Cellular and molecular determinants of all-trans retinoic acid sensitivity in breast cancer: Luminal phenotype and RARα expression

Forty-two cell lines recapitulating mammary carcinoma heterogeneity were profiled for all-trans retinoic acid (ATRA) sensitivity. Luminal and ER+ (estrogen-receptor-positive) cell lines are generally sensitive to ATRA, while refractoriness/low sensitivity is associated with a Basal phenotype and HER2 positivity. Indeed, only 2 Basal cell lines (MDA-MB157 and HCC-1599) are highly sensitive to the retinoid. Sensitivity of HCC-1599 cells is confirmed in xenotransplanted mice. Short-term tissue-slice cultures of surgical samples validate the cell-line results and support the concept that a high proportion of Luminal/ER+ carcinomas are ATRA sensitive, while triple-negative (Basal) and HER2-positive tumors tend to be retinoid resistant. Pathway-oriented analysis of the constitutive gene-expression profiles in the cell lines identifies RARα as the member of the retinoid pathway directly associated with a Luminal phenotype, estrogen positivity and ATRA sensitivity. RARα3 is the major transcript in ATRA-sensitive cells and tumors. Studies in selected cell lines with agonists/antagonists confirm that RARα is the principal mediator of ATRA responsiveness. RARα over-expression sensitizes retinoid-resistant MDA-MB453 cells to ATRA anti-proliferative action. Conversely, silencing of RARα in retinoid-sensitive SKBR3 cells abrogates ATRA responsiveness. All this is paralleled by similar effects on ATRA-dependent inhibition of cell motility, indicating that RARα may mediate also ATRA anti-metastatic effects. We define gene sets of predictive potential which are associated with ATRA sensitivity in breast cancer cell lines and validate them in short-term tissue cultures of Luminal/ER+ and triple-negative tumors. In these last models, we determine the perturbations in the transcriptomic profiles afforded by ATRA. The study provides fundamental information for the development of retinoid-based therapeutic strategies aimed at the stratified treatment of breast cancer subtypes

Improved survival of patients with hypermutation in uterine serous carcinoma

• Uterine serous carcinoma (USC) is an aggressive subtype of endometrial cancer. • Mutations in DNA polymerase ε gene (POLE) are detected in a subset of USC and confer a strong mutator phenotype. • USC patients diagnosed with POLE mutations experience a significantly better prognosis

Novel Axl-driven signaling pathway and molecular signature characterize high-grade ovarian cancer patients with poor clinical outcome

High-grade epithelial ovarian cancer (HGEOC) is a clinically diverse and molecularly heterogeneous disease comprising subtypes with distinct biological features and outcomes. The receptor tyrosine kinases, expressed by EOC cells, and their ligands, present in the microenvironment, activate signaling pathways, which promote EOC cells dissemination. Herein, we established a molecular link between the presence of Gas6 ligand in the ascites of HGEOCs, the expression and activation of its receptor Axl in ovarian cancer cell lines and biopsies, and the progression of these tumors. We demonstrated that Gas6/Axl signalling converges on the integrin \u3b23 pathway in the presence of the adaptor protein p130Cas, thus inducing tumor cell adhesion to the extracellular matrix and invasion. Accordingly, Axl and p130Cas were significantly co-expressed in HGEOC samples. Clinically, we identified an Axl-associated signature of 62 genes able to portray the HGEOCs with the shortest overall survival. These data biologically characterize a group of HGEOCs and could help guide a more effective therapeutic approach to be taken for these patients

Polymerase ε (POLE) ultra-mutation in uterine tumors correlates with T lymphocyte infiltration and increased resistance to platinum-based chemotherapy in vitro

