CD8+ CD226high T cells in liver metastases dictate the prognosis of colorectal cancer patients treated with chemotherapy and radical surgery.

CD226 has been reported to participate in the rescue of CD8+ T cell dysfunction. In this study, we aimed to assess the prognostic value of CD226 in tumor-infiltrating lymphocytes (TILs) derived from colorectal cancer (CRC) liver metastases treated with chemotherapy and radical surgery. TILs from 43 metastases were isolated and analyzed ex vivo using flow cytometry. CD155 and CD3 levels in the tumor microenvironment were assessed by immunohistochemistry. Exploration and validation of biological processes highlighted in this study were performed by bioinformatics analysis of bulk RNA-seq results for 28 CRC liver metastases pretreated with chemotherapy as well as public gene expression datasets. CD226 expression contributes to the definition of the immune context in CRC liver metastases and primary tumors. CD226 on CD8+ T cells was not specifically coexpressed with other immune checkpoints, such as PD1, TIGIT, and TIM3, in liver metastases. Multivariate Cox regression analysis revealed CD226 expression on CD8+ T cells to be an independent prognostic factor (p = 0.003), along with CD3 density at invasion margins (p = 0.003) and TIGIT expression on CD4+ T cells (p = 0.019). CD155 was not associated with the prognostic value of CD226. Gene expression analysis in a validation dataset confirmed the prognostic value of CD226 in CRC liver metastases but not in primary tumors. Downregulation of CD226 on CD8+ TILs in the liver microenvironment was restored by IL15 treatment. Overall, CD226 expression on liver metastasis-infiltrating CD8+ T cells selectively contributes to immune surveillance of CRC liver metastases and has prognostic value for patients undergoing radical surgery.

Chemokine switch regulated by TGF-ß1 in cancer-associated fibroblast subsets determines the efficacy of chemo-immunotherapy.

Combining immunogenic cell death-inducing chemotherapies and PD-1 blockade can generate remarkable tumor responses. It is now well established that TGF-ß1 signaling is a major component of treatment resistance and contributes to the cancer-related immunosuppressive microenvironment. However, whether TGF-ß1 remains an obstacle to immune checkpoint inhibitor efficacy when immunotherapy is combined with chemotherapy is still to be determined. Several syngeneic murine models were used to investigate the role of TGF-ß1 neutralization on the combinations of immunogenic chemotherapy (FOLFOX: 5-fluorouracil and oxaliplatin) and anti-PD-1. Cancer-associated fibroblasts (CAF) and immune cells were isolated from CT26 and PancOH7 tumor-bearing mice treated with FOLFOX, anti-PD-1 ± anti-TGF-ß1 for bulk and single cell RNA sequencing and characterization. We showed that TGF-ß1 neutralization promotes the therapeutic efficacy of FOLFOX and anti-PD-1 combination and induces the recruitment of antigen-specific CD8+ T cells into the tumor. TGF-ß1 neutralization is required in addition to chemo-immunotherapy to promote inflammatory CAF infiltration, a chemokine production switch in CAF leading to decreased CXCL14 and increased CXCL9/10 production and subsequent antigen-specific T cell recruitment. The immune-suppressive effect of TGF-ß1 involves an epigenetic mechanism with chromatin remodeling of CXCL9 and CXCL10 promoters within CAF DNA in a G9a and EZH2-dependent fashion. Our results strengthen the role of TGF-ß1 in the organization of a tumor microenvironment enriched in myofibroblasts where chromatin remodeling prevents CXCL9/10 production and limits the efficacy of chemo-immunotherapy.

Tissue-resident memory T cells in gastrointestinal cancer immunology and immunotherapy: ready for prime time?

Tissue-resident memory T (TRM) cells have emerged as immune sentinels that patrol the tissue microenvironment and orchestrate localized antitumor immunity in various solid cancers. Recent studies have revealed that TRM cells are key players in cancer immunosurveillance, and their involvement has been linked to favorable responses to immunotherapy as well as general better clinical outcome in cancer patients. In this review, we provide an overview of the major advances and recent findings regarding TRM cells phenotype, transcriptional and epigenetic regulation in cancer with a special focus on gastrointestinal tumors. Finally, we highlight the exciting clinical implication of TRM cells in these types of tumors.

Harnessing Antitumor CD4+ T Cells for Cancer Immunotherapy.

Over the past decades, CD4+ T cells have been considered as a supporting actor in the fields of cancer immunotherapy. Until recently, accumulating evidence has demonstrated the critical role of CD4+ T cells during antitumor immunity. CD4+ T cells can either suppress or promote the antitumor cytotoxic CD8+ T cell responses, either in secondary lymphoid organs or in the tumor. In this review, we provide an overview of the multifaceted role of different CD4+ T cell subsets in cancer immune response and their contribution during cancer therapies. Specifically, we focus on the latest progress regarding the impact of CD4+ T cell modulation on immunotherapies and other cancer therapies and discuss the prospect for harnessing CD4+ T cells to control tumor progression and prevent recurrence in patients.

