Improve Immunotherapy Response Research Articles

Reshaping the Tumor Microenvironment of KRASG12D Pancreatic Ductal Adenocarcinoma with combined SOS1 and MEK Inhibition for Improved Immunotherapy Response.

KRAS inhibitors have demonstrated exciting pre-clinical and clinical responses, although resistance occurs rapidly. Here, we investigate the effects of KRAS-targeting therapies on the tumor microenvironment using a library of KRASG12D, p53 mutant, murine PDAC-derived cell lines (KPCY) to leverage immune-oncology combination strategies for long-term tumor efficacy. Our findings show that SOS1 and MEK inhibitors (SOS1i+MEKi) suppressed tumor growth in syngeneic models and increased intra-tumoral CD8+ T cells without durable responses. scRNA-sequencing revealed an increase in inflammatory cancer associated fibroblasts (iCAFs), M2 macrophages, and a decreased dendritic cell quality that ultimately resulted in a highly immunosuppressive microenvironment driven by IL6+ iCAFs. Agonist CD40 treatment was effective to revert macrophage polarization and overcome the lack of mature antigen presenting DCs after SOS1i+MEKi therapy. Treatment increased the overall survival of KPCY tumor-bearing mice. The addition of checkpoint blockade to SOS1i+MEKi combination resulted in tumor free mice with established immune memory. Our data suggests that KRAS inhibition affects myeloid cell maturation and highlights the need for combining KRAS cancer-targeted therapy with myeloid activation to enhance and prolong anti-tumor effects.

Loss of heterozygosity impacts MHC expression on the immune microenvironment in CDK12-mutated prostate cancer

BackgroundIn prostate cancer (PCa), well-established biomarkers such as MSI status, TMB high, and PDL1 expression serve as reliable indicators for favorable responses to immunotherapy. Recent studies have suggested a potential association between CDK12 mutations and immunotherapy response; however, the precise mechanisms through which CDK12 mutation may influence immune response remain unclear. A plausible explanation for immune evasion in this subset of CDK12-mutated PCa may be reduced MHC expression.ResultsUsing genomic data of CDK12-mutated PCa from 48 primary and 10 metastatic public domain samples and a retrospective cohort of 53 low-intermediate risk primary PCa, we investigated how variation in the expression of the MHC genes affected associated downstream pathways. We classified the patients based on gene expression quartiles of MHC-related genes and categorized the tumors into “High” and “Low” expression levels. CDK12-mutated tumors with higher MHC-expressed pathways were associated with the immune system and elevated PD-L1, IDO1, and TIM3 expression. Consistent with an inflamed tumor microenvironment (TME) phenotype, digital cytometric analyses identified increased CD8 + T cells, B cells, γδ T cells, and M1 Macrophages in this group. In contrast, CDK12-mutated tumors with lower MHC expression exhibited features consistent with an immune cold TME phenotype and immunoediting. Significantly, low MHC expression was also associated with chromosome 6 loss of heterozygosity (LOH) affecting the entire HLA gene cluster. These LOH events were observed in both major clonal and minor subclonal populations of tumor cells. In our retrospective study of 53 primary PCa cases from this Institute, we found a 4% (2/53) prevalence of CDK12 mutations, with the confirmation of this defect in one tumor through Sanger sequencing. In keeping with our analysis of public domain data this tumor exhibited low MHC expression at the RNA level. More extensive studies will be required to determine whether reduced HLA expression is generally associated with primary tumors or is a specific feature of CDK12 mutated PCa.ConclusionsThese data show that analysis of CDK12 alteration, in the context of MHC expression levels, and LOH status may offer improved predictive value for outcomes in this potentially actionable genomic subgroup of PCa. In addition, these findings highlight the need to explore novel therapeutic strategies to enhance MHC expression in CDK12-defective PCa to improve immunotherapy responses.

