Integrated mutational landscape analysis of poorly differentiated high-grade neuroendocrine carcinoma of the uterine cervix.

High-grade neuroendocrine cervical cancers (NETc) are exceedingly rare, highly aggressive tumors. We analyzed 64 NETc tumor samples by whole-exome sequencing (WES). Human papillomavirus DNA was detected in 65.6% (42/64) of the tumors. Recurrent mutations were identified in PIK3CA, KMT2D/MLL2, K-RAS, ARID1A, NOTCH2, and RPL10. The top mutated genes included RB1, ARID1A, PTEN, KMT2D/MLL2, and WDFY3, a gene not yet implicated in NETc. Somatic CNV analysis identified two copy number gains (3q27.1 and 19q13.12) and five copy number losses (1p36.21/5q31.3/6p22.2/9q21.11/11p15.5). Also, gene fusions affecting the ACLY-CRHR1 and PVT1-MYC genes were identified in one of the eight samples subjected to RNA sequencing. To resolve evolutionary history, multiregion WES in NETc admixed with adenocarcinoma cells was performed (i.e., mixed-NETc). Phylogenetic analysis of mixed-NETc demonstrated that adenocarcinoma and neuroendocrine elements derive from a common precursor with mutations typical of adenocarcinomas. Over one-third (22/64) of NETc demonstrated a mutator phenotype of C > T at CpG consistent with deficiencies in MBD4, a member of the base excision repair (BER) pathway. Mutations in the PI3K/AMPK pathways were identified in 49/64 samples. We used two patient-derived-xenografts (PDX) (i.e., NET19 and NET21) to evaluate the activity of pan-HER (afatinib), PIK3CA (copanlisib), and ATR (elimusertib) inhibitors, alone and in combination. PDXs harboring alterations in the ERBB2/PI3K/AKT/mTOR/ATR pathway were sensitive to afatinib, copanlisib, and elimusertib (P < 0.001 vs. controls). However, combinations of copanlisib/afatinib and copanlisib/elimusertib were significantly more effective in controlling NETc tumor growth. These findings define the genetic landscape of NETc and suggest that a large subset of these highly lethal malignancies might benefit from existing targeted therapies.

Finding the junction between claudins and endometrial carcinoma.

Endometrial carcinoma (EC) defines a heterogeneous group of neoplastic diseases originating from the transformation of endometrial cells that constitute the internal lining of the uterus. To date several molecular targets have been analysed to describe the natural course of the disease, claudins being among these. Claudins are the main components of tight junctions (TJs), and their main functions are ascribed to the compartmentalization of tissues and cell-cell communication by means of intracellular ions diffusion: these features are typical of epithelial cells. Their overexpression, mis-localization or loss contribute to the malignancy of EC cells. This review collected all available data regarding the expression, regulation and claudin-related signaling pathways to provide a comprehensive view on the influence of claudin in EC progression. Further, the translational potential of claudin differential expression was explored, indicating that their role in personalized medicine could also contribute to EC therapy besides their employment for diagnosis and prognosis.

Functional profiles of curatively treated adenoid cystic carcinoma unveil prognostic features and potentially targetable pathways.

Adenoid cystic carcinoma (ACC) of salivary gland is a slowly growing tumor showing a propensity for delayed recurrence, with decreased survival rates. The identification of poor prognosis patients may help in defining molecular-based targeted strategies in this rare disease orphan of new treatments. Through a gene expression microarray-based approach followed by GSE functional analysis the expression profile of 46 primary untreated ACC samples and of ACC (h-TERT) tumor cells was analyzed. Patients who experienced early relapse showed enrichment in proliferation-related gene sets, including the G2-M checkpoint, E2F and myc targets, and in gene sets related to IFN signaling and aberrant proteostasis (FDR < 0.1), indicating increased mitotic and transcriptional activity in aggressive ACC. Similar functions were enriched in ACC samples classified by immunohistochemical staining as p63-negative, which exhibited increased protein burden and activation of pro-survival stress response pathways compared to p63-positive tumors. Compared to ACC tissues, ACC (h-TERT) cells share transcriptional features of aggressive p63-negative tumors. These data suggest association of specific pathway alterations with histopathological features of ACC, as recapitulated by p63 testing in patient prognostic stratification, anticipating new avenues for therapeutic intervention.

