The prognostic and clinicopathological roles of microsatellite instability, PD-L1 expression and tumor-infiltrating leukocytes in familial adenomatous polyposis.

BACKGROUND: Microsatellite instability, programmed death-ligand 1 and tumor-infiltrating leukocytes are prognostic biomarkers in colorectal cancer but unknown toward familial adenomatous polyposis. AIM: To investigate the prognostic and clinicopathological roles of microsatellite instability, programmed death-ligand 1 and tumor-infiltrating leukocytes in familial adenomatous polyposis. METHODS: Clinical data and paraffin embedded tissues from 45 familial adenomatous polyposis patients were collected. Microsatellite instability was detected by immunohistochemistry and polymerase chain reaction. Programmed death-ligand 1 was detected by immunohistochemistry. Tumor-infiltrating leukocytes comprising CD8+ T cells, M1 and M2 tumor associated macrophages, CD56bright and CD56dim natural killer cells were analyzed using multiple fluorescence immunohistochemistry. RESULTS: Microsatellite instability high was noted in 6 samples but not associated with overall survival or progression-free survival. Programmed death-ligand 1 is negative on tumor cells but positive on tumor-infiltrating leukocytes, and positive programmed death-ligand 1 expression on tumor-infiltrating leucocytes is associated with overall survival. Low CD56bright natural killer cell infiltration was associated with longer progression-free survival and was an independent prognostic factor in FAP. CONCLUSION: For familial adenomatous polyposis, microsatellite instability high can be found but has no correlation with prognosis; programmed death-ligand 1 on tumor-infiltrating leukocytes is related with overall survival; CD56bright natural killer cell is an independent prognostic factor associating with longer progression-free survival.

The Adaptive Immune Landscape of the Colorectal Adenoma-Carcinoma Sequence.

BACKGROUND: The tumor immune microenvironment exerts a pivotal influence in tumor initiation and progression. The aim of this study was to analyze the immune context of sporadic and familial adenomatous polyposis (FAP) lesions along the colorectal adenoma-carcinoma sequence (ACS). METHODS: We analyzed immune cell counts (CD3+, CD4+, CD8+, Foxp3+, and CD57+), tumor mutation burden (TMB), MHC-I expression and PD-L1 expression of 59 FAP and 74 sporadic colorectal lesions, encompassing adenomas with low-grade dysplasia (LGD) (30 FAP; 30 sporadic), adenomas with high-grade dysplasia (22 FAP; 30 sporadic), and invasive adenocarcinomas (7 FAP; 14 sporadic). RESULTS: The sporadic colorectal ACS was characterized by (1) a stepwise decrease in immune cell counts, (2) an increase in TMB and MHC-I expression, and (3) a lower PD-L1 expression. In FAP lesions, we observed the same patterns, except for an increase in TMB along the ACS. FAP LGD lesions harbored lower Foxp3+ T cell counts than sporadic LGD lesions. A decrease in PD-L1 expression occurred earlier in FAP lesions compared to sporadic ones. CONCLUSIONS: The colorectal ACS is characterized by a progressive loss of adaptive immune infiltrate and by the establishment of a progressively immune cold microenvironment. These changes do not appear to be related with the loss of immunogenicity of tumor cells, or to the onset of an immunosuppressive tumor microenvironment.

TCF-1 controls Treg cell functions that regulate inflammation, CD8+ T cell cytotoxicity and severity of colon cancer.