A new workflow combining magnetic cell separation and impedance-based cell dispensing for gentle, simple and reliable cloning of specific CD8+ T cells.

Reverse immunology has open the door to innovative cancer immunotherapy strategies such as immunogenic antigen-based vaccination and transgenic T cell receptor (TCR)-based adoptive cell transfer. This approach enables the identification of immunogenic tumor specific antigen derived peptides. One of the major challenges is the rapid selection of antigen-specific CD8+ T cell clones. Thus, IFNγ-producing CD8+ T cells magnetic sorting combined with limiting dilution cloning approach represents the most common method of specific T cell cloning. However, during plate setup several wells will not contain T cells whereas others will contain mixed population of T cells. In this case, a re-cloning step is required which make limiting dilution based cloning a laborious, inefficient, expensive and a time-consuming method. To address these obstacles, here we present a novel 2-step workflow combining simple, affordable and gentle magnetic cell separation followed by single cell isolation using a device called DispenCell-S1. We aimed to compare this new workflow with the traditional limiting dilution method using in vitro generated antigen-specific CD8+ T cells. Herein, we reported the reliability of DispenCell-S1 method and its efficiency in T cell clones isolation.

Anti-Telomerase CD4+ Th1 Immunity and Monocytic-Myeloid-Derived-Suppressor Cells Are Associated with Long-Term Efficacy Achieved by Docetaxel, Cisplatin, and 5-Fluorouracil (DCF) in Advanced Anal Squamous Cell Carcinoma: Translational Study of Epitopes-HPV01 and 02 Trials.

Docetaxel, cisplatin and 5-fluorouracil (DCF) chemotherapy regimen is highly effective in advanced anal squamous cell carcinoma (SCCA), as demonstrated by the Epitopes-HPV02 study results. Here, we analyzed the impact of DCF regimen and the prognostic value of adaptive immune responses and immunosuppressive cells in SCCA patients included in two prospective studies (Epitopes-HPV01 and HPV02). The presence of T-cell responses against Human papillomavirus (HPV)16-E6/E7 and anti-telomerase (hTERT)-antigens was measured by IFNᵧ-ELISpot. Here, we showed that HPV-adaptive immune responses are increased in SCCA patients. SCCA patients also displayed enhanced circulating TH1 T-cells restricted by hTERT. Exposition to DCF increased hTERT immunity but not HPV or common viruses immune responses. Notably, the correlation of hTERT immune responses with SCCA patients' clinical outcomes highlights that hTERT is a relevant antigen in this HPV-related disease. The influence of peripheral immunosuppressive cells was investigated by flow cytometry. While both regulatory T-cells and monocytic-myeloid-derived suppressive cells (M-MDSC) accumulated in the peripheral blood of SCCA patients, only high levels of M-MDSC were negatively correlated with hTERT adaptive immune responses and predicted poor prognosis. Altogether, our results reveal that hTERT is a relevant antigen in HPV-driven SCCA disease and that M-MDSC levels influence TH1-adaptive immune responses and patients' survival.

FOLFOXIRI vs FOLFIRINOX as first-line chemotherapy in patients with advanced pancreatic cancer: A population-based cohort study.

BACKGROUND: FOLFIRINOX regimen is the first-line reference chemotherapy (L1) in advanced pancreatic ductal adenocarcinoma (aPDAC). FOLFOXIRI, a schedule with a lower dose of irinotecan and no bolus 5-fluorouracil, has demonstrated efficacy and feasibility in colorectal cancer. AIM: To investigate the potential clinical value of FOLFOXIRI in patients with aPDAC in routine clinical practice. METHODS: Analyses were derived from all consecutive aPDAC patients treated in L1 between January 2011 and December 2017 in two French institutions, with either FOLFOXIRI (n = 165) or FOLFIRINOX (n = 124) regimens. FOLFOXIRI consisted of irinotecan (165 mg/m2), oxaliplatin (85 mg/m2), leucovorin (200 mg/m2) and 5-fluorouracil (3200 mg/m2 as a 48-h continuous infusion) every 2 wk. Ninety-six pairs of patients were selected through propensity score matching, and clinical outcomes of the two treatment regimens were compared. RESULTS: Median overall survival was 11.1 mo in the FOLFOXIRI and 11.6 mo in the FOLFIRINOX cohorts, respectively. After propensity score matching, survival rates remained similar between the two regimens in terms of overall survival (hazard ratio = 1.22; P = 0.219) and progression-free survival (hazard ratio = 1.27; P = 0.120). The objective response rate was 37.1% in the FOLFOXIRI group vs 47.8% in the FOLFIRINOX group (P = 0.187). Grade 3/4 toxicities occurred in 28.7% of patients in the FOLFOXIRI cohort vs 19.5% in the FOLFIRINOX cohort (P = 0.079). FOLFOXIRI was associated with a higher incidence of grade 3/4 digestive adverse events. Hematopoietic growth factors were used after each chemotherapy cycle and the low hematological toxicity rates were below 5% with both regimens. CONCLUSION: FOLFOXIRI is feasible in L1 in patients with aPDAC but does not confer any therapeutic benefit as compared with FOLFIRINOX. The low hematological toxicity rates strengthened the relevance of primary prophylaxis with hematopoietic growth factors.

