Single-Cell RNA Sequencing of Ovarian Cancer: Promises and Challenges.

Ovarian cancer remains the leading cause of death from gynecologic malignancy in the Western world. Tumors are comprised of heterogeneous populations of various cancer, immune, and stromal cells; it is hypothesized that rare cancer stem cells within these subpopulations lead to disease recurrence and treatment resistance. Technological advances now allow for the analysis of tumor genomes and transcriptomes at the single-cell level, which provides the resolution to potentially identify these rare cancer stem cells within the larger tumor.In this chapter, we review the evolution of next-generation RNA sequencing techniques, the methodology of single-cell isolation and sequencing, sequencing data analysis, and the potential applications in ovarian cancer. We also summarize the current published work using single-cell sequencing in ovarian cancer.By utilizing this novel technique to characterize the gene expression of rare subpopulations, new targets and treatment pathways may be identified in ovarian cancer to change treatment paradigms.

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.

The impact of a wireless audio system on communication in robotic-assisted laparoscopic surgery: A prospective controlled trial.

BACKGROUND: Robotic surgery presents a challenge to effective teamwork and communication in the operating theatre (OR). Our objective was to evaluate the effect of using a wireless audio headset device on communication, efficiency and patient outcome in robotic surgery. METHODS AND FINDINGS: A prospective controlled trial of team members participating in gynecologic and urologic robotic procedures between January and March 2015. In the first phase, all surgeries were performed without headsets (control), followed by the intervention phase where all team members used the wireless headsets. Noise levels were measured during both phases. After each case, all team members evaluated the quality of communication, performance, teamwork and mental load using a validated 14-point questionnaire graded on a 1-10 scale. Higher overall scores indicated better communication and efficiency. Clinical and surgical data of all patients in the study were retrieved, analyzed and correlated with the survey results. The study included 137 procedures, yielding 843 questionnaires with an overall response rate of 89% (843/943). Self-reported communication quality was better in cases where headsets were used (113.0 ± 1.6 vs. 101.4 ± 1.6; p < .001). Use of headsets reduced the percentage of time with a noise level above 70 dB at the console (8.2% ± 0.6 vs. 5.3% ± 0.6, p < .001), but had no significant effect on length of surgery nor postoperative complications. CONCLUSIONS: The use of wireless headset devices improved quality of communication between team members and reduced the peak noise level in the robotic OR.

Biological Insights into Chemotherapy Resistance in Ovarian Cancer.

The majority of patients with high-grade serous ovarian cancer (HGSOC) initially respond to chemotherapy; however, most will develop chemotherapy resistance. Gene signatures may change with the development of chemotherapy resistance in this population, which is important as it may lead to tailored therapies. The objective of this study was to compare tumor gene expression profiles in patients before and after treatment with neoadjuvant chemotherapy (NACT). Tumor samples were collected from six patients diagnosed with HGSOC before and after administration of NACT. RNA extraction and whole transcriptome sequencing was performed. Differential gene expression, hierarchical clustering, gene set enrichment analysis, and pathway analysis were examined in all of the samples. Tumor samples clustered based on exposure to chemotherapy as opposed to patient source. Pre-NACT samples were enriched for multiple pathways involving cell cycle growth. Post-NACT samples were enriched for drug transport and peroxisome pathways. Molecular subtypes based on the pre-NACT sample (differentiated, mesenchymal, proliferative and immunoreactive) changed in four patients after administration of NACT. Multiple changes in tumor gene expression profiles after exposure to NACT were identified from this pilot study and warrant further attention as they may indicate early changes in the development of chemotherapy resistance.

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.

Single-cell sequencing in ovarian cancer: a new frontier in precision medicine.

