Sunila Pradeep

Hematogenous Metastasis of Ovarian Cancer: Rethinking Mode of Spread

Ovarian cancer has a clear predilection for metastasis to the omentum, but the underlying mechanisms involved in ovarian cancer spread are not well understood. Here, we used a parabiosis model that demonstrates preferential hematogenous metastasis of ovarian cancer to the omentum. Our studies revealed that the ErbB3-neuregulin 1 (NRG1) axis is a dominant pathway responsible for hematogenous omental metastasis. Elevated levels of ErbB3 in ovarian cancer cells and NRG1 in the omentum allowed for tumor cell localization and growth in the omentum. Depletion of ErbB3 in ovarian cancer impaired omental metastasis. Our results highlight hematogenous metastasis as an important mode of ovarian cancer metastasis. These findings have implications for designing alternative strategies aimed at preventing and treating ovarian cancer metastasis.

Hypoxia-mediated downregulation of miRNA biogenesis promotes tumour progression

Cancer-related deregulation of miRNA biogenesis has been suggested, but the underlying mechanisms remain elusive. Here we report a previously unrecognized effect of hypoxia in the downregulation of Drosha and Dicer in cancer cells that leads to dysregulation of miRNA biogenesis and increased tumour progression. We show that hypoxia-mediated downregulation of Drosha is dependent on ETS1/ELK1 transcription factors. Moreover, mature miRNA array and deep sequencing studies reveal altered miRNA maturation in cells under hypoxic conditions. At a functional level, this phenomenon results in increased cancer progression in vitro and in vivo, and data from patient samples are suggestive of miRNA biogenesis downregulation in hypoxic tumours. Rescue of Drosha by siRNAs targeting ETS1/ELK1 in vivo results in significant tumour regression. These findings provide a new link in the mechanistic understanding of global miRNA downregulation in the tumour microenvironment. MicroRNAs play important roles in the maintenance of cellular homeostasis through the post-transcriptional regulation of gene expression. Here, the authors implicate loss of the miRNA biogenesis factor Drosha and altered miRNA maturation in tumour progression under hypoxic conditions.

A Core Human Primary Tumor Angiogenesis Signature Identifies the Endothelial Orphan Receptor ELTD1 as a Key Regulator of Angiogenesis

Summary Limited clinical benefits derived from anti-VEGF therapy have driven the identification of new targets involved in tumor angiogenesis. Here, we report an integrative meta-analysis to define the transcriptional program underlying angiogenesis in human cancer. This approach identified ELTD1, an orphan G-protein-coupled receptor whose expression is induced by VEGF/bFGF and repressed by DLL4 signaling. Extensive analysis of multiple cancer types demonstrates significant upregulation of ELTD1 in tumor-associated endothelial cells, with a higher expression correlating with favorable prognosis. Importantly, ELTD1 silencing impairs endothelial sprouting and vessel formation in vitro and in vivo, drastically reducing tumor growth and greatly improving survival. Collectively, these results provide insight into the regulation of tumor angiogenesis and highlight ELTD1 as key player in blood vessel formation.

Role of Focal Adhesion Kinase in Regulating YB–1–Mediated Paclitaxel Resistance in Ovarian Cancer

