Multiomic Analysis of Subtype Evolution and Heterogeneity in High-Grade Serous Ovarian Carcinoma.

Multiple studies have identified transcriptome subtypes of high-grade serous ovarian carcinoma (HGSOC), but their interpretation and translation are complicated by tumor evolution and polyclonality accompanied by extensive accumulation of somatic aberrations, varying cell type admixtures, and different tissues of origin. In this study, we examined the chronology of HGSOC subtype evolution in the context of these factors using a novel integrative analysis of absolute copy-number analysis and gene expression in The Cancer Genome Atlas complemented by single-cell analysis of six independent tumors. Tumor purity, ploidy, and subclonality were reliably inferred from different genomic platforms, and these characteristics displayed marked differences between subtypes. Genomic lesions associated with HGSOC subtypes tended to be subclonal, implying subtype divergence at later stages of tumor evolution. Subclonality of recurrent HGSOC alterations was evident for proliferative tumors, characterized by extreme genomic instability, absence of immune infiltration, and greater patient age. In contrast, differentiated tumors were characterized by largely intact genome integrity, high immune infiltration, and younger patient age. Single-cell sequencing of 42,000 tumor cells revealed widespread heterogeneity in tumor cell type composition that drove bulk subtypes but demonstrated a lack of intrinsic subtypes among tumor epithelial cells. Our findings prompt the dismissal of discrete transcriptome subtypes for HGSOC and replacement by a more realistic model of continuous tumor development that includes mixtures of subclones, accumulation of somatic aberrations, infiltration of immune and stromal cells in proportions correlated with tumor stage and tissue of origin, and evolution between properties previously associated with discrete subtypes. SIGNIFICANCE: This study infers whether transcriptome-based groupings of tumors differentiate early in carcinogenesis and are, therefore, appropriate targets for therapy and demonstrates that this is not the case for HGSOC.

Molecular characterization of endometrial cancer and therapeutic implications.

PURPOSE OF REVIEW: The present article reviews genomic subtyping of endometrial carcinoma and new molecular markers with therapeutic and prognostic implications. RECENT FINDINGS: Endometrial cancer has historically been classified through histology into endometrioid (type 1) and nonendometrioid (type II, mainly serous) subtypes. Molecular classification through genomic analysis now allows for a major advance in characterization; four distinct subgroups have been identified: polymerase ε (POLE) ultramutated, microsatellite unstable, copy number low/microsatellite stable, and copy number high/'serous-like'. These subtypes have prognostic implications and may aid in the identification of early-stage patients who are at high risk for recurrence. Through analysis of surrogate markers (POLE, MSI, and p53) and other validated molecular alterations (L1CAM), it may be possible to obtain an integrated molecular risk profile. Ongoing studies are utilizing this risk profile in order to identify patients who may benefit from additional treatment for early-stage disease. SUMMARY: Molecular characterization of endometrial cancer into subgroups has prognostic and therapeutic implications. Further development of an integrated molecular risk profile may identify patients who could benefit from additional treatment because of a higher risk of recurrence.

Developing a Clinico-Molecular Test for Individualized Treatment of Ovarian Cancer: The interplay of Precision Medicine Informatics with Clinical and Health Economics Dimensions.

We report recent progress in the development of a precision test for individualized use of the VEGF-A targeting drug bevacizumab for treating ovarian cancer. We discuss the discovery model stage (i.e., past feasibility modeling and before conversion to the production test). Main results: (a) Informatics modeling plays a critical role in supporting driving clinical and health economic requirements. (b) The novel computational models support the creation of a precision test with sufficient predictivity to reduce healthcare system costs up to $30 billion over 10 years, and make the use of bevacizumab affordable without loss of length or quality of life.

Assessment of the Intraoperative Consultation Service Rendered by General Pathologists in a Scenario Where a Well-Defined Decision Algorithm Is Followed.

OBJECTIVES: Intraoperative consultation (IOC) remains an area of general practice even within subspecialized pathology departments. This study assesses the IOCs rendered in a general pathology setting where surgeons integrate these results in a well-defined algorithm, developed with the input of specialized pathologists. METHODS: The surgical decisions to perform lymphadenectomy in patients with endometrial adenocarcinoma operated on at our institution between January 2003 and June 2015 as a result of the IOC assessment of tumor size, histologic grade, and depth of invasion in the hysterectomy specimen were analyzed. RESULTS: Frozen section (FS) was examined in 801 cases. In comparison to permanent section analysis, FS International Federation of Gynecology and Obstetrics (FIGO) grade had an overall accuracy of 0.95 (95% confidence interval [CI], 0.93-0.98). The FS depth of invasion had an overall accuracy of 0.92 (95% CI, 0.89-0.94). FIGO grade was not documented in 47.8%, the depth of myometrial invasion in 45.2%, and tumor size in 41.8% of the pathology reports. CONCLUSIONS: The high omission rate of the needed parameters by the general pathologists would question their overall understanding of the paradigm shift intended by this algorithm. Possible explanations of this phenomenon and potential solutions are discussed.