The transcription factor TCF-1 is essential for the development and function of regulatory T (Treg) cells; however, its function is poorly understood. Here, we show that TCF-1 primarily suppresses transcription of genes that are co-bound by Foxp3. Single-cell RNA-sequencing analysis identified effector memory T cells and central memory Treg cells with differential expression of Klf2 and memory and activation markers. TCF-1 deficiency did not change the core Treg cell transcriptional signature, but promoted alternative signaling pathways whereby Treg cells became activated and gained gut-homing properties and characteristics of the TH17 subset of helper T cells. TCF-1-deficient Treg cells strongly suppressed T cell proliferation and cytotoxicity, but were compromised in controlling CD4+ T cell polarization and inflammation. In mice with polyposis, Treg cell-specific TCF-1 deficiency promoted tumor growth. Consistently, tumor-infiltrating Treg cells of patients with colorectal cancer showed lower TCF-1 expression and increased TH17 expression signatures compared to adjacent normal tissue and circulating T cells. Thus, Treg cell-specific TCF-1 expression differentially regulates TH17-mediated inflammation and T cell cytotoxicity, and can determine colorectal cancer outcome.

Intestinal Epithelial TBK1 Prevents Differentiation of T-helper 17 Cells and Tumorigenesis in Mice.

BACKGROUND & AIMS: Intestinal epithelial cells (IECs) regulate intestinal immune cells, particularly development of T-helper 17 (Th17) cells. Deregulation of this process leads to intestinal inflammation and tumorigenesis, via unknown mechanisms. TANK-binding kinase 1 (TBK1) is expressed by IECs and cells in the innate immune system. We studied the functions of TBK1 in the intestinal immune response and tumorigenesis in mice. METHODS: We performed studies of wild-type mice, mice with conditional disruption of Tbk1 (Tbk1IEC-KO), Tbk1IEC-KO mice crossed with ApcMin/+ mice, and Mt-/- mice crossed with ApcMin/+ mice. Some mice were given intraperitoneal injections of a neutralizing antibody against interleukin 17 (IL17) or IL1ß. Intestine tissues were collected from mice and analyzed by histology, for numbers of adenomas and Th17 cells, and expression of inflammatory cytokines by real-time PCR. IECs were isolated from wild-type and Tbk1IEC-KO mice, stimulated with lipopolysaccharide, co-cultured for with bone marrow-derived macrophages, and analyzed by RNA sequencing and biochemical analyses. RESULTS: Compared to ApcMin/+Tbk1WT mice, ApcMin/+Tbk1IEC-KO mice had significant increases in number and size of intestinal polyps, and significantly more Th17 cells in lamina propria. Administration of an antibody against IL17 reduced the number of intestinal polyps in ApcMin/+Tbk1IEC-KO mice to that observed in ApcMin/+Tbk1WT mice. In culture, TBK1-deficient IECs promoted expression of IL1ß by macrophages, which induced differentiation of naïve CD4+ T cells into Th17 cells. RNA sequencing analysis revealed that the TBK1-deficient IECs had increased expression of metallothionein 1 (MT1), an immune regulator that promotes intestinal inflammation. Intestine tissues from ApcMin/+Mt-/- mice had significant fewer Th17 cells than ApcMin/+Mt+/+ mice, and a significantly lower number of polyps. Analyses of colorectal tumors in the Cancer Genome Atlas found colorectal tumors with high levels of MT1 and IL17 mRNAs to be associated with reduced survival times of patients. CONCLUSIONS: Expression of TBK1 by IECs suppresses expression of MT1 and prevents expression of IL1ß by macrophages and differentiation of Th17 cells, to prevent inflammation and tumorigenesis. Strategies to block this pathway might be developed for colorectal tumorigenesis.

Adenomatous Polyposis Coli Defines Treg Differentiation and Anti-inflammatory Function through Microtubule-Mediated NFAT Localization.

Adenomatous polyposis coli (APC) is a polarity regulator and tumor suppressor associated with familial adenomatous polyposis and colorectal cancer development. Although extensively studied in epithelial transformation, the effect of APC on T lymphocyte activation remains poorly defined. We found that APC ensures T cell receptor-triggered activation through Nuclear Factor of Activated T cells (NFAT), since APC is necessary for NFAT's nuclear localization in a microtubule-dependent fashion and for NFAT-driven transcription leading to cytokine gene expression. Interestingly, NFAT forms clusters juxtaposed with microtubules. Ultimately, mouse Apc deficiency reduces the presence of NFAT in the nucleus of intestinal regulatory T cells (Tregs) and impairs Treg differentiation and the acquisition of a suppressive phenotype, which is characterized by the production of the anti-inflammatory cytokine IL-10. These findings suggest a dual role for APC mutations in colorectal cancer development, where mutations drive the initiation of epithelial neoplasms and also reduce Treg-mediated suppression of the detrimental inflammation that enhances cancer growth.

