Truncating Adenomatous Polyposis Coli Research Articles

APC truncating mutations in Middle Eastern Population: Tankyrase inhibitor is an effective strategy to sensitize APC mutant CRC To 5-FU chemotherapy

Colorectal Cancer (CRC) is highly heterogeneous for which prognosis is dependent mainly on clinical staging. There is a need to stratify subpopulations of CRC on molecular basis to better predict outcome and therapy response. Truncating mutations in adenomatous polyposis coli (APC) are well-described events in CRC carcinogenesis. Clinical and genotypic characterization of Middle Eastern CRC based on presence and type of APC was determined in 412 CRC tumors using modern next generation sequencing. APC truncating mutations were identified in 58.2% (240/412) of CRCs. Overall, mutation was significant predictor of superior overall survival. Further, the type of APC mutations (short or long) did not have impact on clinical outcome. However, in vitro analysis showed difference between CRC cell lines carrying short truncating APC vs CRC cells that carry long truncating APC mutation in response to 5-flourouracil (5-FU). Importantly, we were able to overcome the resistance to 5-FU seen in CRC cells carrying short APC by tankyrase inhibitor, XAV939, thereby inhibiting Wnt/β-catenin signaling cascade. Overall, our results showed that APC mutation status plays an important role in predicting overall survival in Middle Eastern population. Furthermore, in vitro data showed that selective targeting of APC mutated CRC by tankyrase inhibitor can be an effective strategy to overcome 5-FU resistance in CRC cells.

Abstract 3447: APC truncating mutations in Middle Eastern population: Tankyrase inhibitor is an effective strategy to sensitize APC mutant CRC to 5-FU chemotherapy

Abstract Colorectal Cancer (CRC) is the leading cause of cancer related death and the most common cancer affecting Saudi males. Mutation of the adenomatous polyposis coli (APC) gene is considered as the initiating step of transformation in familial and sporadic CRC. The prevalence and prognostic role of APC mutation spectra are not well established in Middle Eastern CRC. Next generation sequencing of 412 CRC tumors was performed to determine mutations in APC. Association with protein expression and clinico-pathological correlation was performed. APC truncating mutation is found at a frequency of 58.2% (240/412) and is significantly associated with superior overall survival, whereas no difference in survival between short truncating and long truncating APC was noted. To test the hypothesis that APC mutations might influence 5- Fluorouracil (5-FU) therapy, which is the first line chemotherapy used in CRC, we compared CRC cell lines and showed that those expressing truncated APC exhibit limited response to 5-FU, unlike cells which were APC wild type. In COLO-320 DM and HCT-15, APC mutated CRC cells, we were able to increase sensitivity of APC truncated cells to 5-FU by inhibiting Tankyrase (TNKs) which are members of PARP family, using XAV939. In conclusion, the study identified the prognostic value of APC truncating mutation in Middle Eastern ethnicity. In vitro data further showed that APC mutation can be used as a molecular biomarker to predict response to 5-FU. Furthermore, Tankyrase inhibitor XAV939 selectively targeting APC-mutated CRC cells can be an effective strategy to overcome 5-FU resistance in these cells. Note: This abstract was not presented at the meeting. Citation Format: Sandeep K. Parvathareddy, Abdul K. Siraj, Rong Bu, Prateeshkumar Poyil, Tariq Masoodi, Divya Sasidharan Padmaja, Rica Concepcion, Rafia Begum, Maria A. Sabido, Roxanne Melosantos, Nasser Al-Sanea, Fouad Al-Dayel, Khawla S. Al-Kuraya. APC truncating mutations in Middle Eastern population: Tankyrase inhibitor is an effective strategy to sensitize APC mutant CRC to 5-FU chemotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3447.

Cenani-Lenz syndrome and other related syndactyly disorders due to variants in LRP4, GREM1/FMN1, and APC: Insight into the pathogenesis and the relationship to polyposis through the WNT and BMP antagonistic pathways.