Enhanced Recovery Program and Length of Stay After Laparotomy on a Gynecologic Oncology Service: A Randomized Controlled Trial.

OBJECTIVE: To estimate whether a rapid recovery program would reduce length of stay among patients undergoing laparotomy on a gynecologic oncology service. METHODS: We conducted a prospective, randomized, controlled trial comparing an enhanced recovery after surgery protocol with routine postoperative care among women undergoing laparotomy on the gynecologic oncology service. Protocol elements included: preoperative counseling, regional anesthesia, intraoperative fluid restriction, and early postoperative ambulation and feeding. A sample size of 50 per group (N=100) was planned to achieve 80% power to detect a two-day difference in our primary outcome, length of hospital stay; secondary outcomes included: total daily narcotics used, time to postoperative milestones, and complications. RESULTS: A total of 112 women were enrolled between 2013 and 2015. Nine patients did not undergo laparotomy and were excluded, leaving 52 and 51 patients in the control and intervention groups, respectively. There was no difference in length of stay between the two groups (median 3.0 in both groups; P=.36). Enhanced recovery after surgery patients used less narcotics on day 0 (10.0 compared with 5.5 morphine equivalents in the control and intervention arms, respectively, P=.09) and day 2 (10.0 compared with 7.5 morphine equivalents, respectively; P=.05); however, there was no statistically significant difference between groups in any of the secondary outcomes. Post hoc analysis based on actual anesthesia received also failed to demonstrate a difference in time to discharge. CONCLUSION: When compared with usual care, introducing a formal enhanced recovery after surgery protocol did not significantly reduce length of stay. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov, https://clinicaltrials.gov, NCT01705288.

Single cell sequencing reveals heterogeneity within ovarian cancer epithelium and cancer associated stromal cells.

OBJECTIVES: The purpose of this study was to determine the level of heterogeneity in high grade serous ovarian cancer (HGSOC) by analyzing RNA expression in single epithelial and cancer associated stromal cells. In addition, we explored the possibility of identifying subgroups based on pathway activation and pre-defined signatures from cancer stem cells and chemo-resistant cells. METHODS: A fresh, HGSOC tumor specimen derived from ovary was enzymatically digested and depleted of immune infiltrating cells. RNA sequencing was performed on 92 single cells and 66 of these single cell datasets passed quality control checks. Sequences were analyzed using multiple bioinformatics tools, including clustering, principle components analysis, and geneset enrichment analysis to identify subgroups and activated pathways. Immunohistochemistry for ovarian cancer, stem cell and stromal markers was performed on adjacent tumor sections. RESULTS: Analysis of the gene expression patterns identified two major subsets of cells characterized by epithelial and stromal gene expression patterns. The epithelial group was characterized by proliferative genes including genes associated with oxidative phosphorylation and MYC activity, while the stromal group was characterized by increased expression of extracellular matrix (ECM) genes and genes associated with epithelial-to-mesenchymal transition (EMT). Neither group expressed a signature correlating with published chemo-resistant gene signatures, but many cells, predominantly in the stromal subgroup, expressed markers associated with cancer stem cells. CONCLUSIONS: Single cell sequencing provides a means of identifying subpopulations of cancer cells within a single patient. Single cell sequence analysis may prove to be critical for understanding the etiology, progression and drug resistance in ovarian cancer.

Molecular characterization of endometrial cancer and therapeutic implications.

PURPOSE OF REVIEW: This article reviews the emerging comprehensive genomic classification of endometrial carcinoma and discusses the therapeutic implications of these subgroups. RECENT FINDINGS: Comprehensive, multiplatform evaluation of endometrial cancers by the Cancer Genome Atlas stratified the molecular aberrations into four distinct subtypes: POLE mutations, microsatellite instability, copy-number low/microsatellite stable, and copy-number high/'serous-like.' POLE-mutant tumors have a favorable prognosis and may often be overtreated. Microsatellite instability hypermutated tumors commonly have alterations in the phosphatidylinositide 3-kinases/AKT/mechanistic target of rapamycin pathway and limiting targeted therapy to this group may lead to greater response rates. Copy-number low/microsatellite stable tumors represent the majority of grade 1 and grade 2 endometrioid cancers and have an intermediate prognosis, few TP53 mutations, but frequent mutations in genes involved with Wingless-related integration site signaling. Approximately 25% of high-grade endometrioid tumors have mutational profiles that classify as copy-number high/'serous-like' and might benefit from treatment approaches similar to those for serous tumors. SUMMARY: Molecular characterization of endometrial cancer classifies tumors into prognostically significant subtypes with a broad range of therapeutic implications.

