CTLA-4 Research Articles

Association between Thyroid Cancer and CTLA-4 Gene Polymorphisms.

The study aimed to explore the relationship between cytotoxic T lymphocyte associate protein-4 (CTLA-4) gene polymorphisms and the onset of thyroid cancer. 200 patients with thyroid cancer were included in the disease group and 200 healthy people were selected as the control group (all of them were admitted to Huashan Hospital (East) of Fudan University). The peripheral blood was collected in both groups, and the polymorphic regions at CTLA-4 gene loci rs3087243 (G>A), rs606231417 (C>T) and rs1553657430 (C>A) were amplified via polymerase chain reaction (PCR). The expression level of the CTLA-4 gene was detected via RT-qPCR. Moreover, the associations of clinical indexes with CTLA-4 genotypes were analyzed. The G allele frequency at CTLA-4 gene locus rs3087243 was raised in the disease group (p=0.000). The frequencies of the GG genotype at rs3087243, TT genotype at rs606231417 and CA genotype at rs1553657430were decreased in the control group (p<0.001, p<0.001, p=0.002). The GA+AAfrequency at rs3087243 and CC+CTfrequency at rs606231417 in the disease group were lower than those in the control group. The linkage disequilibrium was higher at rs606231417 and rs1553657430(D'=0.431). Moreover, the CTLA-4 gene expression was remarkably raised in patients with CC genotype at rs1553657430than that in other genotype patients (p<0.05). The genotype at rs606231417was significantly associated with the calcitonin level in thyroid cancer patients (p=0.039), while the genotype at rs3087243 was significantly associated with the thyroid-stimulating hormone level in thyroid cancer patients (p=0.002).CTLA-4 gene polymorphisms have a significant association with the progression of thyroid cancer, which may be a susceptible factor for it.

Granulomatous and Sarcoid-like Immune-Related Adverse Events following CTLA4 and PD1 Blockade Adjuvant Therapy of Melanoma: A Combined Analysis of ECOG-ACRIN E1609 and SWOG S1404 Phase III Trials and a Literature Review.

Treatment with immune checkpoint inhibitors (ICIs) has been linked to granulomatous and sarcoid-like lesions (GSLs) affecting different organs. This study sought to evaluate GSL incidence in patients with high-risk melanoma treated with cytotoxic T-lymphocyte antigen 4 (CTLA4) or programmed cell death 1 (PD1) blockade adjuvant therapy in two clinical trials: ECOG-ACRIN E1609 and SWOG S1404. Descriptions and GSL severity ratings were recorded. Data were collected from ECOG-ACRIN E1609 and SWOG S1404. Descriptive statistics along with GSL severity grades were reported. Additionally, a literature review for such cases was summarized. A total of 11 GSL cases were reported among 2878 patients treated with either ICI or with High-Dose Interferon Alfa-2b (HDI) in ECOG-ACRIN E1609 and SWOG S1404 trials. Cases were numerically more commonly reported with ipi10, followed by pembrolizumab, ipi3, and HDI, respectively. Most of the cases were grade III. Further, organs involved included lung, mediastinal lymph nodes, skin and subcutaneous tissue, and eye. Furthermore, a summary of 62 reports in the literature was described. GSLs following anti-CTLA4 and anti-PD1 antibody therapy in patients with melanoma were reported unusually. Reported cases ranged in grade from I to III and appeared manageable. Careful attention to these events and their reporting will be essential to better guide practice and management guidelines.

The Tumor Immune Microenvironment in Clear Cell Renal Cell Carcinoma.