Integrated Biomarker Analysis Reveals L1CAM as a Potential Stratification Marker for No Specific Molecular Profile High-Risk Endometrial Carcinoma.

Histopathologic assessment of high-risk endometrial cancer (EC) suffers from intersubject variability and poor reproducibility. The pragmatic classification in four molecular subgroups helps to overcome these limits, showing a significant prognostic value. The "no specific molecular profile" (NSMP) is the most heterogeneous EC subgroup, requiring further characterization to better guide its clinical management. DNA sequencing of POLE exonuclease domain and immunohistochemistry for PMS2, MSH6, and p53 were performed in order to stratify a cohort of 94 high-risk EC patients in the four molecular subgroups. Moreover, a panel of seven additional biomarkers was tested. Patients were found to be 16% POLE-mutated, 36% mismatch repair-deficient, 27% p53-abnormal, and 21% NSMP. In the multivariable model, molecular groups confirmed their significant association with disease-specific survival and progression-free survival, with p53-abnormal and NSMP endometrial cancer characterized by poor outcomes. Among the additional evaluated biomarkers, L1CAM was the only one with a significant prognostic value within the NSMP subgroup. NSMP/L1CAM-positive patients experienced the worst outcome and were "early-relapsing" after platinum-based chemotherapy, with a significantly shorter platinum-free interval compared to L1CAM-negative patients. L1CAM appears to be a promising candidate as a prognostic and predictive biomarker in the high-risk NSMP subgroup, which is actually known to lack specific molecular markers.

L1CAM expression as a predictor of platinum response in high-risk endometrial carcinoma.

For high-risk endometrial cancer (EC) patients, adjuvant chemotherapy is recommended to improve outcome. Yet, predictive biomarkers for response to platinum-based chemotherapy (Pt-aCT) are currently lacking. We tested expression of L1 cell-adhesion molecule (L1CAM), a well-recognised marker of poor prognosis in EC, in tumour samples from high-risk EC patients, to explore its role as a predictive marker of Pt-aCT response. L1CAM expression was determined using RT-qPCR and immunohistochemistry in a cohort of high-risk EC patients treated with Pt-aCT and validated in a multicentric independent cohort. The association between L1CAM and clinicopathologic features and L1CAM additive value in predicting platinum response were determined. The effect of L1CAM gene silencing on response to carboplatin was functionally tested on primary L1CAM-expressing cells. Increased L1CAM expression at both genetic and protein level correlated with high-grade, non-endometrioid histology and poor response to platinum treatment. A predictive model adding L1CAM to prognostic clinical variables significantly improved platinum response prediction (C-index 78.1%, P = .012). In multivariate survival analysis, L1CAM expression was significantly associated with poor outcome (HR: 2.03, P = .019), potentially through an indirect effect, mediated by its influence on response to chemotherapy. In vitro, inhibition of L1CAM significantly increased cell sensitivity to carboplatin, supporting a mechanistic link between L1CAM expression and response to platinum in EC cells. In conclusion, we have demonstrated the role of L1CAM in the prediction of response to Pt-aCT in two independent cohorts of high-risk EC patients. L1CAM is a promising candidate biomarker to optimise decision making in high-risk patients who are eligible for Pt-aCT.

VEGF-D Serum Level as a Potential Predictor of Lymph Node Metastasis and Prognosis in Vulvar Squamous Cell Carcinoma Patients.

