Michael L. Gatza

Parkin targets HIF-1α for ubiquitination and degradation to inhibit breast tumor progression

Mutations in E3 ubiquitin ligase Parkin have been linked to familial Parkinson’s disease. Accumulating evidence suggests that Parkin is a tumor suppressor, but the underlying mechanism is poorly understood. Here we show that Parkin is an E3 ubiquitin ligase for hypoxia-inducible factor 1α (HIF-1α). Parkin interacts with HIF-1α and promotes HIF-1α degradation through ubiquitination, which in turn inhibits metastasis of breast cancer cells. Parkin downregulation in breast cancer cells promotes metastasis, which can be inhibited by targeting HIF-1α with RNA interference or the small-molecule inhibitor YC-1. We further identify lysine 477 (K477) of HIF-1α as a major ubiquitination site for Parkin. K477R HIF-1α mutation and specific cancer-associated Parkin mutations largely abolish the functions of Parkin to ubiquitinate HIF-1α and inhibit cancer metastasis. Importantly, Parkin expression is inversely correlated with HIF-1α expression and metastasis in breast cancer. Our results reveal an important mechanism for Parkin in tumor suppression and HIF-1α regulation.Parkin is an E3 ubiquitin ligase involved in Parkinson’s disease. Parkin has also been linked to cancer suppression but the mechanisms are unclear. Here the authors show that Parkin regulates HIF-1α through ubiquitin-dependent degradation, thus inhibiting metastasis of breast cancer cells.

New mechanisms of resistance to MEK inhibitors in melanoma revealed by intravital imaging

Targeted therapeutics that are initially effective in cancer patients nearly invariably engender resistance at some stage, an inherent challenge in the use of any molecular-targeted drug in cancer settings. In this study, we evaluated resistance mechanisms arising in metastatic melanoma to MAPK pathway kinase inhibitors as a strategy to identify candidate strategies to limit risks of resistance. To investigate longitudinal responses, we developed an intravital serial imaging approach that can directly visualize drug response in an inducible RAF-driven, autochthonous murine model of melanoma incorporating a fluorescent reporter allele (tdTomatoLSL). Using this system, we visualized formation and progression of tumors in situ, starting from the single-cell level longitudinally over time. Reliable reporting of the status of primary murine tumors treated with the selective MEK1/2 inhibitor (MEKi) trametinib illustrated a time-course of initial drug response and persistence, followed by the development of drug resistance. We found that tumor cells adjacent to bundled collagen had a preferential persistence in response to MEKi. Unbiased transcriptional and kinome reprogramming analyses from selected treatment time points suggested increased c-Kit and PI3K/AKT pathway activation in resistant tumors, along with enhanced expression of epithelial genes and epithelial-mesenchymal transition downregulation signatures with development of MEKi resistance. Similar trends were observed following simultaneous treatment with BRAF and MEK inhibitors aligned to standard-of-care combination therapy, suggesting these reprogramming events were not specific to MEKi alone. Overall, our results illuminate the integration of tumor-stroma dynamics with tissue plasticity in melanoma progression and provide new insights into the basis for drug response, persistence, and resistance.Significance: A longitudinal study tracks the course of MEKi treatment in an autochthonous imageable murine model of melanoma from initial response to therapeutic resistance, offering new insights into the basis for drug response, persistence, and resistance. Cancer Res; 78(2); 542-57. ©2017 AACR.

Another Breast Cancer Entity Confirmed: Genomics of Invasive Lobular Breast Cancer

The most frequent signi ficantly altered gene identified in this study was CDH1 in which loss-of-function mutations occurred in 65% of cases and loss of heterozygosity or deletion was identified in 94% of patients. These findings are consistent with other studies and underscore the role of E-cadherin loss in contributing to the characteristic discohesive morphology associated with ILC. 3

Mediator kinase CDK8/CDK19 drives YAP1-dependent BMP4-induced EMT in cancer

CDK8 is a transcription-regulating kinase that controls TGF-β/BMP-responsive SMAD transcriptional activation and turnover through YAP1 recruitment. However, how the CDK8/YAP1 pathway influences SMAD1 response in cancer remains unclear. Here we report that SMAD1-driven epithelial-to-mesenchymal transition (EMT) is critically dependent on matrix rigidity and YAP1 in a wide spectrum of cancer models. We find that both genetic and pharmacological inhibition of CDK8 and its homologous twin kinase CDK19 leads to abrogation of BMP-induced EMT. Notably, selectively blocking CDK8/19 specifically abrogates tumor cell invasion, changes in EMT-associated transcription factors, E-cadherin expression and YAP nuclear localization both in vitro and in vivo in a murine syngeneic EMT model. Furthermore, RNA-seq meta-analysis reveals a direct correlation between CDK8 and EMT-associated transcription factors in patients. Our findings demonstrate that CDK8, an emerging therapeutic target, coordinates growth factor and mechanical cues during EMT and invasion.

