Cancer Mucin Research Articles

The cancer mucin barrier plays a multifunctional role in providing sustained physical resistance to natural killer cell attack

The cancer mucin barrier plays a multifunctional role in providing sustained physical resistance to natural killer cell attack

Clipping cell-surface mucins in cancer by targeted degradation.

Clipping cell-surface mucins in cancer by targeted degradation.

Mucins as Potential Biomarkers for Early Detection of Cancer.

Early detection significantly correlates with improved survival in cancer patients. So far, a limited number of biomarkers have been validated to diagnose cancers at an early stage. Considering the leading cancer types that contribute to more than 50% of deaths in the USA, we discuss the ongoing endeavors toward early detection of lung, breast, ovarian, colon, prostate, liver, and pancreatic cancers to highlight the significance of mucin glycoproteins in cancer diagnosis. As mucin deregulation is one of the earliest events in most epithelial malignancies following oncogenic transformation, these high-molecular-weight glycoproteins are considered potential candidates for biomarker development. The diagnostic potential of mucins is mainly attributed to their deregulated expression, altered glycosylation, splicing, and ability to induce autoantibodies. Secretory and shed mucins are commonly detected in patients' sera, body fluids, and tumor biopsies. For instance, CA125, also called MUC16, is one of the biomarkers implemented for the diagnosis of ovarian cancer and is currently being investigated for other malignancies. Similarly, MUC5AC, a secretory mucin, is a potential biomarker for pancreatic cancer. Moreover, anti-mucin autoantibodies and mucin-packaged exosomes have opened new avenues of biomarker development for early cancer diagnosis. In this review, we discuss the diagnostic potential of mucins in epithelial cancers and provide evidence and a rationale for developing a mucin-based biomarker panel for early cancer detection.

26P Serological evaluation of mucins in ovarian cancer

Ovarian cancer (OC) is one of the most lethal malignancies in females. A biomarker with high sensitivity and specificity has yet not been discovered which can aid in early diagnosis. Mucin proteins (MUC) proteins have been considered promising diagnostic biomarkers in cancer. We analyzed MUC1 and MUC4 for their diagnostic role in OC.

Mucins as anti-cancer targets: perspectives of the glycobiologist.

Mucins are highly O-glycosylated glycoproteins that carry a heterogenous variety of O-glycan structures. Tumor cells tend to overexpress specific mucins, such as the cell surface mucins MUC1 and MUC4 that are engaged in signaling and cell growth, and exhibit abnormal glycosylation. In particular, the Tn and T antigens and their sialylated forms are common in cancer mucins. We review herein methods chosen to use cancer-associated glycans and mucins as targets for the design of anti-cancer immunotherapies. Mucin peptides from the glycosylated and transmembrane domains have been combined with immune-stimulating adjuvants in a wide variety of approaches to produce anti-tumor antibodies and vaccines. These mucin conjugates have been tested on cancer cells in vitro and in mice with significant successes in stimulating anti-tumor responses. The clinical trials in humans, however, have shown limited success in extending survival. It seems critical that the individual-specific epitope expression of cancer mucins is considered in future therapies to result in lasting anti-tumor responses.

Targeting interleukin-17 receptor B enhances gemcitabine sensitivity through downregulation of mucins in pancreatic cancer

Pancreatic cancer is the fourth leading cause of death worldwide due to its poorest prognoses with a 7% 5-year survival rate. Eighty percent of pancreatic cancer patients relapse after chemotherapy and develop early metastasis and drug resistance. Resistance to nucleoside analog gemcitabine frequently used in first-line therapy is an urgent issue in pancreatic cancer treatment. Expression of mucin (MUC) glycoproteins has been shown to enhance chemoresistance via increased cell stemness. Here we show interlukine-17 receptor B (IL-17RB) expression is positively correlated with MUC1 and MUC4 expression in pancreatic cancer cells and tumor tissue. Moreover, IL-17RB transcriptionally up-regulates expression of MUC1 and MUC4 to enhance cancer stem-like properties and resistance to gemcitabine. These results suggest IL-17RB can be a potential target for pancreatic cancer therapy. Indeed, treatment with IL-17RB-neutralizing antibody has a synergistic effect in combination with gemcitabine for killing pancreatic cancer cells. Altogether, these findings provide feasible applications for IL-17RB-targeting therapy in pancreatic cancer treatment.