Background: Radical surgical resection of the primary tumor with mono/bilateral inguinofemoral lymph node dissection is the standard treatment for invasive vulvar squamous cell carcinoma (VSCC) and is frequently related to severe morbidity. Tailoring surgical treatment is of paramount importance, and a comprehensive preoperative evaluation is mandatory. Vascular endothelial growth factor D (VEGF-D) is considered a regulator of lymphangiogenesis involved in tumor spread via lymphatic vessels. The aim of this study was to evaluate the potential of VEGF-D in the prediction of inguinofemoral lymph node metastasis. Methods: We analyzed the preoperative levels of serum VEGF-D (sVEGF-D) from two independent cohorts of patients with VSCC by enzyme-linked immunosorbent assay and its protein expression on tumor tissue by immunohistochemistry. Logistic regression was performed to identify the independent risk factors for lymph node metastasis, and Cox proportional hazard model was used for survival analysis. Results: High levels of sVEGF-D, but not tissue VEGF-D, significantly correlated with positive groin nodes and a more advanced International Federation of Gynecologists and Obstetricians (FIGO) stage. In multivariable analysis, a high sVEGF-D level was an independent predictor of lymph node metastasis and worse prognosis. A prediction model based on sVEGF-D, tumor grade assessed on biopsy, tumor diameter, and lymph node clinical evaluation was able to predict lymph node metastasis, reaching C-index values of 0.79 and 0.73 in the training and validation cohorts, respectively. Conclusions: The preoperative sVEGF-D level might be a reliable biomarker for the prediction of lymph node metastasis and prognosis in patients with VSCC, supporting better clinical/surgical decision. Multicenter prospective studies are required to confirm our findings.

PD-L1 quantification across tumor types using the reverse phase protein microarray: implications for precision medicine.

BACKGROUND: Anti-programmed cell death protein 1 and programmed cell death ligand 1 (PD-L1) agents are broadly used in first-line and second-line treatment across different tumor types. While immunohistochemistry-based assays are routinely used to assess PD-L1 expression, their clinical utility remains controversial due to the partial predictive value and lack of standardized cut-offs across antibody clones. Using a high throughput immunoassay, the reverse phase protein microarray (RPPA), coupled with a fluorescence-based detection system, this study compared the performance of six anti-PD-L1 antibody clones on 666 tumor samples. METHODS: PD-L1 expression was measured using five antibody clones (22C3, 28-8, CAL10, E1L3N and SP142) and the therapeutic antibody atezolizumab on 222 lung, 71 ovarian, 52 prostate and 267 breast cancers, and 54 metastatic lesions. To capture clinically relevant variables, our cohort included frozen and formalin-fixed paraffin-embedded samples, surgical specimens and core needle biopsies. Pure tumor epithelia were isolated using laser capture microdissection from 602 samples. Correlation coefficients were calculated to assess concordance between antibody clones. For two independent cohorts of patients with lung cancer treated with nivolumab, RPPA-based PD-L1 measurements were examined along with response to treatment. RESULTS: Median-center PD-L1 dynamic ranged from 0.01 to 39.37 across antibody clones. Correlation coefficients between the six antibody clones were heterogeneous (range: -0.48 to 0.95) and below 0.50 in 61% of the comparisons. In nivolumab-treated patients, RPPA-based measurement identified a subgroup of tumors, where low PD-L1 expression equated to lack of response. CONCLUSIONS: Continuous RPPA-based measurements capture a broad dynamic range of PD-L1 expression in human specimens and heterogeneous concordance levels between antibody clones. This high throughput immunoassay can potentially identify subgroups of tumors in which low expression of PD-L1 equates to lack of response to treatment.

Infiltration by CXCL10 Secreting Macrophages Is Associated With Antitumor Immunity and Response to Therapy in Ovarian Cancer Subtypes.

