院风湿免疫科住院的抗MDA5阳性DM患者的病例资料进行回顾性分析,收集患者的人口学信息,临床、实验室及影像学检查资料,应用酶联免疫吸附法检测患者血清和BALF中YKL-40水平。绘制受试者工作特征(receiver operating characteristic,ROC)曲线,计算曲线下面积(area under the curve,AUC),评估血清YKL-40对肺损伤的诊断效能。间质性肺病(interstitial lung disease,ILD)由胸部高分辨率CT(high-resolution CT,HRCT)证实。RP-ILD定义为呼吸道症状在3个月内进行性加重,出现呼吸困难和低氧血症,或胸部HRCT显示ILD较之前加重或出现新的ILD。肺部感染经痰、血液、BALF、肺穿刺活检样本检验出病原体确诊。结果:共收集到168例抗MDA5阳性DM患者病例,其中154例合并ILD,66例(39.3%)表现为RP-ILD。经病原学依据证实合并肺部感染患者70例。合并RP-ILD患者中39例(59.1%)合并肺部感染,而非RP-ILD患者仅31例(30.4%)合并肺部感染。RP-ILD合并肺部感染的发生率高于非RP-ILD合并肺部感染者(P<0.001)。血清YKL-40水平在RP-ILD合并肺部感染组高于RP-ILD未合并肺部感染组、非RP-ILD合并肺部感染组和非RP-ILD未合并肺部感染组[83(42~142)vs.42(21~91)vs.43(24~79)vs.38(22~69),P<0.01]。血清YKL-40诊断抗MDA5阳性DM患者RP-ILD合并肺部感染的敏感性、特异性及AUC分别为75%、67%、0.72,其诊断同时存在RP-ILD和肺部感染的抗MDA5阳性DM患者的AUC较诊断仅有RP-ILD和仅有肺部感染者的AUC高,且差异有统计学意义(0.72 vs.0.54和0.55,Z=2.10和2.11,P<0.05)。结论:抗MDA5阳性DM患者合并RP-ILD和肺部感染预后差,血清YKL-40水平对这类患者同时合并RP-ILD和肺部感染有一定的诊断价值。

A multi-center,open-label,randomized,parallel-controlled phase II study comparing pharmacokinetic,pharmacodynamics and safety of ripertamab(SCT400)to rituximab(Mab Thera?)in patients with CD20-positive B-cell non-Hodgkin lymphoma

Objective:This multi-center,open-label,randomized,parallel-controlled phaseⅡstudy aimed to compare the pharmacokinetics(PK),pharmacodynamics(PD)and safety profile of ripertamab(SCT400),a recombinant antiCD20 monoclonal antibody,to rituximab(MabThera^(■))in patients with CD20-positive B-cell non-Hodgkin lymphoma(NHL).Methods:Patients with CD20-positive B-cell NHL who achieved complete remission or unconfirmed complete remission after standard treatment were randomly assigned at a 1:1 ratio to receive a single dose of ripertamab(375mg/m^(2))or rituximab(MabThera^(■),375 mg/m^(2)).PK was evaluated using area under the concentration-time curve(AUC)from time 0 to d 85(AUC_(0-85d)),AUC from time 0 to week 1(AUC0-1 w),AUC from time 0 to week 2(AUC_(0-2 w)),AUC from time 0 to week 3(AUC_(0-3 w)),AUC from time 0 to week 8(AUC_(0-8 w)),maximum serum concentration(C_(max)),terminal half-life(T_(1/2)),time to maximum serum concentration(T_(max))and clearance(CL).Bioequivalence was confirmed if the 90%confidence interval(90%CI)of the geometric mean ratio of ripertamab/rituximab was within the pre-defined bioequivalence range of 80.0%-125.0%.PD,immunogenicity,and safety were also evaluated.Results:From December 30,2014 to November 24,2015,a total of 84 patients were randomized(ripertamab,n=42;rituximab,n=42)and the PK analysis was performed on 76 patients(ripertamab,n=38;rituximab,n=38).The geometric mean ratios of ripertamab/rituximab for AUC_(0-85d),ATC_(0-inf),and Cmaxwere 96.1%(90%CI:87.6%-105.5%),95.9%(90%CI:86.5%-106.4%)and 97.4%(90%CI:91.6%-103.6%),respectively.All PK parameters met the pre-defined bioequivalence range of 80.0%-125.0%.For PD and safety evaluation,there was no statistical difference in peripheral CD 19-positive B-cell counts and CD20-positive B-cell counts at each visit,and no difference in the incidence of anti-drug antibodies was observed between the two groups.The incidences of treatment-emergent adverse events and treatment-related adverse events were also comparable between the two groups.Conclusions:In this study,the PK,PD,immunogenicity,and safety profile of ripertamab(SCT400)were similar to rituximab(MabThera^(■))in Chinese patients with CD20-positive B-cell NHL.

