Emilie Marion Canuto

Association of large noncoding RNA HOTAIR expression and its downstream intergenic CpG island methylation with survival in breast cancer

Large noncoding RNA HOTAIR, transcribed from the antisense strand of HOXC12, interacts with Polycomb Repressive Complex 2 (PRC2) in the regulation of gene activities. Recent work suggests that it may have effects on breast cancer progression and survival. We evaluated HOTAIR expression and the methylation status of its downstream intergenic CpG island in primary breast cancers, and examined associations of these factors with clinical and pathologic features and patient survival. HOTAIR expression and DNA methylation were analyzed in tissue from 348 primary breast cancers with quantitative RT-PCR and methylation-specific PCR, respectively. HOTAIR expression and methylation varied widely in the tissues. A positive correlation was found between DNA methylation and HOTAIR expression. Methylation was associated with unfavorable disease characteristics, whereas no significant associations were found between HOTAIR expression and clinical or pathologic features. In multivariate, but not in univariate, Cox proportional hazard regression models, patients with high HOTAIR expression had lower risks of relapse and mortality than those with low HOTAIR expression. These findings suggest that the intergenic DNA methylation may have important biologic relevance in regulating HOTAIR expression, and that HOTAIR expression may not be an independent prognostic marker in breast cancer, but needs further validation in independent studies.

MicroRNA let-7a: A potential marker for selection of paclitaxel in ovarian cancer management

OBJECTIVES Let-7 is a family of small non-coding RNAs regulating the expression of many genes that control important cellular activities. Let-7 is shown in vitro to sensitize cancer cells to platinum, but induce ovarian cancer resistance to paclitaxel. This study aims to investigate the effect of let-7a expression on survival outcomes of epithelial ovarian cancer (EOC) patients treated with different chemotherapy. METHODS Let-7a expression was measured with qRT-PCR in ovarian tumors of 178 EOC patients who received platinum-based chemotherapy with and without paclitaxel after surgery. Survival analysis was performed to assess the effects of let-7a and chemotherapy on disease outcomes. RESULTS Let-7a expression was detectable in the EOC samples, but the expression was not associated with disease stage, tumor grade, histology and debulking results. Patients who responded to platinum with paclitaxel had significantly lower let-7a than those who did not. Survival analyses showed that patients with high let-7a had better survival compared to those with low let-7a when they were treated with platinum without paclitaxel. The hazards ratios (HRs) for death and disease progression were 0.52 (95% CI: 0.29-0.96) and 0.48 (0.26-0.89) for high let-7a when compared to low let-7a, respectively. However, when patients were treated with platinum and paclitaxel, high let-7a was associated with worse progression-free and overall survival. The HRs for death and disease progression were 3.87 (95% CI: 1.28-11.66) and 3.48 (95% CI: 1.25-9.67) for high let-7a when compared to low let-7a, respectively. Further studies showed that among patients with low let-7a, those treated with paclitaxel in addition to platinum survived better than those treated without paclitaxel [adjusted-HRs were 0.31 (95% CI: 0.15-0.66) for death and 0.40 (95% CI: 0.22-0.75) for disease], while among those with high let-7a, the two types of treatment made no difference in patient survival. CONCLUSIONS The study suggests that the beneficial impact of the addition of paclitaxel on EOC survival was significantly linked to let-7a levels, and that miRNAs such as let-7a may be a useful marker for selection of chemotherapeutic agents in EOC management.

Functional study of risk loci of stem cell-associated gene lin-28B and associations with disease survival outcomes in epithelial ovarian cancer

Several single-nucleotide polymorphisms (SNPs) of the stem cell-associated gene lin-28B have been identified in association with ovarian cancer and ovarian cancer-related risk factors. However, whether these SNPs are functional or might be potential biomarkers for ovarian cancer prognosis remains unknown. The purposes of this study were to investigate the functional relevance of the identified lin-28B SNPs, as well as the associations of genotype and phenotype with epithelial ovarian cancer (EOC) survival. We analyzed five SNPs and mRNA levels of lin-28B in 211 primary EOC tissues using Taqman(®) SNP genotyping assays and SYBR green-based real-time PCR, respectively. The RNA secondary structures at the region of a genome-wide association-identified intronic rs314276 were analyzed theoretically with mfold and experimentally with circular dichroism spectroscopy. We found that rs314276 was a cis-acting expression quantitative trait locus (eQTL) in both additive and dominant models, while rs7759938 and rs314277 were significant or of borderline significance in dominant models only. The rs314276 variant significantly affects RNA secondary structure. No SNPs alone were associated with patient survival. However, we found that among patients initially responding to chemotherapy, those with higher lin-28B expression had higher mortality risk (hazard ratio =3.27, 95% confidence interval: 1.63-6.56) and relapse risk (hazard ratio = 2.53, 95% confidence interval: 1.41-4.54) than those with lower expression, and these associations remained in multivariate analyses. These results suggest that rs314276 alters RNA secondary structure and thereby influences gene expression, and that lin-28B is a cancer stem cell-associated marker, which may be a pharmaceutical target in the management of EOC.

