Differential effects of rapalogues, dual kinase inhibitors on human ovarian carcinoma cells in vitro

Abstract

Ovarian cancer is the second most common gynaecological malignancy and was diagnosed in over 7,000 women in 2011 in the UK. There are currently no reliable biomarkers available for use in a regular screening assay for ovarian cancer and due to characteristic late presentation (78% in stages III and IV) ovarian cancer has a low survival rate (35% after 10 years). The mTOR pathway is a central regulator of growth, proliferation, apoptosis and angiogenesis; providing balance between available resources such as amino acids and growth factors, and stresses such as hypoxia, to control cellular behaviour accordingly. Emerging data links mTOR with the aetiopathogenesis of ovarian cancer. We hypothesised that mTOR inhibitors could play a therapeutic role in ovarian cancer treatment. In this study we began by validating the expression of four main mTOR pathway components, mTOR, DEPTOR, rictor and raptor, at gene and protein level in in vitro models of endometrioid (MDAH-2774) and clear cell (SKOV3) ovarian cancer using qPCR and ImageStream technology. Using a wound healing assay we show that inhibition of the mTOR pathway using rapamycin, rapalogues, resveratrol and NVP BEZ-235 induces a cytostatic and not cytotoxic response up to 18 h in these cell lines. We extended these findings up to 72 h with a proliferation assay and show that the effects of inhibition of the mTOR pathway are primarily mediated by the dephosphorylation of p70S6 kinase. We show that mTOR inhibition does not involve alteration of mTOR pathway components or induce caspase 9 cleavage. Preclinical studies including ovarian tissue of ovarian cancer patients, unaffected controls and patients with unrelated gynaecological conditions show that DEPTOR is reliably upregulated in ovarian cancer.

Surfactant Protein D as a Potential Biomarker and Therapeutic Target in Ovarian Cancer

“…Two microgram of total RNA was used to synthesize cDNA using the Precision nanoScript 2 Reverse Transcription kit (Primer design). Relative expression levels of mTOR, DEPTOR, Rictor and Raptor (19) were assessed by quantitative PCR (Q-PCR) on an ABI FAST HT9000 PCR instrument, using SYBR green mastermix (Primer Design). Gene expression levels were normalized to GAPDH levels.…”

“…Approximately, 70% of patients are diagnosed at advanced stage III (where ovarian cancer has spread into the upper part of the abdomen), or IV (where ovarian cancer cells are outside the abdominal cavity), with nearly 85% expected mortality. If ovarian cancer is diagnosed/detected early (Stage I–in the ovary only), its treatment is much more successful with the survival rates up to 90% (19, 20). In this study, we carried out a bioinformatic analysis to ascertain the potential prognostic role of SP-D in ovarian cancer.…”

Downregulation of DEPTOR inhibits the proliferation, migration, and survival of osteosarcoma through PI3K/Akt/mTOR pathway

“… 14 – 17 In contrast, DEPTOR is overexpressed in multiple myeloma, cervical cancer, ovarian cancer, thyroid carcinoma, and T-cell leukemia, where it is essential for Akt activation and cell survival. 5 , 6 , 18 – 20 Nevertheless, the role of DEPTOR in the tumorigenesis and progression of OS has not been elucidated. Thus, exploring the effect of DEPTOR on OS will be of great benefits and may provide new targets for OS treatment.…”

H2A Histone Family Member X (H2AX) Is Upregulated in Ovarian Cancer and Demonstrates Utility as a Prognostic Biomarker in Terms of Overall Survival

“…Emerging data also links mTOR with the aetiopathogenesis of OC. Our group has previously shown that inhibition of the mTOR pathway using rapamycin, and rapalogues induces a cytostatic effect on preclinical models of OC [24].…”