Immune Gene Expression Signature Research Articles

Abstract PD9-03: Genomic analysis of the CALGB 40603 (Alliance) neoadjuvant trial in TNBC identifies immune features associated with pathological complete response and event-free survival

Abstract Purpose: CALGB (now part of the Alliance for Clinical Trials in Oncology) 40603 was a randomized Phase II study investigating pathological complete response rates (pCR) in triple-negative breast cancer (TNBC) patients receiving neoadjuvant weekly paclitaxel followed by doxorubicin and cyclophosphamide +/- bevacizumab and/or carboplatin for which 5-year event-free survival (EFS) results are now available. This is a valuable resource to characterize clinical and genomic features associated with response and survival. Methods: Clinical parameters and pre-treatment tumor biopsy RNA-sequencing (RNAseq) from 295 TNBC patients were analyzed to identify features associated with pCR and/or EFS. A panel of 689 previously published gene expression signatures were evaluated to provide insights regarding potential associations between clinical endpoints and genomically determined cell types and signaling pathway activities. Additionally, B cell receptor (BCR) and T cell receptor (TCR) sequences were examined, and repertoire abundance, richness, and diversity measures were calculated to investigate correlations with pCR and EFS status. Univariate Mann-Whitney U-Tests were used for continuous variables, and Fisher’s test was used for categorical features, with unadjusted p-values < 0.05 designated as significant. Results: While the addition of bevacizumab and carboplatin each significantly improved pCR rates, neither improved EFS. We examined outcomes according to race and no differences were seen for either pCR rate or EFS. 131 features, including high proliferation and multiple interferon signatures were significantly associated with pCR, but not with EFS. Alternatively, 69 features, including clinical factors for T stage and node status, were prognostic for EFS, but not significantly associated with pCR. Nevertheless, pCR itself was the strongest predictor of EFS, and was the only feature significantly associated with EFS after adjusting for multiple comparisons (Benjamini-Hochberg False Discovery Rate = 6.7e-3). In total, 52 genomic features were significantly correlated with both pCR and EFS, 44 of which were features of the immune microenvironment. Immune associated features included signatures of T cells, B cells and NK cells, immune checkpoint pathways (PD-1, PD-L1, CTLA4), and antigen presentation (dendritic cells, MHC-I, MHC-II). In particular, low BCR evenness, which is a measure of uniformity of unique BCR sequence abundance, was strongly associated with both pCR and EFS, suggesting that an antigen-specific adaptive immune response with clonally selected B cells is occurring in patients that have improved response and survival. Furthermore, a multivariate Cox Proportional Hazards model assessing BCR evenness along with age, T stage, N stage, grade and pCR found BCR evenness to be an independent prognostic feature for EFS in TNBC. Conclusions: Evidence of distinct predictors of pCR and EFS in TNBC patients treated with neoadjuvant chemotherapy suggests that, while pCR is still the strongest prognostic feature, high expression of many immune related gene expression signatures in pretreatment tumor samples are promising biomarkers of improved EFS. In addition to the important role of T cells in an anti-tumor response, these data show high IgG expression and evidence of B cell clonal selection associates with improved response and survival, supporting an important role for B cells in the adaptive response that portends a long-term benefit of chemotherapy in TNBC. Support: U10CA180821, U10CA180882, U24CA196171, P50-CA58223, Genentech, and The Breast Cancer Research Foundation. https://acknowledgments.alliancefound.org; ClinicalTrials.gov Identifier: NCT00861705 Citation Format: Jonathan H Shepherd, Terry Hyslop, Cheng Fan, Aranzazu Fernandez Martinez, Joel Parker, Katherine Hoadley, Zhiyuan Hu, Yun Li, Matthew Soloway, Patricia Spears, Ann Partridge, William Sikov, Lisa A Carey, Charles M Perou. Genomic analysis of the CALGB 40603 (Alliance) neoadjuvant trial in TNBC identifies immune features associated with pathological complete response and event-free survival [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PD9-03.

