Functional malignant cell heterogeneity in pancreatic neuroendocrine tumors revealed by targeting of PDGF-DD.

Intratumoral heterogeneity is an inherent feature of most human cancers and has profound implications for cancer therapy. As a result, there is an emergent need to explore previously unmapped mechanisms regulating distinct subpopulations of tumor cells and to understand their contribution to tumor progression and treatment response. Aberrant platelet-derived growth factor receptor beta (PDGFRß) signaling in cancer has motivated the development of several antagonists currently in clinical use, including imatinib, sunitinib, and sorafenib. The discovery of a novel ligand for PDGFRß, platelet-derived growth factor (PDGF)-DD, opened the possibility of a previously unidentified signaling pathway involved in tumor development. However, the precise function of PDGF-DD in tumor growth and invasion remains elusive. Here, making use of a newly generated Pdgfd knockout mouse, we reveal a functionally important malignant cell heterogeneity modulated by PDGF-DD signaling in pancreatic neuroendocrine tumors (PanNET). Our analyses demonstrate that tumor growth was delayed in the absence of signaling by PDGF-DD. Surprisingly, ablation of PDGF-DD did not affect the vasculature or stroma of PanNET; instead, we found that PDGF-DD stimulated bulk tumor cell proliferation by induction of paracrine mitogenic signaling between heterogeneous malignant cell clones, some of which expressed PDGFRß. The presence of a subclonal population of tumor cells characterized by PDGFRß expression was further validated in a cohort of human PanNET. In conclusion, we demonstrate a previously unrecognized heterogeneity in PanNET characterized by signaling through the PDGF-DD/PDGFRß axis.

Functional subsets of mesenchymal cell types in the tumor microenvironment.

In the field of tumor biology, increasing attention is now focused on the complex interactions between various constituent cell types within the tumor microenvironment as being functionally important for the etiology of the disease. The detailed description of tumor-promoting properties of cancer-associated fibroblasts, endothelial cells, pericytes, and immune cells, introduces novel potential drug targets for improved cancer treatments, as well as a rationale for exploring the tumor stroma as a previously unchartered source for prognostic or predictive biomarkers. However, recent work highlights the fact that cellular identity is perhaps too broadly defined and that subdivision of each cell type may reveal functionally distinct subsets of cells. Here, we will review our current understanding of the diversity of different subsets of mesenchymal cells, i.e., cancer-associated fibroblasts and pericytes, residing within the tumor parenchyma.

Pericytes promote endothelial cell survival through induction of autocrine VEGF-A signaling and Bcl-w expression.

Endothelial cells (ECs) in blood vessels under formation are stabilized by the recruitment of pericytes, both in normal tissues and during angiogenesis in pathologic situations, including neoplasia. In the tumor vasculature, besides supporting the functionality of blood flow, pericytes protect ECs from antiangiogenic therapies, and have thus been implicated in clinical resistance to vascular targeting drugs. However, the molecular nature of the crosstalk between pericytes and ECs is largely unchartered. Herein, we identified pericyte-induced survival signals in ECs by isolation of vascular fragments derived from tumors that had been genetically or pharmacologically engineered to be either pericyte-rich or pericyte-poor. Pericytes induced the antiapoptotic protein Bcl-w in tumor ECs both in vivo and in vitro, thereby conveying protection from cytotoxic damage. The pericyte-dependent survival signaling in ECs was consequential to enforcement of an autocrine loop involving VEGF-A expression in ECs. Through molecular and functional studies, we delineated a signal transduction pathway in ECs downstream of integrin α(v) involving activation of NF-κB as the initiating event of the protective crosstalk from pericytes. Our elucidation of pericyte-derived pro-survival signaling in tumor ECs has potentially important implications for clinical development of antiangiogenic drugs, and suggests new therapeutic targets for rational multitargeting of cancer.

A landscape of response to drug combinations in non-small cell lung cancer.

