LKB1 deficiency in T cells promotes the development of gastrointestinal polyposis.

Germline mutations in STK11, which encodes the tumor suppressor liver kinase B1 (LKB1), promote Peutz-Jeghers syndrome (PJS), a cancer predisposition syndrome characterized by the development of gastrointestinal (GI) polyps. Here, we report that heterozygous deletion of Stk11 in T cells (LThet mice) is sufficient to promote GI polyposis. Polyps from LThet mice, Stk11+/- mice, and human PJS patients display hallmarks of chronic inflammation, marked by inflammatory immune-cell infiltration, signal transducer and activator of transcription 3 (STAT3) activation, and increased expression of inflammatory factors associated with cancer progression [interleukin 6 (IL-6), IL-11, and CXCL2]. Targeting either T cells, IL-6, or STAT3 signaling reduced polyp growth in Stk11+/- animals. Our results identify LKB1-mediated inflammation as a tissue-extrinsic regulator of intestinal polyposis in PJS, suggesting possible therapeutic approaches by targeting deregulated inflammation in this disease.

Significant difference of neutrophil-lymphocyte ratio between colorectal cancer, adenomatous polyp and healthy people.

OBJECTIVE: Tumor was reported to correlate with inflammation and the host's inflammatory response to tumor has been shown to independently predict the outcome. Many measures of the systemic inflammatory response have been studied in recent years. In the present study the full blood count (leukocyte, neutrophil, lymphocyte) of colorectal cancers (CRCs) adenomatous polyps, and healthy people were collected, and the difference of ratios was studied. PATIENTS AND METHODS: A total of 752 individuals (242 colorectal cancers, 248 adenomatous polyps, and 262 healthy people) were randomized enrolled in the present study. The full blood counts (leukocyte, neutrophil, and lymphocyte) of each individual were collected and the NLRs were calculated. RESULTS: The leukocyte count, neutrophil ratio and neutrophil-lymphocyte ratio were the highest in colorectal cancer group, the second in adenomatous polyp group, and the lowest in healthy control (p < 0.001). The lymphocyte ratio was in the reverse order (p < 0.001). The ROC curve analysis showed that sensitivity and specificity levels of NLR were 66.9% and 77.6% for CRCs, 36.7% and 80.9% for adenomatous polyp. The leukocyte count was higher in the advanced adenomatous polyp compared with low-risk group (p = 0.042). Further analyses of the diagnostic value of NLR are warranted in the future. CONCLUSIONS: Difference of leukocyte count, neutrophil ratio and NLR may provide available information in the differential diagnosis of CRC, adenomatous polyp and healthy people.

Association Between Natural Killer Cell Activity and Colorectal Cancer in High-Risk Subjects Undergoing Colonoscopy.

BACKGROUND & AIMS: Low activity of natural killer (NK) cells has been associated with increased risk of cancer and has been reported in patients with colorectal cancer (CRC). Activity of NK cells can be measured in a small volume of whole blood by a commercially available test. We investigated whether this test could be used to identify patients with CRC, using findings from colonoscopy as a reference standard. METHODS: We performed an open-label, prospective, cross-sectional study of 872 high-risk subjects (more than 40 years old) screened for CRC by colonoscopy at a university hospital in Montreal, Canada from October 2014 through January 2016. Blood samples were collected on the day of colonoscopy, prior to the procedure. The test involves stimulation of whole blood with cytokine that induces NK cells to secrete interferon gamma (IFNG), which is quantified by an ELISA. Tissue samples were taken from lesions during the colonoscopy and analyzed histologically; subjects were classified as having no evidence of disease, adenomatous polyps of less than 10 mm, of 10 mm or more, or CRC. We used the non-parametric Mann-Whitney test to compare NK cell activity between subjects with no evidence of CRC and subjects found to have CRC. Receiver operating characteristic curve analysis was used to assess the ability of the test to identify individuals with CRC. The primary objective was to determine the difference in NK cell activity between subjects with vs without CRC. The secondary objective was the test performance, based on receiver operating characteristic analysis, and cut-off value that most accurately identified individuals with CRC. RESULTS: We found a significant difference in NK cell activity between the 23 subjects with CRC (based on pathology analysis) and the 849 subjects without CRC: subjects found to have CRC by colonoscopy had a median level of 86.0 pg IFNG/mL (inter-quartile range, 43.3-151.0 pg IFNG/mL), whereas subjects without CRC had a median level of 298.1 pg IFNG/mL (inter-quartile range, 100.4-920.2 pg IFNG/mL) (P = .0002). The cut-off value that most accurately identified subjects with CRC was 181 pg/mL. The NK cell activity test identified subjects with CRC with 87.0% sensitivity, 60.8% specificity, a positive predictive value of 5.7%, and a negative predictive value of 99.4%. The odds ratio for detection of CRC in subjects with low NK cell activity vs subjects with higher NK cell activity was 10.3 (95% CI, 3.03-34.9). CONCLUSIONS: Using colonoscopy as the reference standard, a test for NK cell activity in whole blood samples identified patients with CRC with 87.0% sensitivity and a negative predictive value of 99.4%. Subjects with low NK cell activity had a 10-fold higher risk of CRC compared with subjects with high NK cell activity. This test might be used in clinical practice to assess patients for risk of CRC. Clinicaltrials.gov number: NCT02291198.