The effect of photobiomodulation on chemotherapy-induced peripheral neuropathy: A randomized, sham-controlled clinical trial.

BACKGROUND: Chemotherapy-induced peripheral neuropathy (CIPN) is a common side effect of cancer therapy with few efficacious treatments. METHODS: We enrolled 70 patients with CIPN in a randomized, double-blinded, sham-controlled, cross-over trial to determine if photobiomodulation (PBM)±physiotherapy reduced the symptoms of neuropathy compared to sham treatment. At the conclusion of follow-up, sham-arm patients could cross-over into a third arm combining PBM and physiotherapy to determine if multimodal treatment had additive effects. Treatment included 30minute sessions 3-times weekly for 6weeks using either PBM or sham therapy. Neuropathy was assessed using the modified total neuropathy score (mTNS) at initiation and 4, 8, and 16weeks after initiating treatment. RESULTS: Sham-treated patients experienced no significant change in mTNS scores at any point during the primary analysis. PBM patients experienced significant reduction in mTNS scores at all time points. Mean changes in mTNS score (and corresponding percent drop from baseline) for sham and PBM-group patients respectively were -0.1 (-0.7%) and -4.2 (-32.4%) at 4weeks (p<0.001), 0.2 (0.0%) and -6.8 (-52.6%) at 8weeks (p<0.001), and 0.0 (0.1%) and -5.0 (-38.8%) at 16weeks (p<0.001). Patients who crossed over into the PBM/PT-group experienced similar results to those treated primarily; changes in mTNS score from baseline were -5.5 (-40.6%) 4weeks (p<0.001), -6.9 (-50.9%) at 8weeks (p<0.001), and -4.9 (-35.9%) at 16weeks (p<0.001). The addition of physiotherapy did not improve outcomes over PBM alone. CONCLUSION AND RELEVANCE: Among patients with CIPN, PBM produced significant reduction in neuropathy symptoms.

PG545 enhances anti-cancer activity of chemotherapy in ovarian models and increases surrogate biomarkers such as VEGF in preclinical and clinical plasma samples.

BACKGROUND: Despite the utility of antiangiogenic drugs in ovarian cancer, efficacy remains limited due to resistance linked to alternate angiogenic pathways and metastasis. Therefore, we investigated PG545, an anti-angiogenic and anti-metastatic agent which is currently in Phase I clinical trials, using preclinical models of ovarian cancer. METHODS: PG545's anti-cancer activity was investigated in vitro and in vivo as a single agent, and in combination with paclitaxel, cisplatin or carboplatin using various ovarian cancer cell lines and tumour models. RESULTS: PG545, alone, or in combination with chemotherapeutics, inhibited proliferation of ovarian cancer cells, demonstrating synergy with paclitaxel in A2780 cells. PG545 inhibited growth factor-mediated cell migration and reduced HB-EGF-induced phosphorylation of ERK, AKT and EGFR in vitro and significantly reduced tumour burden which was enhanced when combined with paclitaxel in an A2780 model or carboplatin in a SKOV-3 model. Moreover, in the immunocompetent ID8 model, PG545 also significantly reduced ascites in vivo. In the A2780 maintenance model, PG545 initiated with, and following paclitaxel and cisplatin treatment, significantly improved overall survival. PG545 increased plasma VEGF levels (and other targets) in preclinical models and in a small cohort of advanced cancer patients which might represent a potential biomarker of response. CONCLUSION: Our results support clinical testing of PG545, particularly in combination with paclitaxel, as a novel therapeutic strategy for ovarian cancer.

Expression and functional significance of HtrA1 loss in endometrial cancer.