Ovarian carcinomas (OCs) are poorly immunogenic and immune checkpoint inhibitors (ICIs) have offered a modest benefit. In this study, high CD3+ T-cells and CD163+ tumor-associated macrophages (TAMs) densities identify a subgroup of immune infiltrated high-grade serous carcinomas (HGSCs) with better outcomes and superior response to platinum-based therapies. On the contrary, in most clear cell carcinomas (CCCs) showing poor prognosis and refractory to platinum, a high TAM density is associated with low T cell frequency. Immune infiltrated HGSC are characterized by the 30-genes signature (OC-IS30) covering immune activation and IFNγ polarization and predicting good prognosis (n = 312, TCGA). Immune infiltrated HGSC contain CXCL10 producing M1-type TAM (IRF1+pSTAT1Y701+) in close proximity to T-cells. A fraction of these M1-type TAM also co-expresses TREM2. M1-polarized TAM were barely detectable in T-cell poor CCC, but identifiable across various immunogenic human cancers. Single cell RNA sequencing data confirm the existence of a tumor-infiltrating CXCL10+IRF1+STAT1+ M1-type TAM overexpressing antigen processing and presentation gene programs. Overall, this study highlights the clinical relevance of the CXCL10+IRF1+STAT1+ macrophage subset as biomarker for intratumoral T-cell activation and therefore offers a new tool to select patients more likely to respond to T-cell or macrophage-targeted immunotherapies.

Comprehensive Profiling of Hypoxia-Related miRNAs Identifies miR-23a-3p Overexpression as a Marker of Platinum Resistance and Poor Prognosis in High-Grade Serous Ovarian Cancer.

The onset of chemo-resistant recurrence represents the principal cause of high-grade serous ovarian carcinoma (HGSOC) death. HGSOC masses are characterized by a hypoxic microenvironment, which contributes to the development of this chemo-resistant phenotype. Hypoxia regulated-miRNAs (HRMs) represent a molecular response of cancer cells to hypoxia and are involved in tumor progression. We investigated the expression of HRMs using miRNA expression data from a total of 273 advanced-stage HGSOC samples. The miRNAs associated with chemoresistance and survival were validated by RT-qPCR and target prediction, and comparative pathway analysis was conducted for target gene identification. Analysis of miRNA expression profiles indicated miR-23a-3p and miR-181c-5p over-expression as associated with chemoresistance and poor PFS. RT-qPCR data confirmed upregulation of miR-23a-3p in tumors from chemoresistant HGSOC patients and its significant association with shorter PFS. In silico miR-23a-3p target prediction and comparative pathway analysis identified platinum drug resistance as the pathway with the highest number of miR-23a-3p target genes. Among them, APAF-1 emerged as the most promising, being downregulated in platinum-resistant patients and in HGSOC chemo-resistant cells. These results highlight miR-23a-3p as a potential biomarker for HGSOC platinum response and prognosis and the miR23a-3p/APAF1 axis as a possible target to overcome platinum-resistance.

Gene Expression Profiling of Olfactory Neuroblastoma Helps Identify Prognostic Pathways and Define Potentially Therapeutic Targets.

Olfactory neuroblastoma (ONB) is a rare sinonasal neoplasm with a peculiar behavior, for which limited prognostic factors are available. Herein, we investigate the transcriptional pathways altered in ONB and correlate them with pathological features and clinical outcomes. We analyze 32 ONB patients treated with curative intent at two independent institutions from 2001 to 2019 for whom there is available pathologic and clinical data. We perform gene expression profiling on primary ONB samples and carry out functional enrichment analysis to investigate the key pathways associated with disease-free survival (DFS). The median age is 53.5 years; all patients undergo surgery and a pure endoscopic approach is adopted in the majority of cases (81.2%). Most patients have advanced disease (stages III-IV, 81.2%) and 84.4% undergo adjuvant (chemo)radiotherapy. The median follow-up is 35 months; 11 (26.8%) patients relapse. Clinical characteristics (gender, stage and Hyams' grade) are not associated with the outcomes. In contrast, TGF-beta binding, EMT, IFN-alpha response, angiogenesis, IL2-STAT5 and IL6-JAK-STAT3 signaling pathways are enriched in patients experiencing recurrence, and significantly associated with shorter DFS. Clustering of transcriptional profiles according to pathological features indicates two distinct molecular groups, defined by either cytokeratin-positive or -negative immunostaining. Definition of the characterizing ONB transcriptomic pathways may pave the way towards tailored treatment approaches.

Immunotherapy for the prevention of high-risk oral disorders malignant transformation: the IMPEDE trial.

