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Promoter methylation of tumor suppressor genes in pre-neoplastic lesions; potential marker of disease recurrence.

Rengucci C, De Maio G, Casadei Gardini A, Zucca M, Scarpi E, Zingaretti C, Foschi G, Tumedei MM, Molinari C, Saragoni L, Puccetti M, Amadori D, Zoli W, Calistri D - J. Exp. Clin. Cancer Res. (2014)

Bottom Line: MS-MLPA results were confirmed by pyrosequencing and immunohistochemistry.Histopathological classification does not permit an accurate evaluation of the risk of recurrence of colorectal lesions.Conversely, results from our methylation analysis suggest that a classification based on molecular parameters could help to define the mechanisms involved in carcinogenesis and prove an effective method for identifying patients at high risk of recurrence.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Background: Epigenetic alterations of specific genes have been reported to be related to colorectal cancer (CRC) transformation and would also appear to be involved in the early stages of colorectal carcinogenesis. Little data are available on the role of these alterations in determining a different risk of colorectal lesion recurrence. The aim of the present study was to verify whether epigenetic alterations present in pre-neoplastic colorectal lesions detected by colonoscopy can predict disease recurrence.

Methods: A retrospective series of 78 adenomas were collected and classified as low (35) or high-risk (43) for recurrence according to National Comprehensive Cancer Network guidelines. Methylation alterations were analyzed by the methylation-specific multiplex ligation probe assay (MS-MLPA) which is capable of quantifying methylation levels simultaneously in 24 different gene promoters. MS-MLPA results were confirmed by pyrosequencing and immunohistochemistry.

Results: Higher levels of methylation were associated with disease recurrence. In particular, MLH1, ATM and FHIT gene promoters were found to be significantly hypermethylated in recurring adenomas. Unconditional logistic regression analysis used to evaluate the relative risk (RR) of recurrence showed that FHIT and MLH1 were independent variables with an RR of 35.30 (95% CI 4.15-300.06, P = 0.001) and 17.68 (95% CI 1.91-163.54, P = 0.011), respectively.

Conclusions: Histopathological classification does not permit an accurate evaluation of the risk of recurrence of colorectal lesions. Conversely, results from our methylation analysis suggest that a classification based on molecular parameters could help to define the mechanisms involved in carcinogenesis and prove an effective method for identifying patients at high risk of recurrence.

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Gene methylation level distribution. A) Hypermethylated genes in the case series subdivided according to the presence or not of disease recurrence. B) Comparison of methylation levels of the three most significant genes in R and NR samples.
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Figure 1: Gene methylation level distribution. A) Hypermethylated genes in the case series subdivided according to the presence or not of disease recurrence. B) Comparison of methylation levels of the three most significant genes in R and NR samples.

Mentions: MS-MLPA analysis was performed for all samples, obtaining a quantification of methylation status for the entire case series. Two probes (GSTP1 and MLH1 CpG 02) were discarded from the analysis because they were negative for methylation (0% methylation level) in 92% and 83% of cases, respectively. We first evaluated the number of hypermethylated promoters in R and NR patients using a methylation level of 20% to define a gene promoter as hypermethylated. Primary lesions that relapsed showed a higher number of hypermethylated markers (median 6, range 2–24) than non recurring lesions (median 4, range 0–12) (Figure 1A).


Promoter methylation of tumor suppressor genes in pre-neoplastic lesions; potential marker of disease recurrence.

Rengucci C, De Maio G, Casadei Gardini A, Zucca M, Scarpi E, Zingaretti C, Foschi G, Tumedei MM, Molinari C, Saragoni L, Puccetti M, Amadori D, Zoli W, Calistri D - J. Exp. Clin. Cancer Res. (2014)

Gene methylation level distribution. A) Hypermethylated genes in the case series subdivided according to the presence or not of disease recurrence. B) Comparison of methylation levels of the three most significant genes in R and NR samples.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4274757&req=5

Figure 1: Gene methylation level distribution. A) Hypermethylated genes in the case series subdivided according to the presence or not of disease recurrence. B) Comparison of methylation levels of the three most significant genes in R and NR samples.
Mentions: MS-MLPA analysis was performed for all samples, obtaining a quantification of methylation status for the entire case series. Two probes (GSTP1 and MLH1 CpG 02) were discarded from the analysis because they were negative for methylation (0% methylation level) in 92% and 83% of cases, respectively. We first evaluated the number of hypermethylated promoters in R and NR patients using a methylation level of 20% to define a gene promoter as hypermethylated. Primary lesions that relapsed showed a higher number of hypermethylated markers (median 6, range 2–24) than non recurring lesions (median 4, range 0–12) (Figure 1A).

Bottom Line: MS-MLPA results were confirmed by pyrosequencing and immunohistochemistry.Histopathological classification does not permit an accurate evaluation of the risk of recurrence of colorectal lesions.Conversely, results from our methylation analysis suggest that a classification based on molecular parameters could help to define the mechanisms involved in carcinogenesis and prove an effective method for identifying patients at high risk of recurrence.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Background: Epigenetic alterations of specific genes have been reported to be related to colorectal cancer (CRC) transformation and would also appear to be involved in the early stages of colorectal carcinogenesis. Little data are available on the role of these alterations in determining a different risk of colorectal lesion recurrence. The aim of the present study was to verify whether epigenetic alterations present in pre-neoplastic colorectal lesions detected by colonoscopy can predict disease recurrence.

Methods: A retrospective series of 78 adenomas were collected and classified as low (35) or high-risk (43) for recurrence according to National Comprehensive Cancer Network guidelines. Methylation alterations were analyzed by the methylation-specific multiplex ligation probe assay (MS-MLPA) which is capable of quantifying methylation levels simultaneously in 24 different gene promoters. MS-MLPA results were confirmed by pyrosequencing and immunohistochemistry.

Results: Higher levels of methylation were associated with disease recurrence. In particular, MLH1, ATM and FHIT gene promoters were found to be significantly hypermethylated in recurring adenomas. Unconditional logistic regression analysis used to evaluate the relative risk (RR) of recurrence showed that FHIT and MLH1 were independent variables with an RR of 35.30 (95% CI 4.15-300.06, P = 0.001) and 17.68 (95% CI 1.91-163.54, P = 0.011), respectively.

Conclusions: Histopathological classification does not permit an accurate evaluation of the risk of recurrence of colorectal lesions. Conversely, results from our methylation analysis suggest that a classification based on molecular parameters could help to define the mechanisms involved in carcinogenesis and prove an effective method for identifying patients at high risk of recurrence.

Show MeSH
Related in: MedlinePlus