Combination of anti-cancer drugs is broadly seen as way to overcome the often-limited efficacy of single agents. The design and testing of combinations are however very challenging. Here we present a uniquely large dataset screening over 5000 targeted agent combinations across 81 non-small cell lung cancer cell lines. Our analysis reveals a profound heterogeneity of response across the tumor models. Notably, combinations very rarely result in a strong gain in efficacy over the range of response observable with single agents. Importantly, gain of activity over single agents is more often seen when co-targeting functionally proximal genes, offering a strategy for designing more efficient combinations. Because combinatorial effect is strongly context specific, tumor specificity should be achievable. The resource provided, together with an additional validation screen sheds light on major challenges and opportunities in building efficacious combinations against cancer and provides an opportunity for training computational models for synergy prediction.

A Case Series of Metastatic Malignant Gastrointestinal Neuroectodermal Tumors and Comprehensive Genomic Profiling Analysis of 20 Cases.

Malignant gastrointestinal neuroectodermal tumor (GNET) is an ultra-rare soft tissue sarcoma, therefore often misdiagnosed and has no available standard treatment. Here, we report 3 cases of metastatic GNET with variable clinical courses. Our small case series as well as extensive literature review, further support that GNET is a spectrum of diseases with variable inherent biology and prognosis. Surgical management in the setting of recurrent/metastatic disease may be appropriate for GNET with indolent nature. Response to systemic treatments including chemotherapy and targeted treatments is variable, likely related to heterogenous biology as well. Furthermore, we retrospectively identified 20 additional GNET cases from Foundation Medicine's genomic database and expanded on their clinicopathological and genomic features. Comprehensive genomic profiling (CGP) with DNA and RNA sequencing of this cohort, in the course of clinical care, demonstrated recurrent EWSR1 chromosomal rearrangements and a sparsity of additional recurrent or driver genomic alterations. All cases had low tumor mutational burden (TMB) and were microsatellite stable.

KRAS G12C NSCLC Models Are Sensitive to Direct Targeting of KRAS in Combination with PI3K Inhibition.

PURPOSE: KRAS-mutant lung cancers have been recalcitrant to treatments including those targeting the MAPK pathway. Covalent inhibitors of KRAS p.G12C allele allow for direct and specific inhibition of mutant KRAS in cancer cells. However, as for other targeted therapies, the therapeutic potential of these inhibitors can be impaired by intrinsic resistance mechanisms. Therefore, combination strategies are likely needed to improve efficacy.Experimental Design: To identify strategies to maximally leverage direct KRAS inhibition we defined the response of a panel of NSCLC models bearing the KRAS G12C-activating mutation in vitro and in vivo. We used a second-generation KRAS G12C inhibitor, ARS1620 with improved bioavailability over the first generation. We analyzed KRAS downstream effectors signaling to identify mechanisms underlying differential response. To identify candidate combination strategies, we performed a high-throughput drug screening across 112 drugs in combination with ARS1620. We validated the top hits in vitro and in vivo including patient-derived xenograft models. RESULTS: Response to direct KRAS G12C inhibition was heterogeneous across models. Adaptive resistance mechanisms involving reactivation of MAPK pathway and failure to induce PI3K-AKT pathway inactivation were identified as likely resistance events. We identified several model-specific effective combinations as well as a broad-sensitizing effect of PI3K-AKT-mTOR pathway inhibitors. The G12Ci+PI3Ki combination was effective in vitro and in vivo on models resistant to single-agent ARS1620 including patient-derived xenografts models. CONCLUSIONS: Our findings suggest that signaling adaptation can in some instances limit the efficacy of ARS1620 but combination with PI3K inhibitors can overcome this resistance.