鸡尾酒抗体D2-40/CD34-CK对淋巴结检出数量不足结直肠癌标本的临床病理评价

目的探讨鸡尾酒抗体D2-40/CD34-CK双重免疫组化染色在淋巴结检出数量不足的结直肠癌(colorectal cancer,CRC)标本脉管侵犯评估中的应用及意义。方法收集133例淋巴结检出数量<12枚的CRC标本,分别行HE染色及鸡尾酒抗体双重免疫组化染色,比较两种方法在脉管侵犯筛查效果中的差异,并分析应用鸡尾酒抗体免疫组化染色证实的脉管侵犯与临床病理特征及患者总生存期(overall survival,OS)的关系。结果 (1)鸡尾酒抗体D2-40/CD34-CK双重免疫组化染色及HE染色对脉管侵犯的检出率分别为42. 9%(57/133)和21. 8%(29/133),差异有显著性(P <0. 001)。(2)鸡尾酒抗体D2-40/CD34-CK双重免疫组化法证实的脉管侵犯与Dukes分期、浸润深度、临床分期、淋巴结转移、肿瘤出芽相关(P <0. 05)。(3)脉管侵犯、侵犯位置深度、程度及脉管侵犯灶数≤2个灶且癌栓细胞数量≥5. 5个与患者OS密切相关(P <0. 05)。结论鸡尾酒抗体D2-40/CD34-CK双重免疫组化染色对脉管侵犯评估优于常规HE染色切片观察,与肿瘤分期、淋巴结转移及出芽情况密切相关,是患者预后的独立影响因素,可作为淋巴结检出数量不足CRC病例的补充检测指标。

CD28/CTLA-4/B7 and CD40/CD40L costimulation and activation of regulatory T cells

Costimulatory signals are crucial for T cell activation. Attempts to block costimulatory pathways have been effective in preventing unwanted immune reactions. In particular, blocking the CD28/cytotoxic T lymphocyte antigen(CTLA)-4/B7 interaction(using CTLA-4Ig) and the CD40/CD40 L interaction(using anti-CD40 L antibodies) prevents T cell mediated autoimmune diseases, transplant rejection and graft vs host disease in experimental models. Moreover, CTLA-4Ig is in clinical use to treat rheumatoid arthritis(abatacept) and to prevent rejection of renal transplants(belatacept). Under certain experimental conditions, this treatment can even result in tolerance. Surprisingly, the underlying mechanisms of immune modulation are still not completely understood. We here discuss the evidence that costimulation blockade differentially affects effector T cells(Teff) and regulatory T cells(Treg). The latter are required to control inappropriate and unwanted immune responses, and their activity often contributes to tolerance induction and maintenance. Unfortunately, our knowledge on the costimulatory requirements of Treg cells is very limited. We therefore summarize the current understanding ofthe costimulatory requirements of Treg cells, and elaborate on the effect of anti-CD40 L antibody and CTLA-4Ig treatment on Treg cell activity. In this context, we point out that the outcome of a treatment aiming at blocking the CD28/CTLA-4/B7 costimulatory interaction can vary with dosing, timing and underlying immunopathology.

Anti-NEP1-40多克隆抗体的制备与检测

目的制备Anti-NEP1-40多克隆抗体。方法合成NEP1-40多肽,采用弗氏佐剂制备免疫抗原,多次多点皮下注射健康雄性家兔,刺激机体产生相应抗体,颈动脉放血分离兔血清。结果获得含NEP1-40抗体的免疫血清,其抗体特异性好,亲和力较高。结论 NEP1-40兔多克隆抗体可以满足NEP1-40相关的科学研究。

Preclinical Evaluation of CD40-Directed Immunotherapy in B-Cell Lymphoma Using [¹⁸F]Fluorothymidine-PET