Study of Muc2 and Muc4 Mucins in Cancer of the Higher Aerodiversity

Objectve: To know the role played by the MUC2 and MUC4 mucins genes in cancers of the upper aerodigestve tract. Material and methods: It was a cross-sectonal retrospectve study of 10 years (July 2004 - July 2013) carried out in the otolaryngology departments of the University Hospital Center of Brazzaville and general hospital Adolph Sice (in Pointe-noire, Congo). All patents with upper aerodigestve tract cancers with histological evidence were included. For the study of mucins “MUC2 et MUC4” the molecular biology technique used was immunohistochemistry on pieces of laryngeal biopsies embedded in parafn. Results: A total of 85 cases of upper aerodigestve tract cancers were collected with a sample comprising the tumor and peri tumoral mucosa. The incidence was 8.5 cases of upper aero-digestve tract cancer per year and a rato of 1.8 in favor of men. The main risk factor was smoking (94.2%) and all patents with squamous cell carcinoma found on the tumor lining. On immunohistochemistry, the mucins MUC2 and MUC4 were present in the normal mucosa of the upper aero-digestve tract and gradually lost their expression from the metaplastc mucosa to the tumor mucosa (P=0.03). These mucins “MUC2 and MUC4” were absent on tumor mucosa in both smoking and non-smoking patents with a statstcally signifcant di?erence (P=0.01). Conclusion: The loss of expression of the MUC2 and MUC4 genes within the tumor mucosa is a key mechanism in the carcinogenesis of the upper aerodigestve tract.

Mucins in pancreatic cancer: A well-established but promising family for diagnosis, prognosis and therapy.

Mucins are a family of multifunctional glycoproteins that mostly line the surface of epithelial cells in the gastrointestinal tract and exert pivotal roles in gut lubrication and protection. Pancreatic cancer is a lethal disease with poor early diagnosis, limited therapeutic effects, and high numbers of cancer‐related deaths. In this review, we introduce the expression profiles of mucins in the normal pancreas, pancreatic precursor neoplasia and pancreatic cancer. Mucins in the pancreas contribute to biological processes such as the protection, lubrication and moisturization of epithelial tissues. They also participate in the carcinogenesis of pancreatic cancer and are used as diagnostic biomarkers and therapeutic targets. Herein, we discuss the important roles of mucins that lead to the lethality of pancreatic adenocarcinoma, particularly MUC1, MUC4, MUC5AC and MUC16 in disease progression, and present a comprehensive analysis of the clinical application of mucins and their promising roles in cancer treatment to gain a better understanding of the role of mucins in pancreatic cancer.

The Histochemical Alterations of Mucin in Colorectal Carcinoma Quantified by Two Efficient Algorithms of Digital Image Analysis.

The practical use of knowledge on the diagnostic-prognostic role of polysaccharide components of mucins in colorectal cancer (CRC) has been difficult, due to the number of histochemical (HC) reaction types, as well as lack of standard methods of computer-assisted analysis of tissue expression of these molecules. Using two algorithms of digital image analysis (by application of Image-Pro Premier and our originally designed program Filter HSV), we evaluated the expression of polysaccharides in tissue samples of CRC patients (n = 33), and fragments of normal colorectal tissue from the same patients (control) using periodic acid Schiff reaction (PAS) (neutral mucins) and alcian blue staining (AB) (acidic mucins). Our results indicate lower expression of the PAS+ and AB+ mucins in CRC, as compared to the control samples. The higher expression of PAS+ polysaccharides was detected in flat tumors than in protruded CRC, while higher AB+ mucins expression was a feature of mucinous CRC subtypes. Positive correlation between mutual PAS+ and AB+ expression, as well as correlations with glucose concentration (PAS+ mucins), and hemoglobin level (AB+ mucins) were observed exclusively in unchanged colorectal samples (control). Both algorithms of digital image analysis (smart segmentation and Filter HSV) work properly and can be used interchangeably in daily practice of pathologists, as useful tools of quantitative evaluation of HC reaction in both normal and cancerous tissues.