Molecular analyses of triple-negative breast cancer in the young and elderly

BackgroundBreast cancer in young adults has been implicated with a worse outcome. Analyses of genomic traits associated with age have been heterogenous, likely because of an incomplete accounting for underlying molecular subtypes. We aimed to resolve whether triple-negative breast cancer (TNBC) in younger versus older patients represent similar or different molecular diseases in the context of genetic and transcriptional subtypes and immune cell infiltration.Patients and methodsIn total, 237 patients from a reported population-based south Swedish TNBC cohort profiled by RNA sequencing and whole-genome sequencing (WGS) were included. Patients were binned in 10-year intervals. Complimentary PD-L1 and CD20 immunohistochemistry and estimation of tumor-infiltrating lymphocytes (TILs) were performed. Cases were analyzed for differences in patient outcome, genomic, transcriptional, and immune landscape features versus age at diagnosis. Additionally, 560 public WGS breast cancer profiles were used for validation.ResultsMedian age at diagnosis was 62 years (range 26–91). Age was not associated with invasive disease-free survival or overall survival after adjuvant chemotherapy. Among the BRCA1-deficient cases (82/237), 90% were diagnosed before the age of 70 and were predominantly of the basal-like subtype. In the full TNBC cohort, reported associations of patient age with changes in Ki67 expression, PIK3CA mutations, and a luminal androgen receptor subtype were confirmed. Within DNA repair deficiency or gene expression defined molecular subgroups, age-related alterations in, e.g., overall gene expression, immune cell marker gene expression, genetic mutational and rearrangement signatures, amount of copy number alterations, and tumor mutational burden did, however, not appear distinct. Similar non-significant associations for genetic alterations with age were obtained for other breast cancer subgroups in public WGS data. Consistent with age-related immunosenescence, TIL counts decreased linearly with patient age across different genetic TNBC subtypes.ConclusionsAge-related alterations in TNBC, as well as breast cancer in general, need to be viewed in the context of underlying genomic phenotypes. Based on this notion, age at diagnosis alone does not appear to provide an additional layer of biological complexity above that of proposed genetic and transcriptional phenotypes of TNBC. Consequently, treatment decisions should be less influenced by age and more driven by tumor biology.

Identification of the immune gene expression signature associated with recurrence of high-grade gliomas.

High-grade gliomas (HGGs), the most common and aggressive primary brain tumors in adults, inevitably recur due to incomplete surgery or resistance to therapy. Intratumoral genomic and cellular heterogeneity of HGGs contributes to therapeutic resistance, recurrence, and poor clinical outcomes. Transcriptomic profiles of HGGs at recurrence have not been investigated in detail. Using targeted sequencing of cancer-related genes and transcriptomics, we identified single nucleotide variations, small insertions and deletions, copy number aberrations (CNAs), as well as gene expression changes and pathway deregulation in 16 pairs of primary and recurrent HGGs. Most of the somatic mutations identified in primary HGGs were not detected after relapse, suggesting a subclone substitution during the tumor progression. We found a novel frameshift insertion in the ZNF384 gene which may contribute to extracellular matrix remodeling. An inverse correlation of focal CNAs in EGFR and PTEN genes was detected. Transcriptomic analysis revealed downregulation of genes involved in messenger RNA splicing, cell cycle, and DNA repair, while genes related to interferon signaling and phosphatidylinositol (PI) metabolism are upregulated in secondary HGGs when compared to primary HGGs. In silico analysis of the tumor microenvironment identified M2 macrophages and immature dendritic cells as enriched in recurrent HGGs, suggesting a prominent immunosuppressive signature. Accumulation of those cells in recurrent HGGs was validated by immunostaining. Our findings point to a substantial transcriptomic deregulation and a pronounced infiltration of immature dendritic cells in recurrent HGG, which may impact the effectiveness of frontline immunotherapies in the GBM management. KEY MESSAGES: Most of the somatic mutations identified in primary HGGs were not detected after relapse. Focal CNAs in EGFR and PTEN genes are inversely correlated in primary and recurrent HGGs. Transcriptomic changes and distinct immune-related signatures characterize HGG recurrence. Recurrent HGGs are characterized by a prominent infiltration of immature dendritic and M2 macrophages.

CTIM-32. PHASE II AND BIOMARKER STUDY OF PEMBROLIZUMAB OR PEMBROLIZUMAB PLUS BEVACIZUMAB FOR RECURRENT GLIOBLASTOMA PATIENTS

