RAC1 plays an essential role in estrogen receptor alpha function in breast cancer cells.

The activity of Rho family GTPase protein, RAC1, which plays important normal physiological functions, is dysregulated in multiple cancers. RAC1 is expressed in both estrogen receptor alpha (ER)-positive and ER-negative breast cancer (BC) cells. However, ER-positive BC is more sensitive to RAC1 inhibition. We have determined that reducing RAC1 activity, using siRNA or EHT 1864 (a small molecule Rac inhibitor), leads to rapid ER protein degradation. RAC1 interacts with ER within the ER complex and RAC1 localizes to chromatin binding sites for ER upon estrogen treatment. RAC1 activity is important for RNA Pol II function at both promoters and enhancers of ER target genes and ER-regulated gene transcription is blocked by EHT 1864, in a dose-dependent manner. Having identified that RAC1 is an essential ER cofactor for ER protein stability and ER transcriptional activity, we report that RAC1 inhibition could be an effective therapeutic approach for ER-positive BC.

Hormone-Dependent Prostate Cancers are Dependent on Rac Signaling for Growth and Survival.

Prostate cancer remains a common cause of cancer mortality in men. Initially, cancers are dependent of androgens for growth and survival. First line therapies reduce levels of circulating androgens or target the androgen receptor (AR) directly. Although most patients show durable responses, many patients eventually progress to castration-resistant prostate cancer (CRPC) creating a need for alternative treatment options. The Rac1 signaling pathway has previously been implicated as a driver of cancer initiation and disease progression. We investigated the role of HACE1, the E3 ubiquitin ligase for Rac1, in prostate cancer and found that HACE1 is commonly lost resulting in hyperactive Rac signaling leading to enhanced cellular proliferation, motility and viability. Importantly, we show that a Rac inhibitor can attenuate the growth and survival of prostate cancer cells. Rac signaling was also found to be critical in prostate cancers that express the AR. Rac inhibition in androgen dependent cells resulted in reduction of AR target gene expression suggesting that targeting Rac1 may be an alternative method for blocking the AR signaling axis. Finally, when used in combination with AR antagonists, Rac inhibition enhanced the suppression of AR target gene expression. Therefore, targeting Rac in prostate cancer has the potential to enhance the efficacy of approved AR therapies.

Spindle assembly checkpoint gene BUB1B is essential in breast cancer cell survival.

PURPOSE: The study aimed to investigate the role of spindle assembly checkpoint (SAC) in cancer cells with compromised genomic integrity. Chromosomal instability (CIN) gives cancer cells an adaptive advantage. However, maintaining the balance of this instability is crucial for the survival of cancer cells as it could lead them to the mitotic catastrophe. Therefore, cancer cells adapt to the detrimental effects of CIN. We hypothesized that changes in SAC might be one such adaptation mechanism. The focus of the study was BUB1B, an integral part of the checkpoint. METHODS: Clinical datasets were analyzed to compare expression levels of SAC genes in normal tissue vs. breast carcinoma. The effects of the reduction of BUB1B expression was examined utilizing RNA interference method with siRNAs. In vitro viability, clonogenicity, apoptosis, and SAC activity levels of a variety of breast cancer (BrCa) cell lines, as well as in vivo tumorigenicity of the triple-negative breast cancer (TNBC) cell line MDA-MB-468, were tested. Additionally, the chromosomal stability of these cells was tested by immunofluorescence staining and flow cytometry. RESULTS: In clinical breast cancer datasets, SAC genes were elevated in BrCa with BUB1B having the highest fold change. BUB1B overexpression was associated with a decreased probability of overall survival. The knockdown of BUB1B resulted in reduced viability and clonogenicity in BrCa cell lines and a significant increase in apoptosis and cell death. However, the viability and apoptosis levels of the normal breast epithelial cell line, MCF12A, were not affected. BUB1B knockdown also impaired chromosome alignment and resulted in acute chromosomal abnormalities. We also showed that BUB1B knockdown on the MDA-MB-468 cell line decreases tumor growth in mice. CONCLUSIONS: A functional spindle assembly checkpoint is essential for the survival of BrCa cells. BUB1B is a critical factor in SAC, and therefore breast cancer cell survival. Impairment of BUB1B has damaging effects on cancer cell viability and tumorigenicity, especially on the more aggressive variants of BrCa.

Rac Signaling Drives Clear Cell Renal Carcinoma Tumor Growth by Priming the Tumor Microenvironment for an Angiogenic Switch.

Clear cell renal cell carcinoma (ccRCC) remains a common cause of cancer mortality. Better understanding of ccRCC molecular drivers resulted in the development of antiangiogenic therapies that block the blood vessels that supply tumors with nutrients for growth and metastasis. Unfortunately, most ccRCC patients eventually become resistant to initial treatments, creating a need for alternative treatment options. We investigated the role of the small GTPase Rac1 in ccRCC. Analysis of ccRCC clinical samples indicates that Rac signaling drives disease progression and predicts patients with poorer outcomes. Investigation of Rac1 identifies multiple roles for Rac1 in the pathogenesis of ccRCC. Rac1 is overexpressed in RCC cell lines and drives proliferation and migratory/metastatic potential. Rac1 is also critical for endothelial cells to grow and form endothelial tubular networks potentiated by angiogenic factors. Importantly, Rac1 controls paracrine signaling of angiogenic factors including VEGF from renal carcinoma cells to surrounding blood vessels. A novel Rac1 inhibitor impaired the growth and migratory potential of both renal carcinoma cells and endothelial cells and reduced VEGF production by RCC cells, thereby limiting paracrine signaling both in vitro and in vivo Lastly, Rac1 was shown to be downstream of VEGF receptor (VEGFR) signaling and required for activation of MAPK signaling. In combination with VEGFR2 inhibitors, Rac inhibition provides enhanced suppression of angiogenesis. Therefore, targeting Rac in ccRCC has the potential to block the growth of tumor cells, endothelial cell recruitment, and paracrine signaling from tumor cells to other cells in the tumor microenvironment.

RAC1b Overexpression Confers Resistance to Chemotherapy Treatment in Colorectal Cancer.

Resistance to chemotherapy represents a major limitation in the treatment of colorectal cancer. Novel strategies to circumvent resistance are critical to prolonging patient survival. Rac1b, a constitutively activated isoform of the small GTPase Rac1, is upregulated with disease progression and promotes cell proliferation and inhibits apoptosis by activation of NF-κB signaling. Here, we show that Rac1b overexpression correlates with cancer stage and confirmed Rac1b expression is associated with increased growth through enhancing NF-κB activity. Rac1b knockdown reduced cellular proliferation and reduced NF-κB activity. Surprisingly, Rac1b expression and NF-κB activity were upregulated in cells treated with chemotherapeutics, suggesting that Rac1b facilitates chemo-resistance through activation of NF-κB signaling. Knockdown of Rac1b or Rac inhibition increases the sensitivity of the cells to oxaliplatin. When used in combination, inhibition of Rac prevents the increase in NF-κB activity associated with chemotherapy treatment and increases the sensitivity of the cells to oxaliplatin. Although Rac inhibition or oxaliplatin treatment alone reduces the growth of colorectal cancer in vivo, combination therapy results in improved outcomes compared with single agents alone. We provide the first evidence that Rac1b expression confers resistance to chemotherapy in colorectal cancer. Additionally, we show that the use of a Rac inhibitor prevents chemoresistance by blocking activation of chemotherapy induced NF-κB signaling, providing a novel strategy to overcome resistance to chemotherapy in colorectal cancer.