Investigating T Cell Immunity in Cancer: Achievements and Prospects

T cells play a key role in tumour surveillance, both identifying and eliminating transformed cells. However, as tumours become established they form their own suppressive microenvironments capable of shutting down T cell function, and allowing tumours to persist and grow. To further understand the tumour microenvironment, including the interplay between different immune cells and their role in anti-tumour immune responses, a number of studies from mouse models to clinical trials have been performed. In this review, we examine mechanisms utilized by tumour cells to reduce their visibility to CD8⁺ Cytotoxic T lymphocytes (CTL), as well as therapeutic strategies trialled to overcome these tumour-evasion mechanisms. Next, we summarize recent advances in approaches to enhance CAR T cell activity and persistence over the past 10 years, including bispecific CAR T cell design and early evidence of efficacy. Lastly, we examine mechanisms of T cell infiltration and tumour regression, and discuss the strengths and weaknesses of different strategies to investigate T cell function in murine tumour models.     

Cell Therapies in Bladder Cancer Management

Currently, bladder cancer (BC) represents a challenging problem in the field of Oncology. The high incidence, prevalence, and progression of BC have led to the exploration of new avenues in its management, in particular in advanced metastatic stages. The recent inclusion of immune checkpoint blockade inhibitors as a therapeutic option for BC represents an unprecedented advance in BC management. However, although some patients show durable responses, the fraction of patients showing benefit is still limited. Notwithstanding, cell-based therapies, initially developed for the management of hematological cancers by infusing immune or trained immune cells or after the engineering of chimeric antigen receptor (CAR) expressing cells, are promising tools to control, or even cure, solid tumors. In this review, we summarize recent cell-based immunotherapy studies, with a special focus on BC.     

Pharmacologic Control of CAR T Cells

Chimeric antigen receptor (CAR) therapy is a promising modality for the treatment of advanced cancers that are otherwise incurable. During the last decade, different centers worldwide have tested the anti-CD19 CAR T cells and shown clinical benefits in the treatment of B cell tumors. However, despite these encouraging results, CAR treatment has also been found to lead to serious side effects and capricious response profiles in patients. In addition, the CD19 CAR success has been difficult to reproduce for other types of malignancy. The appearance of resistant tumor variants, the lack of antigen specificity, and the occurrence of severe adverse effects due to over-stimulation of the therapeutic cells have been identified as the major impediments. This has motivated a growing interest in developing strategies to overcome these hurdles through CAR control. Among them, the combination of small molecules and approved drugs with CAR T cells has been investigated. These have been exploited to induce a synergistic anti-cancer effect but also to control the presence of the CAR T cells or tune the therapeutic activity. In the present review, we discuss opportunistic and rational approaches involving drugs featuring anti-cancer efficacy and CAR-adjustable effect.     

羊布鲁菌O链M抗原提纯及其单克隆抗体的研制

目的研制羊布鲁菌O链M纯化抗原及其单克隆抗体。方法采用冷酚法提纯羊布鲁菌16M的O链抗原,并经琼脂糖凝胶免疫扩散和SDS-PAGE鉴定,用灭活的该抗原免疫BALB/c小鼠,制备单克隆抗体,并进行鉴定。结果建立3株分泌单克隆抗体的细胞株2D10、3C6和1E10,ELISA效价分别为1.380、1.109和1.048,与大肠杆菌0:157、小肠结肠炎耶尔森菌0:9、鼠伤寒沙门菌和猪胸膜肺炎放线杆菌均无交叉反应。3C6和1E10与牛布鲁菌544A发生交叉反应,2D10的亲和常数达5.30×10~7M~(-1)。结论已制备出纯化的羊布鲁菌O链M抗原及其单抗。     

Complementarity determining region residues aspartic acid at H55, serine at H95 and tyrosines at H97 and L96 play important roles in the B72.3 antibody-TAG72 antigen interaction

Structural analysis derived from the crystallographic studyof the chimeric B72.3 antibody illustrated some major atomicinteractions between complementarity determining region (CDR)residues. For example, hydrogen bonds are formed between H35/H95,L50/H97, H53/H55 and H96/L96 respectively. These CDR residuesmay play important roles in the B72.3–TAG72 (antibody-antigen)interaction either by direct interaction with the TAG72 antigenor by maintaining a CDR loop conformation through atomic interactionsbetween CDR residues. In order to confirm these assumptions,we altered these CDR residues by site-directed mutagenesis anddetermined binding affinities of these mutant chimeric antibodiesfor the TAG72 antigen in a solid-phase radioimmunoassay. Wefound that H55, H95, H97 and L96 are important CDR residuesfor the B72.3–TAG72 interaction. Single amino acid substitutionsof aspartic acid and serine by alanine at H55 of CDR2 and atH95 of CDR3 respectively and of tyrosine by phenylalanine atH97 and L96 of CDR3, significantly reduced the binding affinityfor the TAG72 antigen by 20-, 8-, 16- and 45-fold respectively.Therefore, this study reveals some of the requirements for maintainingthe integrity of the B72.3 antibody combining sites.     