Clear cell renal cell carcinoma (ccRCC) is a type of kidney cancer that arises from the cells lining the tubes of the kidney. The tumor immune microenvironment (TIME) of ccRCC is a complex interplay of various immune cells, cytokines, and signaling pathways. One of the critical features of the ccRCC TIME is the presence of infiltrating immune cells, including T cells, B cells, natural killer cells, dendritic cells, and myeloid-derived suppressor cells. Among these cells, CD8+ T cells are particularly important in controlling tumor growth by recognizing and killing cancer cells. However, the TIME of ccRCC is also characterized by an immunosuppressive environment that hinders the function of immune cells. Several mechanisms contribute to the immunosuppressive nature of the ccRCC TIME. For instance, ccRCC cells produce cytokines such as interleukin-10 (IL-10) and transforming growth factor-beta (TGF-β), which suppress immune cell activation and promote the differentiation of regulatory T cells (Tregs). Tregs, in turn, dampen the activity of effector T cells and promote tumor growth. In addition, ccRCC cells can express programmed death-ligand 1 (PD-L1), which interacts with the programmed cell death protein 1 (PD-1) receptor on T cells to inhibit their function. In addition, other immune checkpoint proteins, such as cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and lymphocyte activation gene 3 (LAG-3), also contribute to the immunosuppressive milieu of the ccRCC TIME. Finally, the hypoxic and nutrient-poor microenvironment of ccRCC can stimulate the production of immunosuppressive metabolites, such as adenosine and kynurenine, which further impair the function of immune cells. Understanding the complex interplay between tumor cells and the immune system in the ccRCC TIME is crucial for developing effective immunotherapies to treat this disease.

Effects of SMC1A on immune microenvironment and cancer stem cells in colon adenocarcinoma.

Our previous study suggested that SMC1 has significant functions in colorectal cancer (CRC). However, few reports have shown the effects of structural maintenance of chromosomes 1 (SMC1A) on the immune microenvironment and tumor stem cells. The Cancer Genome Atlas (TCGA) database, CPTAC database, Human Protein Atlas (HPA) database, the Cancer Cell Line Encyclopedia (CCLE) and Tumor Immune Single-cell Hub were used. Flow cytometry and immunohistochemical analysis were checked for immune infiltration on MC38 mice model. Human CRC tissues were tested with RT-qPCR. The mRNA and protein levels of SMC1A were increased in colon adenocarcinoma (COAD) samples.SMC1A was associated with DNA activity. Interestingly, SMC1A was highly expressed in many types of immune cells at single-cell levels. Moreover, the high expression of SMC1A was positively correlated with immune infiltration, and immunohistochemical analysis showed that SMC1A was positively associated with CD45 expression in MC38 mice model. Also, the percentage of IL4+ CD4+ T cells (Th2) and FoxP3+ CD4+ T cells (Tregs) was significantly higher in the SMC1A overexpression group than in control by flow cytometry assay in vivo. SMC1A expression could affect the proliferation of T cells in the mice model. The mutation and somatic cell copy number variation (SCNV) of SMC1A were also associated with immune cell infiltration. In addition to SMC1A in the "hot" T-cell inflammatory microenvironment of colon cancer, SMC1A also positively correlates with the immune checkpoint genes CD274, CTLA4, and PDCD1 in colon adenocarcinoma (COAD) samples. Furthermore, we also found that SMC1A plays a positive correlation with the induction of cancer stem cells (CSCs). Our results also showed that miR-23b-3p binds SMC1A. SMC1A may be a bidirectional target switch that simultaneously regulates the immune microenvironment and tumor stem cells. Moreover, SMC1A may be a biomarker for the prediction of immune checkpoint inhibitor (ICI) therapy.

Dual IL-6 and CTLA-4 blockade regresses pancreatic tumors in a T cell- and CXCR3-dependent manner.

This study aimed to enhance antitumor immune responses to pancreatic cancer via Ab-based blockade of IL-6 and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4). Mice bearing s.c. or orthotopic pancreatic tumors were treated with blocking Abs to IL‑6 and/or CTLA-4. In both tumor models, dual IL-6 and CTLA-4 blockade significantly inhibited tumor growth. Additional investigations revealed that dual therapy induced an overwhelming infiltration of T cells into the tumor as well as changes in CD4+ T cell subsets. Dual blockade therapy elicited CD4+ T cells to secrete increased IFN-γ in vitro. Likewise, in vitro stimulation of pancreatic tumor cells with IFN-γ profoundly increased tumor cell production of CXCR3-specific chemokines, even in the presence of IL-6. In vivo blockade of CXCR3 prevented orthotopic tumor regression in the presence of the combination treatment, demonstrating a dependence on the CXCR3 axis for antitumor efficacy. Both CD4+ and CD8+ T cells were required for the antitumor activity of this combination therapy, as their in vivo depletion via Abs impaired outcomes. These data represent the first report to our knowledge of IL-6 and CTLA‑4 blockade as a means to regress pancreatic tumors with defined operative mechanisms of efficacy.

