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Small cell lung cancer (SCLC) is among the most aggressive and lethal human malignancies. Most patients with SCLC who initially respond to chemotherapy develop disease relapse. Therefore, there is a pressing need to identify novel driver mechanisms of SCLC progression to unlock treatment strategies to improve patient prognosis. SCLC cells comprise subsets of cells possessing progenitor or stem cell properties, while the underlying regulatory pathways remain elusive. Here, we identified the isoform 1 of the neurogenesis-associated protein ASPM (ASPM-I1) as a prominently upregulated stemness-associated gene during the self-renewal of SCLC cells. The expression of ASPM-I1 was found to be upregulated in SCLC cells and tissues, correlated with poor patient prognosis, and indispensable for SCLC stemness and tumorigenesis. A reporter array screening identified multiple developmental signaling pathways, including Hedgehog (Hh) and Wnt pathways, whose activity in SCLC cells depended upon ASPM-I1 expression. Mechanistically, ASPM-I1 stabilized the Hh transcriptional factor GLI1 at the protein level through a unique exon-18–encoded region by competing with the E3 ligases β-TrCP and CUL3. In parallel, ASPM-I1 sustains the transcription of the Hh pathway transmembrane regulator SMO through the Wnt−DVL3−β-catenin signaling axis. Functional studies verified that the ASPM-I1–regulated Hh and Wnt activities significantly contributed to SCLC aggressiveness in vivo. Consistently, the expression of ASPM-I1 positively correlated with GLI1 and stemness markers in SCLC tissues. This study illuminates an ASPM-I1–mediated regulatory module that drives tumor stemness and progression in SCLC, providing an exploitable diagnostic and therapeutic target.Significance:ASPM promotes SCLC stemness and aggressiveness by stabilizing the expression of GLI1, DVL3, and SMO, representing a novel regulatory hub of Hh and Wnt signaling and targetable vulnerability.

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Small cell lung cancer (SCLC) is among the most aggressive and lethal human malignancies. Most patients with SCLC who initially respond to chemotherapy develop disease relapse. Therefore, there is a pressing need to identify novel driver mechanisms of SCLC progression to unlock treatment strategies to improve patient prognosis. SCLC cells comprise subsets of cells possessing progenitor or stem cell properties, while the underlying regulatory pathways remain elusive. Here, we identified the isoform 1 of the neurogenesis-associated protein ASPM (ASPM-I1) as a prominently upregulated stemness-associated gene during the self-renewal of SCLC cells. The expression of ASPM-I1 was found to be upregulated in SCLC cells and tissues, correlated with poor patient prognosis, and indispensable for SCLC stemness and tumorigenesis. A reporter array screening identified multiple developmental signaling pathways, including Hedgehog (Hh) and Wnt pathways, whose activity in SCLC cells depended upon ASPM-I1 expression. Mechanistically, ASPM-I1 stabilized the Hh transcriptional factor GLI1 at the protein level through a unique exon-18–encoded region by competing with the E3 ligases β-TrCP and CUL3. In parallel, ASPM-I1 sustains the transcription of the Hh pathway transmembrane regulator SMO through the Wnt−DVL3−β-catenin signaling axis. Functional studies verified that the ASPM-I1–regulated Hh and Wnt activities significantly contributed to SCLC aggressiveness in vivo. Consistently, the expression of ASPM-I1 positively correlated with GLI1 and stemness markers in SCLC tissues. This study illuminates an ASPM-I1–mediated regulatory module that drives tumor stemness and progression in SCLC, providing an exploitable diagnostic and therapeutic target.Significance:ASPM promotes SCLC stemness and aggressiveness by stabilizing the expression of GLI1, DVL3, and SMO, representing a novel regulatory hub of Hh and Wnt signaling and targetable vulnerability.

Resistance to antitumor treatment contributes to patient mortality. Functional proteomic screening of organoids derived from chemotherapy-treated patients with breast cancer identified nuclear receptor corepressor 2 (NCOR2) histone deacetylase as an inhibitor of cytotoxic stress response and antitumor immunity. High NCOR2 in the tumors of patients with breast cancer predicted chemotherapy refractoriness, tumor recurrence and poor prognosis. Molecular studies revealed that NCOR2 inhibits antitumor treatment by regulating histone deacetylase 3 (HDAC3) to repress interferon regulatory factor 1 (IRF-1)-dependent gene expression and interferon (IFN) signaling. Reducing NCOR2 or impeding its epigenetic activity by modifying its interaction with HDAC3 enhanced chemotherapy responsiveness and restored antitumor immunity. An adeno-associated viral NCOR2-HDAC3 competitor potentiated chemotherapy and immune checkpoint therapy in culture and in vivo by permitting transcription of IRF-1-regulated proapoptosis and inflammatory genes to increase IFN-γ signaling. The findings illustrate the utility of patient-derived organoids for drug discovery and suggest that targeting stress and inflammatory-repressor complexes such as NCOR2-HDAC3 could overcome treatment resistance and improve the outcome of patients with cancer.