重组G145R HBsAg亲和纯化方法的建立及其应用

目的建立重组乙型肝炎病毒G145R变异HBsAg抗体亲和纯化方法。方法用抗-HBs单克隆抗体(D12- McAb)制备亲和层析胶,对2A8细胞(分泌G145R变异HBsAg)培养上清盐析物进行亲和纯化。采用SDS-PAGE、Western blot及ELISA,对纯化产物的纯度、特异性、含量和回收率进行鉴定,并与同法纯化的HBsAg阳性血清及r-wHBsAg提取物进行比较。结果D12-McAb对重组真核表达G145R变异HBsAg、HBsAg阳性血清及r-wHBsAS三者具有相似的亲和性,产物纯度分别为90.3%、95.2%和93.1%,回收率分别为43.3%、72.0%和66.4%。结论已成功地建立了重组G145R变异HBsAg抗体亲和纯化方法,为G145R变异以及其他HBV免疫逃逸变异感染的深入研究奠定了重要的技术基础。     

一个新的不耐热的伤寒沙门氏菌特异性抗原(θ)的发现

我们在研究伤寒沙门氏菌保护性抗原期间,发现了一个新的不耐热的伤寒沙门氏菌特异性抗原,暂时命名为θ抗原。这个抗原存在于各种类型的伤寒沙门氏菌中,并能从Ty2菌苗株细胞提取。以不吸收的抗θ兔血清作试管凝集时,与O90l和H901菌株的凝集效价分别为1:640和1:320。但与Ty2或9,12,Vi:d:-菌株不发生凝集,这一现象以后证实是由于Vi抗原的存在。θ抗原形成凝集较幔(31℃18～24小时),其所形成的凝集块疏松,易摇散、呈半絮状。菌体经100℃30分钟加热后与θ血清不发生凝集。玻片凝集试验显示θ血清与W和V型伤寒沙门氏菌(后者须培养在含酚培养基上)在1分钟内产生明显凝集。而与大多数D群(包括D_1和D_2)沙门氏菌和其他沙门氏菌不凝集,并与下列肠杆菌科细菌亦不凝集,包括志贺氏菌属A、B、C、D群,大肠艾希氏菌O:1～25和致病性大肠艾希氏菌12个不同的血清型,3株变形杆菌以及含有α抗原的Wakefield茵。化学分析显示,粗制θ提取物含有核酸(491μg/ml)。蛋白质(39μg/ml)和KDO(23μg/ml),但不含有O-乙酰基,表明θ抗原不同于Vi抗原。作者建议以θ血清替代现用的O:9,Vi和d因子血清作为伤寒沙门氏菌的血清学诊断。并需继续查明θ抗原的化学本质。     

b型流感嗜血杆菌多糖结合疫苗的研制

［摘 要］目的 研制b型流感嗜血杆菌（Hib）结合疫苗并考察其稳定性。方法 用溴化氰活化提纯Hib荚膜多糖抗原，通过己二酰肼（ADH）与破伤风类毒素（TT）蛋白共价结合，制备b型流感嗜血杆菌结合物，并考察放置在2～8℃及室温9个月和37℃　3个月后的稳定性。结果 用本工艺提取的多糖，合成的多糖衍生物，多糖蛋白结合物的化学组成及结构特性均达到了国外同类产品的质控要求。结合物在2～8℃及室温放置9个月后基本无降解。结论 为临床评价Hib结合疫苗的安全性和保护效果及确定效期提供了实验基础。     

Preparation and Characterization of a Chloroperoxidase-like Catalytic Antibody

The small molecule, meso-tetra(α,α,α,α-o-phenylacetamidophenyl) porphyrin (Mr1147.0) was used as complete antigen to elicit MAb through the immunization and cell fusion techniques. The MAb 1F2 obtained was demonstrated to be very pure by MALDI/TOFMS. The subtype of MAb 1F2 is IgG2a, which has a relative molecular weight of 156,678.8 Da.No significant change in the intensity of absorption peaks in UV and CD spectra was observed over a pH range between 6 and 12. The high stability of the abzyme and the tight binding between Fe porphyrin and antibody were also demonstrated. V_max, K_m, κcat, κcat/K_m for abzyme are 5.18 × 10⁻⁸ Ms⁻¹, 1.50 × 10⁻⁸ M, 0.518 s⁻¹, 3.45 × 10⁷ M⁻¹s⁻¹, respectively. The data obtained indicate that catalytic antibody has high catalytic activity. The chloroperoxidase activity of MAb 1F2-Fe porphyrin complex is stable from 10 °C to 60 °C.     

Molecular modeling of antibody-antigen complexes between the Brucella abortus O-chain polysaccharide and a specific monoclonal antibody

A molecular model of the binding site of an anti-carbohydrateantibody (YsT9.1) has been developed using computer-assistedmodeling techniques and molecular dynamics calculations. Sequencehomologies among YsT9.1 and the Fv regions of McPC603, J539and human Bence-Jones protein REI, all of which have solvedcrystal structures, provided the basis for the modeling. Thegroove-type combining site model had a topography which wascomplementary to low energy confonners of the polysaccharide,a Brucella O-antigen, and the site could be almost completelyfilled by a pentasaccharide epitope in either of two dockingmodes. Putative interactions between this epitope and the antibodyare consistent with the known structural requirements for bindingand lead to the design of oligosaccharide inhibitors that probethe veracity of the modeled docked complex. Ultimately boththe Fv model and the docked complex will be compared with independentcrystal structures of YsT9.1 Fab with and without pentasaccharideinhibitor, currently at the stage of refinement.