PURPOSE: The purpose of this study was to determine if loss of serine protease HtrA1 in endometrial cancer will promote the invasive potential of EC cell lines. EXPERIMENTAL DESIGN: Western blot analysis and immunohistochemistry methods were used to determine HtrA1 expression in EC cell lines and primary tumors, respectively. Migration, invasion assays and in vivo xenograft experiment were performed to compare the extent of metastasis between HtrA1 expressing and HtrA1 knocked down clones. RESULTS: Western blot analysis of HtrA1 in 13 EC cell lines revealed complete loss of HtrA1 expression in all seven papillary serous EC cell lines. Downregulation of HtrA1 in Hec1A and Hec1B cell lines resulted in a three- to fourfold increase in the invasive potential. Exogenous expression of HtrA1 in Ark1 and Ark2 cells resulted in three- to fourfold decrease in both invasive and migration potential of these cells. There was an increased rate of metastasis to the lungs associated with HtrA1 downregulation in Hec1B cells compared to control cells with endogenous HtrA1 expression. Enhanced expression of HtrA1 in Ark2 cells resulted in significantly less tumor nodules metastasizing to the lungs compared to parental or protease deficient (SA mutant) Ark2 cells. Immunohistochemical analysis showed 57% (105/184) of primary EC tumors had low HtrA1 expression. The association of low HtrA1 expression with high-grade endometrioid tumors was statistically significant (P = 0.016). CONCLUSIONS: Collectively, these data indicate loss of HtrA1 may contribute to the aggressiveness and metastatic ability of endometrial tumors.

Loss of HSulf-1 expression enhances autocrine signaling mediated by amphiregulin in breast cancer.

Heparan sulfate (HS) glycosaminoglycans are the oligosaccharide chains of heparan sulfate proteoglycans. The sulfation of HS glycosaminoglycan residues is required for its interaction with various heparin-binding growth factors to promote their biological activities to activate their high affinity receptor tyrosine kinases. We have identified HS glycosaminoglycan-6-O-endosulfatase HSulf-1 as a down-regulated gene in ovarian, breast, and several other cancer cell lines. Here we have shown that HSulf-1 inhibits autocrine activation of the EGFR-ERK (epidermal growth factor receptor-extracellular signal-regulated kinase) pathway induced by serum withdrawal in MDA-MB-468 breast cancer cells. Short hairpin RNA-mediated down-regulation of HSulf-1 in HSulf-1 clonal lines of MDA-MB-468 led to a significant increase in autocrine activation of ERK compared with vector only control. The autocrine signaling was also inhibited with neutralization antibodies against amphiregulin and HB-EGF, the heparin-binding growth factor family of the EGF superfamily. Furthermore, HSulf-1-mediated inhibition of autocrine signaling was associated with reduced cyclin D1 levels, leading to decreased S phase fraction and increased G(2)-M fraction, as well as increased cell death. Finally, evaluation of HSulf-1 expression levels in primary invasive breast tumors by RNA in situ hybridization indicated that HSulf-1 is down-regulated in the majority (60%) of tumors, with a predominant association with lobular histology. These data suggest a potential role of HSulf-1 down-regulation in mammary carcinogenesis.

The utility of computed tomography scans in predicting suboptimal cytoreductive surgery in women with advanced ovarian carcinoma.

BACKGROUND: Computed tomography (CT) scans of the abdomen and pelvis may predict which patients with ovarian carcinoma can undergo optimal cytoreduction at primary surgery. Previous studies have demonstrated that patients with ovarian carcinoma had optimal cytoreduction rates ranging from 50-60%. The authors sought to determine whether these findings applied to a surgical practice with a higher rate of optimal debulking. A predictive model using CT scanning and CA 125 values would allow the authors to determine which patients would be more appropriately treated with neoadjuvant chemotherapy. METHODS: Preoperative CT scans for patients with Stage III/IV ovarian carcinoma (according to the staging system of the International Federation of Gynecology and Obstetrics) who were treated between 1996 and 2001 were evaluated retrospectively by 2 radiologists for 17 criteria evaluating the extent of disease. Clinical data were extracted from medical records. Residual tumors measuring > or = 1 cm were considered suboptimal. Logistic regression was used to evaluate which criteria correlated with optimal cytoreduction. RESULTS: Eighty-seven patients were identified retrospectively who met entry criteria and had preoperative CT scans of sufficient diagnostic quality. Sixty-two patients (71%) received optimal cytoreductive surgery and 45 (52%) required aggressive surgical procedures. In a multivariate model, only diffuse peritoneal thickening (DPT) independently predicted suboptimal surgical resection (P = 0.016). However, a model using both DPT and ascites on most CT scans had a positive predictive value of 68% and a sensitivity of 52% for predicting suboptimal cytoreduction. CONCLUSIONS: The presence of DPT and large-volume ascites was associated with a very low rate of optimal cytoreduction (32%) in a surgical practice. These patients may be more appropriately treated with neoadjuvant chemotherapy followed by surgical cytoreduction.