Abstract PURPOSE Vascular endothelial growth factor (VEGF) is upregulated in glioblastoma and may contribute to immunosuppression. We performed a phase 2 study of pembrolizumab, a programmed death-1 (PD-1) blocking antibody alone or with the anti-VEGF antibody bevacizumab in recurrent glioblastoma with detailed analyses of biomarkers and patient neurologic function. METHODS Eighty bevacizumab-naive, recurrent glioblastoma patients were randomized to receive pembrolizumab with bevacizumab (cohort A, n=50) or pembrolizumab monotherapy (cohort B, n=30). The primary endpoint was six-month progression-free survival (PFS-6). Exploratory endpoints included evaluation of tumor PD-L1 expression, TIL density, immune activation gene expression signature and plasma cytokines with outcome. Changes in neurologic function were prospectively assessed using the Neurologic Assessment in Neuro-Oncology (NANO) scale. RESULTS Pembrolizumab alone or with bevacizumab was well tolerated but of limited benefit. For cohort A, PFS-6 was 26.0% (95% CI: 16.3, 41.5), median OS was 8.8 months (95% CI: 7.7, 14.2), ORR was 20% and median duration of response was 48 weeks. For cohort B, PFS-6 was 6.7% (95% CI: 1.7, 25.4), median OS was 10.3 months (95% CI: 8.5, 12.5) and ORR was 0%. Factors associated with worsened OS included baseline dexamethasone use and increased post-therapy plasma VEGF (cohort A) and wild-type IDH1, unmethylated MGMT and increased baseline PlGF and sVEGFR1 levels (cohort B), but tumor immune markers were not informative. The NANO scale effectively predicted neurologic function. CONCLUSIONS Although well tolerated, pembrolizumab was ineffective both as monotherapy and with bevacizumab for recurrent glioblastoma. Nonetheless, radiographic responses to combinatorial therapy were durable. Baseline dexamethasone use and plasma cytokines but not tumor immunologic biomarkers were associated with outcome. Neurologic function evaluated by the NANO scale contributed to outcome assessment.

EXTH-67. PHARMACOLOGIC INHIBITION OF LYSINE SPECIFIC DEMETHYLASE-1 (LSD1) AS AN ADJUVANT IMMUNE-SENSITIZATION STRATEGY IN DIFFUSE INTRINSIC PONTINE GLIOMA (DIPG)

Abstract Diffuse midline gliomas (DMG), including brainstem DIPG (diffuse intrinsic pontine glioma), are incurable pediatric brain tumors. Mutations in the H3 histone tail (H3.1/3.3-K27M) are a feature of DIPG, highlighting epigenetic aberrations in these tumors. Lysine specific demethylase-1 (LSD1), is an enzyme that acts upon mono and di-methylated histone H3K4/K9, and pharmacological inhibitors of its enzymatic and scaffolding domains are clinically viable. Patient-derived DIPG cell lines, orthotopic mouse models, and pediatric high-grade glioma (pHGG) datasets were used to evaluate effects of LSD1 inhibitors on cytotoxicity and immune gene expression. Because an unique immune signature was sen with enzymatic LSD1 inhibitors in DIPG cells in vitro, immune cell cytotoxicity was assessed in DIPG cells pre-treated with LSD1 inhibitors and informatics platforms were used to determine immune infiltration of pHGG. Selective cytotoxicity and an immunogenic gene expression signature was established in DIPG cell lines and in brainstem tumor-bearing mice using clinically-relevant LSD1 inhibitors. Pediatric HGG patient sequencing data demonstrated survival benefit using this LSD1-dependent gene expression signature. On-target binding of catalytic LSD1 inhibitors was confirmed in DIPG and pre-treatment of DIPG with these inhibitors increased lysis by natural killer (NK) cells. CIBERSORT analysis of patient data confirmed NK infiltration is beneficial to patient survival while CD8 T-cells are negatively prognostic. Catalytic LSD1 inhibitors are non-perturbing to NK cells while scaffolding LSD1 inhibitors are toxic to NK cells and do not induce the gene expression signature in DIPG cells. Treatment of orthotopic xenograft bearing DIPG mouse models with catalytic LSD1 inhibitors and NK cells showed a significant reduction in tumor burden. In summary, we find that LSD1 inhibition using catalytic inhibitors is both selectively cytotoxic and promotes a functional immune gene expression signature that is associated with NK cell killing, representing a therapeutic opportunity for pHGG.

