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  • br C Human FG tumor growth

    2019-10-14


    (C) Human FG tumor growth in mice treated with gemcitabine and either vehicle or SR2211 for 2.5 weeks. Tumor volume fold change is relative to volume at enrollment.
    (DÐF) Primary patient organoid growth in the presence of vehicle or SR2211. Representative images of organoids in Matrigel (D; scale bars represent 100 mm), following recovery from Matrigel (E; scale bars represent 50 mm), and quantiÞcation of organoid circumference (F, left) or volume (F, right) are shown. (G) Growth of primary patient-derived xenografts treated with vehicle or SR2211 for 1.5 weeks; (n = 4).
    (H) RORC ampliÞcation in tumors of patients diagnosed with MIK665 (S-64315) various malignancies.
    (IÐL) Representative TMAs of PDAC and PanINs illustrating scoring for negative, cytoplasmic, and cytoplasmic + nuclear RORg staining (I). Correlation between RORg staining and tumor stage (J), lymphatic invasion (K), and lymph node status (L) is shown. Data represented as mean ± SEM. *p < 0.05; **p < 0.01; ***p < 0.001 by StudentÕs t test or one-way ANOVA. See also Figure S7.
    microarrays from a clinically annotated retrospective cohort of
    116 PDAC patients (Table S6). For 69 patients, matched pancreatic intraepithelial neoplasia (PanIN) lesions were avail-able. RORg protein was detectable (cytoplasmic MIK665 (S-64315) only denoted as low or cytoplasmic and nuclear expression denoted as high; Figure 7I) in 113 PDAC cases and 55 PanIN cases, respectively, and absent in 3 PDAC cases and 14 PanIN cases, respectively. Compared to cytoplasmic expres-sion, nuclear RORg expression in PDAC cases was signiÞ-cantly correlated with higher pathological tumor (pT) stages at diagnosis (Figure 7J). In addition, RORg expression in PanIN lesions was positively correlated with lymphatic vessel inva-sion (L1; Figure 7K) and lymph node metastasis (pN1 and 
    pN2; Figure 7L) by the invasive carcinoma. These results indi-cate that RORg expression in PanIN lesions and nuclear RORg localization in invasive carcinoma could be useful markers to predict PDAC aggressiveness.
    DISCUSSION
    It is an unfortunate truth that the most common outcome for pancreatic cancer patients following a response to cytotoxic therapy is not cure but eventual disease progression and death driven by drug-resistant, stem-cell-enriched populations (Fox et al., 2016; Van den Broeck et al., 2013). The work we report here has allowed us to develop a comprehensive molecular
    map of the core dependencies of pancreatic cancer stem cells by integrating their epigenetic, transcriptomic, and functional genomic landscape. This dataset thus provides a novel resource for understanding therapeutic resistance and relapse and for discovering new vulnerabilities in pancreatic cancer. As an example, the MEGF family of orphan receptors repre-sents a potentially actionable family of adhesion GPCRs, as this class of signaling receptors has been considered drug-gable in cancer and other diseases (Lappano and Maggiolini, 2011). Importantly, our epigenetic analyses revealed a signiÞ-cant relationship between super-enhancer-associated genes and functional dependencies in stem cell conditions; stem-cell-unique, super-enhancer-associated genes were more likely to drop out in the CRISPR screen in stem cell conditions compared to super-enhancer-associated genes in non-stem cells (Figure S7D). This provides additional evidence for the epigenetic and transcriptomic link to functional dependencies in cancer stem cells and further supports previous Þndings that super-enhancer-linked genes may be more important for maintaining cell identity and more sensitive to perturbation (Whyte et al., 2013).
    From the screens presented here, we identiÞed an unex-pected dependence of KPf/fC stem cells on inßammatory and im-mune mediators, such as the CSF1R/IL-34 axis and IL-10R signaling. Although these have been previously thought to act primarily on immune cells in the microenvironment (Guillonneau et al., 2017; Wang et al., 2019), our data suggest that stem cells may have evolved to co-opt this cytokine-rich milieu, allowing them to resist effective immune-based elimination. These Þnd-ings also suggest that agents targeting CSF1R, which are under investigation for pancreatic cancer (Sankhala et al., 2017), may act not only on the tumor microenvironment but also directly on pancreatic epithelial cells themselves. Our studies also raise the possibility that therapies designed to activate the immune system to attack tumors may have effects on tumor cells directly: just as we have learned chemotherapy can kill tumor cells but may also impair the immune system, therapies designed to acti-vate the immune system, such as IL-10, may also promote the growth of tumor cells. This dichotomy of action will need to be considered in order to better optimize immunomodulatory treat-ment strategies.