Obesity and female malignancies

Obesity increases the risk of endometrial and ovarian cancer, and oestrogen receptor (ER)-progesterone receptor (PR)-positive postmenopausal breast cancer. A modest positive association between body mass index (BMI) and cervical cancer has also been found. By contrast, an inverse correlation between BMI and premenopausal breast cancer exists. Endogenous sex hormones, insulin resistance/hyperinsulinaemia, adipokines, cytokines and chronic inflammation, among other factors, may be involved in the promotion of cancer in obese patients. Obesity is also associated with an increased risk of cancer recurrence and mortality most likely due to suboptimal treatment and/or co-morbidities. It is recommended that chemotherapy doses be calculated on the actual body weight and that radical surgery be performed as in non-obese patients. The high risk of peri-operative complications may be reduced by optimizing preoperative clinical conditions. As part of cancer prevention, obese women should be encouraged to adopt healthy lifestyles leading to weight loss and to undergo regular cancer screening.

Anti‐Prothrombin Antibodies are Associated with Adverse Pregnancy Outcome

Citation Marozio L, Curti A, Botta G, Canuto EM, Salton L, Tavella AM, Benedetto C. Anti‐prothrombin antibodies are associated with adverse pregnancy outcome. Am J Reprod Immunol 2011; 66: 404–409

Biological and Clinical Significance of MAD2L1 and BUB1, Genes Frequently Appearing in Expression Signatures for Breast Cancer Prognosis.

To investigate the biologic relevance and clinical implication of genes involved in multiple gene expression signatures for breast cancer prognosis, we identified 16 published gene expression signatures, and selected two genes, MAD2L1 and BUB1. These genes appeared in 5 signatures and were involved in cell-cycle regulation. We analyzed the expression of these genes in relation to tumor features and disease outcomes. In vitro experiments were also performed in two breast cancer cell lines, MDA-MB-231 and MDA-MB-468, to assess cell proliferation, migration and invasion after knocking down the expression of these genes. High expression of these genes was found to be associated with aggressive tumors and poor disease-free survival of 203 breast cancer patients in our study, and the association with survival was confirmed in an online database consisting of 914 patients. In vitro experiments demonstrated that lowering the expression of these genes by siRNAs reduced tumor cell growth and inhibited cell migration and invasion. Our investigation suggests that MAD2L1 and BUB1 may play important roles in breast cancer progression, and measuring the expression of these genes may assist the prediction of breast cancer prognosis.

MicroRNA let-7a modifies the effect of self-renewal gene HIWI on patient survival of epithelial ovarian cancer.

Aberrant expressions of self‐renewal gene HIWI and microRNA (miRNA) let‐7a are observed in epithelial ovarian cancer (EOC). A U‐shape association between HIWI expression and overall survival is seen in several human cancers but unknown in EOC. HIWI directly and/or indirectly interacts with let‐7a, but the clinical relevance of this interaction is yet to be addressed. Here, we analyzed HIWI and let‐7a expressions in 211 primary EOC tissues using quantitative reverse‐transcription PCR to investigate HIWI and its interaction with let‐7a in the prognostic significance of EOC. Associations of HIWI and its interaction with miRNA let‐7a with patient survival were analyzed using the Kaplan–Meier survival curves and Cox proportional hazard regression models. Kaplan–Meier survival curves showed that patients with medium HIWI had poorer overall survival than those with low or high HIWI. An 89% increased death risk (HR = 1.89, 95% CI: 1.29–2.98) was observed in the medium HIWI group in multivariate Cox proportional hazard regression analyses. Among patients with high let‐7a expression, those with medium HIWI had an increased risk of death compared to those with low HIWI (HR = 2.62, 95% CI: 1.30–5.30), whereas among those with low let‐7a, no significant association between HIWI expression and overall survival was observed (HR = 1.63, 95% CI: 0.86–3.08). Moreover, HIWI expression also affected chemotherapy response. The results suggested that miRNA let‐7a could modify the effect of HIWI expression on patient survival of EOC, expanding our understanding of the clinical relevance of HIWI and let‐7a interaction in EOC prognosis.