BACKGROUND We previously found focal adhesion kinase (FAK) inhibition sensitizes ovarian cancer to taxanes; however, the mechanisms are not well understood. METHODS We characterized the biologic response of taxane-resistant and taxane-sensitive ovarian cancer models to a novel FAK inhibitor (VS-6063). We used reverse-phase protein arrays (RPPA) to identify novel downstream targets in taxane-resistant cell lines. Furthermore, we correlated clinical and pathological data with nuclear and cytoplasmic expression of FAK and YB-1 in 105 ovarian cancer samples. Statistical tests were two-sided, and P values were calculated with Student t test or Fisher exact test. RESULTS We found that VS-6063 inhibited FAK phosphorylation at the Tyr397 site in a time- and dose-dependent manner. The combination of VS-6063 and paclitaxel markedly decreased proliferation and increased apoptosis, which resulted in 92.7% to 97.9% reductions in tumor weight. RPPA data showed that VS-6063 reduced levels of AKT and YB-1 in taxane-resistant cell lines. FAK inhibition enhanced chemosensitivity in taxane-resistant cells by decreasing YB-1 phosphorylation and subsequently CD44 in an AKT-dependent manner. In human ovarian cancer samples, nuclear FAK expression was associated with increased nuclear YB-1 expression (χ²) = 37.7; P < .001). Coexpression of nuclear FAK and YB-1 was associated with statistically significantly worse median overall survival (24.9 vs 67.3 months; hazard ratio = 2.64; 95% confidence interval = 1.38 to 5.05; P = .006). CONCLUSIONS We have identified a novel pathway whereby FAK inhibition with VS-6063 overcomes YB-1-mediated paclitaxel resistance by an AKT-dependent pathway. These findings have implications for clinical trials aimed at targeting FAK.

Autocrine Effects of Tumor-Derived Complement

SUMMARY We describe a role for the complement system in enhancing cancer growth. Cancer cells secrete complement proteins that stimulate tumor growth upon activation. Complement promotes tumor growth via a direct autocrine effect that is partially independent of tumor-infiltrating cytotoxic T cells. Activated C5aR and C3aR signal through the PI3K/AKT pathway in cancer cells, and silencing the PI3K or AKT gene in cancer cells eliminates the progrowth effects of C5aR and C3aR stimulation. In patients with ovarian or lung cancer, higher tumoral C3 or C5aR mRNA levels were associated with decreased overall survival. These data identify a role for tumor-derived complement proteins in promoting tumor growth, and they therefore have substantial clinical and therapeutic implications.

Long Noncoding RNA Ceruloplasmin Promotes Cancer Growth by Altering Glycolysis

Long noncoding RNAs (lncRNAs) significantly influence the development and regulation of genome expression in cells. Here, we demonstrate the role of lncRNA ceruloplasmin (NRCP) in cancer metabolism and elucidate functional effects leading to increased tumor progression. NRCP was highly upregulated in ovarian tumors, and knockdown of NRCP resulted in significantly increased apoptosis, decreased cell proliferation, and decreased glycolysis compared with control cancer cells. In an orthotopic mouse model of ovarian cancer, siNRCP delivered via a liposomal carrier significantly reduced tumor growth compared with control treatment. We identified NRCP as an intermediate binding partner between STAT1 and RNA polymerase II, leading to increased expression of downstream target genes such as glucose-6-phosphate isomerase. Collectively, we report a previously unrecognized role of the lncRNA NRCP in modulating cancer metabolism. As demonstrated, DOPC nanoparticle-incorporated siRNA-mediated silencing of this lncRNA in vivo provides therapeutic avenue toward modulating lncRNAs in cancer.

A miR-192-EGR1-HOXB9 regulatory network controls the angiogenic switch in cancer

A deeper mechanistic understanding of tumour angiogenesis regulation is needed to improve current anti-angiogenic therapies. Here we present evidence from systems-based miRNA analyses of large-scale patient data sets along with in vitro and in vivo experiments that miR-192 is a key regulator of angiogenesis. The potent anti-angiogenic effect of miR-192 stems from its ability to globally downregulate angiogenic pathways in cancer cells through regulation of EGR1 and HOXB9. Low miR-192 expression in human tumours is predictive of poor clinical outcome in several cancer types. Using 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC) nanoliposomes, we show that miR-192 delivery leads to inhibition of tumour angiogenesis in multiple ovarian and renal tumour models, resulting in tumour regression and growth inhibition. This anti-angiogenic and anti-tumour effect is more robust than that observed with an anti-VEGF antibody. Collectively, these data identify miR-192 as a central node in tumour angiogenesis and support the use of miR-192 in an anti-angiogenesis therapy.