Systematic investigation of chemo-immunotherapy synergism to shift anti-PD-1 resistance in cancer

Chemo-immunotherapy combinations have been regarded as one of the most practical ways to improve immunotherapy response in cancer patients. In this study, we integrate the transcriptomics data from anti-PD-1-treated tumors and compound-treated cancer cell lines to systematically screen for chemo-immunotherapy synergisms in silico. Through analyzing anti-PD-1 induced expression changes in patient tumors, we develop a shift ability score to measure if a chemotherapy or a small molecule inhibitor treatment can shift anti-PD-1 resistance in tumor cells. By applying shift ability analysis to 41,321 compounds and 16,853 shRNA treated cancer cell lines transcriptomic data, we characterize the landscape of chemo-immunotherapy synergism and experimentally validated a mitochondrial RNA-dependent mechanism for drug-induced immune activation in tumor. Our study represents an effort to mechanistically characterize chemo-immunotherapy synergism and will facilitate future pre-clinical and clinical studies.

Myeloid-T cell interplay and cell state transitions associated with checkpoint inhibitor response in melanoma.

The treatment of melanoma, the deadliest form of skin cancer, has greatly benefited from immunotherapy. However, many patients do not show a durable response, which is only partially explained by known resistance mechanisms. We performed single-cell RNA sequencing of tumor immune infiltrates and matched peripheral blood mononuclear cells of 22 checkpoint inhibitor (CPI)-naive stage III-IV metastatic melanoma patients. After sample collection, the same patients received CPI treatment, and their response was assessed. CPI responders showed high levels of classical monocytes in peripheral blood, which preferentially transitioned toward CXCL9-expressingmacrophages in tumors. Trajectories of tumor-infiltrating CD8+ Tcells diverged at the level of effector memory/stem-like Tcells, with non-responder cells progressing into a state characterized by cellular stress and apoptosis-related gene expression. Consistently, predicted non-responder-enriched myeloid-T/natural killer cell interactions were primarily immunosuppressive, while responder-enriched interactions were supportive of Tcell priming and effector function. Our study illustrates that the tumor immune microenvironment prior to CPI treatment can be indicative of response. In perspective, modulating the myeloid and/or effector cell compartment by altering the described cell interactions and transitions could improve immunotherapy response. This research was funded by Roche Pharma Research and Early Development.

Abstract 1369: PD-1 directly suppresses macrophage metabolism to restrict anti-tumor immunity in an obesity-cancer connection

Abstract Obesity has been established as a leading risk factor for many cancers and can drive tumor progression and metastasis. Paradoxically, obesity has in some cases been associated with better survival and improved response to immune checkpoint blockade therapies. The role of the immune system in the obesity-cancer connection and how obesity affects immunotherapy responses, however, have been unclear. Here we show that obesity enhanced PD-1 expression on macrophages to reduce phagocytosis and antigen presentation to T cells that correlated with reduced T cell expansion and function. This obesity associated immune dysfunction, however, primed for enhanced anti-tumor response to PD-1 blockade. Single cell RNAseq showed obesity remodels myeloid and T cell populations but does not impact non-immune cell populations such as fibroblasts and epithelial cells. Specifically, obesity increased the overall number of tumor-associated macrophages (TAM) while effector T cells were decreased in abundance. Interestingly, the frequency of macrophages expressing PD-1 increased, while the remaining T cells maintained similar or reduced PD-1 expression and appeared less activated. Further, T cell depletion from the tumor microenvironment, enhanced macrophage PD-1 expression and worsened PD1-associated TAM dysfunction in an obesity enhanced manor. Obesity associated cytokines and adipokines including IFN-ã, leptin and insulin induced PD-1 expression on macrophages. This expression is associated with mTOR and c-myc activation signaling pathways; as such inhibiting these signaling pathways blocked PD1 expression. Bulk RNA sequencing analysis revealed PD-1+ TAMs had an altered gene profile compared to PD-1− TAMs. PD1 expressing macrophages exhibited increased mitochondrial respiration and expression of oxidative phosphorylation, increased lipid uptake and increased cell cycling related genes. Conversely PD1- TAMs exhibited increased phagocytosis and antigen presentation. PD-1/PDL1 interaction directly regulated TAMs, as recombinant PDL-1 reduced glycolysis and phagocytosis in purified macrophages, and these effects could be reversed with blocking PD-1 antibody. Conversely, PD-1−/− TAMs had reduced lipid uptake but high rates of glycolysis, phagocytosis, and expression of MHC-II. Myeloid-specific PD-1 deficiency correlated with slower tumor growth and decreased LAG3 and increased CD69 on CD8 T cells. In addition, myeloid specific PD1 deficiency enhanced TAM antigen presentation through OVA specific CD8 T cell activation. These findings identify PD-1 as a metabolic regulator in TAM dysfunction and reveal a unique PD-1 mediated and macrophage-specific mechanism for immune tumor surveillance and checkpoint blockade. This may contribute to improved immunotherapy response in TAM-enriched tumors and obesity. Citation Format: Jackie Bader, Jeffrey Rathmell. PD-1 directly suppresses macrophage metabolism to restrict anti-tumor immunity in an obesity-cancer connection [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1369.