OBJECTIVE: Up to 12 % of all endometrial-carcinomas (EC) harbor DNA-polymerase-ε-(POLE) mutations. It is currently unknown whether the favorable prognosis of POLE-mutated EC is derived from their low metastatic capability, extraordinary number of somatic mutations thus imparting immunogenicity, or a high sensitivity to chemotherapy. METHODS: Polymerase-chain-reaction-amplification and Sanger-sequencing were used to test for POLE exonuclease-domain-mutations (exons 9–14) 131 EC. Infiltration of CD4+ and CD8+ T-lymphocytes (TIL) and PD-1-expression in POLE-mutated vs POLE wild-type EC was studied by immunohistochemistry (IHC) and the correlations between survival and molecular features were investigated. Finally, primary POLE-mutated and POLE-wild-type EC cell lines were established and compared in-vitro for their sensitivity to chemotherapy. RESULTS: Eleven POLE-mutated EC (8.5%) were identified. POLE-mutated tumors were associated with improved progression-free-survival (P<0.05) and displayed increased numbers of CD4+ (44.5 vs 21.8; P = .001) and CD8+ (32.8 vs 13.5; P < .001) TILs when compared to wild-type POLE EC. PD-1 receptor was overexpressed in TILs from POLE-mutated vs wild-type-tumors (81% vs 28%; P < .001). Primary POLE tumor cell lines were significantly more resistant to platinum-chemotherapy in-vitro when compared to POLE-wild-type tumors (P < 0.004). CONCLUSIONS: POLE ultra-mutated EC are heavily infiltrated with CD4+/CD8+ TIL, overexpress PD-1 immune-check-point (i.e., features consistent with chronic antigen-exposure), and have a better prognosis when compared to other molecular subtypes of EC patients. POLE-mutated tumor-cell lines are resistant to platinum-chemotherapy in-vitro suggesting that the better prognosis of POLE-patients is not secondary to a higher sensitivity to chemotherapy but likely linked to enhanced immunogenicity

Polymerase ε (POLE) ultra-mutated tumors induce robust tumor-specific CD4+ T cell responses in endometrial cancer patients

OBJECTIVE: Around 7–10% of endometrial carcinomas are characterized by Polymerase-ε-(POLE) exonuclease-domain-mutations, an ultra-mutated-phenotype and a favorable prognosis. It is currently unknown whether POLE ultra-mutated-tumors are more immunogenic when compared to the other groups of endometrial cancers. METHODS: We used autologous-dendritic-cells (DC) pulsed with whole-tumor-extracts to assess the level of CD8+ and CD4+ T-cell-activation induced by POLE-ultramutated (+) and POLE wild-type (−) endometrial cancer cells in vitro. T-lymphocyte-proliferations were evaluated using CFSE and/or ([3H])thymidine-incorporation-assays while the ability to specifically kill autologous-tumor-cells by cytotoxic-T-lymphocyte (CTL) was tested in standard 4-hr-(51)Cr-cytotoxicity-assays. In order to correlate cytotoxic activity and proliferation by CD4+ and CD8+ T-lymphocytes, respectively, with a particular lymphoid subset, two-color-flow-cytometric analysis of intracellular-cytokine-expression (IFN-γ vs IL-4) at the single cell level, was also performed. RESULTS: DC-pulsed with tumor extracts were able to induce CTL-responses against autologous-tumor-cells in both POLE (+) and POLE (−) cancer patients (P=0.305). These CD8+ T-cell-populations were cytotoxic against tumor-cells but they did not lyse PHA-stimulated-autologous-lymphocytes or autologous-EBV-transformed-lymphoblastoid-control-cell-lines. In contrast, only POLE (+) tumor-lysate-pulsed-DC were able to induce significant proliferation and high IFN-γ expression (i.e., Th1-cytokine-bias) in autologous in vitro DC-stimulated CD4+ T-cells as well as naïve CD4+ and CD8+ T-cells from patients-peripheral-blood (P < 0.05). CONCLUSIONS: POLE ultra-mutated-tumors are significantly more immunogenic when compared to POLE (−) tumors, in particular to the helper arm of the immune system. These data lend support to the hypothesis that the better prognosis of patients with POLE (+) tumors may at least in part be linked to their enhanced immunogenicity

Mean platelet volume is associated with lower risk of overall and non-vascular mortality in a general population: Results from the Moli-sani study