Evaluation of the immune checkpoint factors in idiopathic membranous nephropathy

Idiopathic membranous nephropathy (IMN), a single-organ autoimmune disease, is recognized by autoantibodies to podocyte proteins and identified as the most frequent cause of nephrotic syndrome in adults. T cells are important contributors in autoimmunity since they promote B–cell development, antibody production, direct inflammation, and organ tissue cytotoxicity. This study investigated the inhibitory immune checkpoint (ICP) receptors expressed on T lymphocytes and other immune cells. Thus, PBMCs from IMN patients were obtained before treatment, and the levels of ICPs such as programmed cell death protein 1 (PD-1), cytotoxic T-lymphocyte-associated protein 4 (CTLA4), lymphocyte activation gene-3 (LAG-3), and T cell immunoglobulin-3 (TIM-3) were examined at both gene and protein expression using real time PCR and Western blot tests respectively. The results illustrated that gene expression levels of ICPs reduced significantly in comparison to the control which were verified by related fold changes of protein expression sequentially. Our study revealed that CTLA-4, PD-1, TIM-3, and LAG-3 expression is impaired in IMN patients before treatment which could be a potential target for therapy.

Optimization of CTLA-4 and PD-1 proteins in EMT6 Mouse Mammary Cancer Cells by Western Blot

Targeting the activation of immune checkpoints is recognized as an effective strategy for triggering anti-tumour immune responses in cancer cells. Cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) and programmed cell death protein 1 (PD-1) were identified as potential crucial targets for cancer treatment. Overexpression of CTLA-4 and PD-1 proteins in primary tumour and human cell lines is well documented. In contrary, lack of data was available using animal cell lines. The presence study aims to optimize the expression of CTLA-4 and PD-1 proteins in EMT6 mouse mammary cancer cells using Western blot, and provide basic understanding of their association with breast cancer cell progression. Proteins extracted from EMT6 parental cells were adjusted to 30ng for gel electrophoresis. Afterwards, the protein was transferred to a nitrocellulose membrane for blotting. The membrane was then subjected to chemiluminescent for band detection. Results obtained using beta-actin as a housekeeping gene show that both CTLA-4 (32 kDa) and PD-1 (34 kDa) proteins were expressed by using a 1:1000 dilution for each antibody from the lysate of EMT6 mouse mammary cancer cells. The relative expression of PD-1 (4.0 ± 0.26) is higher compared to CTLA-4 (1.2 ± 1.8). As a conclusion, both CTLA-4 and PD-1 proteins were indeed expressed in EMT6 mouse mammary cancer cells and this outcome provide the platform for extensive in vivo research on the link of both proteins with breast cancer using animal model.

Current application of immunotherapy in melanoma.

Current application of immunotherapy in melanoma.

Genetic variants of CTLA4 are associated with clinical outcome of patients with multiple myeloma.

Immune dysfunction in patients with multiple myeloma (MM) affects both the innate and adaptive immune system. Molecules involved in the immune checkpoint pathways are essential to determine the ability of cancer cells to escape from the immune system surveillance. However, few data are available concerning the role of these molecules in predicting the kinetics of progression of MM. We retrospectively analysed polymorphisms of CTLA4 (rs231775 and rs733618), BTLA (rs9288953), CD28 (rs3116496), PD-1 (rs36084323 and rs11568821) and LAG-3 (rs870849) genes in 239 patients with newly diagnosed MM. Patients with a CTLA4 rs231775 AA/AG genotype showed a median progression-free survival (PFS) significantly lower than those with GG genotype (32.3 months versus 96.8 months respectively; p: 0.008). The 5-year PFS rate was 25% for patients with grouped AA and AG genotype vs 55.4% for patients with GG genotype. Multivariate analysis confirmed the CTLA4 rs231775 genotype as an independent risk factor for PFS (Hazard Ratio (HR): 2.05; 95% CI: 1.0-6.2; p: 0.047). Our results suggest that the CTLA4 genotype may identify patients with earlier progression of MM. This polymorphism could potentially be used as a prognostic biomarker.

Development of a Highly Sensitive Hybrid LC/MS Assay for the Quantitative Measurement of CTLA-4 in Human T Cells.

Cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) is a check point protein expressed on the surface of T cells and plays a central role in regulating the immune response. In recent years, CTLA-4 has become a popular target for cancer immunotherapy in which blocking CTLA-4 can restore T-cell function and enhance the immune response against cancer. Currently, there are many CTLA-4 inhibitors in a variety of modalities, including cell therapies, which are being developed in both preclinical and clinical stages to further harness the potential of the target for the treatment of certain types of cancer. In drug discovery research, measuring the level of CTLA-4 in T cells is important for drug discovery and development because it provides key information for quantitative assessment of the pharmacodynamics, efficacy, and safety of the CTLA-4-based therapies. However, to our best knowledge, there is still no report of a sensitive, specific, accurate, and reliable assay for CTLA-4 measurement. In this work, an LC/MS-based method was developed to measure CTLA-4 in human T cells. The assay demonstrated high specificity with an LLOQ of 5 copies of CTLA-4 per cell when using 2.5 million T cells for analysis. As shown in the work, the assay was successfully used to measure CTLA-4 levels in subtype T-cell samples from individual healthy subjects. The assay could be applied in supporting the studies of CTLA-4-based cancer therapies.

CD86 Is Associated with Immune Infiltration and Immunotherapy Signatures in AML and Promotes Its Progression.

Cluster of differentiation 86 (CD86), also known as B7-2, is a molecule expressed on antigen-presenting cells that provides the costimulatory signals required for T cell activation and survival. CD86 binds to two ligands on the surface of T cells: the antigen CD28 and cytotoxic T lymphocyte-associated protein 4 (CTLA-4). By binding to CD28, CD86-together with CD80-promotes the participation of T cells in the antigen presentation process. However, the interrelationships among CD86, immunotherapy, and immune infiltration in acute myeloid leukemia (AML) are unclear. The immunological effects of CD86 in various cancers (including on chemokines, immunostimulators, MHC, and receptors) were evaluated through a pan-cancer analysis using TCGA and GEO databases. The relationship between CD86 expression and mononucleotide variation, gene copy number variation, methylation, immune checkpoint blockers (ICBs), and T-cell inflammation score in AML was subsequently examined. ESTIMATE and limma packages were used to identify genes at the intersection of CD86 with StromalScore and ImmuneScore. Subsequently, GO/KEGG and PPI network analyses were performed. The immune risk score (IRS) model was constructed, and the validation set was used for verification. The predictive value was compared with the TIDE score. CD86 was overexpressed in many cancers, and its overexpression was associated with a poor prognosis. CD86 expression was positively correlated with the expression of CTLA4, PDCD1LG2, IDO1, HAVCR2, and other genes and negatively correlated with CD86 methylation. The expression of CD86 in AML cell lines was detected by QRT-PCR and Western blot, and the results showed that CD86 was overexpressed in AML cell lines. Immune infiltration assays showed that CD86 expression was positively correlated with CD8 T cell, Dendritic cell, macrophage, NK cell, and Th1_cell and also with immune examination site, immune regulation, immunotherapy response, and TIICs. ssGSEA showed that CD86 was enriched in immune-related pathways, and CD86 expression was correlated with mutations in the genes RB1, ERBB2, and FANCC, which are associated with responses to radiotherapy and chemotherapy. The IRS score performed better than the TIDE website score. CD86 appears to participate in immune invasion in AML and is an important player in the tumor microenvironment in this malignancy. At the same time, the IRS score developed by us has a good effect and may provide some support for the diagnosis of AML. Thus, CD86 may serve as a potential target for AML immunotherapy.

Evidence of Association between CTLA-4 Gene Polymorphisms and Colorectal Cancers in Saudi Patients.