SMAD4 impedes the conversion of NK cells into ILC1-like cells by curtailing non-canonical TGF-ß signaling.

Among the features that distinguish type 1 innate lymphoid cells (ILC1s) from natural killer (NK) cells is a gene signature indicative of 'imprinting' by cytokines of the TGF-ß family. We studied mice in which ILC1s and NK cells lacked SMAD4, a signal transducer that facilitates the canonical signaling pathway common to all cytokines of the TGF-ß family. While SMAD4 deficiency did not affect ILC1 differentiation, NK cells unexpectedly acquired an ILC1-like gene signature and were unable to control tumor metastasis or viral infection. Mechanistically, SMAD4 restrained non-canonical TGF-ß signaling mediated by the cytokine receptor TGFßR1 in NK cells. NK cells from a SMAD4-deficient person affected by polyposis were also hyper-responsive to TGF-ß. These results identify SMAD4 as a previously unknown regulator that restricts non-canonical TGF-ß signaling in NK cells.

Spontaneous Intestinal Tumorigenesis in Apc (/Min+) Mice Requires Altered T Cell Development with IL-17A.

The control of inflammatory diseases requires functional regulatory T cells (Tregs) with significant Gata-3 expression. Here we address the inhibitory role of Tregs on intestinal tumorigenesis in the Apc (/Min+) mouse model that resembles human familial adenomatous polyposis (FAP). Apc (/Min+) mice had a markedly increased frequency of Foxp3+ Tregs and yet decreased Gata-3 expression in the lamina propria. To address the role of heterozygous Apc gene mutation in Tregs, we generated Foxp3-Cre, Apc (flox/+) mice. Tregs from these mice effectively inhibited tumorigenesis comparable to wild type Tregs after adoptive transfer into Apc (/Min+) mice, demonstrating that the heterozygous Apc gene mutation in Tregs does not induce the loss of control over tumor microenvironment. Adoptive transfer of in vitro generated Apc (/Min+) iTregs (inducible Tregs) failed to inhibit intestinal tumorigenesis, suggesting that naïve CD4 T cells generated from Apc (/Min+) mice thymus were impaired. We also showed that adoptively transferred IL-17A-deficient Apc (/Min+) Tregs inhibited tumor growth, suggesting that IL-17A was critical to impair the tumor regression function of Apc (/Min+) Tregs. Taken together, our results suggest that both T cell development in a functional thymus and IL-17A control the ability of Treg to inhibit intestinal tumorigenesis in Apc (/Min+) mice.

Transcriptional changes between uninflamed ulcerative colitis and familial adenomatous polyposis pouch mucosa can be attributed to an altered immune response.

A total proctocolectomy with ileal pouch-anal anastomosis (IPAA) is considered the surgery of choice for definitive management of familial adenomatous polyposis (FAP) and some patients with ulcerative colitis (UC). However, this surgical treatment is often associated with pouchitis, a long-term complication that occurs mostly in UC patients. The purpose of this study was to better define the molecular background of pouchitis. A microarray-based survey was performed using pouch mucosal samples collected from 28 and 8 patients undergoing surgery for UC and FAP, respectively. There were 4,770 genes that significantly differentiated uninflamed from inflamed mucosal samples, and their functional features were represented mostly by metabolic and cell proliferation pathways. In contrast, functional analyses of aberrantly expressed genes between UC and FAP samples, irrespective of mucosal inflammation status, revealed multiple pathways and terms that were linked to changes in immune response. Interestingly, the comparison of uninflamed UC and FAP samples identified a set of 29 altered probe sets, including an inflammation-related transcript encoding a Charcot-Leyden crystal (CLC) protein. The most distinct changes in gene expression profiles differentiating uninflamed UC and FAP pouch mucosal samples were attributed to the Gene Ontology category innate immune response. Our study confirmed that alterations in immune responses can be found between patients who underwent surgery for UC and FAP, independent of the pouch inflammation status. This observation may be important when managing IPAA patients.