Expression of mucins (MUC1, MUC2, MUC5AC and MUC6) in ALK-positive lung cancer: Comparison with EGFR-mutated lung cancer

ALK-positive (ALK+) lung adenocarcinoma usually shows a more advanced-staged disease with frequent nodal metastasis and highly aggressive outcomes compared with EGFR-mutated lung cancers. The aim of this study was to investigate the expression profiles of several mucins in ALK + lung cancers to gain insight into the relationship between the more aggressive biological nature of ALK + lung cancers and the role of mucins.We examined the immunohistochemical profiles of mucins MUC1, MUC2, MUC5AC, and MUC6 in 19 ALK + lung cancers compared with 42 EGFR-mutated lung cancers.ALK + cancers were found to occur in younger patients and were characterized by a solid-predominant histologic subtype with frequent signet ring cells and peritumoral muciphages. By contrast, EGFR-mutated cancers lacked ALK-specific histological patterns. Although all MUC1 and MUC5AC were expressed in both subtypes, MUC1 expression in ALK + cancers was visualized exclusively through cytoplasmic staining, whereas those in EGFR-mutated cancers were predominantly membranous staining in apical area (92.9%) and focally in cytoplasmic staining (7.1%). MUC5AC expression in ALK + cancers was exclusively visualized through cytoplasmic staining (100%), whereas EGFR-mutated cancers showed predominantly perinuclear dot-like patterns (90.5%) and focal cytoplasmic staining (9.5%). MUC2 and MUC6 expression was not detected in either type of lung cancer. ConclusionsThe high frequency of both MUC1 and MUC5AC cytoplasmic expression, coupled with a lack of MUC2 and MUC6 expression in ALK + lung cancer may contribute to the biologically aggressive behavior of ALK + cancer. Inhibitors to these types of mucins may thus act as a barrier to cancerous extension reducing their aggressive behavior.

Differential expression of Cosmc, T-synthase and mucins in Tn-positive colorectal cancers

BackgroundThe Tn neoantigen (GalNAcα1-O-Ser/Thr) is an O-glycan expressed in various types of human cancers. Studies in several Tn-expressing cancer cell lines and pancreatic tumors have identified loss of Cosmc expression caused by either mutations or promoter hypermethylation. In this study, we explored the mechanism(s) for Tn expression in human colorectal cancers (CRC).MethodsTn-expressing cell populations were isolated from CRC cell lines by Fluorescence-associated cell sorting (FACS). The expression of the Tn and sialylated Tn (STn) antigens, Cosmc, T-synthase, and mucins was characterized in paired specimens with CRC and in CRC cell lines by immunostaining, western blot, and qPCR.ResultsUsing well-defined monoclonal antibodies, we confirmed prevalent Tn/STn expression in CRC samples. However, a majority of these tumors had elevated T-synthase activity and expression of both Cosmc and T-synthase proteins. Meanwhile, Tn antigen expression was not caused by mucin overproduction. In addition, we found that Tn-expressing CRC cell lines had either loss-of-function mutations in Cosmc or reversible Tn antigen expression, which was not caused by the deficiency of T-synthase activity.ConclusionsOur results demonstrate multiple mechanisms for Tn expression in CRCs.