Cytotoxic T lymphocyte antigen-4 (CTLA-4) has been identified as an immunosuppressive molecule involved in the negative regulation of T cells. It is highly expressed in several types of autoimmune diseases and cancers including colorectal cancer (CRC). (1) Objective: To explore the association between CTLA-4 single nucleotide polymorphisms (SNP) and risk to (CRC) in the Saudi population. (2) Methods: In this case-control study, 100 patients with CRC and 100 matched healthy controls were genotyped for three CTLA-4 SNPs: rs11571317 (-658C > T), rs231775 (+49A > G) and rs3087243 (CT60 G > A), using TaqMan assay method. Associations were evaluated using odds ratios (ORs) and 95% confidence intervals (95% CIs) for five inheritance models (co-dominant, dominant, recessive, over-dominant and log-additive). Furthermore, CTLA-4 expression levels were evaluated using quantitative real-time PCR (Q-RT-PCR) in colon cancer and adjacent colon tissues. (3) Results: Our result showed a significant association of the G allele (OR = 2.337, p < 0.0001) and GG genotype of the missense SNP +49A > G with increased risk of developing CRC in codominant (OR = 8.93, p < 0.0001) and recessive (OR = 16.32, p < 0.0001) models. Inversely, the AG genotype was significantly associated with decreased risk to CRC in the codominant model (OR = 0.23, p < 0.0001). In addition, the CT60 G > A polymorphism exhibited a strong association with a high risk of developing CRC for the AA genotype in codominant (OR = 3.323, p = 0.0053) and in allele models (OR = 1.816, p = 0.005). No significant association was found between -658C > T and CRC. The haplotype analysis showed that the G-A-G haplotype of the rs11571317, rs231775 and rs3087243 was associated with high risk for CRC (OR = 57.66; p < 0.001). The CTLA-4 mRNA gene expression was found significantly higher in tumors compared to normal adjacent colon samples (p < 0.001). (4) Conclusions: Our findings support an association between the CTLA-4 rs231775 (+49A > G) and rs3087243 (CT60 G > A) polymorphisms and CRC risk in the Saudi population. Further validation in a larger cohort size is needed prior to utilizing these SNPs as a potential screening marker in the Saudi population.

Immune Co-inhibitory Receptors CTLA-4, PD-1, TIGIT, LAG-3, and TIM-3 in Upper Tract Urothelial Carcinomas: A Large Cohort Study.

Programmed cell death 1 ligand 1), programmed cell death protein-1 (PD-1), cytotoxic T-lymphocyte antigen 4 (CTLA-4), T-cell immunoglobulin and mucin-domain containing-3, lymphocyte activation gene-3, and T-cell immunoglobulin and ITIM domain (TIGIT) are considered major immune co-inhibitory receptors (CIRs) and the most promising immunotherapeutic targets in cancer treatment, but they are largely unexplored in upper tract urothelial carcinoma (UTUC). The aim of this Cohort Study was to provide evidence concerning expression profiles and the clinical significance of CIRs among Chinese UTUC patients. A total of 175 UTUC patients who received radical surgery in our center were included. We used immunohistochemistry to evaluate CIR expressions in tissue microarrays (TMAs). Clinicopathological characteristics and prognostic correlations of CIR proteins were retrospectively analyzed. TIGIT, T-cell immunoglobulin and mucin-domain containing-3, PD-1, CTLA-4, Programmed cell death 1 ligand 1, and lymphocyte activation gene-3 high expression was examined in 136(77.7%), 86(49.1%), 57(32.6%), 18(10.3%), 28(16.0%), and 18(10.3%) patients, respectively. Log-rank tests and Multivariate Cox analysis both implied CTLA-4 and TIGIT expression was associated with worse relapse-free survival. In conclusion, this is the largest Chinese UTUC cohort study, and we analyzed the Co-inhibitory receptor expression profiles in UTUC. We identified CTLA-4 and TIGIT expression as promising biomarkers for tumor recurrence. Furthermore, a subset of advanced UTUCs are probably immunogenic, for which single or combined immunotherapy may be potential therapeutic approaches in the future.

Abstract 1098: Combination of a DNA vaccine-induced immune checkpoint blockade and radiation therapy induces anti-tumor immunity

