Anna Måsbäck

Background: Ovarian cancer is often diagnosed in advanced stages, resulting in poor outcomes. There is an unmet need for a sensitive and specific screening tool for early-stage detection of ovarian cancer. Methods: Recognizing that high-grade serous ovarian carcinoma (HGSC) is driven by copy number alterations (CNAs) and that tumor DNA can be detected in cervical samples, we analyzed CNAs from shallow whole genome sequencing of 212 cervical samples from 128 women with/without HGSC, including 29 germline BRCA1/2 mutation carriers. Using a machine-learning classifier, we developed H igh-grade serous ovarian cancer C ervical copy number sig nature (HCsig), a predictor for HGSC detection. Results: HGSC-derived CNAs were detectable with HCsig in cervical samples collected several years before diagnosis. Importantly, HCsig correctly identified HGSC in 79% of archival cervical samples, including 91% stage I-II (0-27 months pre-diagnosis), and 77% stage III-IV (0-65 months pre-diagnosis). Among patients with HGSC who had multiple pre-diagnostic samples collected during the pre-symptomatic phase, 85% had at least one HCpositive cervical sample before surgery (up to 65 months before diagnosis). Detection rates were 90% and 76% for BRCA1/2 -mutated and wildtype HGSC, respectively. Validation in 172 independent samples (0-98 months pre-diagnosis) showed 76% sensitivity and 94% specificity (AUC=0.83), including high sensitivity for early-stage cancers. Conclusions: We show that applying the HCsig classifier to cervical samples, including from non-symptomatic women several years before diagnosis, holds promise for early-stage detection and secondary prevention of HGSC. Moreover, it may serve as a screening tool to aid decision-making regarding timing of risk-reducing surgery in high-risk populations.

Overexpression of the receptor tyrosine kinase MET has been linked to poor survival in several cancer types, and MET has been suggested to interact with stem cell networks. In vitro studies have further suggested a possible benefit of a combined treatment using PARP and MET inhibitors. We used a tissue microarray (TMA) with 130 samples of advanced-stage high-grade serous fallopian tube/ovarian cancer (HGSC) to investigate the prognostic value of MET protein expression alone and in combination with the stem cell factor SOX2. The possible synergistic effects of a PARP and MET inhibitor treatment were evaluated in two cell lines with BRCA1 or BRCA2 deficiency and in their BRCA1/2-proficient counterparts. Patients with tumors positive for MET had worse overall survival (log-rank test, p = 0.015) compared to patients with MET-negative tumors. The prognostic role of MET was even more prominent in the subgroup of patients with SOX2-negative tumors (p = 0.0081). No synergistic effects of the combined treatment with PARP and MET inhibitors were found in the cell lines examined. We conclude that MET expression could be used as a marker for OS in HGSC and that stemness should be taken into consideration when evaluating the mechanisms of this effect.

Serous endometrial cancer (SEC) resembles high-grade serous ovarian cancer (HGSOC) genetically and clinically, with recurrent copy number alterations, TP53 mutations and a poor prognosis. Thus, SEC patients may benefit from targeted treatments used in HGSOC, e.g., PARP inhibitors. However, the preclinical and clinical knowledge about SEC is scarce, and the exact role of defective DNA repair in this tumor subgroup is largely unknown. We aimed to outline the prevalence of homologous recombination repair deficiency (HRD), copy-number alterations, and somatic mutations in SEC. OncoScan SNP arrays were applied to 19 tumors in a consecutive SEC series to calculate HRD scores and explore global copy-number profiles and genomic aberrations. Copy-number signatures were established and targeted sequencing of 27 HRD-associated genes was performed. All factors were examined in relation to HRD scores to investigate potential drivers of the HRD phenotype. Ten of the 19 SEC tumors (53%) had an HRD score > 42, considered to reflect an HRD phenotype. Higher HRD score was associated with loss of heterozygosity in key HRD genes, and copy-number signatures associated with non-BRCA1/2 dependent HRD in HGSOC. A high number of SECs display an HRD phenotype. It remains to be elucidated whether this also confers PARP inhibitor sensitivity.

The transcription factor SOX2 is a well-established and important stem cell marker. Its role in cancer biology remains unclear, but it has been proposed to also be a marker of cancer stem cells. We investigated the role of SOX2 protein expression in women with high-grade serous ovarian cancer (HGSOC) to determine its potential prognostic and treatment predictive value. We constructed a tissue microarray of 130 advanced stage HGSOC tumors with an average of 6 cores each, stained for SOX2 protein expression and evaluated survival outcomes. We also treated two HGSOC cell lines with carboplatin and paclitaxel and measured SOX2 expression by RT-PCR and immunoblotting at different doses and time-points. Among patients with non-radical debulking surgery overall and progression-free survival were shorter for patients with SOX2 positive tumors (mean 26 vs. 39 months, log-rank test: p = .0076, and mean 14 vs. 19 months, p = .055, respectively). Knockdown of SOX2 in cell lines did not affect growth inhibition following chemotherapy treatment. Our results show that SOX2 has a strong prognostic potential among HGSOC patients with residual tumor tissue after debulking surgery and suggest that SOX2 expressing cells remaining after non-radical debulking surgery may constitute a subpopulation of cancer stem cells with greater tumor-initiating potential.