Abstract 1518: Gene expression of trefoil proteins, gastrokines and mucins in Indian gastric cancer patients

Abstract Background: Gastric cancer is the (GC) third most cause of cancer related mortality worldwide. Histopathologically, GC is sub classified into two major histopathological subtypes- diffuse and intestinal. The trefoil factor family - TFF1, TFF2 and TFF3 is known to play a pivotal role in gastrointestinal mucosal defence and repair. Gastrokines (GKN1, GKN2) are stomach specific secretary proteins which play fundamental roles in regulating gastric epithelial homeostasis. Similarly, secreted mucins are large O-glycosylated proteins that participate in the protection/defence of underlying mucosa in normal adults. The loss of expression of trefoil proteins, gastrokines and several mucins in retinoblastoma and colorectal cancer are reported in recent studies. Aims: The molecular underpinnings and gene expression of Indian GC patients are largely unknown. In this study, we evaluated the gene expression of TFF1, TFF2, TFF3, GKN1, GKN2, MUC3A, MUC5AC, MUC13 in gastric cancer patients from India. Additionally, we wanted to see whether these gene expressions are different in two histopathological groups. Materials and Methods: Endoscopic biopsy tissue from gastric tumour and adjacent normal regions were collected with ethical approval and informed consent from 42 patients . After histopathological confirmation by two pathologists, total RNA were isolated from tumour and corresponding normal tissue and gene expression was measured by qRT-PCR. Statistical analysis was performed by R package. Result and Discussion: We identified that TFF1, TFF2, TFF3, GKN1, GKN2 are significantly differentially expressed in gastric tumour tissue with respect to adjacent normal tissue whereas mucins had no significant deregulation. TFF1, TFF2, GKN1 and GKN2 are found to be down regulated in most tumour tissues compared to adjacent normal. Only GKN2 expression is significantly different among diffuse and intestinal subtypes . Our statistical analysis also revealed that gene expression of TFF1, TFF2, GKN1 and GKN2 are highly correlated with the gene expression of MUC5AC. Further study with larger patient sample is required to fully understand the complex interplay of trefoil, gastrokines and mucins in GC. Citation Format: Dipjit Basak, Pronoy Kanti Mondal, Brishti Mondal, Ranajoy Ghosh, Aniket Halder, Gopal Krishna Dhali, Abhijit Chowdhury, Indranil Mukhopadhyay, Shalini Datta. Gene expression of trefoil proteins, gastrokines and mucins in Indian gastric cancer patients [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 1518.

P077 Aberrant Muc1 and Muc13 expression in association with Par polarity complex and Vip dysfunction in a colitis T-cell transfer model

There is emerging evidence that loss of the gastrointestinal mucosal barrier integrity is a significant contributor to the pathophysiology of inflammatory bowel disease (IBD). Mucins represent the major components of the mucus barrier and inappropriate overexpression of transmembrane mucins can affect the barrier integrity by disrupting cell polarity and cell-cell interactions resulting in tight junction dysfunction.1 Aberrant expression of Muc1, Muc4, and Muc13, in particular, is observed in colonic tissue samples of IBD patients.2, 3 Nevertheless, the role of mucins in mucosal barrier disruption in IBD remains elusive. In this study, we investigated the expression patterns of transmembrane mucins and the mediators involved in barrier homeostasis during the progression of colitis in mice. Colitis was induced in immunocompromised SCID mice by the adoptive transfer of CD4+CD25-CD62L+ T cells from the spleens of BALB/c donor mice. At 2, 4, and 6 weeks post-colitis, animals were sacrificed and colonic tissue collected for RNA and protein analysis of membrane-bound mucins (Muc1, Muc4, and Muc13), polarity proteins, and the vasointestinal polypeptide (Vip). This latter protein is implicated in the regulation of tight junction and mucin expression (Hokari et al., 2005). Apoptotic cells in tissue sections were detected using a terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL) assay. Our data showed that Muc1 expression was significantly increased after 4 weeks of colitis (Figure a). Interestingly, a significant decrease of Muc13 expression was observed during acute colitis (2 weeks) whereas a remarkable turnover in expression was noted during chronic colitis (4–6 weeks; Figure b). Normal intestinal MUC13 expression plays crucial roles in epithelial proliferation, differentiation and apoptosis but inappropriate overexpression of this mucin could lower the level of protection. Indeed, a higher number of apoptotic cells was seen in tissue of chronic colitis-induced mice (Figure d). Furthermore, the polarity proteins Par3 and aPKC as well as Vip (Figure c) were significantly reduced in expression from week 2 onwards suggesting a disturbance in cell polarity. Relative expression of Muc1 (a), Muc13 (b), Vip (c) in the colon of mice with acute (2w) and chronic (4–6w) colitis. (d) Number of apoptotic cells in healthy and chronic colitis mice. Asterisk indicates significance (one-way ANOVA; p < 0.05). The findings of this study highlight a potential role for Muc1 and Muc13 in mucosal barrier disruption in IBD and the underlying mechanisms involved necessitate further investigation. 1. Kufe D. Mucins in cancer: function, prognosis and therapy. Nat Rev Cancer, 2009. 2. Sheng YH, Lourie R, Lindén SK, et al.. The MUC13 cell-surface mucin protects against intestinal inflammation by inhibiting epithelial cell apoptosis. Gut, 2011. 3. Maaike V, Tim V, Ricard F, et al. Genetic and transcriptomic bases of intestinal epithelial barrier dysfunction in inflammatory bowel disease. Inflamm Bowel Dis. 2017.