Larger mean platelet volume (MPV) has been associated with adverse health outcomes in high-risk populations or patients with cardiovascular disease (CVD). We tested the association of MPV with mortality in a prospective cohort study including 17,402 subjects randomly recruited from an adult general population within the Moli-sani study (2005-2010). Two distinct subgroups (with or without CVD at baseline) were subsequently analysed. Hazard ratios (HR) were calculated using multivariable Cox-proportional hazard models. Over a median follow up of eight years (137,547 person-years), 925 all-cause deaths occurred (330 vascular, 351 cancer and 244 other deaths). In a multivariable model, the highest MPV quintile (mean MPV=10.0 fL), as compared to the lowest one, was associated with reduced risk of overall mortality (HR=0.79; 95 % confidence interval 0.64-0.98), cancer death (HR=0.70; 0.49-1.00) and death from other non-vascular/non cancer causes (HR=0.55; 0.36-0.84) but not with vascular mortality. The inverse association with overall death appeared even stronger in the subgroup without CVD at baseline (HR=0.64; 0.50-0.81). In contrast, within 920 subjects reporting a previous CVD event, larger MPV was associated with higher risk of total mortality (HR=1.69; 1.05-2.72; p for interaction=0.048) and with a trend of risk for other cause-specific deaths. In conclusion, larger MPV is associated with lower risk of overall and non-vascular death in subjects apparently free from CVD, but appears to be a predictive marker of death in patients with CVD history. The latter is a likely effect modifier of the association between MPV and death

T-DM1, a novel antibody-drug conjugate, is highly effective against uterine and ovarian carcinosarcomas overexpressing HER2

INTRODUCTION: Ovarian and uterine carcinosarcoma (CS) are characterized by their aggressive clinical behavior and poor prognosis. We evaluated the efficacy of trastuzumab-emtansine (T-DM1), against primary HER2 positive and HER2 negative CS cell lines in vitro and in vivo. METHODS: Eight primary CS cell lines were evaluated for HER2 amplification and protein expression by FISH, immunohistochemistry, flow cytometry and qRT-PCR. Sensitivity to T-DM1-induced antibody-dependent-cell-mediated-cytotoxicity (ADCC) was evaluated in 4-hr-chromium-release-assays. T-DM1 cytostatic and apoptotic activities were evaluated using flow cytometry based proliferation assays. In vivo activity of T-DM1 was also evaluated. RESULTS: HER2 protein overexpression and gene amplification were detected in 25% (2/8) of the primary CS cell lines. T-DM1 and T were similarly effective in inducing strong ADCC against CS overexpressing HER2 at 3+ levels. In contrast, T-DM1 was dramatically more effective than T in inhibiting cell proliferation (P<0.0001) and in inducing G2/M phase cell cycle arrest in the HER2 expressing cell lines (shift of G2/M: mean ± SEM from 14.87 ± 1.23% to 66.57 ± 4.56%, P<0.0001). Importantly, T-DM1 was highly active at reducing tumor formation in vivo in CS xenografts overexpressing HER2 (P=0.0001 and P<0.0001 compared to T and vehicle respectively) with a significantly longer survival when compared to T and vehicle mice (P=0.008 and P=0.0001 respectively). CONCLUSION: T-DM1 may represent a novel treatment option for the subset of HER2 positive CS patients with disease refractory to chemotherapy

Reduced mortality risk by a polyphenol-rich diet: An analysis from the Moli-sani study

Objectives: The effect of the polyphenol content of the human diet on mortality risk is not yet fully understood. The aim of this study was to evaluate the association of a polyphenol-rich diet with mortality rate and a possible mediation effect by inflammation, in what we believe to be a novel, holistic approach. Methods: We analyzed 21 302 participants (10 980 women and 10 322 men, aged 6535 y) from the Moli-sani cohort. The participants were followed up for a median of 8.3 y. The European Prospective Investigation into Cancer and Nutrition food frequency questionnaire (FFQ) was used for dietary assessment. Flavonol, flavone, flavanone, flavanol, anthocyanin, isoflavone, and lignan intakes were calculated using European Food Information Resource\u2014Bioactive Substances in Food Information Systems and the polyphenol antioxidant content (PAC)-score was constructed to assess the total content of these nutrients in the diet. Results: Participants included in the highest quintile of intake of various polyphenol classes and subclasses presented a significant lower all-cause mortality risk compared with those in the lowest group of consumption (hazard ratio [HR] &lt; 1; P &lt; 0.05). Cox regression analyses adjusted for potential confounders indicated that participants in higher quintiles of PAC-score had lower all-cause mortality risk (HR &lt;1; P &lt; 0.05). When cause-specific mortality rates were considered, similar effects were observed for cardiocerebrovascular and cancer mortality (HR &lt;1; P &lt; 0.05). Conclusions: The polyphenol content of the diet was associated with reduced mortality risk in a Mediterranean population, possibly through an antiinflammatory mechanism