699O Avelumab first-line (1L) maintenance + best supportive care (BSC) vs BSC alone for advanced urothelial carcinoma (UC): Association between clinical outcomes and exploratory biomarkers

In the phase 3 JAVELIN Bladder 100 trial (NCT02603432), avelumab (anti–PD-L1 IgG1 antibody) 1L maintenance + BSC significantly prolonged overall survival (OS) vs BSC alone in patients (pts) with advanced UC without disease progression with 1L induction chemotherapy in all randomized pts and pts with PD-L1+ tumors. We report exploratory biomarker analyses. Tumor biopsies were collected prior to randomization. Tumor analyses included PD-L1 (Ventana SP263 assay) and CD8 immunohistochemistry, whole-exome sequencing (including FCGR2A and FCGR3A genotypes), whole-transcriptome sequencing, and T-cell receptor (TCR) sequencing. Analyses of peripheral blood included baseline C-reactive protein (CRP) level and neutrophil:lymphocyte ratio (NLR) as well as TCR sequencing before and on treatment. Associations between OS or progression-free survival and biomarkers were evaluated using the Cox proportional hazards model. Hazard ratios and nominal p values were reported, and no multiplicity adjustments were made (5% level for statistical significance). Increased OS benefit with avelumab 1L maintenance + BSC vs BSC alone was positively associated with CD8+ T cells in the invasive margin or tumor center, high tumor mutation burden, and tumor gene expression signatures of innate and adaptive immune response. Increased OS benefit was also positively associated with the number of FCGR2A/FCGR3A alleles encoding high-affinity Fcγ receptors for IgG1. OS benefit with avelumab was negatively associated with tumor epithelial cell gene signatures, mutations affecting fibroblast growth factor receptor signaling, CRP, and NLR. In pts with elevated immune gene expression signatures, lower TGFb expression was associated with greater OS benefit with avelumab. Further analyses will be presented, including outcomes by exploratory PD-L1 cutoffs in tumor vs immune cells. OS benefits of avelumab 1L maintenance in pts with advanced UC are positively associated with biomarkers of immune activity and negatively associated with biomarkers of tumor homeostasis and chronic inflammation.

531MO VHIO immune gene-expression signature (VIGex) to enrich patient selection in immunotherapy (IT) phase I clinical trials

531MO VHIO immune gene-expression signature (VIGex) to enrich patient selection in immunotherapy (IT) phase I clinical trials

DREIMT: a drug repositioning database and prioritization tool for immunomodulation

Drug immunomodulation modifies the response of the immune system and can be therapeutically exploited in pathologies such as cancer and autoimmune diseases. DREIMT is a new hypothesis-generation web tool, which performs drug prioritization analysis for immunomodulation. DREIMT provides significant immunomodulatory drugs targeting up to 70 immune cells subtypes through a curated database that integrates 4960 drug profiles and ∼2600 immune gene expression signatures. The tool also suggests potential immunomodulatory drugs targeting user-supplied gene expression signatures. Final output includes drug-signature association scores, FDRs and downloadable plots and results tables. http://www.dreimt.org. Supplementary data are available at Bioinformatics online.

Abstract 4484: ScRNA-seq of patients with immune-related adverse events and checkpoint inhibitor therapy

Abstract Despite the remarkable success of immune checkpoint inhibitor (ICI) therapy, a significant number of patients develop severe and unpredictable immune-related adverse events (irAEs) affecting a wide variety of organs. Concerns over irAE have led to the exclusion of patients with autoimmune disease from ICI clinical trials. Role of host genetic and immune factors in mediating irAEs remain unclear and it is not clear if the manifestations of irAEs is associated with response to therapy. In this study, we used single cell RNA sequencing (ScRNA-seq) to study the peripheral immune repertoire at a single cell level to identify blood-based signatures predictive of irAEs and response to ICI therapy. We characterized changes in host immune system in 40 patients receiving ICI therapy by performing ScRNA-seq on peripheral blood mononuclear cells (PBMCs) at baseline and post-immunotherapy. In addition, we evaluated serum levels of 40 cytokines/chemokines, antinuclear autoantibodies (ANA) and 124 autoantibodies and performed bulk RNA sequencing on PBMCs at baseline and post immunotherapy in these patients. Our preliminary data analysis identified signatures of autoantibodies and cytokines correlating with response and toxicity. ScRNA-seq data revealed changes in distinct immune cell subsets and gene expression signatures in patient developing irAEs. In this meeting, we will present preliminary ScRNA-seq data combined with immune and genetic correlates of irAEs and response to therapy in our patient cohort. We hope that our studies can help identify blood-based biomarker signatures predictive of irAEs and/or response and reveal novel insights into the mechanisms underlying irAE. Our current genetic data suggests further expanding the genetic studies in larger patient population to identify the role of underlying genetic predisposition to autoimmunity in mediating irAEs. We hope our findings may ultimately help customize therapy, expand use of immunotherapy and prevent toxicities. Citation Format: Shaheen Khan, David E. Gerber. ScRNA-seq of patients with immune-related adverse events and checkpoint inhibitor therapy [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4484.

