New pyrazolo[3,4-d]pyrimidine SRC inhibitors induce apoptosis in mesothelioma cell lines through p27 nuclear stabilization

Abstract

Malignant mesothelioma (MM) is a highly aggressive tumor of the serous membranes for which there is currently no effective curative modality. Recent data suggest that hyperactivation of the tyrosine kinase SRC has a key role in MM development and therefore this kinase represents an important molecular target for MM therapy. We tested new pyrazolo [3,4-d]pyrimidine SRC inhibitors on a panel of MM cell lines expressing the active form of SRC. These SRC inhibitors exerted a significant proapoptotic effect on MM cells without affecting the normal mesothelial cell line MET-5A, supporting a possible use of these SRC inhibitors for a safe treatment of MM. We also showed that SRC inhibitor-induced apoptosis occurred concomitantly with an increase in the nuclear stability of the cyclindependent kinase inhibitor p27. This finding is remarkable considering that loss of nuclear p27 expression is a wellestablished adverse prognostic factor in MM, and p27 nuclear localization is crucial for its tumor-suppressive function. Consistently, SRC inhibition seems to promote the increase in p27 nuclear level also by inactivating the AKT kinase and downregulating cyclin D1, which would otherwise delay p27 nuclear import and provoke its cytoplasmic accumulation. To determine whether p27 stabilization has a direct role in apoptosis induced by SRC inhibition, we stably silenced the CDKN1B gene, encoding p27, in MSTO-211H and REN mesothelioma cells by transduction with lentiviral vectors expressing short hairpin RNAs against the CDKN1B transcript. Strikingly, p27 silencing was able to suppress the apoptosis induced by these SRC inhibitors in both MM cell lines, suggesting that p27 has a crucial proapoptotic role in MM cells treated with SRC inhibitors. Our findings reveal a new mechanism, dependent on p27 nuclear stabilization, by which SRC inhibition can induce apoptosis in MM cells and provide a new rationale for the use of SRC inhibitors in MM therapy.

Antitumor activity of new pyrazolo[3,4-d]pyrimidine SRC kinase inhibitors in Burkitt lymphoma cell lines and its enhancement by WEE1 inhibition

“…This difference in effectiveness is likely to be dependent on a different membrane penetration of the compounds in diverse cell lines, as previously suggested. 35,13 To date, several small molecules targeting SRC have been developed. 12 In the present study, we compared the effectiveness of our compounds with that of the wellknown SRC inhibitor PP2.…”

“…We previously demonstrated that our SRC inhibitors do not affect non-neoplastic cells derived from different tissues, 13,38,41 since these compounds are effective only on cancer cells expressing the active form of SRC. Moreover, it has been shown that G 2 /M checkpoint abrogators, including WEE1 inhibitors, can selectively sensitize cancer cells to anticancer agents without affecting non-cancerous cells.…”

“…Moreover, DAUDI cells also showed an increase in the level of total SRC upon treatment, consistent with previous studies suggesting the existence of a feedback loop trying to compensate for SRC inhibition. 13,14,15 SRC inhibitors induce G 2 /M phase arrest and apoptosis in BL cell lines. To evaluate whether the cytotoxic effects induced by our SRC inhibitors were due to cell cycle arrest or to cell death, we used FACS to analyze the cell cycle profile of BL cell lines after treatment with SI91 and SI163, which were the most effective molecules on RAJI and DAUDI cells, respectively.…”

“…These compounds inhibit SRC activity at nanomolar concentrations, as shown by enzymatic assays, and function as excellent antiproliferative and proapoptotic agents in several tumor types. 12,13 Here, we evaluated the effect of these SRC inhibitors on two BL cell lines (RAJI and DAUDI). We found that these molecules were able to promote apoptosis in both cell lines.…”

Novel 1,5-diphenyl-6-substituted 1H-pyrazolo[3,4-d]pyrimidin-4(5H)-ones induced apoptosis in RKO colon cancer cells

“…The pyrazolo [3,4-d]pyrimidine nucleus is considered as an isostere to the purine nucleus and hence exhibits promising antiproliferative activities. Several members of this family were found to induce apoptosis and/or reduce cell proliferation in many cancer cell lines such as breast, colon, lung and liver cell lines [9][10][11][12][13][14][15][16][17] . Different mechanisms account for the cytotoxic effect of this class of compounds, where they have been reported to act as inhibitors of different enzymes such as CDK 2 14 , epidermal growth factor 15 , Src or dual Src/AbI inhibitors the synthesis of anti-tumor agents.…”