Participação plurissocial para aumentar a inclusão e a assertividade nas tomadas de decisão em saúde relacionadas a tratamentos da osteoporose / Pluri-social participation to increase inclusion and assertiveness in health decision-making related to osteoporosis treatments

Embora as fraturas por fragilidade sejam importantes detratoras de qualidade de vida relacionada à saúde, aumentando a morbimortalidade e acarretando alto impacto clínico, psicossocial e econômico, elas são pouco valorizadas e negligenciadas por médicos e até mesmo por pacientes. Além disso, os critérios de priorização para avaliação de novas tecnologias, em geral, não consideram critérios além dos financeiros para uma tomada de decisão mais inclusiva e assertiva para o tratamento da população de mais alto risco de fratura. Assim, este artigo visa revisitar alguns diferentes pontos de vista e trazer uma reflexão sobre critérios e prioridades na osteoporose. Para isso, foi considerada a perspectiva de múltiplos atores no processo de tomada de decisão em saúde, bem como analisadas as falhas na atenção a uma doença de alta prevalência e que, além do grande impacto econômico gerado para a sociedade, causa repercussões emocionais, incapacidade gerada por fraturas e medo de novas quedas ou pequenos traumas.

Although fragility fractures are important detractors of health-related quality of life, increasing morbidity and mortality and causing a high clinical, psychosocial, and economic impact, they are undervalued and neglected by physicians and even patients. In addition, prioritization criteria for evaluating new technologies, in general, do not consider criteria other than financial ones for a more inclusive and assertive decision-making for the treatment of the population at higher risk of fracture. Thus, this article aims to revisit some different points of view and bring a reflection on criteria and priorities in osteoporosis. For this, the perspective of multiple stakeholders in the health decision-making process was considered, as well as the failures in the care of this highly prevalent disease that, in addition to the great economic impact generated for society, causes emotional repercussions, disability generated by fractures and fear of further falls or minor trauma.

Microenvironmental control of breast cancer subtype elicited through paracrine platelet-derived growth factor-CC signaling.

Breast tumors of the basal-like, hormone receptor-negative subtype remain an unmet clinical challenge, as there is high rate of recurrence and poor survival in patients following treatment. Coevolution of the malignant mammary epithelium and its underlying stroma instigates cancer-associated fibroblasts (CAFs) to support most, if not all, hallmarks of cancer progression. Here we delineate a previously unappreciated role for CAFs as determinants of the molecular subtype of breast cancer. We identified paracrine crosstalk between cancer cells expressing platelet-derived growth factor (PDGF)-CC and CAFs expressing the cognate receptors in human basal-like mammary carcinomas. Genetic or pharmacological intervention of PDGF-CC activity in mouse models of cancer resulted in conversion of basal-like breast cancers into a hormone receptor-positive state that enhanced sensitivity to endocrine therapy in previously resistant tumors. We conclude that specification of breast cancer to the basal-like subtype is under microenvironmental control and is therapeutically actionable.

Differential Effector Engagement by Oncogenic KRAS.

KRAS can bind numerous effector proteins, which activate different downstream signaling events. The best known are RAF, phosphatidylinositide (PI)-3' kinase, and RalGDS families, but many additional direct and indirect effectors have been reported. We have assessed how these effectors contribute to several major phenotypes in a quantitative way, using an arrayed combinatorial siRNA screen in which we knocked down 41 KRAS effectors nodes in 92 cell lines. We show that every cell line has a unique combination of effector dependencies, but in spite of this heterogeneity, we were able to identify two major subtypes of KRAS mutant cancers of the lung, pancreas, and large intestine, which reflect different KRAS effector engagement and opportunities for therapeutic intervention.

Compound genetically engineered mouse models of cancer reveal dual targeting of ALK1 and endoglin as a synergistic opportunity to impinge on angiogenic TGF-ß signaling.