CHARACTERIZATION OF THE MUCIN PHENOTYPE CAN PREDICT GASTRIC CANCER RECURRENCE AFTER ENDOSCOPIC MUCOSAL RESECTION.

Endoscopic mucosal resection is still considered an accepted treatment for early gastric cancer for selected cases. Histopathologic criteria for curative endoscopic resection are intramucosal well-differentiated adenocarcinoma, lateral and deep margins free of tumor, no histological ulceration, and no venous or lymphatic embolism. A 5% local recurrence rate has been described even when all the above-mentioned criteria are met. On the other hand, antigen expression by tumoral cells has been related to the biological behavior of several tumors. To evaluate whether early gastric cancer mucin immunoexpression, p53 and Ki-67, can predict recurrence after endoscopic mucosal resection, even when standard histopathologic criteria for curative measures have been attempted. Twenty-two patients with early gastric cancer were considered to have been completely resected by endoscopic mucosal resection. Local recurrence occurred in 5/22 (22.7%). Immunohistochemical study was possible in 18 (81.8%) resected specimens. Patients were divided in two groups: those with and those without local recurrence. They were compared across demographic, endoscopic, histologic data, and immunohistochemical factors for MUC2, MUC5a, CD10, p53, and Ki-67. Mucin immunoexpression allowed a reclassification of gastric adenocarcinoma in intestinal (10), gastric (2), mixed (4), and null phenotypes (2). Mixed phenotype (positive for both MUC2 and MUC5a) was found in 80% of cases in the local recurrence group, while the intestinal type (positive MUC2 and negative MUC5a) was found in 76.9% of cases without local recurrence (P=0.004). Other observed features did not correlate with neoplastic recurrence. The mixed phenotype of early gastric adenocarcinoma is associated with a higher probability of local recurrence after endoscopic mucosal resection.

SLC3A2, antigen of mAb 3G9, promotes migration and invasion by upregulating of mucins in gastric cancer.

Solute carrier family 3 member 2 (SLC3A2) has been reported to be highly expressed in a variety of carcinomas. However, the function of SLC3A2 in gastric carcinoma (GC) has not been well explored. Monoclonal antibody (mAb) 3G9, generated from immunogen of various human GC cell lines, has been shown to bind to GC tissues specifically. In this study, we identified the target antigen of mAb 3G9 as SLC3A2, and detected the expression profile of SLC3A2 in a panel of gastric cancer cell lines and GC tumor tissues. We found that the increased expression of SLC3A2 was associated with serosal invasion in GC patients. Knockout of SLC3A2 suppressed the migration and invasion of BGC-823 cells in vitro and in vivo, whereas overexpression of SLC3A2 in NCI-N87 cells promoted the migration and invasion in vitro and in vivo. Mechanistic investigations suggested that MUC1, MUC16 and MUC5B were the downstream genes of SLC3A2 in GC cells. Taken together, our data suggested that SLC3A2 promoted the aggressive phenotype of GC by upregulating several mucin genes expression and may serve as a potential biomarker for diagnosis and target therapy.