Abstract 5719: Tumor-suppressive stromal activity of pro-fibrogenic microRNA-21 in initiation and progression of K-Ras-driven mouse models of pancreatic cancer

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a recalcitrant and lethal disease with an overall 5-year survival rate of less than 8%. The abundance and heterogeneity of cancer-associated fibroblasts (CAFs) in a highly desmoplastic tumor microenvironment are unique features of PDAC. Therapeutic strategies aimed at indiscriminately eliminating CAFs have failed in clinical trials. The opposing tumor-restraining or tumor-promoting role of distinct CAFs subpopulations of has emerged as a new paradigm in PDAC basic and translational research. Understanding how the composition of these distinct CAF subpopulations is affected by and interferes with current treatments is a key question to solve in order to obtain more effective and durable clinical responses. Genetic studies in K-Ras-driven genetically engineered mouse models (GEMMs) indicate that α-smooth muscle actin-expressing (SMA+) CAFs have a predominant tumor-restraining role. Here, we studied global loss of pro-fibrotic non-coding regulatory microRNA-21 (miR-21) in PDAC GEMMs. We generated mouse strains carrying wild type or knockout alleles of Mir-21 in a well-characterized K-Ras-driven, p53-deleted PDAC model (LSL-KrasG12D; p53lox/+;Pdx1-Cre, KPC). Even though miR-21 expression was upregulated in cancer cells and cancer-associated fibroblasts in KPC tumors, global loss of miR-21 activity did not inhibit tumor growth. Strikingly, loss of miR-21 accelerated tumor initiation via mucinous cystic neoplastic lesions and progression to locally advanced invasive carcinoma from which animals precipitously succumbed at an early age. We uncovered a cell-autonomous requirement of miR-21 activity for induction of tumor-restraining SMA+ CAFs, whose absence led to a profound remodeling of the stroma and massive infiltrate of tumor-promoting immune cells. Low miR-21 expression negatively correlated with a fibroblast gene expression signature and positively with an immune cell gene expression signature in TCGA PDAC data set (n = 156) mirroring findings in GEMMs. In vitro and in vivo isograft experiments showed that miR-21 activity was dispensable for cancer cell growth, but required for complete execution of TGF-β-mediated programs. Our results suggest that modulation of miR-21 activity may provide a novel therapeutic opportunity to affect the composition of CAF subpopulations. We will present our genetic and pharmacological approaches to modulate stromal activity of miR-21 in established disease in order to enhance current chemotherapy and/or immunotherapy treatments. Citation Format: Josh Schipper, Brooke Jackson, Ian Beddows, Elizabeth Kenyon, Katie Powell, Neil Robertson, Galen Hostetter, Matti Kiupel, Anna Moore, Jose Conejo-Garcia, Lorenzo sempere. Tumor-suppressive stromal activity of pro-fibrogenic microRNA-21 in initiation and progression of K-Ras-driven mouse models of pancreatic cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5719.

Abstract 1705: Identification of an immune gene expression signature for progression of breast ductal carcinoma in situ (dcis)

Abstract Introduction. DCIS is a potential precursor of invasive breast cancer (IBC). Immune escape might play a critical role in the transition from DCIS to IBC. We aim to identify an immune-related gene expression signature of DCIS to distinguish hazardous lesions, and to avoid the burden of overtreatment in women with less harmful DCIS. Since tumor-infiltrating lymphocytes (TILs) occur more frequently in hormone receptor-negative, Her-2 positive DCIS, we focused the analysis on this subtype. Methods. We performed a prospective nested case-control study involving women with pure DCIS (hormone receptor-negative, Her-2 positive, TILs ≥ 20%), treated with breast conservative surgery with or without radiotherapy. Cases were defined as women with DCIS developing subsequent ipsilateral breast event (IBE), controls as women with DCIS without IBE at 8 years of follow-up. Next-generation sequencing gene expression assay (Oncomine™ Immune Response Research Assay) targeting 395 genes associated with tumor-immune systems interactions was performed on RNA extracted from DCIS samples of 25 cases and 25 age- and radiotherapy-matched controls. In order to obtain reliable results, we considered adequate samples with mapped reads > 1 million and valid reads > 800.000. Normalized gene-level count data generated from the run were further processed with Affymetrix™. A Volcano plot was also employed in order to distinguish genes differently expressed between cases and controls according to the log-gene-level fold change and the ANOVA test (p values < 0.05). Univariate (Log-rank) and multivariate Cox proportional hazard models identified genes associated with the disease free survival. Results. Thirty-one samples (15 cases; 16 controls) reached the quality parameters. The analysis identified 11 genes with a significant different expression between cases and controls. The upregulation of three immune-related genes (VTCN1, NT5E and CMKLR1) remained significantly associated with disease free survival. Notably, VTCN1 and NT5E expressions were significantly associated with the risk of invasive local recurrence. Conclusions. This exploratory analysis of hormone receptor-negative, Her-2 positive DCIS with TILs ≥ 20% showed significant differences in immune-related gene expression profiles between women with subsequent IBE and controls. VTCN1 and NT5E may play an important role in immune surveillance mechanisms of DCIS and may be targeted by immune-based therapy to slow or prevent DCIS progression. These results are well worth further validation on future studies. Citation Format: Matteo Lazzeroni, Caternia Fumagalli, Federica Corso, Alberto Ranghiero, Debora Macis, Elena Guerini Rocco, Valentina Aristarco, Harriet Johansson, Aliana Guerrieri-Gonzaga, Davide Serrano, Andrea DeCensi, Sara Gandini, Massimo Barberis, Bernardo Bonanni. Identification of an immune gene expression signature for progression of breast ductal carcinoma in situ (dcis) [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1705.