MARCKS protein overexpression is associated with poor prognosis in male breast cancer.

BACKGROUND: Male breast cancer (MBC) is a rare and aggressive disease. Thus, identification of new therapeutic targets is crucial. OBJECTIVE: Our objective was to evaluate the protein expression of MARCKS (Myristoylated Alanine-Rich C-Kinase Substrate) in MBC and to investigate its prognostic value. MATERIALS AND METHODS: MARCKS protein expression in tumor and stromal cells was analyzed by immunohistochemistry (IHC) in a retrospective series of 96 pre-chemotherapy MBC samples and 80 normal breast samples, from Tunisian patients treated at Salah Azaiez Institute. Correlations were searched between MARCKS expression and clinicopathological features including overall survival (OS). RESULTS: MARCKS was overexpressed in epithelial tumor cells in 66% of the MBC samples versus 26% of normal samples (p= 1.40 × 10-7). Such positive MARCKS expression in epithelial tumor cells was associated with positive HER2 status (p= 4.0 × 10-3). It was associated with shorter OS in uni-and multivariate analysis. By contrast, stromal IHC MARCKS expression was correlated only with tumor grade. CONCLUSION: MARCKS tumor cell overexpression might in part explain the aggressiveness and the poor prognosis of MBC. MARCKS can represent a potential therapeutic target for MBC.

Forkhead box M1 (FOXM1) expression predicts disease free survival and may mediate resistance to chemotherapy and hormonotherapy in male breast cancer.

BACKGROUND: Male breast cancer (MBC) is a rare and neglected disease. Prognostic and predictive factors in MBC are extrapoled from trials conducted on its female counterpart. OBJECTIVE: Since the relationship between the transcription factor Forkhead box M1 (FOXM1) expression and the clinical response to chemotherapy and hormonotherapy in MBC remains unknown, we sought to investigate the predictive value of FOXM1 in MBC. METHODS: FOXM1 expression was assessed in 130 MBC cases. Clinical significance was analyzed by Kaplan Meier curves, log-rank test and multivariate Cox regression analyses. RESULTS: Patients with high FOXM1 expression had a significantly lower response rate to chemotherapy (P = 0.045) and hormonotherapy (P = 0.029) than those with low FOXM1 expression. Multivariate analyses indicated that FOXM1 was an independent prognostic factor for disease free survival in MBC patients (P < 0.001). CONCLUSIONS: FOXM1 may have a reliable predictive significance in male breast cancer and thus may become an important target for male breast cancer therapy in the near future.

Prognostic implications of the intrinsic molecular subtypes in male breast cancer.

PURPOSE: Intrinsic molecular subtyping has been widely used in female breast cancer, and it has proven its significance. In this article, we aimed to study the intrinsic subtypes of male breast cancer (MBC) in correlation with clinicopathological features. METHODS: We retrospectively identified 130 MBC cases from 2004 to 2013. Intrinsic molecular subtypes were determined by immunohistochemistry (IHC). RESULTS: From a total of 130 MBC cases, 45.4% of tumors were luminal A subtype, 44.6% were luminal B, 5% were HER2 positive and 5% were triple negative tumors. There were statistically significant differences between different IHC intrinsic subtypes regarding tumor size (p=0.001), estrogen receptor (ER) status (p=0.001), progesterone receptor (PR) status (p=0.001), HER2 status (p=0.001) and Ki67 proliferation index (p=0.001). CONCLUSION: The distribution of breast cancer intrinsic subtypes in males is different compared to its female counterpart; however, they don't seem to give the same prognostic value.

Overexpression of FOXM1 Is a Potential Prognostic Marker in Male Breast Cancer.

BACKGROUND: Several studies have outlined biological differences between female and male breast cancer (MBC) and concluded that MBC should be considered as an entirely separate disease. Whether FOXM1 has any indication for prognosis in MBC patients remains unknown. We sought to examine the expression levels of FOXM1 in MBC and to identify the relationship between FOXM1 expression and patient survival. PATIENTS AND METHODS: FOXM1 expression was evaluated in a total of 130 MBC specimens. RESULTS: FOXM1 was overexpressed in 37% of the MBC samples. FOXM1 overexpression was significantly associated with tumor size (p = 0.045), histological grade (p = 0.048), lymph node metastasis (p = 0.012), Ki-67 proliferation index (p = 0.016), and molecular subtypes (p < 0.001). Multivariate analyses indicated that FOXM1 was an independent prognostic factor for overall survival in MBC patients (p < 0.001, hazard ratio = 0.69 (0.43-0.96)). CONCLUSIONS: Overexpression of FOXM1 was associated with well-established markers of poor prognosis; thus FOXM1 may represent a potential novel prognostic marker for MBC.