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CRISPR-Cas9 screen reveals a role of purine synthesis for estrogen receptor α activity and tamoxifen resistance of breast cancer cells.

ABSTRACT: In breast cancer, resistance to endocrine therapies that target estrogen receptor α (ERα), such as tamoxifen and fulvestrant, remains a major clinical problem. Whether and how ERα⁺ breast cancers switch from being estrogen-dependent to estrogen-independent remains unclear. With a genome-wide CRISPR-Cas9 knockout screen, we identified previously unknown biomarkers and potential therapeutic targets of endocrine resistance. We demonstrate that high levels of PAICS, an enzyme involved in the de novo biosynthesis of purines, can shift the balance of ERα activity to be more estrogen-independent and tamoxifen-resistant. We find that this may be due to elevated activities of cAMP-activated protein kinase A and mTOR, kinases known to phosphorylate ERα specifically and to stimulate its activity. Genetic or pharmacological targeting of PAICS sensitizes tamoxifen-resistant cells to tamoxifen. Addition of purines renders them more resistant. On the basis of these findings, we propose the combined targeting of PAICS and ERα as a new, effective, and potentially safe therapeutic regimen.

SUBMITTER: Hany D

PROVIDER: S-EPMC10181187 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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CRISPR-Cas9 screen reveals a role of purine synthesis for estrogen receptor α activity and tamoxifen resistance of breast cancer cells.

Hany Dina D Vafeiadou Vasiliki V Picard Didier D

Science advances 20230512 19

In breast cancer, resistance to endocrine therapies that target estrogen receptor α (ERα), such as tamoxifen and fulvestrant, remains a major clinical problem. Whether and how ERα<sup>+</sup> breast cancers switch from being estrogen-dependent to estrogen-independent remains unclear. With a genome-wide CRISPR-Cas9 knockout screen, we identified previously unknown biomarkers and potential therapeutic targets of endocrine resistance. We demonstrate that high levels of PAICS, an enzyme involved in ...[more]

PMID: 37172090

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Project description:BackgroundEstrogen receptor (ER) α is a successful therapeutic target in breast cancer, but patients eventually develop resistance to antiestrogens such as tamoxifen.MethodsTo identify genes whose expression was associated with the development of tamoxifen resistance and metastasis, we used microarrays to compare gene expression in four primary tumors from tamoxifen-treated patients whose breast cancers did not recur vs five metastatic tumors from patients whose cancers progressed during adjuvant tamoxifen treatment. Because Rho guanine dissociation inhibitor (GDI) α was underexpressed in the tamoxifen-resistant group, we stably transfected ERα-positive MCF-7 breast cancer cells with a plasmid encoding a short hairpin (sh) RNA to silence Rho GDIα expression. We used immunoblots and transcription assays to examine the role of Rho GDIα in ER-related signaling and growth of cells in vitro and as xenografts in treated nude mice (n = 8-9 per group) to examine the effects of Rho GDIα blockade on hormone responsiveness and metastatic behavior. The time to tumor tripling as the time in weeks from randomization to a threefold increase in total tumor volume over baseline was examined in treated mice. The associations of Rho GDIα and MTA2 levels with tamoxifen resistance were examined in microarray data from patients. All statistical tests were two-sided.ResultsRho GDIα was expressed at lower levels in ERα-positive tumors that recurred during tamoxifen treatment than in ERα-positive tamoxifen-sensitive primary tumors. MCF-7 breast cancer cells in which Rho GDIα expression had been silenced were tamoxifen-resistant, had increased Rho GTPase and p21-activated kinase 1 activity, increased phosphorylation of ERα at serine 305, and enhanced tamoxifen-induced ERα transcriptional activity compared with control cells. MCF-7 cells in which Rho GDIα expression was silenced metastasized with high frequency when grown as tumor xenografts. When mice were treated with estrogen or estrogen withdrawal, tripling times for xenografts from cells with Rho GDIα silencing were similar to those from vector-containing control cells; however, tripling times were statistically significantly faster than control when mice were treated with tamoxifen (median tripling time for tumors with Rho GDIα small interfering RNA = 2.34 weeks; for control tumors = not reached, hazard ratio = 4.13, 95% confidence interval = 1.07 to 15.96, P = .040 [adjusted for multiple comparisons, P = .119]). Levels of the metastasis-associated protein MTA2 were also increased upon Rho GDIα silencing, and combined Rho GDIα and MTA2 levels were associated with recurrence in 250 tamoxifen-treated patients.ConclusionLoss of Rho GDIα enhances metastasis and resistance to tamoxifen via effects on both ERα and MTA2 in models of ERα-positive breast cancer and in tumors of tamoxifen-treated patients.

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CRISPR-Cas9 screen reveals a role of purine synthesis for estrogen receptor α activity and tamoxifen resistance of breast cancer cells.

Publications

CRISPR-Cas9 screen reveals a role of purine synthesis for estrogen receptor α activity and tamoxifen resistance of breast cancer cells.

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