The Immune Landscapes of Polypoid and Nonpolypoid Precancerous Colorectal Lesions.

Little is known about the immunoediting process in precancerous lesions. We explored this aspect of benign colorectal adenomas with a descriptive analysis of the immune pathways and immune cells whose regulation is linked to the morphology and size of these lesions. Two series of polypoid and nonpolypoid colorectal adenomas were used in this study: 1) 84 samples (42 lesions, each with matched samples of normal mucosa) whose gene expression data were used to quantify the tumor morphology- and size-related dysregulation of immune pathways collected in the Molecular Signature Database, using Gene Set Enrichment Analysis; 2) 40 other lesions examined with immunohistochemistry to quantify the presence of immune cells in the stromal compartment. In the analysis of transcriptomic data, 429 immune pathways displayed significant differential regulation in neoplasms of different morphology and size. Most pathways were significantly upregulated or downregulated in polypoid lesions versus nonpolypoid lesions (regardless of size). Differential pathway regulation associated with lesion size was observed only in polypoid neoplasms. These findings were mirrored by tissue immunostaining with CD4, CD8, FOXP3, MHC-I, CD68, and CD163 antibodies: stromal immune cell counts (mainly T lymphocytes and macrophages) were significantly higher in polypoid lesions. Certain markers displayed significant size-related differences regardless of lesion morphology. Multivariate analysis of variance showed that the marker panel clearly discriminated between precancerous lesions of different morphologies and sizes. Statistical analysis of immunostained cell counts fully support the results of the transcriptomic data analysis: the density of infiltration of most immune cells in the stroma of polypoid precancerous lesions was significantly higher than that observed in nonpolypoid lesions. Large neoplasms also have more immune cells in their stroma than small lesions. Immunoediting in precancerous colorectal tumors may vary with lesion morphology and stage of development, and this variability could influence a given lesion's trajectory to cancer.

Celiac disease and gastric hyperplastic polyps: a case series of an uncommon association.

Gastric polypoid lesions are found in ∼1-4% of patients who undergo esophagogastroduodenoscopy. The hyperplastic lesions are considered non-neoplastic polyps that are distributed randomly in the stomach and they are usually related to chronic gastritis as a result of the exaggerated mucosal healing response. Although several conditions have been associated with celiac disease (CD), such as thyroiditis, Addison's disease, type 1 diabetes mellitus, and autoimmune hepatitis, the association with gastric polyps is rare. In this case series, we present seven newly diagnosed patients (six women) with CD and hyperplastic gastric polyps. Helicobacter pylori infection was ruled out through histology in all the patients and two out of seven had a history of occasional proton pump inhibitor use. An unusual association was found between CD and hyperplastic polyps in this case series.

Immune response against Streptococcus gallolyticus in patients with adenomatous polyps in colon.