Neoadjuvant nivolumab or nivolumab plus LAG-3 inhibitor relatlimab in resectable esophageal/gastroesophageal junction cancer: a phase Ib trial and ctDNA analyses

Gastroesophageal cancer dynamics and drivers of clinical responses with immune checkpoint inhibitors (ICI) remain poorly understood. Potential synergistic activity of dual programmed cell death protein 1 (PD-1) and lymphocyte-activation gene 3 (LAG-3) inhibition may help improve immunotherapy responses for these tumors. We report a phase Ib trial that evaluated neoadjuvant nivolumab (Arm A, n = 16) or nivolumab–relatlimab (Arm B, n = 16) in combination with chemoradiotherapy in 32 patients with resectable stage II/stage III gastroesophageal cancer together with an in-depth evaluation of pathological, molecular and functional immune responses. Primary endpoint was safety; the secondary endpoint was feasibility; exploratory endpoints included pathological complete (pCR) and major pathological response (MPR), recurrence-free survival (RFS) and overall survival (OS). The study met its primary safety endpoint in Arm A, although Arm B required modification to mitigate toxicity. pCR and MPR rates were 40% and 53.5% for Arm A and 21.4% and 57.1% for Arm B. Most common adverse events were fatigue, nausea, thrombocytopenia and dermatitis. Overall, 2-year RFS and OS rates were 72.5% and 82.6%, respectively. Higher baseline programmed cell death ligand 1 (PD-L1) and LAG-3 expression were associated with deeper pathological responses. Exploratory analyses of circulating tumor DNA (ctDNA) showed that patients with undetectable ctDNA post-ICI induction, preoperatively and postoperatively had a significantly longer RFS and OS; ctDNA clearance was reflective of neoantigen-specific T cell responses. Our findings provide insights into the safety profile of combined PD-1 and LAG-3 blockade in gastroesophageal cancer and highlight the potential of ctDNA analysis to dynamically assess systemic tumor burden during neoadjuvant ICI that may open a therapeutic window for future intervention. ClinicalTrials.gov registration: NCT03044613.

Comprehensive Analyses of PANoptosome with Potential Implications in Cancer Prognosis and Immunotherapy.

Cell death resistance significantly contributes to poor therapeutic outcomes in various cancers. PANoptosis, a unique inflammatory programmed cell death (PCD) pathway activated by specific triggers and regulated by the PANoptosome, possesses key features of apoptosis, pyroptosis, and necroptosis, but these cannot be accounted for by any of the three PCD pathways alone. While existing studies on PANoptosis have predominantly centered on infectious and inflammatory diseases, its role in cancer malignancy has been understudied. In this comprehensive investigation, we conducted pan-cancer analyses of PANoptosome component genes across 33 cancer types. We characterized the genetic, epigenetic, and transcriptomic landscapes, and introduced a PANoptosome-related potential index (PANo-RPI) for evaluating the intrinsic PANoptosome assembly potential in cancers. Our findings unveil PANo-RPI as a prognostic factor in numerous cancers, including KIRC, LGG, and PAAD. Crucially, we established a significant correlation between PANo-RPI and tumor immune responses, as well as the infiltration of diverse lymphoid and myeloid cell subsets across nearly all cancer types. Moreover, a high PANo-RPI was consistently associated with improved immunotherapy response and efficacy, as evidenced by re-analysis of multiple immunotherapy cohorts. In conclusion, our study suggests that targeting PANoptosome components and modulating PANoptosis may hold tremendous therapeutic potential in the context of cancer.

