The genetic basis of colorectal cancer in a population-based incident cohort with a high rate of familial disease

Abstract

Background and aims: Colorectal cancer (CRC) is the second most frequent cancer in developed countries. Newfoundland has the highest incidence of CRC in Canada and the highest rate of familial CRC yet reported in the world. To determine the impact of mutations in known CRC susceptibility genes and the contribution of the known pathways to the development of hereditary CRC, an incident cohort of 750 patients with CRC (708 different families) from the Newfoundland population was studied.

Methods: Microsatellite instability (MSI) testing was performed on tumours, together with immunohistochemistry analysis for mismatch repair (MMR) genes. Where indicated, DNA sequencing and multiplex ligation-dependent probe amplifications of MMR genes and APC was undertaken. DNA from all patients was screened for MUTYH mutations. The presence of the BRAF variant, p.V600E, and of MLH1 promoter methylation was also tested in tumours.

Results: 4.6% of patients fulfilled the Amsterdam criteria (AC), and an additional 44.6% fulfilled the revised Bethesda criteria. MSI-high (MSI-H) was observed in 10.7% (n=78) of 732 tumours. In 3.6% (n=27) of patients, CRC was attributed to 12 different inherited mutations in six known CRC-related genes associated with chromosomal instability or MSI pathways. Seven patients (0.9%) carried a mutation in APC or biallelic mutations in MUTYH. Of 20 patients (2.7%) with mutations in MMR genes, 14 (70%) had one of two MSH2 founder mutations. 17 of 28 (61%) AC families did not have a genetic cause identified, of which 15 kindreds fulfilled the criteria for familial CRC type X (FCCTX).

Conclusions: Founder mutations accounted for only 2.1% of cases and this was insufficient to explain the high rate of familial CRC. Many of the families classified as FCCTX may have highly penetrant mutations segregating in a Mendelian-like manner. These families will be important for identifying additional CRC susceptibility loci.

Figure 1

(A) The biological samples obtained from study participants, their derivatives and assays implemented. (*in a few cases RNA was extracted to confirm multiplex ligation-dependent probe amplification (MLPA) results). (B) Testing flow chart to determine mismatch repair (MMR) deficiency and Lynch syndrome. Numbers represent tumours/patients. The number of tumours at the top of the chart are those which had microsatellite instability (MSI) analysis performed. ¹N.D., not determined; ²MLH1-D, MLH1-deficient and includes both MLH1-only and MLH1- and PMS2-deficient tumours; ³Meth or BRAF, methylated MLH1 promoter or presence of p.V600E BRAF mutation.

Figure 2

Summary data from the Newfoundland Colorectal Cancer Registry (NFCCR). (A) Average age of diagnosis of colorectal cancer (CRC) for each risk category. Amsterdam criteria families are excluded in the revised Bethesda criteria category. (B) Participation rates of patients with CRC per age categories. Proxies are included and are assigned age at diagnosis of CRC for their affected relative. (C) Average age at diagnosis of CRC for mutation carriers and for familial CRC type X (FCCTX). (D) Proportion of tumours located in proximal and distal colon in mismatch repair (MMR)-deficient tumours and in tumours from patients in families fulfilling FCCTX. Proximal is defined as proximal to the splenic flexure. MMR refers to tumours from patients which have mutations in MLH1, MSH2, MSH6 or PMS2. Non-MMR refers to tumours from patients with biallelic MUTYH mutations or an APC mutation.

Figure 3

Genes mutated in patients from the Newfoundland Colorectal Cancer Registry (NFCCR) according to clinical risk criteria (*revised Bethesda criteria and excluding high risk families).

Figure 4

Summary of microsatellite instability (MSI) and immunohistochemistry (IHC) results from the 772 tumours collected through the Newfoundland Colorectal Cancer Registry (NFCCR). ‘IHC-I’, IHC-intact; ‘IHC-D’, IHC-deficient; ‘IHC-ND’, IHC not done.

Figure 5

Proportion of patients (n=41) from high risk families indicating the genetic aetiology of their colorectal cancer predisposition. CIN, chromosomal instability; FCCTX, familial colorectal cancer type X; MSI, microsatellite instability.