Cenani-Lenz (C-L) syndrome is characterized by oligosyndactyly, metacarpal synostosis, phalangeal disorganization, and other variable facial and systemic features. Most cases are caused by homozygous and compound heterozygous missense and splice mutations of the LRP4 gene. Currently, the syndrome carries one OMIM number (212780). However, C-L syndrome-like phenotypes as well as other syndactyly disorders with or without metacarpal synostosis/phalangeal disorganization are also known to be associated with specific LRP4 mutations, adenomatous polyposis coli (APC) truncating mutations, genomic rearrangements of the GREM1-FMN1 locus, as well as FMN1 mutations. Surprisingly, patients with C-L syndrome-like phenotype caused by APC truncating mutations have no polyposis despite the increased levels of β catenin. The LRP4 and APC proteins act on the WNT (wingless-type integration site family) canonical pathway, whereas the GREM-1 and FMN1 proteins act on the bone morphogenetic protein (BMP) pathway. In this review, we discuss the different mutations associated with C-L syndrome, classify its clinical features, review familial adenomatous polyposis caused by truncating APC mutations and compare these mutations to the splicing APC mutation associated with syndactyly, and finally, explore the pathophysiology through a review of the cross talks between the WNT canonical and the BMP antagonistic pathways.

Dietary acrylamide intake and the risk of colorectal cancer with specific mutations in KRAS and APC

Acrylamide, a probable human carcinogen, is present in heat-treated carbohydrate-rich foods. Epidemiological studies have not shown a clear association between acrylamide intake and colorectal cancer (CRC) risk. This may be due to the molecular heterogeneity in colorectal tumors, which was not taken into consideration before. Since the acrylamide metabolite glycidamide induces specific DNA mutations in rodents, we investigated whether acrylamide is associated with CRC risk characterized by mutations in Kirsten-ras (KRAS) and adenomatous polyposis coli (APC); key genes in colorectal carcinogenesis. This case-cohort analysis, within the Netherlands Cohort Study on diet and cancer, was based on 7.3 years of follow-up. Acrylamide intake was assessed with a food frequency questionnaire. Mutation analysis of codons 1286-1520 in exon 15 in APC and codons 12 and 13 in exon 1 in KRAS was performed on tumor tissue of 733 cases. Hazard ratios (HR) were calculated using Cox proportional hazards analysis. Among men, acrylamide intake was statistically significantly associated with an increased risk of particularly tumors with an activating KRAS mutation {HR fourth versus first quartile: 2.12 [95% confidence interval (CI): 1.16-3.87], P trend: 0.01}. Among women, acrylamide intake was statistically significantly associated with a decreased risk of particularly tumors with a truncating APC mutation (fourth versus first quartile: 0.47 (95% CI: 0.23-0.94), P trend: 0.02), but only in the highest quartile of intake. This is the first study to show that acrylamide might be associated with CRC with specific somatic mutations, differentially in men and women. More research is needed to corroborate or refute these findings.

Familial adenomatous polyposis and hypertension

In October, 2008, a 36-year-old man was referred to our unit with new onset of hypertension and adrenal hyperplasia. He had been diagnosed with familial adenomatous polyposis (FAP) in childhood, and had undergone a total colectomy at the age of 17 years. He had been under endoscopic surveillance since, with a recent polyp count of 50–60 (March). The next month, as part of his routine FAP screening programme, he had an abdominal CT scan, which showed bilateral macronodular adrenal hyperplasia. The largest nodule measured 4 cm by 2 cm on the right and 2·7 cm by 2·9 cm on the left (fi gure). On assessment in our unit, the patient was hypertensive at 150/100 mm Hg on lisinopril and amlodipine. Examination showed no clinical features suggestive of Cushing’s syndrome. Serum potassium concentration was normal at 3·8 mmol/L. Other biochemical investi gations excluded Cushing’s syndrome and phaeo chromocytoma. With confounding antihypertensive medications discontinued, plasma aldosterone measured 590 pmol/L, plasma renin activity (PRA) 0·2 pmol/mL·h, giving a ratio of aldosterone to PRA of 2950 (>2000 is highly suggestive of primary hyperaldosteronism). Saline infusion did not suppress aldosterone to below 280 pmol/L, further supporting this diagnosis. Selective adrenal vein sampling showed that the cortisol-corrected aldosterone ratio in the left adrenal vein was 3·05 (dominant) compared with a ratio of 0·74 in the right adrenal vein (non-dominant), with a ratio of 0·80 in the low inferior vena cava (peripheral). The dominant to non-dominant ratio at 4·15 was above the recommended cut-off of 4·0 for diagnosing unilateral aldosterone hypersecretion with 95% sensitivity and 100% specifi city. Furthermore, the non-dominant to peripheral ratio of >1 confi rmed that autonomous secretion of aldosterone was confi ned to the left. There were no radiological features of the right adrenal mass suggesting malignancy, apart from its size. With the fi nding of dominant left-sided aldosterone secretion, we decided that the patient would benefi t from laproscopic left adrenalectomy. When last seen in March, 2010, he was awaiting surgery, delayed pending investigation into duodenal polyps, another manifestation of FAP. FAP is associated with many extra-intestinal manifestations. Adrenal gland enlargement is even more common in patients with FAP than would be expected from the ascertainment bias due to frequent abdominal imaging. In a series of 738 FAP patients, 14 (7·4%) were found to have adrenal enlargement, compared with 0·6–3·4% in the general population. Of these, two were hypertensive and surgical removal of cortisol-secreting tumours was necessary. In a study of 30 FAP patients with adrenal adenomas, 3 needed adrenalectomy: one for a phaeochromocytoma; one for a non-functioning, malignant tumour; and one for an enlarging, non-functioning, benign adenoma. Thus, the majority of adrenal tumours associated with FAP are non-functioning and no treat ment is needed unless they enlarge. Diagnosis of those rare cases that are associated with hormone hypersecretion is important, because they are surgically treatable. Our case report describes aldosterone-secreting adrenal enlargement in a hypertensive patient with FAP. Recently, an association between tumour-suppressor APC (adenomatous polyposis coli) gene mutations and hyperaldosteronism has been proposed. Mice heterozygous for a truncating APC mutation had higher aldosterone concentrations than controls, with corresponding increases in urinary potassium excretion, despite the presence of hypertension and plasma volume expansion. These fi ndings suggest a potential pathophysiological link between the causative APC mutation and hyper aldosteronism in FAP. All FAP patients with incidentally discovered adrenal masses should have regular blood pressure measurements and be screened for possible Cushing’s syndrome, Conn’s syndrome, and phaeo chromo cytoma.

