Department of Pharmacology and Toxicology, Medical College of Wisconsin, Milwaukee, WI 53226.
Cancer Center, Medical College of Wisconsin, Milwaukee, WI 53226.
Proc Natl Acad Sci U S A. 2020 Feb 18;117(7):3627-3636. doi: 10.1073/pnas.1914153117. Epub 2020 Feb 4.
The chaperone protein SmgGDS promotes cell-cycle progression and tumorigenesis in human breast and nonsmall cell lung cancer. Splice variants of SmgGDS, named SmgGDS-607 and SmgGDS-558, facilitate the activation of oncogenic members of the Ras and Rho families of small GTPases through membrane trafficking via regulation of the prenylation pathway. SmgGDS-607 interacts with newly synthesized preprenylated small GTPases, while SmgGDS-558 interacts with prenylated small GTPases. We determined that cancer cells have a high ratio of SmgGDS-607:SmgGDS-558 (607:558 ratio), and this elevated ratio is associated with reduced survival of breast cancer patients. These discoveries suggest that targeting SmgGDS splicing to lower the 607:558 ratio may be an effective strategy to inhibit the malignant phenotype generated by small GTPases. Here we report the development of a splice-switching oligonucleotide, named SSO Ex5, that lowers the 607:558 ratio by altering exon 5 inclusion in SmgGDS pre-mRNA (messenger RNA). Our results indicate that SSO Ex5 suppresses the prenylation of multiple small GTPases in the Ras, Rho, and Rab families and inhibits ERK activity, resulting in endoplasmic reticulum (ER) stress, the unfolded protein response, and ultimately apoptotic cell death in breast and lung cancer cell lines. Furthermore, intraperitoneal (i.p.) delivery of SSO Ex5 in MMTV-PyMT mice redirects SmgGDS splicing in the mammary gland and slows tumorigenesis in this aggressive model of breast cancer. Taken together, our results suggest that the high 607:558 ratio is required for optimal small GTPase prenylation, and validate this innovative approach of targeting SmgGDS splicing to diminish malignancy in breast and lung cancer.
伴侣蛋白 SmgGDS 促进人乳腺癌和非小细胞肺癌的细胞周期进程和肿瘤发生。SmgGDS 的剪接变体，称为 SmgGDS-607 和 SmgGDS-558，通过调节 prenylation 途径，通过膜运输促进 Ras 和 Rho 家族致癌成员的小 GTPase 的激活。SmgGDS-607 与新合成的 prenylated 小 GTPase 相互作用，而 SmgGDS-558 与 prenylated 小 GTPase 相互作用。我们确定癌细胞具有高比例的 SmgGDS-607：SmgGDS-558（607:558 比率），并且这种升高的比率与乳腺癌患者的生存率降低相关。这些发现表明，靶向 SmgGDS 剪接以降低 607:558 比率可能是抑制小 GTPase 产生的恶性表型的有效策略。在这里，我们报告了一种剪接转换寡核苷酸的开发，该寡核苷酸命名为 SSO Ex5，通过改变 SmgGDS pre-mRNA（信使 RNA）中外显子 5 的包含来降低 607:558 比率。我们的结果表明，SSO Ex5 抑制 Ras、Rho 和 Rab 家族中小 GTPase 的 prenylation 并抑制 ERK 活性，导致内质网（ER）应激、未折叠蛋白反应，最终导致乳腺癌和肺癌细胞系中的细胞凋亡。此外，SSO Ex5 在 MMTV-PyMT 小鼠中的腹腔内（i.p.）给药在乳腺中重新定向 SmgGDS 剪接并减缓这种侵袭性乳腺癌模型中的肿瘤发生。总之，我们的结果表明高 607:558 比率是小 GTPase prenylation 的最佳要求，并验证了靶向 SmgGDS 剪接以减少乳腺癌和肺癌恶性程度的这种创新方法。
