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A series of boronic-chalcone derivatives were synthesized and tested for antitumor activity against human breast cancer cell lines. The results show the boronic-chalcones are more toxic to breast cancer cells compared to normal breast cells than other known chalcones.

Epigenetic mechanisms, such as DNA methylation and histone deacetylation, may play a role in loss of estrogen receptor alpha (ER) expression in ER negative human breast cancer cells. Our previous studies showed that pharmacologic inhibition of these mechanisms using the DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine (AZA), and the histone deacetylase (HDAC) inhibitor, Trichostatin A (TSA), resulted in expression of functional ER mRNA and protein. Therefore, we sought to characterize the effects of a recently described HDAC inhibitor, Scriptaid, on cell growth and ER expression and function in ER negative human breast cancer cell lines. Scriptaid treatment of three ER negative cell lines, MDA-MB-231, MDA-MB-435 and Hs578t, resulted in significant growth inhibition and increased acetylation of H3 and H4 histone tails. Quantitative Real Time PCR showed 2000-20,000-fold increase of ER mRNA transcript in all three cell lines after 48 h of Scriptaid treatment. Further, dose dependent re-expression of an estrogen responsive gene, the progesterone receptor (PR), indicated that induced ER is functional. As seen with TSA and AZA, Scriptaid and AZA co-treatment was more effective in inducing ER than Scriptaid or AZA alone. In vivo analysis using a xenograft mouse model bearing MDA-MB-231 tumors showed decreased tumor growth following Scriptaid or TSA treatment. Our results indicate that the novel HDAC inhibitor, Scriptaid, inhibits tumor growth in vitro and in vivo and, in conjunction with AZA, acts to re-express functional ER. These data suggest that Scriptaid or related HDAC inhibitors are candidates for further study in breast cancer.

Protein phosphatase 2A (PP2A) is a ubiquitously expressed member of the serine-threonine phosphatase family that is involved in regulation of many cellular processes including transcription, translation, cellular metabolism, and apoptosis. Because of a correlation between PP2A and estrogen receptor ␣ (ER) expression in several human breast cancer cell lines, the effect of PP2A on regulation of ER expression in the human breast cancer cell line MCF-7 was studied. Inhibition of PP2A using the pharmacologic inhibitor okadaic acid at 250 nM for 16 h resulted in a 60% reduction in PP2A activity in MCF-7 cells concurrent with a 75% reduction in ER mRNA and protein expression. Similar results were obtained with a small interfering RNA probe that specifically inhibited PP2A expression. ER promoter studies showed that regulation of ER through the PP2A pathway did not occur through transcriptional activation. Rather, PP2A mediated ER expression through modulation of ER mRNA stability through degradation of ER mRNA, reversible with concomitant treatment with the proteasomal inhibitor MG 132. These data suggest a novel pathway controlling ER expression resulting from the activation of PP2A, potentially providing a novel therapeutic target.

Absence of the estrogen receptor alpha (ER) in human breast cancer cells is an indicator of poor prognosis, and predictive of lack of response to hormonal therapy. Previous studies in our laboratory and others have shown that epigenetic regulation, including DNA methylation and histone deacetylation, are common mechanisms leading to ER gene silencing. Through the use of pharmacologic inhibitors, 5-aza 2'deoxycytidine (AZA) and Trichostatin A (TSA), we have shown that alterations in both of these mechanisms results in synergistic reexpression of ER mRNA and functional protein. These alterations may play a larger role in stimulation of cell signaling pathways leading to ER expression. We have utilized newly developed genome wide screening microarray techniques to identify gene(s) contributing to the hormone independent phenotype and AZA/TSA mediated ER expression. From this screen, we identified and confirmed expression of 4 candidate genes (PP2A, XCL1, THY1 and NBC4) as potential regulators of the hormone independent phenotype. Expression of two genes, XCL1 and PP2A, appeared to be correlated with ER expression. PP2A expression was not changed with ER degradation using ICI 182,780 whereas XCL1 expression decreased in the presence of AZA/TSA and ICI 182,780. This suggests that PP2A may be a determinant of ER expression while XCL1 appears to be ER responsive and downstream of ER expression. These gene products may be novel targets to be further explored in the development of new therapeutics for ER negative breast cancer.

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2016-11-02T18:49:00

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