Improved chemosensitivity following mucolytic therapy in patient-derived models of mucinous appendix cancer.

Abundant intraperitoneal (IP) accumulation of extracellular mucus in patients with appendiceal mucinous carcinoma peritonei (MCP) causes compressive organ dysfunction and prevents delivery of chemotherapeutic drugs to cancer cells. We hypothesized that reducing extracellular mucus would decrease tumor-related symptoms and improve chemotherapeutic effect in patient-derived models of MCP. Mucolysis was achieved using a combination of bromelain (BRO) and N-acetylcysteine (NAC). Ex vivo experiments of mucolysis and chemotherapeutic drug delivery/effect were conducted with MCP and non-MCP tissue explants. In vivo experiments were performed in mouse and rat patient-derived xenograft (PDX) models of early and late (advanced) MCP. MCP tumor explants were less chemosensitive than non-MCP explants. Chronic IP administration of BRO + NAC in a mouse PDX model of early MCP and a rat PDX model of late (advanced) MCP converted solid mucinous tumors into mucinous ascites (mucolysis) that could be drained via a percutaneous catheter (rat model only), significantly reduced solid mucinous tumor growth and improved the efficacy of chemotherapeutic drugs. Combination of BRO + NAC efficiently lyses extracellular mucus in clinically relevant models of MCP. Conversion of solid mucinous tumors into mucinous ascites decreases tumor bulk and allows for minimally invasive drainage of liquified tumors. Lysis of extracellular mucus removes the protective mucinous coating surrounding cancer cells and improves chemotherapeutic drug delivery/efficacy in cancer cells. Our data provide a preclinical rationale for the clinical evaluation of BRO + NAC as a therapeutic strategy for MCP.

Rational application of targeted therapeutics in mucinous colon/appendix cancers with positive predictive factors.

Molecular-targeted therapies have demonstrated disappointing results against most advanced solid cancers. This may largey be attributed to irrational drug use against unselected cancers. We investigated the efficacy of dual MEK-PI3K drug therapy against KRAS mutated mucin 2 (MUC2)-secreting LS174T cells and patient-derived ex vivo and in vivo models of KRAS mutated mucinous colon/appendix cancers. These tumors demonstrate unique phenotypic and genotypic features that likely predict sensitivity to this targeted co-therapy. Co-treatment with MEK inhibitor (trametinib) and PI3K inhibitor (pictilisib)-induced synergistic cytotoxicity and intrinsic mitochondrial-mediated apoptosis in LS174T cells and tumor explants in vitro. Dual drug therapy also induced endoplasmic reticulum stress (ERS)-associated proteins (GRP78/BiP, ATF4, and CHOP). However, CHOP knock-down assays demonstrated that mitochondrial-mediated apoptosis in LS174T cells was not ERS-dependent. Dual drug therapy also significantly decreased MUC2 expression, MUC2 post-translational modification (palmitoylation) and secretion in LS174T cells, suggesting a simultaneous cytotoxic and mucin suppressive mechanism of action. We also demonstrated effective mucinous tumor growth suppression in ex vivo epithelial organoid (colonoid) cultures and in in vivo intraperitoneal patient-derived xenograft models derived from mucinous colon/appendix cancer. These promising preclinical data support a role for dual MEK-PI3K inhibitor therapy in mucinous colon/appendix cancers. We postulate that mucinous KRAS mutated cancers are especially vulnerable to this co-treatment based on their unique phenotypic and genotypic characteristics.

Synergistic apoptosis following endoplasmic reticulum stress aggravation in mucinous colon cancer.

BACKGROUND: Mucinous colon cancers (MCC) are characterized by abundant production of mucin 2 (MUC2) protein and are less sensitive to standard systemic chemotherapy. We postulated that severe/persistent endoplasmic reticulum stress (ERS) aggravation in MCC would overwhelm compensatory cytoprotective pathways and induce apoptosis. RESULTS: Basal levels of ERS markers were higher in MCC and dnTCF-LS174T cells than non-mucinous tumors and these levels were significantly increased by combinatorial treatment with ERS aggravators celecoxib + orlistat. Combination treatment inhibited cell viability and synergistically induced apoptosis. Treatment-induced cell death was ERS-dependent, apoptotic pathways were not activated following knockdown of ERS protein CHOP. Dual drug treatment significantly reduced mucinous tumor growth in vivo and induced ERS and apoptosis, consistent with in vitro experiments. CONCLUSIONS: Novel therapies are needed since MCC are more resistant to standard systemic chemotherapy. This study suggests ERS aggravation is a viable therapeutic strategy to reduce tumor growth in MCC.

Targeting G-protein coupled receptor-related signaling pathway in a murine xenograft model of appendiceal pseudomyxoma peritonei.

Cancer cells aberrantly express mucins to enhance their survival. Relative chemoresistance of appendiceal pseudomyxoma peritonei (PMP) is attributed to abundant extracellular mucin 2 (MUC2) protein production. We hypothesized that simultaneous MUC2 inhibition and apoptosis induction would be effective against mucinous tumors. In vitro studies were conducted using LS174T cells (MUC2-secreting human colorectal cancer cells), PMP explant tissue, and epithelial organoid cultures (colonoids) derived from mucinous appendix cancers. In vivo studies were conducted using murine intraperitoneal patient-derived xenograft model of PMP. We found COX-2 over-expression in PMP explant tissue, which is known to activate G-protein coupled EP4/cAMP/PKA/CREB signaling pathway. MUC2 expression was reduced in vitro by small molecule inhibitors targeting EP4/PKA/CREB molecules and celecoxib (COX-2 inhibitor), and this was mediated by reduced CREB transcription factor binding to the MUC2 promoter. While celecoxib (5-40 µM) reduced MUC2 expression in vitro in a dose-dependent fashion, only high-dose celecoxib (≥ 20 µM) decreased cell viability and induced apoptosis. Chronic oral administration of celecoxib decreased mucinous tumor growth in our in vivo PMP model via a combination of MUC2 inhibition and induction of apoptosis. We provide a preclinical rationale for using drugs that simultaneously inhibit MUC2 production and induce apoptosis to treat patients with PMP.