Our aim was to examine the humoral immune response against Streptococcus gallolyticus subspecies gallolyticus antigens in individuals subjected to a routine colonoscopy in which colon adenomatous polyps were present or not. Serum samples from 133 individuals with adenomatous polyps and serum samples from 53 individuals with a normal colonoscopy were included. Western blot was performed in all subjects using a whole cell antigen from S. gallolyticus ATCC 9809, and rabbit antisera against the whole cell bacteria was prepared as a control. By analyzing the immune profile of the rabbit-immunized sera by Western-blot, at least 22 proteins were identified as immunogenic in S. gallolyticus. When we evaluated sera from human subjects, two proteins of approximately 30 and 22 kDa were most prominent. Based on this 2-protein band pattern, Western-blot profiles from human subjects were compared. The detection of a protein band of 22 kDa was associated with the presence of adenomatous polyps in colon [odds ratios (OR) 7.98, 95% confidence intervals (CI): 3.54-17.93], p < 0.001. When the presence of the 30 kDa protein alone or both the 22 and 30 kDa proteins were analyzed, the OR increased to 22.37 (95% CI: 3.77-131.64), p < 0.001. The specificity was 84.9 for the presence of the 22 kDa protein, and 98.1 for the presence of the 30 kDa protein alone or both 22 and 30 kDa bands. Serum from individuals with adenomatous polyps recognized two proteins from S. gallolyticus. This result confirmed the possible association of S. gallolyticus with adenomatous polyps in the colon.

Study of the expression of toll-like receptors in different histological types of colorectal polyps and their relationship with colorectal cancer.

PURPOSE: Dysregulation of toll-like receptors (TLR) signaling can result in chronic inflammatory and over-exuberant repair responses. The aim of this study was to investigate the expression and clinical relevance of TLR in colorectal polyps. METHODS: The expression levels of six TLR were analyzed in 70 patients with different histological types of colorectal polyps, 38 of which developed colorectal carcinoma (CC). These analyses were performed by real-time polymerase chain reaction, western blot, and immunohistochemistry. RESULTS: TLR9 expression was higher in hyperplastic or adenomatous polyps compared to other polyp types. Hyperplastic polyps also showed increased TLR7 levels compared to the other polyp types. TLR7 expression was lower in both hyperplastic and tubulovillous adenoma polyps from patients who developed CC. TLR9 expression was decreased in hyperplastic and villous polyps from patients who developed CC. CONCLUSIONS: Our findings suggest a possible protective role of TLR expression against malignant transformation in the colorectal mucosa. TLR may represent a pathological marker of CC risk in colorectal polyps. The role of these factors in the pathology of colorectal polyps deserves further investigation.

A therapeutic cancer vaccine targeting carcinoembryonic antigen in intestinal carcinomas.

A genetic vaccine platform based on DNA electroporation (DNA-EP) and adenovirus (Ad) was used to generate immune response against human carcinoembryonic antigen (CEA) and antitumor effects in murine models with spontaneous tumors arising in an orthotopic location. CEA transgenic (CEA.Tg) mice treated with the carcinogen 1,2-dimethylhydrazine developed CEA-overexpressing tumors that resembled human sporadic colorectal cancer. APC1638N/CEA hybrid mice, generated by crossing mice carrying the adenomatous polyposis coli (Apc1638N) gene mutation with CEA.Tg mice, are representative of human familial polyposis and develop polyps that overexpress the antigen. In both models, the DNA-EP/Ad vaccine succeeded in breaking immune tolerance and achieved significant antitumor effects in therapeutic settings. Our data suggest that genetic vaccines targeting CEA may be feasible strategies against gut tumors that overexpress the antigen. In addition, these models are powerful systems for evaluating antigen-specific tumor immunity and assessing therapeutic vaccine strategies for human colorectal cancer.

Colorectal polyposis and immune-based therapies.