Abstract Background: Immune checkpoint blockade (ICB) immunotherapy improves survival for many cancer patients, but those with immunologically ‘cold’ tumors do not derive benefit. Radiation therapy (RT) has been demonstrated to augment antitumor immunity in preclinical and clinical studies and may enhance response to ICB in immunologically cold tumors. We have developed DNA-based, cost-effective, and versatile ICB immunotherapies. Here, we test the use of these alone and in combination with RT as a novel treatment approach in syngeneic murine melanoma models. Materials and methods: We constructed ICB DNA vaccines targeting cytotoxic T lymphocyte antigen 4 (CTLA-4) and programmed cell death 1 (PD-1), CTLA-4 Vax and PD-1 Vax, respectively. These were delivered to mice by intramuscular or intratumoral electroporation. The effects of combinations of ICB DNA vaccines and RT were monitored for generation of endogenous anti-CTLA-4 and anti-PD-1 antibodies, modification of tumor microenvironment (TME), and tumor response in mice bearing syngeneic B78 or B16 melanoma. Results: Intramuscular vaccination with CTLA-4 Vax and PD-1 Vax induced endogenous anti-CTLA-4 and anti-PD-1 antibodies, respectively, and the titers increased with additional vaccinations. Vaccination with CTLA-4 Vax and PD-1 Vax alone or in combination (Dual Vax) did not inhibit the murine B16 melanoma model; however, the combination of CTLA-4 Vax and RT (CTLA-4 Vax + RT) led to enhanced infiltration of the TME with effector T cells. Dual Vax combined with RT enabled systemic anti-tumor immunity in mice bearing two B16 melanoma tumors in which only one was radiated. With intra-tumoral route of vaccination, the Dual Vax + RT treatment induced a trend toward systemic anti-tumor response in the immunologically cold B78 tumor model. Conclusions: DNA vaccines that stimulate endogenous production of antibodies against immune checkpoints may serve as an alternative form of ICB. The combination of ICB DNA vaccine and RT enabled anti-tumor immunity and tumor response in preclinical models of melanoma. Citation Format: Keng-Hsueh Lan, Ying-Chun Sheng, Keng-Li Lan, KyungMann Kim, Sung-Hsin Kuo, Zachary Morris. Combination of a DNA vaccine-induced immune checkpoint blockade and radiation therapy induces anti-tumor immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1098.

Abstract 3165: ImmuneSig: A deep learning framework to develop gene expression signature of immune checkpoint inhibitor therapy

Abstract Immune checkpoint inhibitor (ICI) therapy targeting cytotoxic T lymphocyte-associated protein 4 (CTLA-4), programmed death 1 (PD1), and programmed death-ligand 1 (PDL1) induces durable remission in some patients with various cancer types. Given the variable patient outcomes, many studies have been conducted to identify prognostic biomarkers associated with ICI response. However, due to intra- and inter-tumor heterogeneity and the complexity of the immune system, those biomarkers have diverse efficacy for predicting outcomes in clinical trials. Here we propose a deep learning framework with unsupervised self-learning autoencoders that identifies predictive features from patient transcriptomics data. This approach allows us to determine the contribution of each gene to the response classifier and identifies potential gene signatures for predicting immunotherapy response by using the learned weights of trained autoencoders. We applied our framework to 20+ ICI clinical cohorts across different cancer types and compared the prediction performance of our method with other well-known immune response markers evaluated using a suite of machine learning algorithms (random forest, logistic regression, and XGboosting) with cross-validation. We found that in some cohorts, the learned features from autoencoders improve the predictive power of ICI response when evaluated with the area under the operating characteristic curve. Our framework uncovers potential immune gene signatures associated with patient outcomes. Our work will help advance cancer immunology and immuno-oncology research, provide insights into tumor immunity, and lead to the discovery of progressively more immune markers and novel immune targets. Citation Format: Yang Liu, Aashna Jhaveri, Sudhenshna Bodapati, Cheryl Wong, Franziska Michor. ImmuneSig: A deep learning framework to develop gene expression signature of immune checkpoint inhibitor therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3165.

Abstract 6366: NB002, a novel therapeutic antibody targeting unique epitope on TIM-3 presenting potent antitumor activity