Abstract 381: Genomic alterations in mucins in cancer

Abstract Mucins promote the growth, aggressiveness, and invasiveness of various cancer types. Changes in the expression or localization of mucins can significantly alter patient survival. Due to their predominance in multiple cancers, biomarkers and therapeutic agents have been developed to improve patient care. However, an extensive investigation across multiple cancer subtypes examining the genomic status of the cohort of mucins has not been performed and could yield significant leads into new roles of mucins in different cancers. Utilizing The Cancer Genome Atlas, the cancers of 11 mucin expressing tissues were examined for the genomic alterations in mutations, mRNA expression levels, copy numbers, methylation status, and the impact on patient survival due to these genomic features. It was discovered that different mucins across multiple tissue origins and cancer subtypes might have a different rate, pattern, and survival impact. MUC1 was not mutated in the majority of cancers; however, we did observe a frequent T112P mutation in pancreatic tumors that was also present in other cancers. MUC4 was frequently mutated at H4205 and MUC4 mutations were seen to have survival differences. Multiple significant differences were discovered in mRNA expression, which varied distinctly between cohorts. One such pattern was observed with de novo expression of MUC21 in colorectal cancers, in which mRNA expression was suddenly induced in cancer and increased in a stage dependent manner. Copy number alterations were seen for only a few cancer subtypes and mucins, including MUC4 in lung squamous cell carcinomas. Global promoter demethylation was generally observed across the cancers. MUC15 in renal cancers was seen to be one of the most demethylated mucins, in which mRNA was seen to be substantially decreased or expunged. This extensive study investigated the status of multiple mucins in multiple cancer settings and highlights a few noticeable mucins that are suggested to be further studied for roles in biomarkers, survival, and etiology for cancer progression. Citation Format: Ryan J. King, Fang Yu, Pankaj K. Singh. Genomic alterations in mucins in cancer [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 381. doi:10.1158/1538-7445.AM2017-381

Abstract 5587: Novel monoclonal antibodies block N-glycosylation Sites of the MUC16 ectodomain in ovarian carcinoma

Abstract The MUC16 gene encodes the CA125 antigen that imparts a negative survival impact on patients with high-grade serous ovarian cancer. Short elements from the C-terminal region of MUC16 transform 3T3 cells. Introduction of the 114 amino acid C-terminal portion of MUC16 promotes invasive growth in xenograft models. This effect is dependent on N-glycosylation sites of MUC16 and interactions present on the cell surface. Mechanisms by which the ectodomain promotes MUC16 oncogenic behaviors, the function of N-glycosylation and the implicated cell surface receptors are not fully delineated. To unravel the complex MUC16 cancer biology, in this study we investigated the glycosylation-based mechanisms and potential partners involved in MUC16 oncogenic effects, and show that site-specific glycosylation of MUC16 plays a key role in mucin-related transformation by mediating complex cell surface interactions. These effects are mediated through MGAT5-dependent N-glycosylation of two proximal N- glycosylation sites within the 58 amino acid retained MUC16 ectodomain acting in combination with Galectin-3 and growth factor receptors. Neither N- nor O-glycosylation sites in the more distally located MUC16 regions can functionally substitute for those two sites. The tumor-enhancing properties of MUC16 depend on co-localization of MUC16, Galectin-3, and growth factors receptors on lipid rafts. Loss of either Galectin-3 expression or the glycosylation enzyme MGAT5 completely abrogates MUC16 tumor promotion. Using synthetic glycopeptides, we have developed novel monoclonal antibodies (mAbs) directed at the crucial ectodomain N-glycosylation sites to inhibit the glycosylation- dependent effects of MUC16 on metastasis and invasion. These novel antibodies were characterized in vitro and in vivo studies using various ovarian cancer cell lines. All of the antibodies against the N- glycosylation sites closest to the cell membrane of MUC16 block Galectin-3-mediated polymerization to cell surface signaling molecules and inhibit the tumor-promoting effects of MUC16. The implications are broad: both for the biological understanding of cancer mucin biology and for potential therapeutic strategies. Citation Format: Dharmarao Thapi, Alberto Fernandez-Tejada, Abraham J. Axelrod, Nestor Rosales, Xiujun Yan, Marina Stasenko, Sahityasri Thapi, Amy Wang, Samuel J. Danishefsky, David R. Spriggs. Novel monoclonal antibodies block N-glycosylation Sites of the MUC16 ectodomain in ovarian carcinoma [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 5587. doi:10.1158/1538-7445.AM2017-5587