LIN-28B/let-7a/IGF-II axis molecular subtypes are associated with epithelial ovarian cancer prognosis

OBJECTIVES Aberrant expressions of LIN-28B, let-7a and IGF-II occur in epithelial ovarian cancer, and the LIN-28B/let-7a/IGF-II axis is associated with human disease. The purpose of this study was to investigate the associations between LIN-28B/let-7a/IGF-II axis molecular subtypes and epithelial ovarian cancer prognosis. METHODS Using quantitative reverse transcription PCR, we analyzed LIN-28B, let-7a and IGF-II mRNA in 211 primary epithelial ovarian cancer tissues, and also performed Classification and Regression Tree (CART) and survival analyses. RESULTS Four terminal subtypes were identified in the CART analysis in combination with survival analysis. Kaplan-Meier survival curves showed that subtypes LIN-28B(low)let-7a(low) and LIN-28B(low)let-7a(high)IGF-II(low) had significantly better survival than subtypes LIN-28B(high) or LIN-28B(low)let-7a(high)IGF-II(high) (p<0.0001 for overall, p=0.017 for progression-free survival, respectively). Multivariate Cox regression models showed that compared to subtype LIN-28B(high), subtypes LIN-28B(low)let-7a(low) and LIN-28B(low)let-7a(high)IGF-II(low) had significantly reduced mortality and reduced relapse risks. Moreover, subtype LIN-28B(low)let-7a(low) had better response to chemotherapy than subtype LIN-28B(high). CONCLUSIONS These results suggest that molecular subtypes of the LIN-28B/let-7a/IGF-II axis associate with heterogeneous progression and may have clinical implications in predicting epithelial ovarian cancer prognosis.

Second-line chemotherapy in recurrent clear cell ovarian cancer: Results from the Multicenter Italian Trials in Ovarian cancer (MITO-9)

Background and Aims: Ovarian clear cell carcinoma (CCC) has a poorer prognosis than other subtypes of ovarian cancer. In this study, we evaluated the responsiveness to second-line chemotherapy in recurrent ovarian CCC. Methods: The MITO-9 project investigated a cohort of patients observed between 1991 and 2007 in 20 centers. We identified 72 out of 240 patients with recurrent disease (28% stage I-II and 72% stage III-IV at diagnosis). Results: In 56% of patients, the clear cell histology was pure. Twenty-five patients were platinum-resistant, 18 were platinum-sensitive with a platinum-free interval (PFI) of 6-12 months, and 29 had a PFI >12 months. Upon recurrence, 47% of patients were treated with platinum chemotherapy according to the PFI. The overall response rate (RR) to platinum was 80%, with 55, 100, and 80% RR in patients with PFI of 6-12, >12, and >24 months. The RR to nonplatinum agents in resistant patients was 33%. Among the nonplatinum agents used in primary and secondary resistant cases, gemcitabine, administered in 12 cases, had a higher activity (RR = 66%) compared to topotecan or liposomal doxorubicin (n = 31; RR = 33 and 10%, respectively). Conclusions: This study showed that the treatment of recurrent ovarian CCC should be based on the PFI as in the other subtypes. Data in platinum-resistant patients suggest gemcitabine as the drug with the highest activity. We recommend that gemcitabine be studied prospectively in a phase 2 trial.

Paclitaxel is necessary for improved survival in epithelial ovarian cancers with homologous recombination gene mutations

PURPOSE To investigate the impact of somatic mutations in homologous recombination (HR) genes on the chemotherapeutic response and survival of patients with epithelial ovarian cancer (EOC). EXPERIMENTAL DESIGN We performed targeted massively parallel sequencing of tumor DNA from 158 patients with EOC. We associated adjuvant chemotherapy and clinical outcome with mutations in selected genes, focusing on those encoding HR proteins. RESULTS HR mutations were found in 47 (30%) tumors. We did not detect an overall survival (OS) difference in advanced stage patients whose tumors had HR mutations compared to those without (median OS of 49.6 months (95% CI 29.9-57.7) vs. 43.3 months (95% CI 31.9-75.47), p = 0.87). However, when stratified by chemotherapy regimen, patients whose tumors had TP53 and HR mutations demonstrated a marked survival advantage when treated with platinum and paclitaxel vs. platinum +/− cyclophosphamide (median OS of 90 months (95% CI 50-NA) vs. 29.5 months (95% CI 17.7-50.5), p = 0.0005). CONCLUSIONS Previous studies demonstrating a survival advantage for EOC patients with somatic HR mutations have been conducted with almost universal use of both platinum and paclitaxel. Our study is the first to our knowledge to compare cohorts with somatic HR gene mutations treated with and without paclitaxel containing platinum regimens. The survival benefit attributed to the platinum sensitivity of HR deficient ovarian cancers may depend upon the combined use of paclitaxel.