L-DOS47 Elevates Pancreatic Cancer Tumor pH and Enhances Response to Immunotherapy.

Acidosis is an important immunosuppressive mechanism that leads to tumor growth. Therefore, we investigated the neutralization of tumor acidity to improve immunotherapy response. L-DOS47, a new targeted urease immunoconjugate designed to neutralize tumor acidity, has been well tolerated in phase I/IIa trials. L-DOS47 binds to CEACAM6, a cell-surface protein that is highly expressed in gastrointestinal cancers, allowing urease to cleave endogenous urea into two NH4+ and one CO2, thereby raising local pH. To test the synergetic effect of neutralizing tumor acidity with immunotherapy, we developed a pancreatic orthotopic murine tumor model (KPC961) expressing human CEACAM6. Using chemical exchange saturation transfer-magnetic resonance imaging (CEST-MRI) to measure the tumor extracellular pH (pHe), we confirmed that L-DOS47 raises the tumor pHe from 4 h to 96 h post injection in acidic tumors (average increase of 0.13 units). Additional studies showed that combining L-DOS47 with anti-PD1 significantly increases the efficacy of the anti-PD1 monotherapy, reducing tumor growth for up to 4 weeks.

CircMAPK1 promoted CD8 + T cell infiltration in LUAD by improving the IGF2BP1 dependent CCL5 upregulation

Lung adenocarcinoma (LUAD) is the most common pathological subtype of lung cancer and has a poor prognosis. Immune Checkpoint Blockage (ICB) have been shown to improve the survival of LUAD in the last decade. CD8 + T cell infiltration is significantly related to LUAD prognosis and plays a critical role in ICB response efficiency. Chemokines expressed and secreted by tumor and microenvironment cells regulate the recruitment of CD8 + T cells. A cytoplasm-dominant circRNA, termed circMAPK1, was found to be down-regulated in LUAD and dramatically suppressed the growth of LUAD upon circMAPK1 overexpression in immunocompetent mice. Meanwhile, it was found that circMAPK1 significantly promoted the CD8 + T cell intratumoral infiltration in vitro and in vivo. CircMAPK1 was identified as binding IGF2BP1 in the cytoplasm and inducing IGF2BP1 to occupy the 3′UTR of CCL5 mRNA, resulting in retained stability of CCL5 mRNA. In general, circMAPK1 is a microenvironment-associated circRNA that recruits CD8 + T cells in LUAD. CircMAPK1 is an effective microenvironment regulator and a potential nucleic acid drug that can be combined with ICB to improve immunotherapy response efficiency.

Regorafenib alone or in combination with high/low-dose radiotherapy plus toripalimab as third-line treatment in patients with metastatic colorectal cancer: protocol for a prospective, randomized, controlled phase II clinical trial (SLOT).

https://clinicaltrials.gov/study/NCT05963490?cond=NCT05963490&rank=1, identifier NCT05963490.

134P TERT-associated DNA polymerases genes link CAF and CD8+ T cells to improve immunotherapy response rate across multiple cancers

134P TERT-associated DNA polymerases genes link CAF and CD8+ T cells to improve immunotherapy response rate across multiple cancers

Decoding the Complexity of Immune-Cancer Cell Interactions: Empowering the Future of Cancer Immunotherapy.