T-cell receptor sequencing reveals the clonal diversity and overlap of colonic effector and FOXP3+ T cells in ulcerative colitis.

BACKGROUND: FOXP3 regulatory T cell prevent inflammation but are paradoxically increased in ulcerative colitis (UC). Local T-cell activation has been hypothesized to account for increased FOXP3 expression in colon lamina propria (LP) T cells. METHODS: To see if human FOXP3 LP T cells are an activated fraction of otherwise FOXP3 effector T cells and explore their clonal diversity in health and disease, we deep sequenced clonally unique T-cell receptor hypervariable regions of FOXP3 and FOXP3CD4 T-cell subpopulations from inflamed versus noninflamed colon LP or mesenteric lymph nodes of patients with or without UC. RESULTS: The clonal diversity of each LP T-cell population was not different between patients with versus without UC. Repertoire overlap was only seen between a minority of FOXP3 and FOXP3 cells, including recently activated CD38 cells and Th17-like CD161 effector T cells, but this repertoire overlap was not different between patients with versus without UC and was no larger than the overlap between Helios and Helios FOXP3 cells. CONCLUSIONS: Thus, at steady state, only a minority of FOXP3, and particularly Helios, T cells share a T-cell receptor sequence with FOXP3 effector populations in the colon LP, even in UC, revealing distinct clonal origins for LP regulatory T cell and effector T cells in humans.

Efficient drug delivery and induction of apoptosis in colorectal tumors using a death receptor 5-targeted nanomedicine.

Death Receptor 5 (DR5) is a pro-apoptotic cell-surface receptor that is a potential therapeutic target in cancer. Despite the potency of DR5-targeting agents in preclinical models, the translation of these effects into the clinic remains disappointing. Herein, we report an alternative approach to exploiting DR5 tumor expression using antibody-targeted, chemotherapy-loaded nanoparticles. We describe the development of an optimized polymer-based nanotherapeutic incorporating both a functionalized polyethylene glycol (PEG) layer and targeting antibodies to limit premature phagocytic clearance whilst enabling targeting of DR5-expressing tumor cells. Using the HCT116 colorectal cancer model, we show that following binding to DR5, the nanoparticles activate caspase 8, enhancing the anti-tumor activity of the camptothecin payload both in vitro and in vivo. Importantly, the combination of nanoparticle-induced DR5 clustering with camptothecin delivery overcomes resistance to DR5-induced apoptosis caused by loss of BAX or overexpression of anti-apoptotic FLIP. This novel approach may improve the clinical activity of DR5-targeted therapeutics while increasing tumor-specific delivery of systemically toxic chemotherapeutics.

ASC-associated inflammation promotes cecal tumorigenesis in aryl hydrocarbon receptor-deficient mice.