The progression from precancerous (adenomatous) colon polyps to malignant colorectal cancer involves the complex actions of various cytokines on T cell proliferation, cell-cell adhesion, apoptosis and host immunity. A broad spectrum of new treatments, including innovative molecular therapies such as gene therapy and treatment with cytokines, is under experimental and preclinical investigation. Nonsteroidal anti-inflammatory drugs and selective cyclooxygenase-2 inhibitors have traditionally been used as inflammation-reducing agents in cases of colon adenoma. Currently, adjuvant immunotherapies such as recombinant gene therapy and antibody-cytokine fusion proteins are assuming a more significant role in the management of colorectal neoplasia. Furthermore, advances in antitumour necrosis factor antibodies for the treatment of ulcerative colitis and Crohn's disease may have potential as chemoprotective agents for the treatment of colon polyposis. The present review aims to discuss the immunological mechanisms underlying colon tumour progression and the molecular and immune-based therapies that are leading to new methods of prognosis and treatment.

Colon polyps and cytokines: emerging immunological mechanisms.

Cell-mediated immune responses are an essential aspect of tumour-host interactions in colorectal cancers. The progression from precancerous (adenomatous) colon polyps to malignant colorectal cancer depends on a complex pathway involving the activities of activated T lymphocytes. The immune response is initiated when either cytotoxic T lymphocyte (CTL) CD8+ cells or CD4+ T-helper cells recognize the antigen from a human cancer cell. The cell-mediated response is largely initiated and controlled by the actions of various cytokines, which exert profound effects on T cell proliferation, cell-cell adhesion, apoptosis, and host immunity. The existence of an immune response to colon cancer is supported by studies of immunological treatments in humans and transplantable murine cancer models in animals. IL-2, IL-12, IFN-gamma, TNF-alpha, and TRAIL are implicated in enhancing cytotoxic and apoptotic effects in response to colon adenomas. In addition, growth factors, oncogenic cytokines and immunosuppressive factors may play a crucial role in the growth and survival of premaligant colonic tissue. This review aims to increase knowledge of the immunological mechanisms underlying colon tumour progression in the hopes that a greater understanding of the molecular pathways will lead to improved methods of prognosis and treatment.

The prognostic value of cell proliferation in colorectal adenomas assessed with tritiated thymidine and anti-proliferating cell nuclear antigen.

We analyze the cell kinetics of colorectal adenomas by tritiated thymidine (3HTdR) autoradiographic method and anti-proliferating cell nuclear antigen (PCNA) antibodies. A total of 46 patients who underwent prior endoscopic polypectomy for colorectal adenomas were reevaluated by colonoscopy for 4 years. Thymidine labeling index (T-LI) in index adenomas ranged from 1.40 to 38.0% (median value: 10. 75%); PCNA labeling index (PCNA-LI) in index adenomas ranged from 0 to 27.0% (median value: 1.95%). Among the 46 patients studied, 16 developed recurrent adenomas (Group A) and 30 were free of recurrent adenomas (Group B). The T-LI and PCNA-LI comparisons between Groups A and B were statistically significant (p < 0.0001, chi2 test). These results demonstrate that T-LI and PCNA-LI in colorectal adenomas might be helpful to predict the development of metachronous adenomas and hence to plan the follow-up of patients with adenomatous polyps after polypectomy.

Increased prevalence of colonic polyps and altered lymphocyte subset pattern in the colonic lamina propria in acromegaly.

OBJECTIVE: The balance of evidence suggests that acromegaly is a risk factor for colonic neoplasia. We have evaluated the prevalence of colonic polyps in acromegalics from Southern Italy and characterized the lymphocyte subsets in the colonic lamina propria in order to analyze differences in the colonic immunological environment. DESIGN: All the patients and controls were submitted to pancolonoscopy. Ten per-endoscopic biopsies of the intestinal mucosa surrounding polyps were carried out to evaluate lymphocyte subsets. PATIENTS: Fifty acromegalics and 318 sex- and age-matched controls entered this study. Colonic lamina propria lymphocyte subsets were studied in 34 patients and 34 controls. RESULTS: Colonic polyps were resected in 23 acromegalics (46%) and 42 controls (13.2%; P < 0.0001); hyperplastic polyps were found in 24% and 6.3%, adenomatous polyps in 22 and 6.9%, (P < 0.01), adenocarcinoma in 2 and 1.2% while synchronous polyps occurred in 18% and 2.5% (P < 0.01), respectively. The number of polyps was significantly correlated with age both in acromegalics (r = 0.422, P < 0.005) and in controls (r = 0.865, P < 0.001). However, polyp prevalence was greater in patients aged below 40 yrs (r.r = 1.9) and in patients with two or more skin tags (r.r = 1.2). A significant decrease of CD20, CD19, CD16, gamma/delta, CD4@leu8- and increase of CD3 and CD4+/leu8+ was found in the lamina propria lymphocyte subsets. CONCLUSIONS: The results of this study confirm that acromegalics are at increased risk of colonic polyps compared to the healthy population. The increased prevalence of premalignant polyps, namely the adenomatous type, suggests that acromegalics should undergo a careful screening and follow-up by pancolonoscopy. An impairment of mucosal immune surveillance seems to exist in acromegaly although a causal effect in the polyp formation cannot be ruled out.