The tumor and tumor microenvironment (TME) consist of a complex network of cells, including malignant, immune, fibroblast, and vascular cells, which communicate with each other. Disruptions in cell-cell communication within the TME, caused by a multitude of extrinsic and intrinsic factors, can contribute to tumorigenesis, hinder the host immune system, and enable tumor evasion. Understanding and addressing intercellular miscommunications in the TME are vital for combating these processes. The effectiveness of immunotherapy and the heterogeneous response observed among patients can be attributed to the intricate cellular communication between immune cells and cancer cells. To unravel these interactions, various experimental, statistical, and computational techniques have been developed. These include ligand-receptor analysis, intercellular proximity labeling approaches, and imaging-based methods, which provide insights into the distorted cell-cell interactions within the TME. By characterizing these interactions, we can enhance the design of cancer immunotherapy strategies. In this review, we present recent advancements in the field of mapping intercellular communication, with a particular focus on immune-tumor cellular interactions. By modeling these interactions, we can identify critical factors and develop strategies to improve immunotherapy response and overcome treatment resistance.

PD-14 TERT-associated DNA polymerases genes link CAF and CD8+ T cells to improve immunotherapy response rate across multiple cancers

PD-14 TERT-associated DNA polymerases genes link CAF and CD8+ T cells to improve immunotherapy response rate across multiple cancers

Comprehensive analysis of 33 human cancers reveals clinical implications and immunotherapeutic value of the solute carrier family 35 member A2.

Solute carrier family 35 member A2 (SLC35A2), which belongs to the SLC35 solute carrier family of human nucleoside sugar transporters, has shown regulatory roles in various tumors and neoplasms. However, the function of SLC35A2 across human cancers remains to be systematically assessed. Insights into the prediction ability of SLC35A2 in clinical practice and immunotherapy response remains limited. We obtained the gene expression and protein levels of SLC35A2 in a variety of tumors from Molecular Taxonomy of Breast Cancer International Consortium, The Cancer Genome Atlas, Gene Expression Omnibus, Chinese Glioma Genome Atlas, and Human Protein Atlas databases. The SLC35A2 level was validated by immunohistochemistry. The predictive value for prognosis was evaluated by Kaplan-Meier survival and Cox regression analyses. Correlations between SLC35A2 expression and DNA methylation, genetic alterations, tumor mutation burden (TMB), microsatellite instability (MSI), and tumor microenvironment were performed using Spearman's correlation analysis. The possible downstream pathways of SLC35A2 in different human cancers were explored using gene set variation analysis. The potential role of SLC35A2 in the tumor immune microenvironment was evaluated via EPIC, CIBERSORT, MCP-counter, CIBERSORT-ABS, quanTIseq, TIMER, and xCell algorithms. The difference in the immunotherapeutic response of SLC35A2 under different expression conditions was evaluated by the tumor immune dysfunction and exclusion (TIDE) score as well as four independent immunotherapy cohorts, which includes patients with bladder urothelial carcinoma (BLCA, N = 299), non-small cell lung cancer (NSCLC, N = 72 and N = 36) and skin cutaneous melanoma (SKCM, N = 25). Potential drugs were identified using the CellMiner database and molecular docking. SLC35A2 exhibited abnormally high or low expression in 23 cancers and was significantly associated with the prognosis. In various cancers, SLC35A2 expression and mammalian target of rapamycin complex 1 signaling were positively correlated. Multiple algorithmic immune infiltration analyses suggested an inverse relation between SLC35A2 expression and infiltrating immune cells, which includes CD4+T cells, CD8+T cells, B cells, and natural killer cells (NK) in various tumors. Furthermore, SLC35A2 expression was significantly correlated with pan-cancer immune checkpoints, TMB, MSI, and TIDE genes. SLC35A2 showed significant predictive value for the immunotherapy response of patients with diverse cancers. Two drugs, vismodegib and abiraterone, were identified, and the free binding energy of cytochrome P17 with abiraterone was higher than that of SLC35A2 with abiraterone. Our study revealed that SLC35A2 is upregulated in 20 types of cancer, including lung adenocarcinoma (LUAD), breast invasive carcinoma (BRCA), colon adenocarcinoma (COAD), and lung squamous cell carcinoma (LUSC). The upregulated SLC35A2 in five cancer types indicates a poor prognosis. Furthermore, there was a positive correlation between the overexpression of SLC35A2 and reduced lymphocyte infiltration in 13 cancer types, including BRCA and COAD. Based on data from several clinical trials, patients with LUAD, LUSC, SKCM, and BLCA who exhibited high SLC35A2 expression may experience improved immunotherapy response. Therefore, SLC35A2 could be considered a potential predictive biomarker for the prognosis and immunotherapy efficacy of various tumors. Our study provides a theoretical basis for further investigating its prognostic and therapeutic potentials.