PURPOSE OF REVIEW: This article discusses the advances, applications and challenges of using single-cell RNA sequencing data in guiding treatment decisions for ovarian cancer. RECENT FINDINGS: Genetic heterogeneity is a hallmark of ovarian cancer biology and underlies treatment resistance. Defining the different cell types present within a single ovarian cancer is difficult, but could ultimately lead to improvements in diagnosis and treatment. Next-generation sequencing technologies have rapidly increased our understanding of the molecular landscape of epithelial ovarian cancers, but the majority of these studies are conducted on bulk samples, resulting in data that represents an 'average' of all cells present. Single-cell sequencing provides a means to characterize heterogeneity with a tumor tissue in ovarian cancer patients and opens up opportunity to determine key molecular properties that influence clinical outcomes, including prognosis and treatment response. SUMMARY: Single-cell sequencing provides a powerful tool in improving our understanding of tumor cell heterogeneity for the purpose of informing personalized cancer treatment.

The prognostic significance of histologic type in early stage cervical cancer - A multi-institutional study.

BACKGROUND: Cervical adenocarcinomas (ADC) have been viewed as more aggressive than squamous cell carcinoma (SCC). We analyzed an international cohort of early stage cervical cancer to determine the impact of histologic type. METHODS: Retrospective analysis of patients with SCC (148 patients) and ADC (130 patients) stages IA1-IB2 who underwent surgery at our three institutions (two from Detroit, one from Mexico) from 2000-2010 was performed for: age, stage, tumor size, lymphovascular invasion (LVI), invasion depth, lymph node status (LN), recurrence and survival. Pathologic review proceeded inclusion. RESULTS: In the Latino population, ADC's tended to be higher grade (p=0.01), while SCC's were larger with deeper invasion (p<0.001). LVI and LN were not significantly different. Recurrence rate (RR) was 8% (8/101) in ADC and 11.8% (9/76) in SCCs. 5 year survival (OS) was equivalent (98.2% and 95.2% for ADC and SCC respectively, p=0.369). In the Detroit cohort, we noted no difference in size, grade, depth of invasion, LVI, LN. RR was 8/72 (13.7%) for SCC and 4/29 (13.7%) but not statistically different between the tumor types (p=0.5). 5 year survival was 91% and 92% for ADC and SCC, respectively. In this population 33% of the patients with SCC and 34% of the patients with ADC received adjuvant chemo-radiation (p=0.4). Histologic type demonstrated no significant outcome difference for any type of adjuvant therapy. CONCLUSION: Comparing early stage disease cervical ADC and SCC suggests equivalent recurrence and survival. Therefore, the paradigm of more aggressive management of early stage cervical ADC warrants further investigation.

Neo-adjuvant chemotherapy in the treatment of advanced malignant germ cell tumors of ovary.

OBJECTIVE: In order to preserve fertility, we attempted neo-adjuvant chemotherapy (NACT) in patients of malignant ovarian germ cell tumor (MOGCT) with advance and bulky disease. PATIENTS AND METHODS: Between January 1988 and December 2009, 23 patients received NACT. Patient's median age was 19 years, ranging from 14 to 28 years. FIGO stages III - 20 and IV - 3. Histology subtypes were: dysgerminoma, n = 14, mixed GCT, n=6 and 3 had endodermal sinus tumor. Patients were planned for four cycles of BEP (bleomycin, etoposide and cisplatin) chemotherapy followed by fertility sparing surgery (unilateral salpingo-oophorectomy+omentectomy ± lymphadenectomy). RESULTS: Following NACT - 21 patients responded; complete (CR) - 16 and partial response (PR) - 5. One patient progressed and another was lost to follow-up after 2 cycles. 18 of 21 responders underwent surgery; 13/18 had pathological CR, 5/18 had residual disease and achieved CR following 2 more cycles of BEP. 3 patients refused for surgery; 2 relapsed at 9 and 12 months, and achieved second CR following salvage chemotherapy and surgery, third patient continues to be disease-free. Currently, 21 of 23 patients are alive and disease-free at a median follow-up of 74 months. 18/21 patients have resumed menstruation and 10 eligible patients have delivered 13 full term healthy babies. These results are comparable to patients with advanced disease (n = 43) treated with standard approach (initial surgery and adjuvant chemotherapy) during the same period. CONCLUSION: NACT followed by fertility sparing surgery could be a reasonable option for patients of advanced MOGCT, not suitable for optimal cyto-reduction.