Immune Checkpoint Inhibitors in the Treatment of Renal Cancer: Current State and Future Perspective.

Systemic treatment of renal cancer (RCC) has undergone remarkable changes over the past 20 years with the introduction of immunotherapeutic agents targeting programmed cell death (PD-1)/programmed death-ligand 1 (PD-L1) axis, as a single-agent or combined with anti-CTLA-4 monoclonal antibodies (MoAbs) or a multi-target vascular endothelial growth factor-(VEGF) tyrosine kinase inhibitor (TKI). In this paper, we review the main evidence on the use of Immune Checkpoint Inhibitors (ICIs) for RCC treatment from the first demonstration of activity of a nivolumab single agent in a phase I trial to the novel combination strategies (anti-PD-1 plus anti-CTLA4 or anti-PD-1 plus TKI). In addition, we discuss the use of anti-PD-1/PD-L1 agents in patients with non-clear cells and rare histological subtype RCC. Then, we critically examine the current findings in biomarkers that have been proposed to be prognostic or predictive to the response of immunotherapy including immune gene expression signature, B7-H1 expression, PBRM1 loss of function, PD-L1 expression, frame shift indel count, mutations in bromodomain-containing genes in patients with MiT family translocation RCC (tRCC), high expression of the T-effector gene signature, and a high myeloid inflammation gene expression pattern. To date, a single biomarker as a predictor of response has not been established. Since the dynamic behavior of the immune response and the different impact of ICI treatment on patients with specific RCC subtypes, the integration of multiple biomarkers and further validation in clinical trials are needed.

Identification of an Immune Gene Expression Signature for Predicting Lung Squamous Cell Carcinoma Prognosis.

Growing evidence indicates that immune-related biomarkers play an important role in tumor processes. This study investigates immune-related gene expression and its prognostic value in lung squamous cell carcinoma (LUSC). A cohort of 493 samples of patients with LUSC was collected and analyzed from data generated by the TCGA Research Network and ImmPort database. The R coxph package was employed to mine significant immune-related genes using univariate analysis. Lasso and stepwise regression analyses were used to construct the LUSC prognosis prediction model, and clusterProfiler was used for gene functional annotation and enrichment analysis. The Kaplan-Meier analysis and ROC were used to evaluate the model efficiency in predicting and classifying LUSC case prognoses. We identified 14 immune-related genes to incorporate into our prognosis model. The patients were divided into two subgroups (Risk-H and Risk-L) according to their risk score values. Compared to Risk-L patients, Risk-H patients showed significantly improved overall survival (OS) in both training and testing sets. Functional annotation indicated that the 14 identified genes were mainly enriched in several immune-related pathways. Our results also revealed that a risk score value was correlated with various signaling pathways, such as the JAK-STA signaling pathway. Establishment of a nomogram for clinical application demonstrated that our immune-related model exhibited good predictive prognostic performance. Our predictive prognosis model based on immune signatures has potential clinical implications for assessing the overall survival and precise treatment for patients with LUSC.

Loss of microRNA-21 leads to profound stromal remodeling and short survival in K-Ras-driven mouse models of pancreatic cancer.