Abstract 3277: In vivo CRISPR screening identifies PTDSS1 as a potential combinatorial target to improve immune checkpoint therapy response

Abstract Immune checkpoint therapy (ICT) has recently revolutionized the treatment paradigm for various types of cancer. However, despite the unprecedented safety and efficacy profile compared to traditional treatment options, many patients do not respond to ICT, highlighting the urgent need for novel strategies to improve ICT efficacy. Here, we used a pooled in vivo CRISPR knockout screening in a mouse bladder cancer model treated with immune checkpoint inhibitor to discover targets that may improve immunotherapy response. Our data identified Ptdss1, which encodes phosphatidylserine synthase 1, as a potential regulator of anti-PD-1 response. Depletion of Ptdss1 not only increased tumor cell immunogenicity and response to interferon gamma, but also facilitated development of an anti-tumor microenvironment. Ptdss1 depletion in tumor cells increased effector CD8+ T cells cytolytic function, decreased regulatory T cells activation and inhibitory function, increased frequency of iNOS+ myeloid cells and induced macrophage M2-to-M1 polarization. Ptdss1 knockdown induced changes in the tumor microenvironment were further amplified when combined with anti-PD-1 treatment, which translates into better tumor control and prolonged animal survival in mouse bladder cancer model. Together, our results provide insights on a new therapeutic strategy for overcoming resistance to immune checkpoint therapy through combination of PTDSS1 inhibition and PD-1 blockade. Citation Format: Jielin Liu, Swetha Anandhan, Sreyashi Basu, Yulong Chen, Sangeeta Goswami, Ashwat Nagarajan, Shelley Herbrich, Liangwen Xiong, Baoxiang Guan, Padmanee Sharma. In vivo CRISPR screening identifies PTDSS1 as a potential combinatorial target to improve immune checkpoint therapy response [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 3277.

Abstract 6646: Inhibition of HuR/CD147/IL-6 axis enhances anti-PD1 immunotherapy response in cancer