Abstract Antibody-based Immune checkpoint inhibitor (ICI) therapies targeting cytotoxic T lymphocyte antigen-4 (CTLA-4) or programmed death receptor-1 (PD-1) have significantly improved clinical outcomes in cancer treatment. Nonetheless, a large portion of the population has limited benefit from these ICI treatments due to lack of efficacy, resistance and intolerance. T-cell immunoglobulin and mucin domain containing-3 (TIM-3) has been proposed as a target for cancer immunotherapy. It is an inhibitory receptor modulating immune tolerance, and its expression has been broadly reported to be closely correlated to T cell exhaustion and pessimistic prognosis. Besides negatively modulating T cell status, TIM-3 is also expressed on natural killer (NK) cells, dendritic cells (DCs), macrophages and regulates immune responses bridging innate immunity and adaptive immunity. NB002 is a novel humanized Fc-engineered IgG1κ antibody, developed via NeologicsBio’s integrated target validation &amp; antibody screening Tier-A platform. It has been characterized in a serial of function and efficacy studies and completed IND-enabling preclinical development. NB002 binds to human TIM3 with high affinity in the low nanomolar range and antagonize the TIM-3 pathway effectively in reporter cell and primary cell assays. NB002 increased the secretion of IFN-γ by activated PBMC cells and enhanced activation and killing activity of NK cells. Moreover, it exhibited a distinctive activity of restoring myeloid cells and enhancing antigen presentation and processing during anti-tumor immune responses, and superior efficacy in multiple humanized mouse models of cancer as monotherapy regimen. Importantly, X-ray crystallography demonstrated that NB002 recognized a unique epitope on TIM-3 and was confirmed by epitope binning assay and structural alignment. These data could potentially explain novel features of NB002, restoring both acquired and innate immunity which could lead cold tumor to hot tumor.In summary, these data showed that NB002 is a potent anti-TIM-3 antibody with pre-clinical properties. A phase 1 study is planned and patients with advanced metastatic solid tumors will be recruited. Citation Format: Yu Zhang, Jinyu Dong, Binbin Wang, Jie Ni, Dong Wang, Liegang Shao, Meiguang Xiong, Ting Wang, Florencia McAllister, Xin Dong. NB002, a novel therapeutic antibody targeting unique epitope on TIM-3 presenting potent antitumor activity. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6366.

Abstract 1867: Pre-clinical characterization of anti-CEACAM1 antibody

Abstract Targeting checkpoint molecules cytotoxic T lymphocyte antigen-4 (CTLA-4) or programmed death receptor-1 (PD-1) expressed on immune cells has shown promising results in the treatment of cancer patients. Nevertheless only a small portion of population benefited from current ICI treatments, so there should be other mechanisms for immune evasion potentially. The carcinoembryonic antigen cell adhesion molecule 1 (CEACAM1) is an activation-induced inhibitory molecule on T and NK cells. In a wide range of cancers, it’s also expressed on tumor cells and other immune cells including macrophages and neutrophils. Homophilic interaction between CEACAM1 expressed on tumor cells and immune cells dampens NK and T cell function, mediates tumor migration, and promotes tumor angiogenesis, which implies that this homophilic interaction is an important mechanism for tumor progression. Here, we describe the identification and characterization of a novel humanized anti-human CEACAM1 therapeutic antibody developed via Neologics’ novel target validation and antibody screening Tier-A platform. This anti-CEACAM1 antibody specifically binds to recombinant human CEACAM1 with nM level affinity and significantly enhances T cell activities and NK killing on CEACAM1 expressing tumor cells, inhibits tumor cell migration in vitro as well. In humanized mouse models, this antibody evidently reduces tumor growth alone or combined with other immune checkpoint blockers. Taken together, we have demonstrated this anti-CEACAM1 antibody specifically enhance NK cell activity, modulate T cell immune activation and inhibit tumor migration supporting further clinical investigation for various indications. Citation Format: Jinyu Dong, Yu Zhang, Baiyang Wang, Tingting Pu, Liegang Shao, Binbin Wang, Dong Wang, Jie Ni, Sunan Li, Xin Dong. Pre-clinical characterization of anti-CEACAM1 antibody [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1867.

Immune checkpoint inhibitor-associated acute kidney injury in patients with cancer: a systematic review and meta-analysis of risk factors.

As a novel antineoplastic drug, immune checkpoint inhibitors (ICIs) are associated with a spectrum of autoimmune-related side effects, including acute kidney injury (AKI). Understanding the risk factors for immune-associated acute kidney injury will inform future symptom management measures to reduce this risk. This study aims to identify the risk factors for ICIs-AKI in cancer patients through a systematic review and meta-analysis. The systematic search was conducted in The Cochrane Library, Pubmed, Embase, Web of Science, China National Knowledge Infrastructure (CNKI), Wanfang Data, and VIP Database. The related studies published since the establishment of the database to Aug 22, 2022, were screened, data was extracted following the inclusion and exclusion criteria, and the quality of the selected studies was assessed by Newcastle-Ottawa Scale (NOS). The above was performed independently by the two reviewers. The estimated pooled odds ratios (ORs) for risk factors of developing ICIs-AKI were conducted by random-effects meta-analysis. A total of 8 publications, including 5267 patients, were included. Meta-analysis results showed that extrarenal immune-related adverse events (irAEs), therapy with the cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), male, hypertension, pre-existent use of a diuretic, and a proton pump inhibitor (PPI) were significantly correlated to ICIs-AKI. We identified extrarenal irAEs, CTLA-4 treatments, males, hypertension, previous use of diuretics, and PPIs are essential predictors of ICIs-AKI. These findings are helpful for healthcare providers to monitor ICIs-AKI for management and timely interventions.

