No abstract

SUMMARY MutS protein homolog 2 (MSH2) is a key DNA mismatch repair protein. It forms the MSH2-MSH6 (MutSα) and MSH2-MSH3 (MutSβ) heterodimers, which help to ensure genomic integrity. MutSα not only recognizes and repairs mismatched nucleotides but also recognizes DNA adducts induced by DNA-damaging agents, and triggers cell-cycle arrest and apoptosis. Loss or depletion of MutSα from cells leads to microsatellite instability (MSI) and resistance to DNA damage. Although the level of MutSα can be reduced by the ubiquitin-proteasome pathway, the detailed mechanisms of this regulation remain elusive. Here we report that histone deacetylase 6 (HDAC6) sequentially deacetylates and ubiquitinates MSH2, leading to MSH2 degradation. In addition, HDAC6 significantly reduces cellular sensitivity to DNA-damaging agents and decreases cellular DNA mismatch repair activities by downregulation of MSH2. Overall, these findings reveal a mechanism by which proper levels of MutSα are maintained.

No abstract

Although hypoxia has been shown to reprogram cancer cells toward glycolytic shift, the identity of extrinsic stimuli that induce metabolic reprogramming independent of hypoxia, especially in ovarian cancer, is largely unknown. In this study, we use patient-derived ovarian cancer cells and high-grade serous ovarian cancer cell lines to demonstrate that lysophosphatidic acid (LPA), a lipid growth factor and GPCR ligand whose levels are substantially increased in ovarian cancer patients, triggers glycolytic shift in ovarian cancer cells. Inhibition of the G protein α-subunit Gαi2 disrupted LPA-stimulated aerobic glycolysis. LPA stimulated a pseudohypoxic response via Rac-mediated activation of NADPH oxidase and generation of reactive oxygen species, resulting in activation of HIF1α. HIF1α in turn induced expression of glucose transporter-1 and the glycolytic enzyme hexokinase-2 (HKII). Treatment of mice bearing ovarian cancer xenografts with an HKII inhibitor, 3-bromopyruvate, attenuated tumor growth and conferred a concomitant survival advantage. These studies reveal a critical role for LPA in metabolic reprogramming of ovarian cancer cells and identify this node as a promising therapeutic target in ovarian cancer. These findings establish LPA as a potential therapeutic target in ovarian cancer, revealing its role in the activation of HIF1α-mediated metabolic reprogramming in this disease. .

Cancer Associated fibroblasts (CAFs) play a critical role in cancer progression, metastasis, and therapy resistance. Molecular events that confer CAF-phenotype to predecessor-cells are not fully understood. We demonstrate here that the ovarian cancer cell-conditioned medium (OCC-CM) induces CAF-phenotype in MRC5 lung-fibroblasts and it can be mimicked by LPA. While OCC-CM and LPA stimulated the expression of cellular CAF-markers by 3-days, they induced aerobic glycolysis, a metabolic marker for CAF, by 6 hrs. OCC-CM/LPA-induced glycolysis in lung (MRC5) as well as ovarian fibroblasts (NOF151) was inhibited by the LPA-receptor antagonist, Ki16425. Ovarian cancer patient-derived ascitic fluid-induced aerobic glycolysis in both NFs and Ovarian CAFs and it was inhibited by Ki16425. Further analysis indicated that LPA upregulated HIF1α-levels and the silencing of HIF1α attenuated LPA-induced glycolysis in both NOFs and

Recent studies have identified a critical role for lysophosphatidic acid (LPA) in the progression of ovarian cancer. Using a transcription factor activation reporter array, which analyzes 45 distinct transcription factors, it has been observed that LPA observed robustly activates the transcription factor hypoxia-induced factor-1α (HIF1α) in SKOV3.ip ovarian cancer cells. HIF1α showed 150-fold increase in its activation profile compared to the untreated control. Validation of the array analysis indicated that LPA stimulates a rapid increase in the levels of HIF1α in ovarian cancer cells, with an observed maximum level of HIF1α-induction by 4 hours. Our report demonstrates that LPA stimulates the increase in HIF1α levels via Gαi2. Consistent with the role of HIF1α in epithelial to mesenchymal transition (EMT) of cancer cells, LPA stimulates EMT and associated invasive cell migration along with an increase in the expression levels N-cadherin and Slug/Snail2. Using the expression of Slug/Snail2 as a marker for EMT, we demonstrate that the inhibition of Gαi2, HIF1α or Src attenuates this response. In line with the established role of EMT in promoting invasive cell migration, our data demonstrates that the inhibition of HIF1α with the clinically used HIF1α inhibitor, PX-478, drastically attenuates LPA-stimulates invasive migration of SKOV3.ip cells. Thus, our present study demonstrates that LPA utilizes a Gαi2-mediated signaling pathway via Src kinase to stimulate an increase in HIF1α levels and downstream EMT-specific factors such as Slug, leading to invasive migration of ovarian cancer cells.

Histone deacetylase 9 (HDAC9), like most Class II HDACs, catalyzes the removal of acetyl moieties from the ⑀-amino groups of conserved lysine residues in the N-terminal tail of histones. Biologically, HDAC9 regulates a wide variety of normal and abnormal physiological functions, including cardiac growth, T-regulatory cell function, neuronal disorders, muscle differentiation, development, and cancer. In a biochemical approach to identify non-histone substrates of HDAC9, we found that HDAC9 co-purifies specifically with the ataxia telangiectasia group D-complementing (ATDC; also called TRIM29) protein. HDAC9 deacetylates ATDC, alters the ability of ATDC to associate with p53, and consequently inhibits the cell proliferation-promoting activity of ATDC. These results implicate the importance of non-histone deacetylation by HDAC9 and confirm and further extend the multifunctions of this Class II deacetylase.

In this multicenter, randomized, double-blind, placebo-controlled trial of autologous bone marrow cell therapy for CLI, the therapy was well tolerated without significant adverse events. The BMAC group demonstrated trends toward improvement in amputation, pain, quality of life, Rutherford classification, and ABI when compared with controls. This pilot allowed us to identify several areas for improvement for future trials and CLI studies. These recommendations include elimination of treadmill testing, stratification by Rutherford class, and more liberal inclusion of patients with renal insufficiency. Our strongest recommendation is that CLI studies that include Rutherford 4 patients should incorporate a composite endpoint reflecting pain and quality of life.