Despite significant progress in diagnostics and treatments, such as targeted endocrine therapy, 30% of patients with hormone-dependent breast cancer eventually develop disease recurrence predominantly due to drug resistance. Resistance to hormone deprivation therapy is multifactorial and involves several molecular events. An increasing body of research has identified an emerging hallmark of cancer describing the capability of modifying and reprogramming cellular metabolism in order to fuel neoplastic proliferation. Our group has previously uncovered how the specific type of treatment plays a significant role in this process. In particular, breast cancer (BCa) cells developing resistance to aromatase inhibitors (AI) endogenously trigger cholesterol biosynthesis (CB) through sterol regulatory element binding protein 1 (SREBP1) regulation leading to a sustained oestrogen independent, oestrogen receptor alpha (ERα) activation. Cellular lipid metabolism is controlled by SREBP1. The altered lipid metabolism, also known as “lipogenic phenotype”, has been linked with prostate cancer (PCa) pathogenesis: the expression of SREBP1 in prostate cancer is strongly correlated with Gleason grade (pathological grade) and its overexpression is sufficient to increase tumorigenicity and invasion of prostate cancer cells. Furthermore, de novo lipid biosynthesis has been associated with cancer progression, poorer prognosis and shorter patient survival. Considering what is known about the pathobiology of lipids in cancer, it is plausible that invading cells evolve mechanisms to bypass the tight homeostatic regulation of intracellular cholesterol adapting to their new environmental conditions. With this idea in mind, we sought to identify the molecular mechanisms of activation of SREBP1 as key regulator of de novo cholesterol biosynthesis in hormone-dependent cancers resistant to endocrine therapy.

Firstly, we wanted to investigate SREBP1 regulation in hormone-dependent cancer cells. We found that SREBP1-driven lipogenesis is consistently upregulated after long-term steroid deprivation, thus when cells become hormone independent. In vivo immunohistochemistry (IHC) data support the hypothesis that SREBP1 might be pivotal in driving de novo cholesterol biosynthesis in endocrine therapy resistant BCa cells. Moreover, the switch of metabolic dependency upon resistance development identified by metabolic profiling, is associated with increased de novo cholesterol and fatty acid synthesis.
In order to examine whether cholesterol biosynthesis may be upregulated by modulating SREBP1 signalling, we investigated SREBP1 recruitment to the chromatin. Optimization of ChIP protocol allowed for a genome-wide profiling of SREBP1 binding in BCa and PCa cancer cell lines. Downstream analysis showed a difference in SREBP1 recruitment between parental and long-term starved derived cell lines in MCF7. Furthermore, SREBP1 binding profiles distinguished cancer cells based on the tissue of origin (breast versus prostate cancer). Our data also confirmed a significant co-occurrence between AR and SREBP1 binding sites in PCa and suggested a possible crosstalk between SREBP1 and the ERα in BCa on chromatin.
Thirdly, we asked what the targets of SREBP1 are and if they can promote invasive potential. ChIP-seq differential binding analysis unexpectedly revealed non-canonical targets for SREBP1. In particular, we showed that cells acquiring resistance to AI undergo active cytoskeleton re-organisation via Keratin 80 (KRT80) and actin remodelling. This process is driven by epigenetic reprogramming at the type II keratin locus dependent on de novo SREBP1 binding to a single enhancer that is activated upon chronic AI treatment and leading to KRT80 upregulation. Our data strongly suggest that therapy plays a direct role in shaping the biophysical properties and invasive potential of breast cancer cells, by inducing epigenetic rearrangements leading to KRT80 upregulation and concomitant cytoskeletal reorganization.

In summary, our study investigates the role of SREBP1 as a key player in endogenous cholesterol accumulation and autonomous activation of the nuclear receptor signalling, leading to the hormone independent tumour proliferation and invasion via global cytoskeletal re-arrangements in hormone-dependent cancers.