Tumor microenvironment signaling and therapeutics in cancer progression.

Tumor development and metastasis are facilitated by the complex interactions between cancer cells and their microenvironment, which comprises stromal cells and extracellular matrix (ECM) components, among other factors. Stromal cells can adopt new phenotypes to promote tumor cell invasion. A deep understanding of the signaling pathways involved in cell-to-cell and cell-to-ECM interactions is needed to design effective intervention strategies that might interrupt these interactions. In this review, we describe the tumor microenvironment (TME) components and associated therapeutics. We discuss the clinical advances in the prevalent and newly discovered signaling pathways in the TME, the immune checkpoints and immunosuppressive chemokines, and currently used inhibitors targeting these pathways. These include both intrinsic and non-autonomous tumor cell signaling pathways in the TME: protein kinase C (PKC) signaling, Notch, and transforming growth factor (TGF-β) signaling, Endoplasmic Reticulum (ER) stress response, lactate signaling, Metabolic reprogramming, cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) and Siglec signaling pathways. We also discuss the recent advances in Programmed Cell Death Protein 1 (PD-1), Cytotoxic T-Lymphocyte Associated Protein 4 (CTLA4), T-cell immunoglobulin mucin-3 (TIM-3) and Lymphocyte Activating Gene 3 (LAG3) immune checkpoint inhibitors along with the C-C chemokine receptor 4 (CCR4)- C-C class chemokines 22 (CCL22)/ and 17 (CCL17), C-C chemokine receptor type 2 (CCR2)- chemokine (C-C motif) ligand 2 (CCL2), C-C chemokine receptor type 5 (CCR5)- chemokine (C-C motif) ligand 3 (CCL3) chemokine signaling axis in the TME. In addition, this review provides a holistic understanding of the TME as we discuss the three-dimensional and microfluidic models of the TME, which are believed to recapitulate the original characteristics of the patient tumor and hence may be used as a platform to study new mechanisms and screen for various anti-cancer therapies. We further discuss the systemic influences of gut microbiota in TME reprogramming and treatment response. Overall, this review provides a comprehensive analysis of the diverse and most critical signaling pathways in the TME, highlighting the associated newest and critical preclinical and clinical studies along with their underlying biology. We highlight the importance of the most recent technologies of microfluidics and lab-on-chip models for TME research and also present an overview of extrinsic factors, such as the inhabitant human microbiome, which have the potential to modulate TME biology and drug responses.

Comprehensive analysis of alternative polyadenylation regulators of CTLA4 and immune infiltration in clear cell renal cell carcinoma.

Clear cell renal cell carcinoma (ccRCC) is the most common renal cancer. Alternative polyadenylation (APA) plays an important role in the progression and immunity of multiple tumors. Although immunotherapy has emerged as an important treatment option for metastatic renal cell carcinoma, whether APA affects the tumor immune microenvironment (TIME) in ccRCC remains unclear. Patients with ccRCC were classified into two groups by performing a consensus clustering analysis of APA factor expression profiles. The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO) databases were used to assess the association between APA regulators and ccRCC prognosis. Through the use of the R package, GSVA, the correlation between SNRNP70 expression and tumor immune features were analyzed. The TCGA data revealed that APA regulators were associated with Cytotoxic T-Lymphocyte Associated Protein 4 (CTLA4) expression. Cluster 1 exhibited a higher grade and histological tumor stage, as well as a worse prognosis compared to Cluster 2. A ssGSEA analysis demonstrated that Cluster 2 possessed an extensively higher level of immune infiltration. Moreover, high SNRNP70 expression was found to be positively correlated with CTLA4 expression and a poor prognosis in ccRCC. Thus, SNRNP70 might represent a novel immune-related prognostic biomarker in ccRCC. A pan-cancer analysis suggested that SNRNP70 may also play a role in other types of cancer by affecting the TIME. The data from this study indicate that APA regulators play a key role in immune infiltration in ccRCC. SNRNP70 is a promising prognostic biomarker and a potential target for ccRCC's immunotherapy.