Angiogenesis occurs early in tumor development, sustains primary tumor growth and provides a route for metastatic escape. The TGF-ß family receptors modulate angiogenesis via endothelial-cell specific pathways. Here we investigate the interaction of two such receptors, ALK1 and endoglin, in pancreatic neuroendocrine tumors (PanNET). Independently, ALK1 and endoglin deficiencies exhibited genetically divergent phenotypes, while both highly correlate to an endothelial metagene in human and mouse PanNETs. A concurrent deficiency of both receptors synergistically decreased tumor burden to a greater extent than either individual knockdown. Furthermore, the knockout of Gdf2 (BMP9), the primary ligand for ALK1 and endoglin, exhibited a mixed phenotype from each of ALK1 and endoglin deficiencies; overall primary tumor burden decreased, but hepatic metastases increased. Tumors lacking BMP9 display a hyperbranching vasculature, and an increase in vascular mesenchymal-marker expression, which may be implicit in the increase in metastases. Taken together, our work cautions against singular blockade of BMP9 and instead demonstrates the utility of dual blockade of ALK1 and endoglin as a strategy for anti-angiogenic therapy in PanNET.

Image-based 3D modeling study of the influence of vessel density and blood hemoglobin concentration on tumor oxygenation and response to irradiation.

PURPOSE: Hypoxia is one of the most important factors influencing clinical outcome after radiotherapy. Improved knowledge of factors affecting the levels and distribution of oxygen within a tumor is needed. The authors constructed a theoretical 3D model based on histological images to analyze the influence of vessel density and hemoglobin (Hb) concentration on the response to irradiation. METHODS: The pancreases of a Rip-Tag2 mouse, a model of malignant insulinoma, were excised, cryosectioned, immunostained, and photographed. Vessels were identified by image thresholding and a 3D vessel matrix assembled. The matrix was reduced to functional vessel segments and enlarged by replication. The steady-state oxygen tension field of the tumor was calculated by iteratively employing Green's function method for diffusion and the Michaelis-Menten model for consumption. The impact of vessel density on the radiation response was studied by removing a number of randomly selected vessels. The impact of Hb concentration was studied by independently changing vessel oxygen partial pressure (pO(2)). For each oxygen distribution, the oxygen enhancement ratio (OER) was calculated and the mean absorbed dose at which the tumor control probability (TCP) was 0.99 (D(99)) was determined using the linear-quadratic cell survival model (LQ model). RESULTS: Decreased pO(2) shifted the oxygen distribution to lower values, whereas decreased vessel density caused the distribution to widen and shift to lower values. Combined scenarios caused lower-shifted distributions, emphasising log-normal characteristics. Vessel reduction combined with increased blood pO(2) caused the distribution to widen due to a lack of vessels. The most pronounced radiation effect of increased pO(2) occurred with tumor tissue with 50% of the maximum vessel density used in the simulations. A 51% decrease in D(99), from 123 to 60 Gy, was found between the lowest and highest pO(2) concentrations. CONCLUSIONS: Our results indicate that an intermediate vascular density region exists where enhanced blood oxygen concentration may be beneficial for radiation response. The results also suggest that it is possible to distinguish between diffusion-limited and anemic hypoxia from the characteristics of the pO(2) distribution.

Deficiency for endoglin in tumor vasculature weakens the endothelial barrier to metastatic dissemination.

Therapy-induced resistance remains a significant hurdle to achieve long-lasting responses and cures in cancer patients. We investigated the long-term consequences of genetically impaired angiogenesis by engineering multiple tumor models deprived of endoglin, a co-receptor for TGF-ß in endothelial cells actively engaged in angiogenesis. Tumors from endoglin-deficient mice adapted to the weakened angiogenic response, and refractoriness to diminished endoglin signaling was accompanied by increased metastatic capability. Mechanistic studies in multiple mouse models of cancer revealed that deficiency for endoglin resulted in a tumor vasculature that displayed hallmarks of endothelial-to-mesenchymal transition, a process of previously unknown significance in cancer biology, but shown by us to be associated with a reduced capacity of the vasculature to avert tumor cell intra- and extravasation. Nevertheless, tumors deprived of endoglin exhibited a delayed onset of resistance to anti-VEGF (vascular endothelial growth factor) agents, illustrating the therapeutic utility of combinatorial targeting of multiple angiogenic pathways for the treatment of cancer.