Migration and invasion of cancer cells constitute fundamental processes in tumor progression and metastasis. Migratory cancer cells commonly upregulate expression of plasminogen activator inhibitor 1 (PAI1), and PAI1 correlates with poor prognosis in breast cancer. However, mechanisms by which PAI1 promotes migration of cancer cells remain incompletely defined. Here we show that increased PAI1 drives rearrangement of the actin cytoskeleton, mitochondrial fragmentation, and glycolytic metabolism in triple-negative breast cancer (TNBC) cells. In two-dimensional environments, both stable expression of PAI1 and treatment with recombinant PAI1 increased migration, which could be blocked with the specific inhibitor tiplaxtinin. PAI1 also promoted invasion into the extracellular matrix from coculture spheroids with human mammary fibroblasts in fibrin gels. Elevated cellular PAI1 enhanced cytoskeletal features associated with migration, actin-rich migratory structures, and reduced actin stress fibers. In orthotopic tumor xenografts, we discovered that TNBC cells with elevated PAI1 show collagen fibers aligned perpendicular to the tumor margin, an established marker of invasive breast tumors. Further studies revealed that PAI1 activates ERK signaling, a central regulator of motility, and promotes mitochondrial fragmentation. Consistent with known effects of mitochondrial fragmentation on metabolism, fluorescence lifetime imaging microscopy of endogenous NADH showed that PAI1 promotes glycolysis in cell-based assays, orthotopic tumor xenografts, and lung metastases. Together, these data demonstrate for the first time that PAI1 regulates cancer cell metabolism and suggest targeting metabolism to block motility and tumor progression.Implications: We identified a novel mechanism through which cancer cells alter their metabolism to promote tumor progression.

Bone constitutes the most common site of breast cancer metastases either at time of presentation or recurrent disease years after seemingly successful therapy. Bone metastases cause substantial morbidity, including life-threatening spinal cord compression and hypercalcemia. Given the high prevalence of patients with breast cancer, health-care costs of bone metastases (>$20,000 per episode) impose a tremendous economic burden on society. To investigate mechanisms of bone metastasis, we developed femoral artery injection of cancer cells as a physiologically relevant model of bone metastasis. Comparing young (~6 weeks), skeletally immature mice to old (~6 months) female mice with closed physes (growth plates), we showed significantly greater progression of osteolytic metastases in young animals. Bone destruction increased in the old mice following ovariectomy, emphasizing the pathologic consequences of greater bone turnover and net loss. Despite uniform initial distribution of breast cancer cells throughout the hind limb after femoral artery injection, we observed preferential formation of osteolytic bone metastases in the proximal tibia. Tropism for the proximal tibia arises in part because of TGF-β, a cytokine abundant in both physes of skeletally immature mice and matrix of bone in mice of all ages. We also showed that age-dependent effects on osteolytic bone metastases did not occur in male mice with disseminated breast cancer cells in bone. These studies establish a model system to specifically focus on pathophysiology and treatment of bone metastases and underscore the need to match biologic variables in the model to relevant subsets of patients with breast cancer.

Mutations in the epidermal growth factor receptor (EGFR) have been implicated in nearly one-third of nonsmall-cell lung cancers. For patients harboring non-traditional mutations, genomic and transcriptomic sequencing can help direct treatment. As cancer genomics evolves, novel driver mutations continue to be uncovered. We report on a unique EGFR-GRB2 fusion in a 48-year-old female never-smoker. This patient presented with stage IV lung adenocarcinoma (T2aN3M1) with metastatic disease in the iliac wing and liver. Despite systemic treatment, this patient continued to progress. On whole transcriptome sequencing, this patient was found to have a novel EGFR-GRB2 RNA fusion transcript similar to other EGFR fusions described in the literature. After treatment with osimertinib, this patient experienced remarkable clinical and radiological improvements. We believe that, especially for patients with metastatic lung cancer, the presence of novel driver mutations should be investigated. Potentially, patients harboring similar mutations may demonstrate analogous improvements with targeted treatment using the most recent generation of tyrosine kinase inhibitors.

70 Background: Loeb et al. recently highlighted that academic physicians were statistically more likely to recommend genetic testing to prostate cancer patients per NCCN guidelines compared to physicians at community practices. Several articles have outlined the rate of pathogenic mutations and metastasis free survival (MFS) for prostate cancer patients treated at academic institutions. However, there is a relative paucity of equivalent data regarding patients treated at community urology clinics. We felt it was important to retrospectively review data from our large community based uro-oncology practice and present our findings in an effort to clarify this topic. Methods: We collected data on 562 prostate cancer patients treated at our multidisciplinary uro-oncology clinic between 2016 and 2018. We found 363 patients that satisfied the inclusion criteria of having germline genetic mutation testing and at least 1 year of follow-up. Patients were stratified into three categories based on the results of their germline genetic test: negative for germline mutations, positive for germline pathogenic mutations, or positive for VUS (variant of uncertain significance) mutations. Analysis of variances (ANOVA) was conducted to assess for any differences in age, Gleason score, metastasis rate, and MFS across the groups. A significance level of.05 was used. Results: All patients were treated according to guideline recommendations as was standard for the practice. There was no statistically significant difference in average age or Gleason score between the groups. There was also no statistically significant difference between the MFS across the three groups. Although the metastasis rate and MFS in the group without any mutations (9.8%, 18.3 months) was clinically significant compared to the groups with pathogenic mutations (7.5%, 16.5 months) and VUS mutations (7%, 16.6 months), the differences were not statistically significant (p=.754 and.127 respectively). Conclusions: In our community based uro-oncology practice, we found no statistically significant difference in MFS between patients with pathogenic germline mutations, patients without germline mutations, and patients with VUS mutations at 1 year of follow-up. This does not exclude the possibility of an impact of germline mutations on MFS as the data matures to reach 5 or more years of follow-up.[Table: see text]