PAX2 and tamoxifen function as partners inducing the expression of pro-apoptotic genes
Tamoxifen is the most prescribed Selective Estrogen Receptor Modulator in patients with Estrogen Receptor (ER) positive breast cancers. Previously, several groups including ours have reported that Paired Box 2 protein (PAX2) is a dependent factor for tamoxifen to repress the transcription of ERBB2/HER2 in ER positive breast cancer1,2. Now, we have characterized the role of PAX2 in ER+/HER2- breast cancers, specifically under tamoxifen treatment3. We have showed that the combined action of PAX2 and tamoxifen induces both coding gene transcripts and enhancer RNAs (eRNAs) nearby genes mainly involved in cell death processes. PAX2 ChIP-seq revealed that tamoxifen boosts the binding of PAX2 towards the promoter regions of pro-apoptotic genes, while PAX2 induced-eRNAs, contribute to increase the expression of these target genes. In our study, the high expression of a subset of these PAX-related genes is associated with better patients’ survival regardless of tamoxifen treatment, whereas their expression is found to be significantly reduced in tumors relapsing to adjuvant tamoxifen. As examples of gene targets, the PAX2-tamoxifen team induces the expression of the Interferon Regulatory Factor 1 (IRF1), the E3 ubiquitin ligase RNF114B and the protein BDH14, which is associated with cell autophagy, considered as a preliminary step for apoptosis. Mechanistically, IRF1 leads to the initiation of apoptosis by activating cell cycle inhibitors such as p21cip1 or p27kip1, caspases, and/or Fas ligand5. In addition, IRF1 could also exert its function by facilitating p53 acetylation and, as a consequence, its stabilization6. Our work also indicates that tamoxifen enhances the upregulation of RNF114B. This molecule functions as an E3 ubiquitin ligase for Pirh2287, a negative regulator of p53. Hence, RNF114B, by promoting Pirh228 ubiquitination and proteosomal degradation, contributes to maintain p53 levels. Altogether, our findings show that tamoxifen and PAX2 promote the expression of genes essential for apoptosis. Furthermore, our study supports the concept that the expression of PAX2-induced genes in luminal tumors might function as a biomarker of good prognosis and also as a predictive factor of benefit from tamoxifen.
Neodjuvant setting is the best scenario to test the role of PAX2 and/or their transcripts
The Selective ER Modulator tamoxifen has been the mainstay of endocrine treatment in early and advanced breast cancer for decades. Since early 2000’s, Aromatase Inhibitors (AI) has progressively replaced tamoxifen, first in postmenopausal metastatic breast cancer patients8–11 then in postmenopausal adjuvant setting12, in the postmenopausal neoadjuvant scenario13 and more recently, in the premenopausal population with early breast cancer on GnRH analogs14,15. In both the neoadjuvant and adjuvant scenarios, tamoxifen has shown to be slightly inferior to Aromatase Inhibitors (AI). However, AI are associated to a non-meaningless toxicity (arthromyalgies, asthenia, bone loss, sexual dysfunction among others) leading to endocrine therapy discontinuation in a substantial percentage of women, for whom tamoxifen is still a suitable option). Moreover, tamoxifen is still considered an appropriate therapy in certain populations, as for men, for women with premalignant lesions, and for patients with unclear ovarian reserve, as those with peri-menopausal status or chemoinduced amenorrhea, since in these latter subgroups AI may have an incomplete effectiveness or even a stimulatory effect16.
Based on our results, we hypothesize that the poorer tamoxifen results observed in phase III studies in comparison with AI might be due to the fact that only 40–60% of luminal tumors are positive for the expression of PAX21, which might be specifically predictive of tamoxifen sensitivity. If that was the case, it is reasonable to think that tamoxifen might provide at least similar benefit than AI in a well-selected population for PAX2 positivity. To rapidly test this hypothesis in this subgroup of PAX positive patients, a neoadjuvant study comparing AI and tamoxifen, and even other endocrine therapies such a fulvestrant or new oral Selective ER Degraders, would be required. In this hypothetical study a strong translational research on baseline and post-therapy biopsies should be included. In that way, the impact of each drugs on proliferation, but also their effect on apoptosis17 along with the putative specific predictive role of PAX 2 for tamoxifen may be appropriately addressed. In relation with this, a similar o higher proportion of PEPI score equal to zero in the tamoxifen arm compared to AI/other arms would support a synergistic effect of the PAX2-tamoxifen combination and would predict an improved disease-free survival18. Finally, to reinforce this hypothesis, and besides that neoadjuvant research, to conduct a retrospective analysis of PAX2 and/or their transcripts, as IRF1, in the adjuvant scenario would be extremely useful. This could be performed in some of the phase trials that compared tamoxifen vs AI in the adjuvant scenario and in a wider population with early breast cancer endocrine naïve. This work would be highly informative of PAX and/or PAX-induced genes predictive and prognostic role, respectively, and if proven, may stablish the basis for additional prospective studies in early luminal breast cancer.
- Hurtado, A. et al. Regulation of ERBB2 by oestrogen receptor–PAX2 determines response to tamoxifen. Nature 456, 663–666 (2008).
- Beauchemin, D., Lacombe, C. & Van Themsche, C. PAX2 is activated by estradiol in breast cancer cells of the luminal subgroup selectively, to confer a low invasive phenotype. Mol. Cancer 10, 148 (2011).
- Wang, S. et al. The proapoptotic gene interferon regulatory factor-1 mediates the antiproliferative outcome of paired box 2 gene and tamoxifen. Oncogene (2020) doi:10.1038/s41388-020-01435-4.
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- Ito, A. et al. p300/CBP-mediated p53 acetylation is commonly induced by p53-activating agents and inhibited by MDM2. EMBO J. 20, 1331–1340 (2001).
- Sane, S. & Rezvani, K. Essential Roles of E3 Ubiquitin Ligases in p53 Regulation. Int. J. Mol. Sci. 18, 442 (2017).
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- Ellis, M. J. et al. Ki67 Proliferation Index as a Tool for Chemotherapy Decisions During and After Neoadjuvant Aromatase Inhibitor Treatment of Breast Cancer: Results From the American College of Surgeons Oncology Group Z1031 Trial (Alliance). J. Clin. Oncol. 35, 1061–1069 (2017).