High-grade serous ovarian cancer (HGSOC) is the most common subtype of epithelial ovarian cancer and early detection is challenging. TP53 mutations are a hallmark of HGSOC and detection of these mutations in liquid-based Pap samples could provide a method for early diagnosis. Here we evaluate the use of IBSAFE, an ultra-sensitive droplet digital PCR (ddPCR) method, for detecting TP53 mutations in liquid-based Pap samples collected from fifteen women at the time of diagnosis (diagnostic samples) and/or up to seven years prior to diagnosis (archival samples). We analysed tumours for somatic TP53 mutations with next generation sequencing and were able to detect the corresponding mutations in diagnostic samples from six of eight women, while one patient harboured a germline mutation. We further detected a mutation in an archival sample obtained 20 months prior to the ovarian cancer diagnosis. The custom designed IBSAFE assays detected minor allele frequencies (MAFs) with very high assay sensitivity (MAF = 0.0068%) and were successful despite low DNA abundance (0.17–206.14 ng, median: 17.27 ng). These results provide support for further evaluation of archival liquid-based Pap samples for diagnostic purposes and demonstrate that ultra-sensitive ddPCR should be evaluated for ovarian cancer screening in high-risk groups or in the recurrent setting.

Purpose: For primary melanomas, tumor thickness, mitotic rate, and ulceration are well-laid cornerstones of prognostication. However, a molecular exposition of melanoma aggressiveness is critically missing. We recently uncovered a four-class structure in metastatic melanoma, which predicts outcome and informs biology. This raises the possibility that a molecular structure exists even in the early stages of melanoma and that molecular determinants could underlie histophenotype and eventual patient outcome. Experimental Design: We subjected 223 archival primary melanomas to a horizontally integrated analysis of RNA expression, oncogenic mutations at 238 lesions, histomorphometry, and survival data. Results: Our previously described four-class structure that was elucidated in metastatic lesions was evident within the expression space of primary melanomas. Because these subclasses converged into two larger prognostic and phenotypic groups, we used the metastatic lesions to develop a binary subtype-based signature capable of distinguishing between “high” and “low” grade forms of the disease. The two-grade signature was subsequently applied to the primary melanomas. Compared with low-grade tumors, high-grade primary melanomas were significantly associated with increased tumor thickness, mitotic rate, ulceration (all P < 0.01), and poorer relapse-free (HR = 4.94; 95% CI, 2.84–8.59), and overall (HR = 3.66; 95% CI, 2.40–5.58) survival. High-grade melanomas exhibited elevated levels of proliferation and BRCA1/DNA damage signaling genes, whereas low-grade lesions harbored higher expression of immune genes. Importantly, the molecular-grade signature was validated in two external gene expression data sets. Conclusions: We provide evidence for a molecular organization within melanomas, which is preserved across all stages of disease. Clin Cancer Res; 18(15); 4026–36. ©2012 AACR.

Primary cutaneous malignant melanomas (CMMs) from 26 individuals belonging to nine families with an identified CDKN2A mutation were clinically and histopathologically compared with 78 matched CMM controls and with a population-based series of CMMs (n = 667). All tumours were histopathologically re-examined. CDKN2A-associated cases were significantly less invasive compared with the matched controls, with an adjusted odds ratio (adjOR) of 2.9 and a 95% confidence interval (CI) of 1.0–8.1 (P = 0.04). According to the odds ratio (OR) values, CDKN2A-associated cases seemed to have tumours more often located on the head and neck (adjOR 2.9, 95% CI 0.6–13.7), with less inflammation (adjOR 0.7, 95% CI 0.3–1.8) and regression (adjOR 0.6, 95% CI 0.2–1.8) but more frequent histological ulceration (adjOR 1.9, 95% CI 0.6–5.8). In comparison with the population-based material, CDKN2A-associated cases were significantly younger at diagnosis (crude OR 3.5, 95% CI 1.6–7.5, divided at 50 years) and had less regressive reaction in their tumours (crude OR 0.35, 95% CI 0.2–0.8). No significant differences were seen for tumour thickness between the different groups. On multivariate analysis, the overall survival was significantly worse for thicker tumours and older age (P = 0.04 for both). To our knowledge this is the first description of the histopathological features of CMMs from families with mutations in the CDKN2A gene.

A population-based study from Sweden identified 711 patients with cutaneous malignant melanoma diagnosed in 1965, 1975, 1985 and 1989. Prognostic factors were evaluated and a review of the literature was performed. On univariate analysis, thick tumours (> 0.8 mm) (odds ratio [OR] 1.8, 95% confidence interval [CI] 1.6–2.1), increasing Clark level (OR 1.8, 95% CI 1.6–2.0), ulceration (OR 1.8, 95% CI 1.6–2.0), nodular melanoma (OR 1.5, 95% CI 1.3–1.6) and increasing age (continuous variable, P < 0.0001) were associated with a shorter survival. Location on extremities (OR 0.8, 95% CI 0.7–0.9), inflammation (OR 0.8, 95% CI 0.7–0.9) and female gender (OR 0.8, 95% CI 0.8–0.9) were associated with improved survival. On multivariate analysis, thick tumours (> 0.8 mm) (OR 1.5, 95% CI 1.2–1.7) and ulceration (OR 1.4, 95% CI 1.2–1.6) were independently related to a poor prognosis, while location on extremities (OR 0.8, 95% CI 0.7–0.9), inflammation (OR 0.8, 95% CI 0.7–0.9) and female gender (OR 0.8, 95% CI 0.8–1.0) were associated with improved survival. No difference in mean tumour thickness was seen over time, but there was a significant increase in the percentage of thin melanomas (< 0.8 mm) in 1985 (P = 0.01) and 1989 (P = 0.002) compared with 1965. The incidence of melanomas with inflammation increased significantly (P = 0.04), as did age at diagnosis (P = 0.005).