Tumor mutational burden is a determinant of immune-mediated survival in breast cancer

ABSTRACT Mounting evidence supports a role for the immune system in breast cancer outcomes. The ability to distinguish highly immunogenic tumors susceptible to anti-tumor immunity from weakly immunogenic or inherently immune-resistant tumors would guide development of therapeutic strategies in breast cancer. Genomic, transcriptomic and clinical data from The Cancer Genome Atlas (TCGA) and Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) breast cancer cohorts were used to examine statistical associations between tumor mutational burden (TMB) and the survival of patients whose tumors were assigned to previously-described prognostic immune subclasses reflecting favorable, weak or poor immune-infiltrate dispositions (FID, WID or PID, respectively). Tumor immune subclasses were associated with survival in patients with high TMB (TMB-Hi, P < 0.001) but not in those with low TMB (TMB-Lo, P = 0.44). This statistical relationship was confirmed in the METABRIC cohort (TMB-Hi, P = 0.047; TMB-Lo, P = 0.39), and also found to hold true in the more-indolent Luminal A tumor subtype (TMB-Hi, P = 0.011; TMB-Lo, P = 0.91). In TMB-Hi tumors, the FID subclass was associated with prolonged survival independent of tumor stage, molecular subtype, age and treatment. Copy number analysis revealed the reproducible, preferential amplification of chromosome 1q immune-regulatory genes in the PID immune subclass. These findings demonstrate a previously unappreciated role for TMB as a determinant of immune-mediated survival of breast cancer patients and identify candidate immune-regulatory mechanisms associated with immunologically cold tumors. Immune subtyping of breast cancers may offer opportunities for therapeutic stratification.

Differentiation among prostate cancer patients with Gleason score of 7 using histopathology image and genomic data

Prostate cancer is the most common non-skin related cancer affecting 1 in 7 men in the United States. Treatment of patients with prostate cancer still remains a difficult decision-making process that requires physicians to balance clinical benefits, life expectancy, comorbidities, and treatment-related side effects. Gleason score (a sum of the primary and secondary Gleason patterns) solely based on morphological prostate glandular architecture has shown as one of the best predictors of prostate cancer outcome. Significant progress has been made on molecular subtyping prostate cancer delineated through the increasing use of gene sequencing. Prostate cancer patients with Gleason score of 7 show heterogeneity in recurrence and survival outcomes. Therefore, we propose to assess the correlation between histopathology images and genomic data with disease recurrence in prostate tumors with a Gleason 7 score to identify prognostic markers. In the study, we identify image biomarkers within tissue WSIs by modeling the spatial relationship from automatically created patches as a sequence within WSI by adopting a recurrence network model, namely long short-term memory (LSTM). Our preliminary results demonstrate that integrating image biomarkers from CNN with LSTM and genomic pathway scores, is more strongly correlated with patients recurrence of disease compared to standard clinical markers and engineered image texture features. The study further demonstrates that prostate cancer patients with Gleason score of 4+3 have a higher risk of disease progression and recurrence compared to prostate cancer patients with Gleason score of 3+4.

SMAD4 Suppresses WNT-Driven Dedifferentiation and Oncogenesis in the Differentiated Gut Epithelium

The cell of origin of colon cancer is typically thought to be the resident somatic stem cells, which are immortal and escape the continual cellular turnover characteristic of the intestinal epithelium. However, recent studies have identified certain conditions in which differentiated cells can acquire stem-like properties and give rise to tumors. Defining the origins of tumors will inform cancer prevention efforts as well as cancer therapies, as cancers with distinct origins often respond differently to treatments. We report here a new condition in which tumors arise from the differentiated intestinal epithelium. Inactivation of the differentiation-promoting transcription factor SMAD4 in the intestinal epithelium was surprisingly well tolerated in the short term. However, after several months, adenomas developed with characteristics of activated WNT signaling. Simultaneous loss of SMAD4 and activation of the WNT pathway led to dedifferentiation and rapid adenoma formation in differentiated tissue. Transcriptional profiling revealed acquisition of stem cell characteristics, and colabeling indicated that cells expressing differentiated enterocyte markers entered the cell cycle and reexpressed stem cell genes upon simultaneous loss of SMAD4 and activation of the WNT pathway. These results indicate that SMAD4 functions to maintain differentiated enterocytes in the presence of oncogenic WNT signaling, thus preventing dedifferentiation and tumor formation in the differentiated intestinal epithelium.Significance: This work identifies a mechanism through which differentiated cells prevent tumor formation by suppressing oncogenic plasticity. Cancer Res; 78(17); 4878-90. ©2018 AACR.

Amplification of SOX4 promotes PI3K/Akt signaling in human breast cancer

PurposeThe PI3K/Akt signaling axis contributes to the dysregulation of many dominant features in breast cancer including cell proliferation, survival, metabolism, motility, and genomic instability. While multiple studies have demonstrated that basal-like or triple-negative breast tumors have uniformly high PI3K/Akt activity, genomic alterations that mediate dysregulation of this pathway in this subset of highly aggressive breast tumors remain to be determined.MethodsIn this study, we present an integrated genomic analysis based on the use of a PI3K gene expression signature as a framework to analyze orthogonal genomic data from human breast tumors, including RNA expression, DNA copy number alterations, and protein expression. In combination with data from a genome-wide RNA-mediated interference screen in human breast cancer cell lines, we identified essential genetic drivers of PI3K/Akt signaling.ResultsOur in silico analyses identified SOX4 amplification as a novel modulator of PI3K/Akt signaling in breast cancers and in vitro studies confirmed its role in regulating Akt phosphorylation.ConclusionsTaken together, these data establish a role for SOX4-mediated PI3K/Akt signaling in breast cancer and suggest that SOX4 may represent a novel therapeutic target and/or biomarker for current PI3K family therapies.