Dietary fat and risk of colon and rectal cancer with aberrant MLH1 expression, APC or KRAS genes

ObjectiveTo investigate baseline fat intake and the risk of colon and rectal tumors lacking MLH1 (mutL homolog 1, colon cancer, nonpolyposis type 2) repair gene expression and harboring mutations in the APC (adenomatous polyposis coli) tumor suppressor gene and in the KRAS (v-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog) oncogene.MethodsAfter 7.3 years of follow-up of the Netherlands Cohort Study (n = 120,852), adjusted incidence rate ratios (RR) and 95% confidence intervals (CI) were computed, based on 401 colon and 130 rectal cancer patients.ResultsTotal, saturated and monounsaturated fat were not associated with the risk of colon or rectal cancer, or different molecular subgroups. There was also no association between polyunsaturated fat and the risk of overall or subgroups of rectal cancer. Linoleic acid, the most abundant polyunsaturated fatty acid in the diet, was associated with increased risk of colon tumors with only a KRAS mutation and no additional truncating APC mutation or lack of MLH1 expression (RR = 1.41, 95% CI 1.18–1.69 for one standard deviation (i.e., 7.5 g/day) increase in intake, p-trend over the quartiles of intake <0.001). Linoleic acid intake was not associated with risk of colon tumors without any of the gene defects, or with tumors harboring aberrations in either MLH1 or APC.ConclusionLinoleic acid intake is associated with colon tumors with an aberrant KRAS gene, but an intact APC gene and MLH1 expression, suggesting a unique etiology of tumors with specific genetic aberrations.

Adenomatous polyposis coli‐deficient zebrafish are susceptible to digestive tract neoplasia

Truncation of the tumour suppressor adenomatous polyposis coli (APC) constitutively activates the Wnt/beta-catenin signalling pathway. This event constitutes the primary transforming event in sporadic colorectal cancer in humans. Moreover, humans or mice carrying germline truncating mutations in APC develop large numbers of intestinal adenomas. Here, we report that zebrafish that are heterozygous for a truncating APC mutation spontaneously develop intestinal, hepatic and pancreatic neoplasias that are highly proliferative, accumulate beta-catenin and express Wnt target genes. Treatment with the chemical carcinogen 7,12-dimethylbenz[a]anthracene accelerates the induction of these lesions. These observations establish apc-mutant zebrafish as a bona fide model for the study of digestive tract cancer.