The microenvironment of pancreatic cancer adenocarcinoma (PDAC) is highly desmoplastic with distinct tumor-restraining and tumor-promoting fibroblast subpopulations. Re-education rather than indiscriminate elimination of these fibroblasts has emerged as a new strategy for combination therapy. Here, we studied the effects of global loss of profibrotic noncoding regulatory microRNA-21 (miR-21) in K-Ras-driven p53-deleted genetically engineered mouse models of PDAC. Strikingly, loss of miR-21 accelerated tumor initiation via mucinous cystic neoplastic lesions and progression to locally advanced invasive carcinoma from which animals precipitously succumbed at an early age. The absence of tumor-restraining myofibroblasts and a massive infiltrate of immune cells were salient phenotypic features of global miR-21 loss. Stromal miR-21 activity was required for induction of tumor-restraining myofibroblasts in in vivo isograft transplantation experiments. Low miR-21 expression negatively correlated with a fibroblast gene expression signature and positively with an immune cell gene expression signature in The Cancer Genome Atlas PDAC data set (n = 156) mirroring findings in the mouse models. Our results exposed an overall tumor-suppressive function of miR-21 in in vivo PDAC models. These results have important clinical implications for anti-miR-21-based inhibitory therapeutic approaches under consideration for PDAC and other cancer types. Mechanistic dissection of the cell-intrinsic role of miR-21 in cancer-associated fibroblasts and other cell types will be needed to inform best strategies for pharmacological modulation of miR-21 activity to remodel the tumor microenvironment and enhance treatment response in PDAC.

Mathematical Modelling Using Predictive Biomarkers for the Outcome of Canine Leishmaniasis upon Chemotherapy.

Prediction parameters of possible outcomes of canine leishmaniasis (CanL) therapy might help with therapeutic decisions and animal health care. Here, we aimed to develop a diagnostic method with predictive value by analyzing two groups of dogs with CanL, those that exhibited a decrease in parasite load upon antiparasitic treatment (group: responders) and those that maintained high parasite load despite the treatment (group: non-responders). The parameters analyzed were parasitic load determined by q-PCR, hemogram, serum biochemistry and immune system-related gene expression signature. A mathematical model was applied to the analysis of these parameters to predict how efficient their response to therapy would be. Responder dogs restored hematological and biochemical parameters to the reference values and exhibited a Th1 cell activation profile with a linear tendency to reach mild clinical alteration stages. Differently, non-responders developed a mixed Th1/Th2 response and exhibited markers of liver and kidney injury. Erythrocyte counts and serum phosphorus were identified as predictive markers of therapeutic response at an early period of assessment of CanL. The results presented in this study are highly encouraging and may represent a new paradigm for future assistance to clinicians to interfere precociously in the therapeutic approach, with a more precise definition in the patient’s prognosis.

An immune gene expression signature distinguishes central nervous system metastases from primary tumours in non–small-cell lung cancer

BackgroundDissemination of non–small-cell lung cancer (NSCLC) in the central nervous system is a frequent and challenging clinical problem. Systemic or local therapies rarely prolong survival and have modest activity regarding local control. Alterations in gene expression in brain metastasis versus primary tumour may increase aggressiveness and impair therapeutic efforts. MethodsWe identified 25 patients with surgically removed NSCLC brain metastases in two different patient cohorts. For 13 of these patients, primary tumour samples were available. Gene expression analysis using the nCounter® PanCancer Immune Profiling gene expression panel (nanoString technologies Inc.) was performed in brain metastases and primary tumour samples. Identification of differentially expressed genes was conducted on normalized data using the nSolver analysis software. ResultsWe compared gene expression patterns in brain metastases with primary tumours. Brain metastasis samples displayed a distinct clustering pattern compared to primary tumour samples with a statistically significant downregulation of genes related to immune response and immune cell activation. Results from KEGG term analysis on differentially expressed genes revealed a concomitant enrichment of multiple KEGG terms associated with the immune system. We identified a 12-gene immune signature that clearly separated brain metastases from primary tumours. ConclusionsWe identified a unique gene downregulation pattern in brain metastases compared with primary tumours. This finding may explain the lower intracranial efficacy of systemic therapy, especially immunotherapy, in brain metastasis of patients with NSCLC.

Characterization of 7A5: A Human CD137 (4-1BB) Receptor Binding Monoclonal Antibody with Differential Agonist Properties That Promotes Antitumor Immunity.