Differential platelet levels affect response to taxane-based therapy in ovarian cancer

Purpose: We hypothesized that platelet levels during therapy could serve as a biomarker for response to therapy and that manipulation of platelet levels could impact responsiveness to chemotherapy. Experimental Design: The medical records of patients with recurrent or progressive ovarian cancer were retrospectively queried for changes in platelet and CA-125 levels during primary therapy. In vitro coculture experiments and in vivo orthotopic models of human ovarian cancer in mice were used to test the effect of modulating platelet levels on tumor growth and responsiveness to docetaxel. Results: Thrombocytosis at the diagnosis of ovarian cancer was correlated with decreased interval to progression (P = 0.05) and median overall survival (P = 0.007). Mean platelet levels corrected during primary therapy and rose at recurrence. Contrary to treatment-responsive patients, in a cohort of patients refractory to primary therapy, platelet levels did not normalize during therapy. In A2780, HeyA8, and SKOV3-ip1 ovarian cancer cell lines, platelet coculture protected against apoptosis (P < 0.05). In orthotopic models of human ovarian cancer, platelet depletion resulted in 70% reduced mean tumor weight (P < 0.05). Compared with mice treated with docetaxel, mice treated with both docetaxel and platelet-depleting antibody had a 62% decrease in mean tumor weight (P = 0.04). Platelet transfusion increased mean aggregate tumor weight 2.4-fold (P < 0.05), blocked the effect of docetaxel on tumor growth (P = 0.55) and decreased tumor cell apoptosis. Pretransfusion aspirinization of the platelets blocked the growth-promoting effects of transfusion. Conclusions: Platelet-driven effects of chemotherapy response may explain clinical observations. Clin Cancer Res; 21(3); 602–10. ©2014 AACR.

Calcium Dependent FAK/CREB/TNNC1 Signaling Mediates the Effect of Stromal MFAP5 on Ovarian Cancer Metastatic Potential

Ovarian cancer is the most lethal gynecologic malignancy in the United States, and advanced serous ovarian adenocarcinoma is responsible for most ovarian cancer deaths. However, the stroma-derived molecular determinants that modulate patient survival have yet to be characterized. Here we identify a stromal gene signature for advanced high-grade serous ovarian cancer using microdissected stromal ovarian tumor samples and find that stromal microfibrillar-associated protein 5 (MFAP5) is a prognostic marker for poor survival. Further functional studies reveal that FAK/CREB/TNNC1 signaling pathways mediate the effect of MFAP5 on ovarian cancer cell motility and invasion potential. Targeting stromal MFAP5 using MFAP5 specific siRNA encapsulated in chitosan nanoparticles significantly decreases ovarian tumor growth and metastasis in vivo, suggesting that it may be a new modality of ovarian cancer treatment.

Hypoxia-upregulated microRNA-630 targets Dicer, leading to increased tumor progression.

MicroRNAs (miRNAs) are small RNA molecules that affect cellular processes by controlling gene expression. Recent studies have shown that hypoxia downregulates Drosha and Dicer, key enzymes in miRNA biogenesis, causing a decreased pool of miRNAs in cancer and resulting in increased tumor growth and metastasis. Here we demonstrate a previously unrecognized mechanism by which hypoxia downregulates Dicer. We found that miR-630, which is upregulated under hypoxic conditions, targets and downregulates Dicer expression. In an orthotopic mouse model of ovarian cancer, delivery of miR-630 using 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) nanoliposomes resulted in increased tumor growth and metastasis, and decreased Dicer expression. Treatment with the combination of anti-miR-630 and anti-vascular endothelial growth factor antibody in mice resulted in rescue of Dicer expression and significantly decreased tumor growth and metastasis. These results indicate that targeting miR-630 is a promising approach to overcome Dicer deregulation in cancer. As demonstrated in the study, use of DOPC nanoliposomes for anti-miR delivery serves as a better alternative approach to cell line-based overexpression of sense or antisense miRNAs, while avoiding potential in vitro selection effects. Findings from this study provide a new understanding of miRNA biogenesis downregulation observed under hypoxia and suggest therapeutic avenues to target this dysregulation in cancer.