Cressey, L. | Search bioRxiv

bioRxiv2024

Endocrine persistence in ER+ breast cancer is accompanied by metabolic vulnerability in oxidative phosphorylation

Despite adjuvant treatment with endocrine therapies, estrogen receptor-positive (ER+) breast cancers recur in a significant proportion of patients. Recurrences are attributable to clinically undetectable endocrine-tolerant persister cancer cells that retain tumor-forming potential. Therefore, strategies targeting such persister cells may prevent recurrent disease. Using CRISPR-Cas9 genome-wide knockout screening in ER+ breast cancer cells, we identified a survival mechanism involving metabolic reprogramming with reliance upon mitochondrial respiration in endocrine-tolerant persister cells. Quantitative proteomic profiling showed reduced levels of glycolytic proteins in persisters. Metabolic tracing of glucose revealed an energy-depleted state in persisters where oxidative phosphorylation was required to generate ATP. A phase II clinical trial was conducted to evaluate changes in mitochondrial markers in primary ER+/HER2-breast tumors induced by neoadjuvant endocrine therapy (NCT04568616). In an analysis of tumor specimens from 32 patients, tumors exhibiting residual cell proliferation after aromatase inhibitor-induced estrogen deprivation with letrozole showed increased mitochondrial content. Genetic profiling and barcode lineage tracing showed that endocrine-tolerant persistence occurred stochastically without genetic predisposition. Mice bearing cell line- and patient-derived xenografts were used to measure the anti-tumor effects of mitochondrial complex I inhibition in the context of endocrine therapy. Pharmacological inhibition of complex I suppressed the tumor-forming potential of persisters and synergized with the anti-estrogen fulvestrant to induce regression of patient-derived xenografts. These findings indicate that mitochondrial metabolism is essential in endocrine-tolerant persister ER+ breast cancer cells and warrant the development of treatment strategies to leverage this vulnerability in the context of endocrine-sensitive disease. Statement of SignificanceEndocrine-tolerant persister cancer cells that survive endocrine therapy can cause recurrent disease. Persister cells exhibit increased energetic dependence upon mitochondria for survival and tumor re-growth potential.

cancer biology↗

bioRxiv2023

FAM122A ensures cell cycle interphase progression and checkpoint control as a SLiM-dependent substrate-competitive inhibitor to the B55/PP2A phosphatase

The Ser/Thr protein phosphatase 2A (PP2A) is a highly conserved collection of heterotrimeric holoenzymes responsible for the dephosphorylation of many regulated phosphoproteins. Substrate recognition and the integration of regulatory cues are mediated by B regulatory subunits that are complexed to the catalytic subunit (C) by a scaffold protein (A). PP2A/B55 substrate recruitment was thought to be mediated by charge-charge interactions between the surface of B55 and its substrates. Challenging this view, we recently discovered a conserved SLiM [RK]-V-x-x-[VI]-R in a range of proteins, including substrates such as the retinoblastoma-related protein p107 and TAU (Fowle et al. eLife 2021;10:e63181). Here we report the identification of this SLiM in FAM122A, an inhibitor of B55/PP2A. This conserved SLiM is necessary for FAM122A binding to B55 in vitro and in cells. Computational structure prediction with AlphaFold2 predicts an interaction consistent with the mutational and biochemical data and supports a mechanism whereby FAM122A uses the SLiM in the form of a short -helix to dock to the B55 top groove. In this model, FAM122A spatially constrains substrate access by occluding the catalytic subunit with a second -helix immediately adjacent to helix 1. Consistently, FAM122A functions as a competitive inhibitor as it prevents binding of substrates in in vitro competition assays and the dephosphorylation of CDK substrates by B55/PP2A in cell lysates. Ablation of FAM122A in human cell lines reduces the rate of proliferation, progression through cell cycle transitions and abrogates G1/S and intra-S phase cell cycle checkpoints. FAM122A-KO in HEK293 cells results in attenuation of CHK1 and CHK2 activation in response to replication stress. Overall, these data strongly suggest that FAM122A is a SLiM-dependent, substrate-competitive inhibitor of B55/PP2A that suppresses multiple functions of B55 in the DNA damage response and in timely progression through the cell cycle interphase.

molecular biology↗