Triple-negative breast cancer (TNBC) remains the most challenging breast cancer subtype to treat. CoA synthase (CoAsy) is a bifunctional enzyme, encoded by the COASY gene, which catalyzes the last two steps of CoA biosynthesis. COASY has been reported as a hit in several large RNAi library screens for cancer. Therefore, we sought to investigate the dependency of TNBC cell line proliferation on CoAsy expression. Initially, knockdown of CoAsy expression was achieved by RNAi and reduced proliferation was observed in two TNBC cell lines, HCC1806 and MDA-MB-231. To further investigate the role of CoAsy, we established stable inducible shRNA cell lines from the same TNBC cell lines as well as the normal-like breast cell line MCF10A. Three separate cell lines, each expressing one of three different shRNA constructs targeting COASY, and a non-targeted shRNA control cell line were generated from each parent cell line. The induction of COASY shRNA for 4 days resulted in >99% knockdown of CoAsy for all three COASY shRNA constructs. However, this robust knockdown of CoAsy protein expression had no detectable impact on cell growth with 4-day induction times. Even 8-day induction times resulted in no apparent impact on cell growth. There was also no effect of CoAsy knockdown on the rate of cell migration. Measurement of CoA levels in cell lysates indicated that CoAsy knockdown reduced CoA to approximately half the normal level. Thus, CoAsy knockdown showed no detectable effect on the in vitro proliferation and migration of these cell lines possibly due to the cell’s ability to maintain adequate levels of CoA through some unknown mechanism.

三阴性乳腺癌 (TNBC) 仍然是最具挑战性的乳腺癌亚型治疗。CoA 合酶 (CoAsy) 是一种双功能酶，由 COASY 基因编码，可催化 CoA 生物合成的最后两个步骤。据报道，COASY 在几个大型的癌症 RNAi 文库筛选中很受欢迎。因此，我们试图研究 TNBC 细胞系增殖对 CoAsy 表达的依赖性。最初，通过 RNAi 实现 CoAsy 表达的敲低，并在两种 TNBC 细胞系 HCC1806 和 MDA-MB-231 中观察到增殖减少。为了进一步研究 CoAsy 的作用，我们从相同的 TNBC 细胞系以及正常样乳腺细胞系 MCF10A 建立了稳定的诱导型 shRNA 细胞系。三个独立的细胞系，每个细胞系表达针对 COASY 的三种不同 shRNA 构建体中的一种，和非靶向 shRNA 对照细胞系由每个亲本细胞系产生。COASY shRNA 诱导 4 天导致所有三种 COASY shRNA 构建体的 CoAsy 敲低 > 99%。然而，这种对 CoAsy 蛋白表达的强烈抑制在 4 天的诱导时间内对细胞生长没有可检测到的影响。即使是 8 天的诱导时间也不会对细胞生长产生明显影响。CoAsy 敲低对细胞迁移率也没有影响。细胞裂解物中 CoA 水平的测量表明 CoAsy 敲低将 CoA 降低到正常水平的大约一半。因此，CoAsy 敲低对这些细胞系的体外增殖和迁移没有显示出可检测的影响，这可能是由于细胞通过一些未知机制维持足够水平的 CoA 的能力。