Abstract MIP-068: TUMOR–PROMOTING EFFECTS OF THE OVARIAN CANCER MUCIN, MUC16, ARE ASSOCIATED WITH SUPPRESSION OF INNATE IMMUNITY

Abstract The membrane-spanning mucin MUC16 promotes proliferation and metastasis of ovarian cancer cells. Here, we demonstrate that mice implanted with MUC16-knockdown human ovarian tumors show greater than two-fold increase in survival as compared to controls with tumors that express this mucin. While the lower survival of mice bearing MUC16-positive tumors is reportedly attributable to the ability of MUC16 to increase proliferation and metastasis, we also show that murine NK cells and macrophages are more efficient at lysing MUC16-knockdown cells. Macrophages showed infiltration in the MUC16-knockdown tumors but were restricted to the surrounding stroma in MUC16-positive tumors. In vitro cytotoxicity assays with NK cells and macrophages isolated from mice stimulated with agonistic anti-CD40 antibody showed 2-3-fold increased activity against the MUC16-knockdown cells as compared to matching target cells expressing this mucin. Finally, knockdown of MUC16 increased the susceptibility of cancer cells to ADCC and lysis by KS-IL2, an immunocytokine investigated as a treatment for ovarian cancer. Collectively our results support a role for MUC16 in promoting tumor growth. The ability of murine innate cells to selectively target MUC16-knockdown cells indicates that in vivo experiments investigating the biology of this mucin should be interpreted taking into account its immunomodulating activity as well as its growth promoting properties. Citation Format: Mildred Felder, Arvinder Kapur, Alexander L. Rakhmilevich, Xiaoyi Qu2, Joseph Connor, and Manish S. Patankar. TUMOR–PROMOTING EFFECTS OF THE OVARIAN CANCER MUCIN, MUC16, ARE ASSOCIATED WITH SUPPRESSION OF INNATE IMMUNITY [abstract]. In: Proceedings of the 11th Biennial Ovarian Cancer Research Symposium; Sep 12-13, 2016; Seattle, WA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(11 Suppl):Abstract nr MIP-068.

Genomic alterations in mucins across cancers.

The significance of mucins in cancers has led to the development of novel biomarkers and therapeutic agents against cancers. Despite significant advances in the understanding of mucins, systemic investigations into the role of mucins in cancer biology focusing particularly on the histological subtypes and stages, along with other variables, are yet to be carried out to discover potential novel functions and cancer-specific roles. Here, we investigated 11 mucin expressing cancers for DNA mutations, mRNA expression, copy number, methylation, and the impacts these genomic features may have on patient survival by utilizing The Cancer Genome Atlas dataset. We demonstrate that mucin DNA mutations have a significant rate, pattern, and impact on cancer patient survival depending on the tissue of origin. This includes a frequent T112P mutation in MUC1 that is seen in half of the pancreatic MUC1 mutations, as well as being present in other cancers. We also observed a very frequent MUC4 mutation at H4205, which correlated with survival outcomes in patients. Furthermore, we observed significant alterations in mucin mRNA expression in multiple tumor types. Our results demonstrate de novo expression of certain mucins in cancer tissues, including MUC21 in colorectal cancers. We observed a general decrease in promoter methylation for mucins, which correlated with decreased expression of many genes, such as MUC15 in kidney cancers. Lastly, several mucin gene loci demonstrated copy number increase in multiple histological subtypes. Thus, our study presents a comprehensive analysis of genomic alterations in mucins and their corresponding roles in cancer progression.