Gastric cancer (GC) is the third leading cause of cancer mortality globally and a molecularly heterogeneous disease. Identifying the driver pathways in GC progression is crucial to improving the clinical outcome. Recent studies identified ASPM (abnormal spindle-like microcephaly-associated) and FOXM1 (Forkhead box protein M1) as novel Wnt and cancer stem cell (CSC) regulators; their pathogenetic roles and potential crosstalks in GC remain unclarified. The expression patterns of ASPM isoforms and FOXM1 were profiled in normal gastric epithelial and GC tissues. The functional roles of ASPM and FOXM1 in Wnt activity, cancer stemness and GC progression, and the underlying signaling processes were investigated. Approximately one third of GC cells upregulate the expression of ASPM isoform I (ASPMiI) in their cytoplasm; the tumors with a high ASPMiI positive score (≥ 10%) are associated with a poor prognosis of the patients. Mechanistically, the molecular interplay among FOXM1, ASPMiI and DVL3 was found to converge on β-catenin to control the Wnt activity and the stemness property of GC cells. This multi-mode Wnt-regulatory module serves to reinforce Wnt signals in CSCs by transcriptional regulation (FOXM1–ASPM), protein–protein interactions (ASPMiI–DVL3–β-catenin), and nuclear translocation (FOXM1–β-catenin). This study illuminates a novel Wnt- and stemness-regulatory mechanism in GC cells and identifies a novel subset of FOXM1highASPMiIhigh GC with potential to guide Wnt- and stemness-related diagnostics and therapies.

Resistance to antitumor treatment contributes to patient mortality. Functional proteomic screening of organoids derived from chemotherapy-treated patients with breast cancer identified nuclear receptor corepressor 2 (NCOR2) histone deacetylase as an inhibitor of cytotoxic stress response and antitumor immunity. High NCOR2 in the tumors of patients with breast cancer predicted chemotherapy refractoriness, tumor recurrence and poor prognosis. Molecular studies revealed that NCOR2 inhibits antitumor treatment by regulating histone deacetylase 3 (HDAC3) to repress interferon regulatory factor 1 (IRF-1)-dependent gene expression and interferon (IFN) signaling. Reducing NCOR2 or impeding its epigenetic activity by modifying its interaction with HDAC3 enhanced chemotherapy responsiveness and restored antitumor immunity. An adeno-associated viral NCOR2-HDAC3 competitor potentiated chemotherapy and immune checkpoint therapy in culture and in vivo by permitting transcription of IRF-1-regulated proapoptosis and inflammatory genes to increase IFN-γ signaling. The findings illustrate the utility of patient-derived organoids for drug discovery and suggest that targeting stress and inflammatory-repressor complexes such as NCOR2-HDAC3 could overcome treatment resistance and improve the outcome of patients with cancer.

Recurrent and hormone-refractory prostate cancer (PCA) exhibits aggressive behaviors while current therapeutic approaches show little effect of prolonging the survival of patients with PCA. Thus, a deeper understanding of the patho-molecular mechanisms underlying the disease progression in PCA is crucial to identify novel diagnostic and/or therapeutic targets to improve the outcome of patients. Recent evidence suggests that activation of Wnt signaling in cancer stem cells (CSCs) contributes to cancer progression in malignant tumors. Here, we report that a novel Wnt co-activator ASPM (abnormal spindle-like microcephaly associated) maintains the prostate CSC subpopulation by augmenting the Wnt-β-catenin signaling in PCA. ASPM expression is incrementally upregulated in primary and metastatic PCA, implicating its potential role in PCA progression. Consistently, downregulation of ASPM expression pronouncedly attenuated the proliferation, colony formation, and the invasive behavior of PCA cells, and dramatically reduced the number of ALDH

Background & Aims Many patients with pancreatic ductal adenocarcinoma (PDAC) develop recurrent or metastatic diseases after surgery, so it is important to identify those most likely to benefit from aggressive therapy. Disruption of tissue microarchitecture is an early step in pancreatic tumorigenesis and a parameter used in pathology grading of glandular tumors. We investigated whether changes in gene expression during pancreatic epithelial morphogenesis were associated with outcomes of patients with PDAC after surgery. Methods We generated architectures of human pancreatic duct epithelial cells in a 3-dimensional basement membrane matrix. We identified gene expression profiles of the cells during different stages of tubular morphogenesis (tubulogenesis) and of PANC-1 cells during spheroid formation. Differential expression of genes was confirmed by immunoblot analysis. We compared the gene expression profile associated with pancreatic epithelial tubulogenesis with that of PDAC samples from 27 patients, as well as with their outcomes after surgery. Results We identified a gene expression profile associated with tubulogenesis that resembled the profile of human pancreatic tissue with differentiated morphology and exocrine function. Patients with PDACs with this profile fared well after surgery. Based on this profile, we established a 6−28 gene tubulogenesis-specific signature that accurately determined the prognosis of independent cohorts of patients with PDAC (total n = 128; accuracy = 81.2%−95.0%). One gene, ASPM , was down-regulated during tubulogenesis but up-regulated in human PDAC cell lines and tumor samples; up-regulation correlated with patient outcomes (Cox regression P = .0028). Bioinformatic, genetic, biochemical, functional, and clinical correlative studies showed that ASPM promotes aggressiveness of PDAC by maintaining Wnt-β-catenin signaling and stem cell features of PDAC cells. Conclusions We identified a gene expression profile associated with pancreatic epithelial tubulogenesis and a tissue architecture−specific signature of PDAC cells that is associated with patient outcomes after surgery.

