Robert L. Dood

Platelets reduce anoikis and promote metastasis by activating YAP1 signaling

Thrombocytosis is present in more than 30% of patients with solid malignancies and correlates with worsened patient survival. Tumor cell interaction with various cellular components of the tumor microenvironment including platelets is crucial for tumor growth and metastasis. Although it is known that platelets can infiltrate into tumor tissue, secrete pro-angiogenic and pro-tumorigenic factors and thereby increase tumor growth, the precise molecular interactions between platelets and metastatic cancer cells are not well understood. Here we demonstrate that platelets induce resistance to anoikis in vitro and are critical for metastasis in vivo. We further show that platelets activate RhoA-MYPT1-PP1-mediated YAP1 dephosphorylation and promote its nuclear translocation which induces a pro-survival gene expression signature and inhibits apoptosis. Reduction of YAP1 in cancer cells in vivo protects against thrombocytosis-induced increase in metastasis. Collectively, our results indicate that cancer cells depend on platelets to avoid anoikis and succeed in the metastatic process.Platelets have been associated with increased tumor growth and metastasis but the mechanistic details of this interaction are still unclear. Here the authors show that platelets improve anoikis resistance of cancer cells and increase metastasis by activating Yap through a RhoA/MYPT-PP1 pathway.

Metabolic Markers and Statistical Prediction of Serous Ovarian Cancer Aggressiveness by Ambient Ionization Mass Spectrometry Imaging

Ovarian high-grade serous carcinoma (HGSC) results in the highest mortality among gynecological cancers, developing rapidly and aggressively. Dissimilarly, serous borderline ovarian tumors (BOT) can progress into low-grade serous carcinomas and have relatively indolent clinical behavior. The underlying biological differences between HGSC and BOT call for accurate diagnostic methodologies and tailored treatment options, and identification of molecular markers of aggressiveness could provide valuable biochemical insights and improve disease management. Here, we used desorption electrospray ionization (DESI) mass spectrometry (MS) to image and chemically characterize the metabolic profiles of HGSC, BOT, and normal ovarian tissue samples. DESI-MS imaging enabled clear visualization of fine papillary branches in serous BOT and allowed for characterization of spatial features of tumor heterogeneity such as adjacent necrosis and stroma in HGSC. Predictive markers of cancer aggressiveness were identified, including various free fatty acids, metabolites, and complex lipids such as ceramides, glycerophosphoglycerols, cardiolipins, and glycerophosphocholines. Classification models built from a total of 89,826 individual pixels, acquired in positive and negative ion modes from 78 different tissue samples, enabled diagnosis and prediction of HGSC and all tumor samples in comparison with normal tissues, with overall agreements of 96.4% and 96.2%, respectively. HGSC and BOT discrimination was achieved with an overall accuracy of 93.0%. Interestingly, our classification model allowed identification of three BOT samples presenting unusual histologic features that could be associated with the development of low-grade carcinomas. Our results suggest DESI-MS as a powerful approach for rapid serous ovarian cancer diagnosis based on altered metabolic signatures. Cancer Res; 77(11); 2903-13. ©2017 AACR.

Sustained Adrenergic Signaling Promotes Intratumoral Innervation through BDNF Induction

Mounting clinical and preclinical evidence supports a key role for sustained adrenergic signaling in the tumor microenvironment as a driver of tumor growth and progression. However, the mechanisms by which adrenergic neurotransmitters are delivered to the tumor microenvironment are not well understood. Here we present evidence for a feed-forward loop whereby adrenergic signaling leads to increased tumoral innervation. In response to catecholamines, tumor cells produced brain-derived neurotrophic factor (BDNF) in an ADRB3/cAMP/Epac/JNK-dependent manner. Elevated BDNF levels in the tumor microenvironment increased innervation by signaling through host neurotrophic receptor tyrosine kinase 2 receptors. In patients with cancer, high tumor nerve counts were significantly associated with increased BDNF and norepinephrine levels and decreased overall survival. Collectively, these data describe a novel pathway for tumor innervation, with resultant biological and clinical implications.Significance: Sustained adrenergic signaling promotes tumor growth and metastasis through BDNF-mediated tumoral innervation. Cancer Res; 78(12); 3233-42. ©2018 AACR.