BACKGROUND: Oral Potentially Malignant Disorders (OPMD) have a non-negligible malignant transformation rate of up to 8%. Loss of heterozygosity (LOH) in critical chromosomal loci has proven to be the most effective marker in defining the risk of transformation and it is found in about 28% of OPMD and may therefore identify patients carrying higher risk. To date, clinical management of OPMD is limited to surgical excision and clinical surveillance, which however do not fully prevent oral cancer development. Immune system has been shown to play a key role in transformation surveillance mechanism and an immunosuppressive imbalance may be responsible for progression to cancer. Given all these considerations, we designed a clinical trial with the aim to prevent OPMD neoplastic transformation and revert the LOH status. METHODS: This is a phase II, open label, single arm, multicentric trial involving Italian referral centres and expected to enrol 80 patients out of a total of 175 screened. Patients who meet all inclusion criteria and test positive for LOH after an incisional biopsy of the OPMD will undergo a short course of immunotherapy with 4 administration of avelumab. After 6 months since treatment start, resection of the entire OPMD will be performed and LOH assessment will be repeated. The follow-up for malignant transformation and safety assessment will last 30 months from the end of treatment, for a total planned study duration of approximately 5.5 years. DISCUSSION: Restoring the activity of immune system through checkpoint inhibitor may play a crucial role against malignant transformation of OPMD by reverting the balance in favour of immune control and preventing cancer occurrence. TRIAL REGISTRATION: This trial was prospectively registered in ClinicalTrials.gov as NCT04504552 on 7th August 2020.

Integrated mutational landscape analysis of uterine leiomyosarcomas.

Uterine leiomyosarcomas (uLMS) are aggressive tumors arising from the smooth muscle layer of the uterus. We analyzed 83 uLMS sample genetics, including 56 from Yale and 27 from The Cancer Genome Atlas (TCGA). Among them, a total of 55 Yale samples including two patient-derived xenografts (PDXs) and 27 TCGA samples have whole-exome sequencing (WES) data; 10 Yale and 27 TCGA samples have RNA-sequencing (RNA-Seq) data; and 11 Yale and 10 TCGA samples have whole-genome sequencing (WGS) data. We found recurrent somatic mutations in TP53, MED12, and PTEN genes. Top somatic mutated genes included TP53, ATRX, PTEN, and MEN1 genes. Somatic copy number variation (CNV) analysis identified 8 copy-number gains, including 5p15.33 (TERT), 8q24.21 (C-MYC), and 17p11.2 (MYOCD, MAP2K4) amplifications and 29 copy-number losses. Fusions involving tumor suppressors or oncogenes were deetected, with most fusions disrupting RB1, TP53, and ATRX/DAXX, and one fusion (ACTG2-ALK) being potentially targetable. WGS results demonstrated that 76% (16 of 21) of the samples harbored chromoplexy and/or chromothripsis. Clinically actionable mutational signatures of homologous-recombination DNA-repair deficiency (HRD) and microsatellite instability (MSI) were identified in 25% (12 of 48) and 2% (1 of 48) of fresh frozen uLMS, respectively. Finally, we found olaparib (PARPi; P = 0.002), GS-626510 (C-MYC/BETi; P < 0.000001 and P = 0.0005), and copanlisib (PIK3CAi; P = 0.0001) monotherapy to significantly inhibit uLMS-PDXs harboring derangements in C-MYC and PTEN/PIK3CA/AKT genes (LEY11) and/or HRD signatures (LEY16) compared to vehicle-treated mice. These findings define the genetic landscape of uLMS and suggest that a subset of uLMS may benefit from existing PARP-, PIK3CA-, and C-MYC/BET-targeted drugs.

The Claudin-Low Subtype of High-Grade Serous Ovarian Carcinoma Exhibits Stem Cell Features.