Cigarette Smoking and Colorectal Cancer: APC Mutations, hMLH1 Expression, and GSTM1 and GSTT1 Polymorphisms

The contribution of cigarette smoking to sporadic colorectal cancer may differ according to molecular aspects of the tumor or according to glutathione S-transferase M1 (GSTM1) or glutathione S-transferase T1 (GSTT1) genotype. In the prospective Netherlands Cohort Study on Diet and Cancer, adjusted incidence rate ratios for 1986-1993 were computed for overall colorectal cancer, tumors with and without adenomatous polyposis coli (APC) mutations, and tumors with and without human mut-L homologue 1 (hMLH1) expression, according to cigarette smoking characteristics (661 cases, 2,948 subcohort members). Case-only analyses were performed to estimate odds ratios for interaction between cigarette smoking and GSTM1 and GSTT1 genotypes. In comparison with never smokers, a high smoking frequency increased the risk of colorectal cancer (for a five-cigarette/day increment, incidence rate ratio (IRR) = 1.07, 95% confidence interval (CI): 1.03, 1.12), and this association was stronger in 371 tumors without a truncating APC mutation (IRR = 1.11, 95% CI: 1.05, 1.17). Long-term smoking was associated with lack of hMLH1 expression in 56 tumors (for a 10-year increment, IRR = 1.17, 95% CI: 1.00, 1.37). No statistically significant interactions between smoking and GSTM1 or GSTT1 genotype were observed. These results indicate that cigarette smoking is associated with risk of colorectal cancer, and this association may depend on molecular characteristics of the tumor as defined by APC mutation and hMLH1 expression status.

TruncatingAPCmutations have dominant effects on proliferation, spindle checkpoint control, survival and chromosome stability

The majority of human tumour cells are aneuploid owing to an underlying chromosome instability phenotype. While the genetic lesions that cause chromosome instability remain undefined, mouse ES cells harbouring homozygous adenomatous polyposis coli (APC) mutations are frequently tetraploid. In addition, colon cancer cells with APC mutations have weakened kinetochore-microtubule interactions. Furthermore, mitotic spindles assembled in APC-depleted Xenopus egg extracts are aberrant. Therefore, to determine whether APC mutations can initiate chromosome instability in human cells, we expressed N-terminal APC fragments in HCT-116 cells, a near diploid colon cancer cell line with two wild-type APC alleles. We show that cells expressing N-APC mutants exit mitosis prematurely in the presence of spindle toxins, consistent with a spindle checkpoint defect. In addition, N-APC cells show enhanced survival following prolonged spindle damage. In contrast to controls, the N-APC survivors frequently contain dicentric chromosomes and then go on to become highly aneuploid. These observations suggest that truncating APC mutations can exert dominant effects which in turn can initiate chromosome instability. As such, APC mutation not only compromises tumour suppressor function but may also have oncogenic properties. We suggest therefore that the initial APC mutation acts as a 'double whammy', destabilising the genome and setting the stage for deregulated proliferation upon loss of the second APC allele.

Dietary factors and the occurrence of truncating APC mutations in sporadic colon carcinomas: a Dutch population-based study.

The interactions between environmental factors and the genetic and epigenetic changes that drive colon carcinogenesis are not clear. Dietary factors reported previously to be associated with colon cancer risk may well influence the occurrence of specific somatic alterations in colon tumors. To explore this idea, data from a Dutch population-based case-control study (184 cases, 259 controls) on sporadic colon cancer were used to assess associations between dietary factors and the occurrence of truncating mutations in the adenomatous polyposis coli (APC) gene in carcinomas. Single-strand conformation polymorphism analysis and DNA sequencing were used to screen tumors for mutations in the mutation cluster region of APC. Usual dietary habits were assessed by an interview-based questionnaire. Truncating APC mutations were detected in 63 (34%) of the tumors. Vegetable consumption was inversely associated with APC(+) (with mutation) tumors [odds ratio (OR) and 95% confidence interval (CI) for highest versus lowest tertile, OR: 0.6, 95% CI: 0.3-1.3] as well as APC(-) (without mutation) tumors (OR: 0.3, 95% CI: 0.2-0.5). Alcohol intake was positively associated with APC(-) tumors (OR: 1.7, 95% CI: 1.0-3.0) and inversely associated with APC(+) tumors (OR: 0.5, 95% CI: 0.3-1.1). Positive associations were observed for meat, fish and fat with APC(+) tumors (OR: 1.7, 95% CI: 0.8-3.6; OR: 1.4, 95% CI: 0.7-2.8; OR: 4.5, 95% CI: 1.6-12.8, respectively). Of the dietary factors examined, vegetable consumption and alcohol intake were significantly different related to APC(+) tumors than to APC(-) tumors (APC(+) versus APC(-), OR: 2.3, 95% CI: 1.0-5.3; OR: 0.3, 95% CI: 0.2-0.7, respectively). Our data suggest that vegetables play a protective role in the etiology of both tumor subsets, although this role appears to be less influential in the APC(+) group. Alcohol seems to especially promote the development of APC(-) tumors whereas meat, fish and fat appear to enhance the development of APC(+) tumors.