The CD137 receptor plays a key role in mediating immune response by promoting T cell proliferation, survival, and memory. Effective agonism of CD137 has the potential to reinvigorate potent antitumor immunity either alone or in combination with other immune-checkpoint therapies. In this study, we describe the discovery and characterization of a unique CD137 agonist, 7A5, a fully human IgG1 Fc effector-null monoclonal antibody. The biological properties of 7A5 were investigated through in vitro and in vivo studies. 7A5 binds CD137, and the binding epitope overlaps with the CD137L binding site based on structure. 7A5 engages CD137 receptor and activates NF-κB cell signaling independent of cross-linking or Fc effector function. In addition, T cell activation measured by cytokine IFNγ production is induced by 7A5 in peripheral blood mononuclear cell costimulation assay. Human tumor xenograft mouse models reconstituted with human immune cells were used to determine antitumor activity in vivo. Monotherapy with 7A5 inhibits tumor growth, and this activity is enhanced in combination with a PD-L1 antagonist antibody. Furthermore, the intratumoral immune gene expression signature in response to 7A5 is highly suggestive of enhanced T cell infiltration and activation. Taken together, these results demonstrate 7A5 is a differentiated CD137 agonist antibody with biological properties that warrant its further development as a cancer immunotherapy. GRAPHICAL ABSTRACT: http://mct.aacrjournals.org/content/molcanther/19/4/988/F1.large.jpg.

Integrated immune gene expression signature and molecular classification in gastric cancer: New insights.

Gastric cancer (GC) is characterized by extreme heterogeneity due to histopathological differences, molecular characteristics, and immune gene expression signature. Until recently, several targeted therapies failed due to this complexity. The recent immunotherapy resulted in more effective and safe approaches in several malignancies. All tumors could be considered potentially immunogenic and the new knowledge regarding the interactions among tumor cells, immune cells, and tumor microenvironment (TME) allowed to reverse possible immune resistance. The immune response is a complex multisteps process that finely regulates the balance between the recognition of non-self and the prevention of autoimmunity. Cancer cells can use these pathways to suppress tumor immunity as a major mechanism of immune resistance. The recent molecular classifications of GCs by The Cancer Genome Atlas (TCGA) and by the Asian Cancer Research (ACRG) networks, together with the identification of multiple biomarkers, open new perspectives for stratification of patients who might benefit from a long-term immune checkpoint therapy. One of the major processes that contribute to an immunosuppressive microenvironment is represented by tumor angiogenesis. The cellular mechanisms inducing both angiogenesis and immunosuppressive responses are often reached by the same cell types and soluble factors, such as vascular endothelial growth factor A (VEGFA). Recent studies point out that combinatorial strategies should be adapted as useful therapeutic approach to reverse the immunosuppressive status of microenvironment occurring in a relevant percentage of gastric tumors.

3O VHIO immune gene-expression signature (VIGex) to enrich patient selection in immunotherapy phase I clinical trials

3O VHIO immune gene-expression signature (VIGex) to enrich patient selection in immunotherapy phase I clinical trials

Molecular characterization of neuroendocrine bladder cancer.

561 Background: Neuroendocrine bladder cancer (NEBC) is a rare and aggressive variant that is associated with poor survival outcomes. Because NEBC is rare, the molecular features of NEBC remain poorly characterized. Therefore, we characterized NEBC at the molecular level to understand the underlying biology and identify novel therapeutic targets. Methods: Whole transcriptome RNAseq was performed on FFPE cores from 24 NEBCs and 51 conventional muscle-invasive bladder cancers (MIBCs) from Tenon Hospital in Paris. Results: Unsupervised cluster analysis of 75 tumors generated 2 distinct clusters that separated NEBCs from MIBCs. The NEBC tumors were strongly enriched with biomarkers for the characteristic of neuroendocrine or small cell malignancies, including DLL3, SOX2, and EZH2. In addition, E2F1 pathway is significantly enriched due to the impair of RB/p53 pathways. Further, the NEBCs were enriched with the TCGA’s neuronal differentiation genes that were associated with high response rates in patients treated in atezolizumab (anti-PDL1) within the context of the ImVigor 210 trial. Nevertheless, the NEBCs were characterized by suppressed immune pathway gene expression signatures, such as the Th1 pathway, effector T cell lymphocyte, and IFNg that are usually highly enriched in tumors that are sensitive to immune checkpoint blockade. Of candidate mechanisms, the suppressed TGFbpathway activity observed in the NEBCs was the most obvious explanation for sensitivity to checkpoint blockade. Conclusions: NEBCs are distinct from conventional MIBCs by gene expression signature. They are also characterized by overexpression of canonical neuroendocrine markers and inhibition of TGFb pathway activity.