The aryl hydrocarbon receptor (AhR) plays a suppressive role in cecal carcinogenesis by CUL4B/AhR-mediated ubiquitylation and degradation of ß-catenin, which is activated by xenobiotics and natural ligands. AhR-deficient (AhR(-)(/-)) mice develop cecal tumors with severe inflammation. To elucidate whether the tumors develop autonomously in AhR(-/-) mice due to impaired ß-catenin degradation or in association with accelerated inflammation, we performed two kinds of experiments using germ-free (GF) AhR(-/-) mice and compound mutant mice lacking genes for AhR and apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), which plays an essential role in caspase-1 activation in inflammasomes. Both GF AhR(-/-) and AhR(-/-)•ASC(-/-) mice showed considerably reduced tumor development compared with that in AhR(-/-) mice albeit in a 'cancer-prone' state with aberrant ß-catenin accumulation. Blocking of the interleukin (IL)-1ß signaling pathway by treatment with a caspase-1 inhibitor, YVAD, reduced cecal tumorigenesis in AhR(-/-) mice. Signal transducers and activators of transcription 3 (STAT3) activation was detected in the cecal epithelium of the AhR(-/-) mice due to enhanced IL-6 production. An inhibitor of the STAT3 signaling pathway, AG490 suppressed the tumor formation. ASC-mediated inflammation was also found to play a critical role in tumor development in Apc(Min/+) mice, a mouse model of familial adenomatous polyposis. Collectively, these results revealed an important role of the bacteria-triggered or ASC-mediated inflammation signaling pathway in the intestinal tumorigenesis of mice and suggest a possible chemical therapeutic intervention, including AhR ligands and inhibitors of the inflammation pathway.

Synergistic effect of interleukin-10-receptor variants in a case of early-onset ulcerative colitis.

AIM: To investigated the molecular cause of very early-onset ulcerative colitis (UC) in an 18-mo-old affected child. METHODS: We analysed the interleukin-10 (IL10) receptor genes at the DNA and RNA level in the proband and his relatives. Beta catenin and tumor necrosis factor-α (TNFα) receptors were analysed in the proteins extracted from peripheral blood cells of the proband, his relatives and familial adenomatous polyposis (FAP) and PTEN hamartoma tumor syndrome (PHTS) patients. Samples were also collected from the proband's inflamed colorectal mucosa and compared to healthy and tumour mucosa collected from a FAP patient and patients affected by sporadic colorectal cancer (CRC). Finally, we examined mesalazine and azathioprine effects on primary fibroblasts stabilised from UC and FAP patients. RESULTS: Our patient was a compound heterozygote for the IL10RB E47K polymorphism, inherited from his father, and for a novel point mutation within the IL10RA promoter (the -413G->T), inherited from his mother. Beta catenin and tumour necrosis factor α receptors-I (TNFRI) protein were both over-expressed in peripheral blood cells of the proband's relatives more than the proband. However, TNFRII was over-expressed only in the proband. Finally, both TNFα-receptors were shown to be under-expressed in the inflamed colon mucosa and colorectal cancer tissue compared to healthy colon mucosa. Consistent with this observation, mesalazine and azathioprine induced, in primary fibroblasts, IL10RB and TNFRII over-expression and TNFRI and TNFα under-expression. We suggest that ß-catenin and TNFRI protein expression in peripheral blood cells could represent molecular markers of sub-clinical disease in apparently healthy relatives of patients with early-onset UC. CONCLUSION: A synergistic effect of several variant alleles of the IL10 receptor genes, inherited in a Mendelian manner, is involved in UC onset in this young child.

Abating colon cancer polyposis by Lactobacillus acidophilus deficient in lipoteichoic acid.

An imbalance of commensal bacteria and their gene products underlies mucosal and, in particular, gastrointestinal inflammation and a predisposition to cancer. Lactobacillus species have received considerable attention as examples of beneficial microbiota. We have reported previously that deletion of the phosphoglycerol transferase gene that is responsible for lipoteichoic acid (LTA) biosynthesis in Lactobacillus acidophilus (NCK2025) rendered this bacterium able to significantly protect mice against induced colitis when delivered orally. Here we report that oral treatment with LTA-deficient NCK2025 normalizes innate and adaptive pathogenic immune responses and causes regression of established colonic polyps. This study reveals the proinflammatory role of LTA and the ability of LTA-deficient L. acidophilus to regulate inflammation and protect against colonic polyposis in a unique mouse model.

Inherited Syndromes Predisposing to Inflammation and GI Cancer.