Claudin-low cancer (CL) represents a rare and biologically aggressive variant of epithelial tumor. Here, we identified a claudin-low molecular profile of ovarian high-grade serous carcinoma (HGSOC), which exhibits the main characteristics of the homonym breast cancer subtype, including low epithelial differentiation and high mesenchymal signature. Hierarchical clustering and a centroid based algorithm applied to cell line collection expression dataset labeled 6 HGSOC cell lines as CL. These have a high energy metabolism and are enriched in CD44+/CD24- mesenchymal stem-like cells expressing low levels of cell-cell adhesion molecules (claudins and E-Cadherin) and high levels of epithelial-to-mesenchymal transition (EMT) induction transcription factors (Zeb1, Snai2, Twist1 and Twist2). Accordingly, the centroid base algorithm applied to large retrospective collections of primary HGSOC samples reveals a tumor subgroup with transcriptional features consistent with the CL profile, and reaffirms EMT as the dominant biological pathway functioning in CL-HGSOC. HGSOC patients carrying CL profiles have a worse overall survival when compared to others, likely to be attributed to its undifferentiated/stem component. These observations highlight the lack of a molecular diagnostic in the management of HGSOC and suggest a potential prognostic utility of this molecular subtyping.

Genome-wide study of salivary miRNAs identifies miR-423-5p as promising diagnostic and prognostic biomarker in oral squamous cell carcinoma.

Survival rates of oral squamous cell carcinoma (OSCC) remained substantially unchanged over the last decades; thus, additional prognostic tools are strongly needed. Salivary miRNAs have emerged as excellent non-invasive cancer biomarker candidates, but their association with OSCC prognosis has not been investigated yet. In this study, we analyzed global salivary miRNA expression in OSCC patients and healthy controls, with the aim to define its diagnostic and prognostic potential. Methods: Saliva was collected from patients with newly diagnosed untreated primary OSCC and healthy controls. Global profiling of salivary miRNAs was carried out through a microarray approach, while signature validation was performed by quantitative real-time PCR (RT-qPCR). A stringent statistical approach for microarray and RT-qPCR data normalization was applied. The diagnostic performance of miRNAs and their correlation with OSCC prognosis were comprehensively analyzed. Results: In total, 25 miRNAs emerged as differentially expressed between OSCC patients and healthy controls and, among them, seven were significantly associated with disease-free survival (DFS). miR-106b-5p, miR-423-5p and miR-193b-3p were expressed at high levels in saliva of OSCC patients and their combination displays the best diagnostic performance (ROC - AUC = 0.98). Moreover, high expression of miR-423-5p was an independent predictor of poor DFS, when included in multivariate survival analysis with the number of positive lymph nodes - the only significant clinical prognosticator. Finally, we observed a significant decrease in miR-423-5p expression in matched post-operative saliva samples, suggesting its potential cancer-specific origin. Conclusion: Salivary miRNAs identified in our cohort of patients show to be accurate in OSCC detection and to effectively stratify patients according to their likelihood of relapse. These results, if validated in an independent set of patients, could be particularly promising for screening/follow-up of high-risk populations and useful for preoperative prognostic assessment.

Pre-treatment Serum HE4 Level as a Novel Independent Prognostic Biomarker for Uterine Cervical Carcinoma Patients.

In spite of the effective implementation of screening programs, uterine cervical carcinoma (UCC) remains one of the major causes of cancer death among women around the world. The aim of this study was to investigate the prognostic value of serum human epididymis protein 4 (HE4) in UCC. Pre-treatment serum samples from 109 UCC patients and 99 healthy women were analyzed for HE4 levels by a quantitative chemiluminescent microparticle immunoassay on the automated ARCHITECT instrument. HE4 serum (sHE4) levels were significantly higher in UCC patients, regardless of tumor stage, compared with healthy controls. Elevated sHE4 levels were significantly associated with advanced FIGO stage and absence of disease-free interval after treatment. In univariable analysis, higher sHE4 levels were significantly correlated with shorter overall survival and progression-free survival. In multivariable analysis, sHE4 retained its significance as independent adverse prognostic factor for both survival endpoints. This study indicates that sHE4 is associated with a more aggressive tumor phenotype and a worse patient's prognosis. These results suggest the potential role of sHE4 as a novel prognostic marker and as an indicator of high-risk UCC patients for a tailored surgical and adjuvant therapy.