Chan et al. report that treatment of tumor-bearing mice with low-dose metronomic chemotherapy prevents stromal secretion of ELR+ chemokines and induction of tumor-initiating cells usually observed with administration of drugs at maximum tolerated dose., Although traditional chemotherapy kills a fraction of tumor cells, it also activates the stroma and can promote the growth and survival of residual cancer cells to foster tumor recurrence and metastasis. Accordingly, overcoming the host response induced by chemotherapy could substantially improve therapeutic outcome and patient survival. In this study, resistance to treatment and metastasis has been attributed to expansion of stem-like tumor-initiating cells (TICs). Molecular analysis of the tumor stroma in neoadjuvant chemotherapy–treated human desmoplastic cancers and orthotopic tumor xenografts revealed that traditional maximum-tolerated dose chemotherapy, regardless of the agents used, induces persistent STAT-1 and NF-κB activity in carcinoma-associated fibroblasts. This induction results in the expression and secretion of ELR motif–positive (ELR+) chemokines, which signal through CXCR-2 on carcinoma cells to trigger their phenotypic conversion into TICs and promote their invasive behaviors, leading to paradoxical tumor aggression after therapy. In contrast, the same overall dose administered as a low-dose metronomic chemotherapy regimen largely prevented therapy-induced stromal ELR+ chemokine paracrine signaling, thus enhancing treatment response and extending survival of mice carrying desmoplastic cancers. These experiments illustrate the importance of stroma in cancer therapy and how its impact on treatment resistance could be tempered by altering the dosing schedule of systemic chemotherapy.

Deregulation of transforming growth factor (TGF)-β function is a common feature of pancreatic cancer, rendering these cancers unresponsive to TGF-β–stimulated growth inhibition. Recent findings have supported a primary role for Kruppel-like factor 10 (KLF10) as an important transcription factor involved in mediating TGF-β1 signaling. The aim of this study was to evaluate the correlation between KLF10 expression and the clinical and pathologic features of pancreatic cancer. Tissue specimens from patients with pancreatic adenocarcinoma were retrospectively collected for immunohistochemical analysis. To demonstrate that Klf10 expression was primarily regulated by methylation status, the Klf10 promoter was examined by methylation-specific PCR using a pancreatic cancer cell line (Panc-1). DNA methyltransferase (DNMT) inhibitor and small-interfering RNA depletion of DNMT genes were used to reverse KLF10 expression in the Panc-1 cells. In parallel, DNMT1 expression was evaluated in the pancreatic cancer tissue specimens. In 95 pancreatic cancer tissue specimens, KLF10 expression was inversely correlated with pancreatic cancer stage ( P = 0.01). Multivariable analysis revealed that, in addition to the presence of distant metastasis at diagnosis ( P = 0.001 and 0.001, respectively), KLF10 was another independent prognostic factor related to progression-free and overall survival ( P = 0.018 and 0.037, respectively). The loss of KLF10 expression in advanced pancreatic cancer is correlated with altered methylation status, which seems to be regulated by DNMT1. Our results suggest that KLF10 is a potential clinical predictor for progression of pancreatic cancer.

Metastasis is the major cause of cancer mortality in androgen-independent prostate cancer (PCA) and understanding it is crucial to improving the outcome of patients. Recent evidence suggests that activation of Wnt signaling in cancer stem cells (CSCs) contributes to cancer progression in malignant tumors. Here we report that a novel Wnt activator ASPM (Abnormal spindle-like microcephaly associated) maintains the prostate CSC subpopulation through augmenting the Wnt-β-catenin signaling in PCA. Functional studies showed that downregulation of ASPM expression attenuated the proliferation and invasive of PCA cells and reduced the number of prostate CSCs and inhibited cancer stemness and tumorigenesis. Mechanistically, ASPM interacts with dishevelled-3 (Dvl-3) and inhibits its proteasome-dependent degradation, thereby increasing the protein stability of Dvl-3 and enabling the β-catenin transcriptional activity in PCA cells. The clinical significance of the above findings was credentialed by the profound up-regulation of ASPM in metastatic PCA and the strong correlation of cytoplasmic ASPM expression with poor metastasis-free survival of patients with resected PCA. Collectively, our data points to ASPM as a novel oncoprotein and an essential regulator of cancer stemness in PCA, which has important clinical and therapeutic significance (supported by Ministry of Science and Technology grants MOST 104-2314-B-400-022 and MOST 105-2314-B-400-018 and National Health Research Institutes NHRIs grant CA-106-PP-09 to K.K.T). Citation Format: Kelvin K. Tsai, Vincent C. Pai, Chung-Chi Hsu, Tze-Sian Chan. ASPM promotes prostate cancer tumorigenesis by bolstering cancer stemness through Wnt-Dvl-3-b-catenin signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 538.