Cancers arising within the gastrointestinal (GI) tract are commonly associated with an immune component at their inception and later in their maintenance. While many of the immune factors and immune cell types surrounding these lesions have been highlighted, the underlying pre-dispositions in immunesupported carcinogenesis are not well characterised. Inherited Mendelian GI disorders such as polyposis syndromes, while classically due to germline mutations in non-immune genes, commonly demonstrate alterations in key immune and inflammatory genes. In some cases immune based therapies have been shown to provide at least some benefit in animal models of these syndromes. The advent of genome wide association studies has begun to powerfully examine the genetic nature of complex non-Mendelian GI diseases highlighting polymorphisms within immune related genes and their potential to provide the niche in which GI cancers may originate. Here in the role in which Mendelian and non-Mendelian genetics of immune related factors supporting GI malignancy will be presented and discussed.

Activation of signal transducer and activator of transcription-1 (STAT-1) and differential expression of interferon-gamma and anti-inflammatory proteins in pelvic ileal pouches for ulcerative colitis and familial adenomatous polyposis.

Pouchitis after total rectocolectomy is the most common complication of ulcerative colitis (UC). The immunological mechanisms involved in the genesis of pouchitis are unclear. Therefore, we evaluated the inflammatory activity in normal ileal pouch mucosa by determining signal transducers and activators of transcription (STAT-1) activation and cytokine expression in patients operated for UC and familial adenomatous polyposis (FAP). Eighteen asymptomatic patients, who underwent total rectocolectomy and J pouch, were evaluated: nine with UC and nine with FAP. The activation of STAT-1 and cytokine expression were determined by immunoblot of total protein extracts from pouch mucosal biopsies. The absence of pouchitis was assessed by clinical, histological and endoscopic parameters, according to the Pouchitis Disease Activity Index. The patients were not receiving any medication. Analysis of variance (anova) and Tukey-Kramer's test were applied. The local ethical committee approved the study and informed consent was signed by all participants. STAT-1 activation was increased in UC when compared to FAP and controls (P < 0.05). Higher levels of interferon (IFN)-gamma expression were observed in UC patients when compared to the control group (P < 0.05), but were similar to FAP. In contrast, cytokine signalling (SOCS-3) and interleukin (IL)-10 expression were similar in all groups (P > 0.05). These findings could explain the higher susceptibility to this inflammatory complication in UC when compared to FAP. A tendency towards increased levels of IFN-gamma and STAT-1 in patients with UC, even without clinical and endoscopic evidence of pouchitis, was observed; studying inflammatory activity in asymptomatic ileal pouches may help understanding of the pathogenesis of pouchitis.

T-regulatory cells shift from a protective anti-inflammatory to a cancer-promoting proinflammatory phenotype in polyposis.

T-regulatory (Treg) cells play a major role in cancer by suppressing protective antitumor immune responses. A series of observations (from a single laboratory) suggest that Treg cells are protective in cancer by virtue of their ability to control cancer-associated inflammation in an interleukin (IL)-10-dependent manner. Here, we report that the ability of Treg cells to produce IL-10 and control inflammation is lost in the course of progressive disease in a mouse model of hereditary colon cancer. Treg cells that expand in adenomatous polyps no longer produce IL-10 and instead switch to production of IL-17. Aberrant Treg cells from polyp-ridden mice promote rather than suppress focal mastocytosis, a critical tumor-promoting inflammatory response. The cells, however, maintain other Treg characteristics, including their inability to produce IL-2 and ability to suppress proliferation of stimulated CD4 T cells. By promoting inflammation and suppressing T-helper functions, these cells act as a double-edged knife propagating tumor growth.

Polyps wrap mast cells and Treg within tumorigenic tentacles.

Gounaris and colleagues describe a previously unrecognized cross-talk between mast cells and Treg in colon adenomatous polyposis (Gounaris et al., Cancer Res 2009;69:5490-7). Adoptively transferred Treg suppress the focal mastocytosis that fosters tumor initiation and progression. In contrast, endogenous Treg, which abundantly infiltrate polyps, show proinflammatory activity under unknown microenvironmental cues that promote mast cell differentiation and expansion. Compartmentalized Treg plasticity seems to be a key factor in establishing the optimal milieu for cancer development in the intestines. Treg partnership with mast cells recapitulates the complexity of innate-adaptive networks characterizing gut inflammation and represents a novel target for cancer immunotherapy.

