3D chromatin remodeling potentiates transcriptional programs driving cell invasion.

The contribution of deregulated chromatin architecture, including topologically associated domains (TADs), to cancer progression remains ambiguous. CCCTC-binding factor (CTCF) is a central regulator of higher-order chromatin structure that undergoes copy number loss in over half of all breast cancers, but the impact of this defect on epigenetic programming and chromatin architecture remains unclear. We find that under physiological conditions, CTCF organizes subTADs to limit the expression of oncogenic pathways, including phosphatidylinositol 3-kinase (PI3K) and cell adhesion networks. Loss of a single CTCF allele potentiates cell invasion through compromised chromatin insulation and a reorganization of chromatin architecture and histone programming that facilitates de novo promoter-enhancer contacts. However, this change in the higher-order chromatin landscape leads to a vulnerability to inhibitors of mTOR. These data support a model whereby subTAD reorganization drives both modification of histones at de novo enhancer-promoter contacts and transcriptional up-regulation of oncogenic transcriptional networks.

p66ShcA functions as a contextual promoter of breast cancer metastasis.

BACKGROUND: The p66ShcA redox protein is the longest isoform of the Shc1 gene and is variably expressed in breast cancers. In response to a variety of stress stimuli, p66ShcA becomes phosphorylated on serine 36, which allows it to translocate from the cytoplasm to the mitochondria where it stimulates the formation of reactive oxygen species (ROS). Conflicting studies suggest both pro- and anti-tumorigenic functions for p66ShcA, which prompted us to examine the contribution of tumor cell-intrinsic functions of p66ShcA during breast cancer metastasis. METHODS: We tested whether p66ShcA impacts the lung-metastatic ability of breast cancer cells. Breast cancer cells characteristic of the ErbB2+/luminal (NIC) or basal (4T1) subtypes were engineered to overexpress p66ShcA. In addition, lung-metastatic 4T1 variants (4T1-537) were engineered to lack endogenous p66ShcA via Crispr/Cas9 genomic editing. p66ShcA null cells were then reconstituted with wild-type p66ShcA or a mutant (S36A) that cannot translocate to the mitochondria, thereby lacking the ability to stimulate mitochondrial-dependent ROS production. These cells were tested for their ability to form spontaneous metastases from the primary site or seed and colonize the lung in experimental (tail vein) metastasis assays. These cells were further characterized with respect to their migration rates, focal adhesion dynamics, and resistance to anoikis in vitro. Finally, their ability to survive in circulation and seed the lungs of mice was assessed in vivo. RESULTS: We show that p66ShcA increases the lung-metastatic potential of breast cancer cells by augmenting their ability to navigate each stage of the metastatic cascade. A non-phosphorylatable p66ShcA-S36A mutant, which cannot translocate to the mitochondria, still potentiated breast cancer cell migration, lung colonization, and growth of secondary lung metastases. However, breast cancer cell survival in the circulation uniquely required an intact p66ShcA S36 phosphorylation site. CONCLUSION: This study provides the first evidence that both mitochondrial and non-mitochondrial p66ShcA pools collaborate in breast cancer cells to promote their maximal metastatic fitness.

p66ShcA potentiates the cytotoxic response of triple-negative breast cancers to PARP inhibitors.

Triple-negative breast cancers (TNBCs) lack effective targeted therapies, and cytotoxic chemotherapies remain the standard of care for this subtype. Owing to their increased genomic instability, poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) are being tested against TNBCs. In particular, clinical trials are now interrogating the efficacy of PARPi combined with chemotherapies. Intriguingly, while response rates are low, cohort of patients do respond to PARPi in combination with chemotherapies. Moreover, recent studies suggest that an increase in levels of ROS may sensitize cells to PARPi. This represents a therapeutic opportunity, as several chemotherapies, including doxorubicin, function in part by producing ROS. We previously demonstrated that the p66ShcA adaptor protein is variably expressed in TNBCs. We now show that, in response to therapy-induced stress, p66ShcA stimulated ROS production, which, in turn, potentiated the synergy of PARPi in combination with doxorubicin in TNBCs. This p66ShcA-induced sensitivity relied on the accumulation of oxidative damage in TNBCs, rather than genomic instability, to potentiate cell death. These findings suggest that increasing the expression of p66ShcA protein levels in TNBCs represents a rational approach to bolster the synergy between PARPi and doxorubicin.