Adenomatous polyposis coli tumor suppressor protein has signaling activity in Xenopus laevis embryos resulting in the induction of an ectopic dorsoanterior axis.

Mutations in the adenomatous polyposis coli (APC) tumor suppressor gene are linked to both familial and sporadic human colon cancer. So far, a clear biological function for the APC gene product has not been determined. We assayed the activity of APC in the early Xenopus embryo, which has been established as a good model for the analysis of the signaling activity of the APC-associated protein beta-catenin. When expressed in the future ventral side of a four-cell embryo, full-length APC induced a secondary dorsoanterior axis and the induction of the homeobox gene Siamois. This is similar to the phenotype previously observed for ectopic beta-catenin expression. In fact, axis induction by APC required the availability of cytosolic beta-catenin. These results indicate that APC has signaling activity in the early Xenopus embryo. Signaling activity resides in the central domain of the protein, a part of the molecule that is missing in most of the truncating APC mutations in colon cancer. Signaling by APC in Xenopus embryos is not accompanied by detectable changes in expression levels of beta-catenin, indicating that it has direct positive signaling activity in addition to its role in beta-catenin turnover. From these results we propose a model in which APC acts as part of the Wnt/beta-catenin signaling pathway, either upstream of, or in conjunction with, beta-catenin.

Cranial desmoid tumor associated with homozygous inactivation of the adenomatous polyposis coli gene in a 2‐year‐old girl with familial adenomatous polyposis

Familial adenomatous polyposis (FAP) is a dominantly inherited disorder characterized by the presence of more than 100 adenomatous polyps in the colon and rectum starting in the second decade of life. FAP is associated with extra colonic manifestations, including desmoid tumors. A 2-year-old girl presented with a rapidly enlarging tumor of the forehead and a family history of FAP. The tumor was cultured for cytogenetic studies. A DNA linkage study using flanking and intragenic polymorphisms of the adenomatous polyposis coli (APC) gene was performed to identify the allele loss in the tumor. Germline mutation identification was by single strand conformation polymorphism analysis of exon 15 of the APC gene, with subsequent double stranded sequencing of fragments with conformational changes. A mutation-induced loss of a restriction site was used to confirm allele loss in the tumor. Microscopically, the tumor had desmoid features. Cytogenetic analysis of the tumor demonstrated loss of chromosome region 5(q21q22). A truncating adenomatous polyposis coli (APC) gene mutation was identified in the leukocyte DNA from the child and her affected father. Linked DNA markers suggested that the tumor had lost the maternal, wild-type allele. A mutation-induced restriction endonuclease site alteration demonstrated hemizygosity of the mutant sequence in the tumor DNA. These findings are compatible with the presence of a "second hit" inactivation of the APC gene and implicate this gene in the pathogenesis of desmoid tumors.

Cranial desmoid tumor associated with homozygous inactivation of the adenomatous polyposis coli gene in a 2‐year‐old girl with familial adenomatous polyposis

BACKGROUND Familial adenomatous polyposis (FAP) is a dominantly inherited disorder characterized by the presence of more than 100 adenomatous polyps in the colon and rectum starting in the second decade of life. FAP is associated with extra colonic manifestations, including desmoid tumors. METHODS A 2-year-old girl presented with a rapidly enlarging tumor of the forehead and a family history of FAP. The tumor was cultured for cytogenetic studies. A DNA linkage study using flanking and intragenic polymorphisms of the adenomatous polyposis coli (APC) gene was performed to identify allele loss in the tumor. Germline mutation identification was by single strand conformation polymorphism analysis of exon 15 of the APC gene, with subsequent double stranded sequencing of fragments with conformational changes. A mutation-induced loss of a restriction site was used to confirm allele loss in the tumor. RESULTS Microscopically, the tumor had desmoid features. Cytogenetic analysis of the tumor demonstrated loss of chromosome region 5(q21q22). A truncating adenomatous polyposis coli (APC) gene mutation was identified in the leukocyte DNA from the child and her affected father. Linked DNA markers suggested that the tumor had lost the maternal, wild-type allele. A mutation-induced restriction endonuclease site alteration demonstrated hemizygosity of the mutant sequence in the tumor DNA. CONCLUSIONS These findings are compatible with the presence of a “second hit” inactivation of the APC gene and implicate this gene in the pathogenesis of desmoid tumors. Cancer 1996;77:972-6.