Colorectal carcinomas in MUTYH-associated polyposis display histopathological similarities to microsatellite unstable carcinomas.

BACKGROUND: MUTYH-associated polyposis (MAP) is a recessively inherited disorder which predisposes biallelic carriers for a high risk of polyposis and colorectal carcinoma (CRC). Since about one third of the biallelic MAP patients in population based CRC series has no adenomas, this study aimed to identify specific clinicopathological characteristics of MAP CRCs and compare these with reported data on sporadic and Lynch CRCs. METHODS: From 44 MAP patients who developed > or = 1 CRCs, 42 of 58 tumours were analyzed histologically and 35 immunohistochemically for p53 and beta-catenin. Cell densities of CD3, CD8, CD57, and granzyme B positive lymphocytes were determined. KRAS2, the mutation cluster region (MCR) of APC, p53, and SMAD4 were analyzed for somatic mutations. RESULTS: MAP CRCs frequently localized to the proximal colon (69%, 40/58), were mucinous in 21% (9/42), and had a conspicuous Crohn's like infiltrate reaction in 33% (13/40); all of these parameters occurred at a higher rate than reported for sporadic CRCs. Tumour infiltrating lymphocytes (TILs) were also highly prevalent in MAP CRCs. Somatic APC MCR mutations occurred in 14% (5/36) while 64% (23/36) had KRAS2 mutations (22/23 c.34G>T). G>T transversions were found in p53 and SMAD4, although the relative frequency compared to other mutations was low. CONCLUSION: MAP CRCs show some similarities to micro-satellite unstable cancers, with a preferential proximal location, a high rate of mucinous histotype and increased presence of TILs. These features should direct the practicing pathologist towards a MAP aetiology of CRC as an alternative for a mismatch repair deficient cause. High frequent G>T transversions in APC and KRAS2 (mutated in early tumour development) but not in P53 and SMAD4 (implicated in tumour progression) might indicate a predominant MUTYH effect in early carcinogenesis.

Intrarectal vaccination with recombinant vaccinia virus expressing carcinoembronic antigen induces mucosal and systemic immunity and prevents progression of colorectal cancer.

The gastrointestinal mucosa contains an intact immune system that protects the host from pathogens and communicates with the systemic immune system. Absorptive epithelial cells in the mucosa give rise to malignant tumors although the interaction between tumor cells and the mucosal immune system is not well defined. The pathophysiology of colorectal cancer has been elucidated through studies of hereditary syndromes, such as familial adenomatous polyposis, a cancer predisposition syndrome caused by germline mutations in the adenomatous polyposis coli tumor suppressor gene. Patients with FAP develop adenomas and inevitably progress to invasive carcinomas by the age of 40. To better delineate the role of mucosal immunity in colorectal cancer, we evaluated the efficacy of intrarectal recombinant vaccinia virus expressing the human carcinoembryonic Ag (CEA) in a murine FAP model in which mice are predisposed to colorectal cancer and also express human CEA in the gut. Mucosal vaccination reduced the incidence of spontaneous adenomas and completely prevented progression to invasive carcinoma. The therapeutic effects were associated with induction of mucosal CEA-specific IgA Ab titers and CD8(+) CTLs. Mucosal vaccination was also associated with an increase in systemic CEA-specific IgG Ab titers, CD4(+) and CD8(+) T cell responses and resulted in growth inhibition of s.c. implanted CEA-expressing tumors suggesting communication between mucosal and systemic immune compartments. Thus, intrarectal vaccination induces mucosal and systemic antitumor immunity and prevents progression of spontaneous colorectal cancer. These results have implications for the prevention of colorectal cancer in high-risk individuals.