Macrophage depletion through colony stimulating factor 1 receptor pathway blockade overcomes adaptive resistance to anti-VEGF therapy

Anti-angiogenesis therapy has shown clinical benefit in patients with high-grade serous ovarian cancer (HGSC), but adaptive resistance rapidly emerges. Thus, approaches to overcome such resistance are needed. We developed the setting of adaptive resistance to anti-VEGF therapy, and performed a series of in vivo experiments in both immune competent and nude mouse models. Given the pro-angiogenic properties of tumor-associated macrophages (TAMs) and the dominant role of CSF1R in macrophage function, we added CSF1R inhibitors following emergence of adaptive resistance to anti-VEGF antibody. Mice treated with a CSF1R inhibitor (AC708) after anti-VEGF antibody resistance had little to no measurable tumor burden upon completion of the experiment while those that did not receive a CSF1R inhibitor still had abundant tumor. To mimic clinically used regimens, mice were also treated with anti-VEGF antibody and paclitaxel until resistance emerged, and then a CSF1R inhibitor was added. The addition of a CSF1R inhibitor restored response to anti-angiogenesis therapy, resulting in 83% lower tumor burden compared to treatment with anti-VEGF antibody and paclitaxel alone. Collectively, our data demonstrate that the addition of a CSF1R inhibitor to anti-VEGF therapy and taxane chemotherapy results in robust anti-tumor effects.

Adrenergic-mediated increases in INHBA drive CAF phenotype and collagens

Adrenergic signaling is known to promote tumor growth and metastasis, but the effects on tumor stroma are not well understood. An unbiased bioinformatics approach analyzing tumor samples from patients with known biobehavioral profiles identified a prominent stromal signature associated with cancer-associated fibroblasts (CAFs) in those with a high biobehavioral risk profile (high Center for Epidemiologic Studies Depression Scale [CES-D] score and low social support). In several models of epithelial ovarian cancer, daily restraint stress resulted in significantly increased CAF activation and was abrogated by a nonspecific β-blocker. Adrenergic signaling-induced CAFs had significantly higher levels of collagen and extracellular matrix components than control tumors. Using a systems-based approach, we found INHBA production by cancer cells to induce CAFs. Ablating inhibin β A decreased CAF phenotype both in vitro and in vivo. In preclinical models of breast and colon cancers, there were increased CAFs and collagens following daily restraint stress. In an independent data set of renal cell carcinoma patients, there was an association between high depression (CES-D) scores and elevated expression of ACTA2, collagens, and inhibin β A. Collectively, our findings implicate adrenergic influences on tumor stroma as important drivers of CAFs and establish inhibin β A as an important regulator of the CAF phenotype in ovarian cancer.

Defining Survivorship Trajectories Across Patients With Solid Tumors: An Evidence-Based Approach

Importance Survivorship involves a multidisciplinary approach to surveillance and management of comorbidities and secondary cancers, overseen by oncologists, surgeons, and primary care physicians. Optimal timing and coordination of care, however, is unclear and often based on arbitrary 5-year cutoffs. Objective To determine high- and low-risk periods for all tumor types that could define when survivorship care might best be overseen by oncologists and when to transition to primary care physicians. Design, Setting, and Participants In this pan-cancer, longitudinal, observational study, excess mortality hazard, calculated as an annualized mortality risk above a baseline population, was plotted over time. The time this hazard took to stabilize defined a high-risk period. The percent morality elevation above age- and sex-matched controls in the latter low-risk period was reported as a mortality gap. The US population–based Surveillance, Epidemiology, and End Results database defined the cancer population, and the US Census life tables defined controls. Incident cases of patients with cancer were separated into tumor types based on International Classification of Diseases for Oncology definitions. Exposures Population-level data on incident cancer cases was compared with the general US population. Main Outcomes and Measures Overall mortality and cause of death were reported on observed cancer cases. Results A total of 2 317 185 patients (median age, 63 years; 49.8% female) with 66 primary tumor types were evaluated. High-risk surveillance period durations ranged from less than 1 year (breast, prostate, lip, ocular, and parathyroid cancers) up to 19 years (unspecified gastrointestinal cancers). The annualized mortality gap, representing the excess mortality in the stable period, ranged from a median 0.26% to 9.33% excess annual mortality (thyroid and hypopharyngeal cancer populations, respectively). Cluster analysis produced 6 risk cluster groups: group 1, with median survival of 16.2 (5th to 95th percentile range [PR], 10.7-40.2) years and median high-risk period of 2.5 (PR, 0-5.0) years; group 2, 8.3 (PR, 5.1-23.3) and 2.5 (PR, 4.0-8.0) years; group 3, 2.8 (PR, 1.4-3.7) and 7.0 (PR, 6.0-11.1) years; group 4, 1.6 (PR, 1.5-1.8) and 6.0 (PR, 5.1-11.4) years; group 5, 0.8 (PR, 0.5-1.2) and 0.8 (PR, 0.5-1.2) years; and group 6, 0.5 (PR, 0.4-0.8) and 12.0 (PR, 9.3-12.9) years, respectively. Subanalyses of selected tumor types in these groups revealed that stratifying on stage and histologic type can change the risk cluster and guidance for care. Conclusions and Relevance These findings indicate that a standardized 5-year surveillance period is inadequate for some cancers while excessive for others. High-risk cancers require the most resources with the longest high-risk period, highest persistent baseline mortality risk, and longest period of primary cancer mortality, all arguing for longer follow-up with an oncologist in these cancers.