Low Expression of Claudin-7 as Potential Predictor of Distant Metastases in High-Grade Serous Ovarian Carcinoma Patients.

High-grade serous ovarian carcinoma (HGSOC) usually spreads directly into the peritoneal cavity following a transcoelomic dissemination route, although distant hematogenous metastasis exist and have been reported. However, no tumor markers can currently predict the risk of distant metastases in HGSOC. Claudins, belonging to tight-junction proteins, are dysregulated in HGSOC and functionally related to cancer progression. Here we analyzed claudin-3, -4, and -7 expression as potential markers of distant metastases. Using quantitative RT-PCR and immunohistochemistry we assessed the expression of claudins in primary HGSOC tissues, normal ovarian, and normal fallopian tube epithelia and correlated it with clinicopathological features, including the site of metastasis and the route of dissemination. Gene set enrichment analysis was performed on microarray-generated gene expression data to investigate key pathways in patients with distant metastases. We found the overall expression level of claudin-3, -4, and -7 mRNA decreased in HGSOC compared to normal tubal epithelium, currently considered the potential site of origin of many HGSOC. The reduced expression of claudin-7 is significantly associated with the development of distant metastases (p = 0.016), mainly by hematogenous route (p = 0.025). In patients with diminished expression of claudin-7, immunohistochemical staining revealed a heterogeneous pattern of membranous staining with discontinuous expression of claudin-7 along the cell border, indicative of a dischoesive architecture. The estimated reduction in the probability of distant disease is of 39% per unit increase in the level of claudin-7 (p = 0.03). Genes involved in epithelial to mesenchymal transition, hypoxia, and angiogenesis processes resulted strongly associated to hematogenous recurrence. Our data suggest a potential role of claudin-7 in discriminating distant metastatic events in HGSOC patients. The quantification of its expression levels could be a useful tool to identify patient deserving a personalized follow-up in terms of clinical and radiological assessment.

Expression profiles of PRKG1, SDF2L1 and PPP1R12A are predictive and prognostic factors for therapy response and survival in high-grade serous ovarian cancer.

High-grade serous ovarian cancer (HGS-EOCs) is generally sensitive to front-line platinum (Pt)-based chemotherapy although most patients at an advanced stage relapse with progressive resistant disease. Clinical or molecular data to identify primary resistant cases at diagnosis are not yet available. HGS-EOC biopsies from 105 Pt-sensitive (Pt-s) and 89 Pt-resistant (Pt-r) patients were retrospectively selected from two independent tumor tissue collections. Pathway analysis was done integrating miRNA and mRNA expression profiles. Signatures were further validated in silico on a cohort of 838 HGS-EOC cases from a published dataset. In all, 131 mRNAs and 5 miRNAs belonging to different functionally related molecular pathways distinguish Pt-s from Pt-r cases. Then, 17 out of 23 selected elements were validated by orthogonal approaches (SI signature). As resistance to Pt is associated with a short progression-free survival (PFS) and overall survival (OS), the prognostic role of the SI signature was assessed, and 14 genes associated with PFS and OS, in multivariate analyses (SII signature). The prognostic value of the SII signature was validated in a third extensive cohort. The expression profiles of SDF2L1, PPP1R12A and PRKG1 genes (SIII signature) served as independent prognostic biomarkers of Pt-response and survival. The study identified a prognostic molecular signature based on the combined expression profile of three genes which had never been associated with the clinical outcome of HGS-EOC. This may lead to early identification, at the time of diagnosis, of patients who would not greatly benefit from standard chemotherapy and are thus eligible for novel investigational approaches.

Transcriptional Characterization of Stage I Epithelial Ovarian Cancer: A Multicentric Study.