Abstract Immune checkpoint blockade (ICB) targeting PD-1/PD-L1 has emerged as a revolutionizing treatment modality for multiple cancers. However, durable remission and objective response rates with ICB monotherapy remain low. Several immune resistance mechanisms have been revealed and used to develop combinational therapy. In consideration of the durable and impressive responses observed in ‘hot’ tumors, it will be valuable to make an endeavor to elucidate the tumor evasion mechanisms. Despite the well-established mechanisms in immunotherapy resistance, our study uncovered a new mechanism of tumor immune evasion through the RNA-binding protein HuR/CD147 axis. The RNA-binding protein Hu antigen R (HuR) is overexpressed in a variety of cancers and promotes tumorigenesis by interacting with a subset of oncogenic mRNAs. The tumor-associated antigen CD147, a highly glycosylated transmembrane immunoglobulin, is widely and specifically expressed in multiple malignancies. We phenotypically observed the downregulation of CD147 in HuR-deficient cells and elucidated that HuR regulates CD147 mRNA via binding to the AU-rich elements of its 3' UTR. The direct binding of HuR to CD147 mRNA in multiple types of cells was validated by Ribonucleoprotein immunoprecipitation (RNP-IP) and RNA pulldown assay. These data together clarify HuR is engaged in CD147 post-transcription regulation. KH-39 is a HuR-specific small molecule inhibitor, which disrupts the HuR function by competing for the binding site with its target mRNAs. KH-39 treatment reduced CD147 level in a time-/dose-dependent manner. We also tested the anti-tumor action of HuR small molecular inhibitor KH-39 in tumor proliferation, clonogenicity, cell death, cytokine secretion, and in vivo tumor growth. The cytokine array assay indicated multiple immune-related cytokines changed after HuR inhibition and CD147 knockdown, including IL-6, which is proven to act as an immunomodulating agent in the tumor microenvironment. These data demonstrated the potential mechanism of overcoming immunoresistance by inhibiting the HuR/CD147/IL-6 axis. To test the combination strategy using HuR inhibitor and anti-PD-1 antibody in vivo, we established the EMT6 orthotopic mouse breast cancer model. A combination of HuR inhibitor KH-39 and anti-PD-1 antibody enhanced the anti-PD1 immunotherapy response and prolonged survival compared to the single agent group. Taken together, we preliminarily explored the HuR/CD147/IL-6 axis in tumor immune evasion, which might be a promising therapeutic target to overcome immune evasion and improve immunotherapy response in cancer. This finding may have reference significance for clinical cancer therapy. Citation Format: Qi Zhang, Lauren Dandreo, Xiaoqing Wu, Liang Xu. Inhibition of HuR/CD147/IL-6 axis enhances anti-PD1 immunotherapy response in cancer [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 6646.

Abstract 5841: Exploring the role of versican in immune exclusion within triple negative breast cancer

Abstract Immunotherapy has found limited success in solid tumors such as triple negative breast cancer (TNBC). The poor response can be attributed to the presence of the extracellular matrix (ECM) that can act biochemically or physically to interfere with cytotoxic T cell migration, leading to a limitation in tumor cell killing. Through previous work on deconstructing this matrix and outlining the spatial arrangement of proteins, we were able to identify versican as a key protein in T cell location. Versican is a chondroitin sulfate proteoglycan and has multiple functions in the ECM due to its ability to bind many ECM components. Using immunohistochemistry in this study, we analyzed the localization of CD8+ T cells within human TNBC tissues to classify them as inflamed (tumorhi, stromahi), desert (tumorlo, stromalo) or immune excluded (tumorlo, stromahi). From the analysis of 26 tissues, nearly half were classified as immune excluded with most of the CD8+ T cells restricted to the stroma, whilst only a quarter were inflamed. When comparing this to versican accumulation we noticed that immune excluded tissues have significantly higher levels of versican in the tumor epithelium compared to inflamed tissues where versican is mostly found within the stroma. To investigate the function of versican’s structure, we carried out transwell migration assays using versican enriched protein treated with or without chondroitinase ABC and showed that the chondroitin sulfates attached to versican’s peptide backbone are an important factor towards T cell trafficking with a reduction in migration following chondroitinase ABC treatment. Comparison of versican levels to chondroitin sulfate via immunohistochemistry showed a positive correlation within inflamed tissues, however no correlation was observed in immune excluded tissues. Versican isoforms expression in TNBC tissues and cell lines was examined with qRT-PCR which showed that within all tissues and cell lines the V0 and V1 isoform are the highest expressed out of the 5 isoforms. Overall, in this work, we have found that versican is differentially expressed in excluded tissues in comparison to inflamed tissues. With chondroitin sulfate shown to play a major role in immune cell trafficking, understanding how it differs between inflamed and excluded tumors can help identify potential targets to improve immunotherapy response. Citation Format: Priyanka Hirani, Kimberly M. Alonge, Daniel J. Pennington, Pedro R. Cutillas, Thomas N. Wight, Oliver M. Pearce. Exploring the role of versican in immune exclusion within triple negative breast cancer. [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 5841.

