Standardized Approach for Microsatellite Instability Detection in Colorectal Carcinomas

González-Garcı́a, Isabel; Moreno, Vı́ctor; Navarro, Matilde; Martí-Ragué, Joan; Marcuello, Eugenio; Benasco, C; Campos, Olga; Capellà, Gabriel; Peinado, Miguel A.

doi:10.1093/jnci/92.7.544

Cited by 73 publications

(43 citation statements)

References 21 publications

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“…Annotated, anonimized tissues were obtained after approval by the human investigations committee at the Ciutat Sanitària i Universitària de Bellvitge, Barcelona. The six tumors were part of a group of colorectal neoplasms previously studied for MSI (9). All were adenocarcinomas with variable amounts of mucin production.…”

Section: Methodsmentioning

confidence: 99%

Metapopulation dynamics and spatial heterogeneity in cancer

González-Garcı́a

Solé

Costa

2002

Proc. Natl. Acad. Sci. U.S.A.

189

146

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T he standard model invoked to explain tumor progression, the increase of biological malignancy with time, is clonal evolution, first proposed by Nowell in 1976 (1). Successive acquisition of mutations generates diverse clones, and it is postulated that the emergence of a dominant clone underlies the biological and clinical properties of a tumor at a point in its natural history (2). The study of structural genetic alterations found in human tumors provides further support for the clonal evolution model (3, 4), and limited observations suggest that the tempo of tumor progression may follow a punctuated pattern with long periods of stasis and periods of rapid genetic change in the tumor cells (5). In contrast to the concept of the emergence of a dominant clone, it is becoming increasingly apparent not only that microheterogeneity of tumor cells can be demonstrated at the phenotypic level, but that genetic heterogeneity is present in many advanced tumor types. Recent reports demonstrating genetic diversity within loci that play a key mechanistic role in tumor formation [e.g., K-ras, p53, transforming growth factor-␤RII (TGF␤RII), and BAX] present a paradox (6-8). How can the prolonged coexistence of genetically diverse populations and the concomitant lack of dominance be explained? After studying microdiversity in tumors exhibiting a microsatellite instability (MSI), we present a possible answer to this question. In silico modeling of the geography of clonal diversity suggests that the same fundamental principles that explain lack of dominance in complex ecosystems underlie the maintenance of clonal diversity in human tumors. Materials and MethodsHuman Tissues and Microdissection. Annotated, anonimized tissues were obtained after approval by the human investigations committee at the Ciutat Sanitària i Universitària de Bellvitge, Barcelona. The six tumors were part of a group of colorectal neoplasms previously studied for MSI (9). All were adenocarcinomas with variable amounts of mucin production. Four tumors were staged as Duke's B (T-2, T-3, T-5, and T-6), and two tumors belonged to the Duke's C with visceral metastases (T-1 and T-4). For microdissection, archival paraffin tissue blocks were cut at a thickness of 6 m and stained with methyl green. Microdissection was carried out by laser capture (PixCell; Arcturus Engineering, Mountain View, CA), by using a ϫ40 lens and paying attention to the exclusion of stromal cells. Intraepithelial lymphocytes in tumor glands were not identified. The micrographs in Fig. 1a show how a tumoral glandular structure is acquired: from the field shown at Left, a single structure is lifted from the tissue (Center). The result (Right) verifies the purity of the microdissected cell population. Each pool of cells analyzed represents the population of tumor cells enclosed in a contiguous surface of 100 m 2 .Genotyping. DNA extraction with proteinase K, as manufacturers recommend, was followed by PCR amplification by using 40 cycles of three loci: bat26 (a 26 A tract within an intron of ...

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Section: Methodsmentioning

confidence: 99%

Metapopulation dynamics and spatial heterogeneity in cancer

González-Garcı́a

Solé

Costa

2002

Proc. Natl. Acad. Sci. U.S.A.

