第二代抗雄激素藥物 - 安可坦 (Enzalutamid) 是去勢抗性前列腺癌（CRPC）患者的治療新選擇。但是幾乎所有的CRPC患者在給予安可坦治療的6到12個月後，開始對安可坦產生嚴重耐藥性。根據我們之前的發現，在前列腺癌樣本獲得安可坦耐藥性後，有觀察到3'UTR使用情形的改變，並且在安可坦抗藥性（EnzaR） 細胞中也有觀察到參與聚腺苷酸化的重要因子 - Nudix 水解酶 21 （NUDT21） 的過量表達，這個結果便表明NUDT21或許藉由調控3'UTR使用情形進而影響到EnzaR。 然而在造成安可坦抗藥性中，我們仍不清楚NUDT21是如何調節3'UTR使用情形的?
有趣的是，我們在NUDT21蛋白質中鑑定到了精氨酸15的單甲基化位點（R15me1），並且之前從未有文章對其進行過研究。因此，我們假設NUDT21的R15me1可能會去調節3'UTR使用情形進而參與到EnzaR的發生，為了去驗證這一想法，我們製造了一隻能夠專一識別NUDT21 R15me1的特異性抗體，我們的結果表明，不論在體外和體內，NUDT21 R15me1的抗體都能專一的識別精氨酸15處的單甲基化狀態。並且也發現到與前列腺癌細胞 - LNCaP細胞相比，EnzaR細胞中的NUDT21 R15me1表現量是降低的，然而NUDT21總量卻是不減反增的。並且我們也發現到蛋白質精氨酸甲基轉移酶7（PRMT7）會負責去完成NUDT21蛋白質的精氨酸15單甲基化修飾。此外，當前列腺癌細胞經過雄激素處理後，會通過依賴雄激素受體（AR）的方式增加了LNCaP細胞中NUDT21 R15me1和PRMT7的表達量，同時降低了NUDT21總量的表達量。接下來我們還將了LNCaP細胞共同處理兩種藥物 - PRMT7抑製劑（SGC3027）與安可坦，結果大大增強了細胞對安可坦藥物的抗藥性。更重要的是，在獲得安可坦抗藥性後，NUDT21結合到COUP-TFII基因序列中確實從遠端的多聚腺苷酸化位點 (PAS) 轉而結合到近端PAS的位點。然而NUDT21 R15me1卻減弱了這種現象，並且在缺少NUDT21 R15me1後，會去增加NUDT21蛋白質的穩定性。最後我們也在前列腺癌的臨床樣本中分別驗證了NUDT21總量和R15me1形式的表達量。結果顯示，NUDT21的總量形式在前列腺癌中是過表達的，而NUDT21 R15me1的結果則是相反的。總而言之，我們的結果表明，安可坦藥物處理後會去抑制 PRMT7 進而導致降低了NUDT21 R15me1表現量，最終通過調節3'UTR使用情形而促進安可坦的抗藥性。

Enzalutamide, a second-generation anti-androgen drug, is a new treatment option for castration resistant prostate cancer (CRPC) patient. However, almost all of CRPC patients develop drug resistance for enzalutamide after six to twelve months of treatment. According to our previous findings, 3'UTR usage switch was observed in the PCa specimen after acquiring enzalutamide resistance and nudix hydrolase 21 (NUDT21), an alternative polyadenylation (APA) regulator, was overexpressed in enzalutamide-resistant (EnzaR) cells, suggesting its potential role involved in 3'UTR usage switch. However, the underlying mechanism of how NUDT21 regulates 3'UTR usage switch during the development of enzalutamide resistance is still unclear.
Interestingly, a potential mono-methylation site of arginine 15 (R15me1) is identified in NUDT21 protein by us and it has never been examined before. Thus, we hypothesize that R15me1 of NUDT21 may regulate 3'UTR usage switch and involve in the development of EnzaR. To test the idea, an antibody recognized NUDT21 R15me1 was generated by us and our results indicated that NUDT21 R15me1 antibody specifically recognized mono-methylation status of NUDT21 at arginine 15 in vitro and in vivo. Moreover, NUDT21 R15me1 level was decreased while NUDT21 total form was increased in EnzaR cells compared to its parental LNCaP cells. We also identified that protein arginine methyltransferase 7 (PRMT7) was responsible for adding R15me1 modification to NUDT21. In addition, androgen treatment increased both expression levels of NUDT21 R15me1 and PRMT7 while decreased the expression level of NUDT21 total form in LNCaP cells via androgen receptor (AR) dependent manner. Next, we also showed that LNCaP cells co-treated enzalutamide with PRMT7 inhibitor (SGC3027) greatly enhanced the resistance for enzalutamide treatment. Importantly, binding profile of NUDT21 total form indeed had 3'UTR usage switch from distal APA site to proximal APA site in COUP-TFII transcript after acquiring enzalutamide resistance. However, NUDT21 R15me1 attenuated this phenomena. In addition, lack of NUDT21 R15me1 increased protein stability of NUDT21. Finally, expression profile of NUDT21 total and R15me1 forms were respectively verified in the clinical specimens of PCa. Results showed that NUDT21 total form were overexpressed in the prostate cancer while NUDT21 R15me1 form had the reversed pattern. Taken together, our results suggest that down-regulation of PRMT7 by enzalutamide may lead to decrease of NUDT21 R15me1 level and contribute to develop enzalutamide resistance by causing the 3'UTR usage switch of NUDT21 downstream transcript.

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