DNA barcoding reveals ongoing immunoediting of clonal cancer populations during metastatic progression and immunotherapy response

Cancers evade the immune system through the process of cancer immunoediting. While immune checkpoint inhibitors are effective for reactivating tumour immunity in some cancer types, many other solid cancers, including breast cancer, remain largely non-responsive. Understanding how non-responsive cancers evade immunity and whether this occurs at the clonal level will improve immunotherapeutic design. Here we use DNA barcoding to track murine mammary cancer cell clones during immunoediting and determine clonal transcriptional profiles that allow immune evasion following anti-PD1 plus anti-CTLA4 immunotherapy. Clonal diversity is significantly restricted by immunotherapy treatment in both primary tumours and metastases, demonstrating selection for pre-existing breast cancer cell populations and ongoing immunoediting during metastasis and treatment. Immunotherapy resistant clones express a common gene signature associated with poor survival of basal-like breast cancer patient cohorts. At least one of these genes has an existing small molecule that can potentially be used to improve immunotherapy response.

Cancer stem/progenitor signatures refine the classification of clear cell renal cell carcinoma with stratified prognosis and decreased immunotherapy efficacy

Recent studies suggest that cancer stemness drives the acquired drug resistance process in cancer therapy. The complementary information provided by multi-omics data can help us to gain a deeper understanding of this process. This study aims to elucidate the impact of cancer stemness on the frontline treatment of clear cell renal cell carcinoma (ccRCC). Consensus clustering based on stem/progenitor signatures refined 3 subgroups in 1,730 tumor samples. We identified master regulons that regulate cancer stemness phenotypes, including key transcription factors, DNA methyltransferases, and promoter methylation probes. In addition, we compared the clinicopathological traits, genomic heterogeneity, cancer hallmarks, tumor microenvironment (TME), and oncological prognosis of the stemness subgroups. Cancer stemness was correlated with reduced efficiency of immune checkpoint blockade therapy. Cancer stemness profoundly affects the prognosis and treatment outcome of ccRCC by increasing genomic instability, tumor-associated malignant events, and immunosuppressive factors. For clinical application, we established and validated a 243-gene signature from stem/progenitor-related genes to distinguish anti-PD-1 outcomes. Overall, this presented study suggested that cancer stemness leads to adaptive resistance to anti-PD-1 treatment in CD8+ T-infiltrated ccRCC and provides a new reference for strategy development to further improve immunotherapy response rates.

Hippo Signaling Pathway Regulates Cancer Cell-Intrinsic MHC-II Expression.

MHC-II is known to be mainly expressed on the surface of antigen-presenting cells. Evidence suggests MHC-II is also expressed by cancer cells and may be associated with better immunotherapy responses. However, the role and regulation of MHC-II in cancer cells remain unclear. In this study, we leveraged data mining and experimental validation to elucidate the regulation of MHC-II in cancer cells and its role in modulating the response to immunotherapy. We collated an extensive collection of omics data to examine cancer cell-intrinsic MHC-II expression and its association with immunotherapy outcomes. We then tested the functional relevance of cancer cell-intrinsic MHC-II expression using a syngeneic transplantation model. Finally, we performed data mining to identify pathways potentially involved in the regulation of MHC-II expression, and experimentally validated candidate regulators. Analyses of preimmunotherapy clinical samples in the CheckMate 064 trial revealed that cancer cell-intrinsic MHC-II protein was positively correlated with more favorable immunotherapy outcomes. Comprehensive meta-analyses of multiomics data from an exhaustive collection of data revealed that MHC-II is heterogeneously expressed in various solid tumors, and its expression is particularly high in melanoma. Using a syngeneic transplantation model, we further established that melanoma cells with high MHC-II responded better to anti-PD-1 treatment. Data mining followed by experimental validation revealed the Hippo signaling pathway as a potential regulator of melanoma MHC-II expression. In summary, we identified the Hippo signaling pathway as a novel regulator of cancer cell-intrinsic MHC-II expression. These findings suggest modulation of MHC-II in melanoma could potentially improve immunotherapy response.