Lactobacillus reuteri decreases adherent tumor mucins and enhances chemotherapeutic susceptibilities via ERK/JNK pathways

BackgroundColorectal cancer (CRC) is the second leading cause of cancer related deaths in the United States. Although CRC is commonly treated with the chemotherapeutict Fluorouracil (5‐FU), only 10–15% of advanced CRC tumors respond. Mucins, a hallmark of carcinomas derived from epithelia, have long been implicated in the pathogenesis of cancer including CRC. The cancer mucin complex created by adherent mucins (MUC1, MUC3, MUC4) and secreted mucins (MUC2, MUC5AC) have been hypothesized to limit the uptake of the chemotherapy drugs, thus promoting chemoresistance. Several members of the gut microbiota are capable of altering mucin production and may represent potential treatments to increase the chemotherapeutic susceptibility in resistant cancers.Methods &amp; ResultsThe mucus producing adenocarcinoma cell line HT‐29‐MTX‐E12, which aberrantly expresses both adherent and secreted mucins, was incubated with conditioned media (CM) from probiotic human gut microbes. Analysis of MUC expression by qRT‐PCR revealed that several Bifidobacterium spp. and Lactobacillus spp. were capable of decreasing adherent and secreted mucin expression. Of the Lactobacillus members, L. reuteri ATCC 6475 was capable of altering total adherent mucins, as determined by immunofluorescence (IF), alcian blue (AB), and biotinylation. L. reuteri was able to stimulate expulsion of stored mucin granules as determined by media AB and MUC5AC protein as determined by IF. To assess whether bacterial‐specific changes in mucin expression correlated with increased cancer susceptibility to drug treatment, HT‐29‐MTX‐E12 cells were incubated with CM followed by the chemotherapeutic agent 5‐FU. L. reuteri CM increased susceptibility to 5‐FU treatment resulting in significant decreases in cancer cell viability as determined by resazurin. This effect was independent of lactate. The L. reuteri secreted factor was found to be a &gt;10 kDa, heat stable protein. 130 L. reuteri mutants were screened from a secretome library and three mutants were identified that lacked the ability to promote 5‐FU susceptibility. Two mutants encoded uncharacterized proteins, and one mutant encoded a bacterial ABC transporter. Magpix multiplex assay revealed that WT L. reuteri, but not the mutants, were able to promote ERK, CREB, and JNK phosphorylation. No changes were observed in p7056, STAT3, STAT5, p38 or NF‐κB. This correlated with no alteration in 5‐FU susceptibility when the cells were pre‐treated with an NF‐κB inhibitor. To establish the ability of L. reuteri to alter mucins and promote tumor 5‐FU susceptibility in vivo, C57BL/6J‐ApcMin/J mice were colonized with L. reuteri bacteria or treated with un‐inoculated bacterial media. L. reuteri colonized ApcMin mice had significantly decreased adherent Muc1, Muc4, and Muc5ac tumor masses compared with media‐treated mice, consistent with the HT29‐MTX cell line data. To examine 5‐FU susceptibility ApcMin mouse tumors were excised and ex‐vivo treated with L. reuteri CM followed by 5‐FU. Addition of CM resulted in enhanced susceptibility of male and female intestinal tumors to 5‐FU treatment as assessed by resazurin.ConclusionTogether this data suggests that bacterial‐induced modifications of cancer‐driven mucus in combination with traditional chemotherapeutic agents may provide new therapeutic strategies for the treatment of chemoresistant cancer.Support or Funding InformationT32 DK007664