189

146

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“…This series was part of a larger collection of patients prospectively included in a study designed to evaluate the prognostic value of genetic and epigenetic alterations (Gonzalez-Garcia et al, 2000;Vendrell et al, 2005). DNA methylation profiles were analysed in all 108 tumours and their normal tissue counterpart.…”

Section: Samplesmentioning

confidence: 99%

Long-range epigenetic silencing at 2q14.2 affects most human colorectal cancers and may have application as a non-invasive biomarker of disease

et al. 2009

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Large chromosomal regions can be suppressed in cancer cells as denoted by hypermethylation of neighbouring CpG islands and downregulation of most genes within the region. We have analysed the extent and prevalence of long-range epigenetic silencing at 2q14.2 (the first and best characterised example of coordinated epigenetic remodelling) and investigated its possible applicability as a non-invasive diagnostic marker of human colorectal cancer using different approaches and biological samples. Hypermethylation of at least one of the CpG islands analysed (EN1, SCTR, INHBB) occurred in most carcinomas (90%), with EN1 methylated in 73 and 40% of carcinomas and adenomas, respectively. Gene suppression was a common phenomenon in all the tumours analysed and affected both methylated and unmethylated genes. Detection of methylated EN1 using bisulfite treatment and melting curve (MC) analysis from stool DNA in patients and controls resulted in a predictive capacity of, 44% sensitivity in positive patients (27% of overall sensitivity) and 97% specificity. We conclude that epigenetic suppression along 2q14.2 is common to most colorectal cancers and the presence of a methylated EN1 CpG island in stool DNA might be used as biomarker of neoplastic disease.

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“…Indeed, in 2002, it was shown that MSI-L could be detected in the vast majority of CRCs simply by expanding the number of tested microsatellites (Laiho et al, 2002), strengthening the idea that increasing the number of tested markers is not of any help in the MS-status assessment. The possibility that the distribution of MSI can be bimodal (MSI vs MSS) or trimodal (MSI-H vs MSI-L vs MSS) found no evidence for a separate subset comprising MSI-L CRCs (Gonzalez-Garcia et al, 2000). Accordingly, the MSI-L phenotype was not associated with any clinico-pathological or molecular feature (such as, K-RAS mutations or APC LOH) (Laiho et al, 2002).…”

Section: The Operative Definition Of Msimentioning

confidence: 99%

Differences and evolution of the methods for the assessment of microsatellite instability

2008

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Microsatellite instability (MSI) originates from the systematic accumulation of uncorrected deletion/insertion in repetitive DNA tracts in cancer cells with a deficient mismatch repair system. Among colorectal cancers, the MSI signature identifies hereditary cases arising in patients with germline mutations in hMLH1, hMSH2, PMS2 and a fraction of those with hMSH6 mutations, as well as sporadic cancers with epigenetic hMLH1 promoter hypermethylation. Considering the specific pathogenesis, pathological features, natural history and response to 5-fluoro-uracil-based chemotherapy of the MSI cancers, confusion about the genetic markers for MSI recognition seems surprising. In this clinically relevant field, an agreement has not been reached concerning the use of di-or mononucleotide markers for MSI assessment. The Revised Bethesda Guidelines still recommend a panel of markers consisting of mono-and dinucleotides, despite being questioned whether it is congruous to continue to use dinucleotide markers for MSI identification. In any event, no single marker is accurate enough for MSI testing, and an awareness of their pros and cons is required for proper interpretation of results. In recent years, several papers have reported different prevalence of MSI in unrelated series, largely depending on the detection and classification method, suggesting that MSI test interpretation also requires the understanding of the phenomenon rather than simply the crude satisfaction of panel recommendations. Inaccuracies can otherwise lead to under-or overdiagnosis and inaccurate disease classification, which always have a negative impact on the clinical practice of medicine.

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Standardized Approach for Microsatellite Instability Detection in Colorectal Carcinomas

Cited by 73 publications

References 21 publications

Metapopulation dynamics and spatial heterogeneity in cancer

Metapopulation dynamics and spatial heterogeneity in cancer

Long-range epigenetic silencing at 2q14.2 affects most human colorectal cancers and may have application as a non-invasive biomarker of disease

Differences and evolution of the methods for the assessment of microsatellite instability

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