Stage I epithelial ovarian cancer (EOC) represents about 10% of all EOCs. It is characterized by a complex histopathological and molecular heterogeneity, and it is composed of five main histological subtypes (mucinous, endometrioid, clear cell and high, and low grade serous), which have peculiar genetic, molecular, and clinical characteristics. As it occurs less frequently than advanced-stage EOC, its molecular features have not been thoroughly investigated. In this study, using in silico approaches and gene expression data, on a multicentric cohort composed of 208 snap-frozen tumor biopsies, we explored the subtype-specific molecular alterations that regulate tumor aggressiveness in stage I EOC. We found that single genes rather than pathways are responsible for histotype specificities and that a cAMP-PKA-CREB1 signaling axis seems to play a central role in histotype differentiation. Moreover, our results indicate that immune response seems to be, at least in part, involved in histotype differences, as a higher immune-reactive behavior of serous and mucinous samples was observed with respect to other histotypes.

Whole-exome sequencing of cervical carcinomas identifies activating ERBB2 and PIK3CA mutations as targets for combination therapy.

The prognosis of advanced/recurrent cervical cancer patients remains poor. We analyzed 54 fresh-frozen and 15 primary cervical cancer cell lines, along with matched-normal DNA, by whole-exome sequencing (WES), most of which harboring Human-Papillomavirus-type-16/18. We found recurrent somatic missense mutations in 22 genes (including PIK3CA, ERBB2, and GNAS) and a widespread APOBEC cytidine deaminase mutagenesis pattern (TCW motif) in both adenocarcinoma (ACC) and squamous cell carcinomas (SCCs). Somatic copy number variants (CNVs) identified 12 copy number gains and 40 losses, occurring more often than expected by chance, with the most frequent events in pathways similar to those found from analysis of single nucleotide variants (SNVs), including the ERBB2/PI3K/AKT/mTOR, apoptosis, chromatin remodeling, and cell cycle. To validate specific SNVs as targets, we took advantage of primary cervical tumor cell lines and xenografts to preclinically evaluate the activity of pan-HER (afatinib and neratinib) and PIK3CA (copanlisib) inhibitors, alone and in combination, against tumors harboring alterations in the ERBB2/PI3K/AKT/mTOR pathway (71%). Tumors harboring ERBB2 (5.8%) domain mutations were significantly more sensitive to single agents afatinib or neratinib when compared to wild-type tumors in preclinical in vitro and in vivo models (P = 0.001). In contrast, pan-HER and PIK3CA inhibitors demonstrated limited in vitro activity and were only transiently effective in controlling in vivo growth of PIK3CA-mutated cervical cancer xenografts. Importantly, combinations of copanlisib and neratinib were highly synergistic, inducing long-lasting regression of tumors harboring alterations in the ERBB2/PI3K/AKT/mTOR pathway. These findings define the genetic landscape of cervical cancer, suggesting that a large subset of cervical tumors might benefit from existing ERBB2/PIK3CA/AKT/mTOR-targeted drugs.

FXYD5 (Dysadherin) upregulation predicts shorter survival and reveals platinum resistance in high-grade serous ovarian cancer patients.

BACKGROUND: High-grade serous ovarian carcinoma (HGSOC) is generally associated with a very dismal prognosis. Nevertheless, patients with similar clinicopathological characteristics can have markedly different clinical outcomes. Our aim was the identification of novel molecular determinants influencing survival. METHODS: Gene expression profiles of extreme HGSOC survivors (training set) were obtained by microarray. Differentially expressed genes (DEGs) and enriched signalling pathways were determined. A prognostic signature was generated and validated on curatedOvarianData database through a meta-analysis approach. The best prognostic biomarker from the signature was confirmed by RT-qPCR and by immunohistochemistry on an independent validation set. Cox regression model was chosen for survival analysis. RESULTS: Eighty DEGs and the extracellular matrix-receptor (ECM-receptor) interaction pathway were associated to extreme survival. A 10-gene prognostic signature able to correctly classify patients with 98% of accuracy was identified. By an 'in-silico' meta-analysis, overexpression of FXYD domain-containing ion transport regulator 5 (FXYD5), also known as dysadherin, was confirmed in HGSOC short-term survivors compared to long-term ones. Its prognostic and predictive power was then successfully validated, both at mRNA and protein level, first on training than on validation sample set. CONCLUSION: We demonstrated the possible involvement of FXYD5 and ECM-receptor interaction signal pathway in HCSOC survival and prognosis.