Inhibiting nuclear phospho-progesterone receptor enhances antitumor activity of onapristone in uterine cancer

Although progesterone receptor (PR)–targeted therapies are modestly active in patients with uterine cancer, their underlying molecular mechanisms are not well understood. The clinical use of such therapies is limited because of the lack of biomarkers that predict response to PR agonists (progestins) or PR antagonists (onapristone). Thus, understanding the underlying molecular mechanisms of action will provide an advance in developing novel combination therapies for cancer patients. Nuclear translocation of PR has been reported to be ligand-dependent or -independent. Here, we identified that onapristone, a PR antagonist, inhibited nuclear translocation of ligand-dependent or -independent (EGF) phospho-PR (S294), whereas trametinib inhibited nuclear translocation of EGF-induced phospho-PR (S294). Using orthotopic mouse models of uterine cancer, we demonstrated that the combination of onapristone and trametinib results in superior antitumor effects in uterine cancer models compared with either monotherapy. These synergistic effects are, in part, mediated through inhibiting the nuclear translocation of EGF-induced PR phosphorylation in uterine cancer cells. Targeting MAPK-dependent PR activation with onapristone and trametinib significantly inhibited tumor growth in preclinical uterine cancer models and is worthy of further clinical investigation. Mol Cancer Ther; 17(2); 464–73. ©2017 AACR.

Abstract 788: Less is more: macrophage depletion via CSF1/CSF1R pathway improves anti-VEGF therapy after adaptive resistance

Objectives: Anti-angiogenesis therapy shows clinical benefit in patients with high-grade serous ovarian cancer (HGSC), but adaptive resistance typically occurs. Thus, approaches to overcome such resistance are needed. Here, we systematically assessed immune cell populations enriched during adaptive resistance and identify novel therapeutic avenues. Methods: A series of in vitro and in vivo (immune competent and nude mice) experiments were carried out. Animals were treated with anti-VEGF antibody continuously until resistance emerged, at which point full immune profiling was performed. Based on these results, efficacy of AC708 (CSF1R inhibitor to target tumor-associated macrophages) was tested in the adaptive-resistance models. F4/80 antibody was used as a macrophage marker. Results: On the basis of full immune profiling, we detected significantly increased macrophage infiltration in tumors with anti-VEGF antibody resistance compared to tumors from sensitive mice (p Conclusions: The addition of CSf1R inhibitor to anti-VEGF therapy and taxane chemotherapy results in robust anti-tumor effects. To confirm these findings, a clinical trial at our institution is nearing activation. The REDIRECT (RandomizEd Induction DIscontinuation TRial of EmaCTuzumab) trial will randomize patients to continue weekly paclitaxel and biweekly bevacizumab with or without a CSF1R inhibitor (emactuzumab) following an induction phase. Citation Format: Yasmin A. Lyons, Sunila Pradeep, Jean M. Hansen, Michael J. Wagner, Robert L. Dood, Sherry Y. Wu, Rebecca A. Previs, Wei Hu, Robert L. Coleman, Anil K. Sood. Less is more: macrophage depletion via CSF1/CSF1R pathway improves anti-VEGF therapy after adaptive resistance [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 788. doi:10.1158/1538-7445.AM2017-788