A new murine monoclonal antibody, CMU10, as a marker for colonic carcinoma and precancerous conditions.

OBJECTIVE: To investigate the usefulness of a cancer-associated mucin antigen using a new monoclonal antibody, CMU10, as a tumor marker. Expression was assessed in normal adult, fetal, hyperplastic, preneoplastic, and neoplastic tissues of colon. DESIGN: CMU10 monoclonal antibody recognizes a mucin antigen that is not expressed in normal fetal and adult large intestine, but is rather commonly expressed in cancerous and precancerous lesions. Immunocytochemical analysis was performed on human tissues obtained at surgery or at autopsy. RESULTS: Expression of this mucin antigen was observed in 27 (98.4%) of 28 cases of colorectal carcinoma, 24 (96%) of 25 cases of adenomatous polyps, 9 (75%) of 12 cases of ulcerative colitis, and 9 (100%) of 9 cases of hyperplastic polyps. In cancerous specimens, the antigen distribution was mainly localized in the secretory mucin and surface membrane, whereas in precancerous lesions it was located in dysplastic, dilated, and distorted crypts. CONCLUSIONS: Because CMU10 differentially recognizes an antigen in cancerous and precancerous tissues, but not in normal tissues, it may be useful as a tumor marker for immunodiagnosis and, hence, early detection.

HLA-DR antigen expression in colonic adenomas and adenocarcinomas.

Previous studies have shown that malignant transformation is sometimes associated with the aberrant expression of HLA class II antigens. The functional significance of such aberrant expression is not known. Since HLA-DR antigen is expressed in normal colonic mucosa, it would be interesting to see if malignant transformation could result in the aberrant suppression of this antigen. Sixteen colonic adenocarcinomas, 29 colonic adenomatous polyps and 23 samples of normal colonic mucosa, including 9 cases of colonic mucosa adjacent to carcinoma, were stained immunohistochemically for HLA-DR antigen. The intensity and distribution of the antigen staining in the cytoplasm and luminal surface of the epithelial elements were analysed semiquantitatively. The lymphoplasmacytic infiltrate in the lamina propria was also evaluated. Cytoplasmic HLA-DR antigen expression was found to be significantly diminished in moderately and poorly differentiated adenocarcinomas but not in adenomas or well-differentiated adenocarcinomas. This suggests that the change in HLA-DR expression is not intrinsic to the neoplastic process but may merely be due to the fact that malignant cells, as they become less differentiated, tend to show alterations in their antigenic phenotype.

CD44 is associated with proliferation in normal and neoplastic human colorectal epithelial cells.

Flash-frozen biopsies obtained from surgical specimens of three adenomatous polyps and 22 colorectal adenocarcinomas (19 primary and three metastatic) were tested by immunohistochemistry for CD44 expression using F10-44-2 monoclonal antibody. CD44 positivity was correlated with proliferative status defined by Ki-67 monoclonal antibody reactivity. In normal colonic mucosa, CD44 was expressed in the proliferative zone of crypts. In tumours, CD44 expression was associated with proliferative areas irrespective of tumour stage or differentiation. Non-proliferating areas of the carcinomatous epithelium did not express CD44 although non-proliferating stromal lymphoid tissue did. There was no apparent association with tumour progression. F10-44-2-defined CD44 is consistently expressed during proliferation by normal colorectal epithelial cells and by both benign and malignant colorectal tumour cells.