Background: Inhibition of the lymphoma surface antigen CD40 by the antagonistic CD40 antibody NVP-HCD122 (HCD122) demonstrates activity in various lymphoma subtypes. In this preclinical in vivo study we examined the suitability of positron emission tomography (PET) using the thymidine analogue 3’-deoxy-3’-[18F]fluorothymidine (FLT) for early response assessment upon HCD122 treatment in diffuse large B cell lymphoma (DLBCL). Methods: Immunodeficient mice bearing human DLBCL xenografts (SU-DHL-4) received weekly intraperitoneal injections of HCD122. Tumor growth was followed up until Day 14. Molecular imaging with FLT-PET was performed before (Day 0) and after start of therapy (Day 2 and Day 7). On Day 14 lymphoma xenografts were explanted for immunohistochemical analysis to correlate PET findings with CD40 surface expression on tumor tissue. Results: Treatment with HCD122 significantly delayed tumor growth resulting in a tumor growth inhibition of 45% on Day 14. Significant reduction of tumor-to-background ratio (TBR) of FLT-PET was seen in treated animals on Day 7 and preceded change of tumor volume, thus predicting therapy response to HCD122. Immunohistochemical analysis of xenografts revealed significantly higher CD40 expression on treated than on untreated tissue. Moreover, we found a significant correlation between CD40 expression and FLT-PET response for xenograft tumor treated with HCD122. Conclusions: Treatment of DLBCL with the antagonistic CD40 antibody HCD122 can be monitored with FLT-PET as early as seven days after commencement of therapy and seems to increase CD40 expression on tumor tissue.

Induction of specific immunosuppression of cardiac allograft rejection withmonoclonal antibodies to CD44, LFA-1 and ICAM-1

Objective:To evaluate the immunosuppressive effect of monoclonal antibodies (McAb) against cell surface adhesion molecules on transplant rejection. Methods: C57BL/6 (H-2b) mouse cardiac grafts were transplanted into BALB/c(H- 2d) mice. This model was used to investigate the possibility of immunosuppressive induction with CD44 McAb, leukocyte function associated antigen (LFA-1) and intercellular adhesion molecule (ICAM-1). Results: Treatment of the allograft recipients with CD44 McAb alone, or both LFA-1 and ICAM-1 or combination of these 3 McAb significantly prolonged the cardiac allografts survival as compared with PBS controls (P<0.01). The combination of anti-CD44 and ICAM-1 and LFA-1 McAb was shown to produce more significant prolongation of grafts survival than anti-CD44 McAb alone or both anti-ICAM- 1 and LFA-1 McAb (P < 0.01). Histological examination of the grafts treated with the McAb displayed greatly reduced mononuclear cell infiltration. The proliferation of spleen cells from recipient BALB/c with McAb treatment was significantly inhibited in response to the stimulators of C57BL/6 spleen cells, but increased upon the stimulation of C3H/He (H-2k) spleen cells, as demonstrated by mixed lymphocyte reaction. Similarly, the cytotoxic activity against donor H-2-compatible (H-2b) target cells, EL-4 cells, was significantly suppressed. The spleen cells from allografted recipient BALB/c mice with McAb treatment induced specific tolerance for C57BL/6 cardiac grafts in allografted recipients, whereas those from allografted BALB/c mice without McAb treatment induced acute rejection. Conclusion: These results indicate that antiadhesion therapy using a combination of McAb to adhesion molecules can induce specific immunosuppression of transplant rejection.

BGB-A445,a novel non-ligand-blocking agonistic anti-OX40 antibody,exhibits superior immune activation and antitumor effects in preclinical models

t OX40 is a costimulatory receptor that is expressed primarily on activated CD4+,CD8+,and regulatory T cells.The ligation of OX40 to its sole ligand OX40L potentiates T cell expansion,differentiation,and activation and also promotes dendritic cells to mature to enhance their cytokine production.Therefore,the use of agonistic anti-Ox40 antibodies for cancer immunotherapy has gained great interest.However,most of the agonistic anti-OX40 antibodies in the clinic are OX40L-competitive and show limited efficacy.Here,we discovered that BGB-A445,a non-ligand-competitive agonistic anti-OX40 antibody currently under clinical investigation,induced optimal T cell activation without impairing dendritic cell function.In addition,BGB-A445 dose-dependently and significantly depleted regulatory T cells in vitro and in vivo via antibody-dependent cellular cytotoxicity.In the MC38 syngeneic model established in humanized OX40 knock-in mice,BGB-A445 demonstrated robust and dose-dependent antitumor efficacy,whereas the ligand-competitive anti-Ox40 antibody showed antitumor efficacy characterized by a hook effect.Furthermore,BGB-A445 demonstrated a strong combination antitumor effect with an anti-PD-1 antibody.Taken together,our findings show that BGB-A445,which does not block OX40-OX40L interaction in contrast to clinical-stage anti-OX40 antibodies,shows superior immune-stimulating effects and antitumor efficacy and thus warrants further clinical investigation.