Redston, Mark

4052 Background: 18q LOH may be a useful marker in stage II colon carcinoma to assess who needs adjuvant chemotherapy. The objective of this study was to evaluate the intra- and inter-laboratory reproducibility of an assay for 18q LOH that is similar to the integral assay used in E5202, the current Intergroup phase III stage II colon carcinoma clinical trial. Methods: Specimens from 130 stage II/III colon cancer patients were obtained from the Cooperative Human Tissue Network and evaluated at three different laboratories. Samples were divided into subsamples, coded and distributed. Each laboratory used similar equipment in an established PCR protocol with 5 dinucleotide repeats. Starting materials used both histologic sections and extracted DNA samples of tumor and non-neoplastic tissue obtained only by scalpel microdissection. LOH was defined as tumor/normal peak ratio that was >1.35 or <0.67 without microsatellite instability (MSI). Tumors were categorized as either MSI+, LOH + or -, uninformative/monoallelic, or unevaluable (DNA not amplifiable). Standard tests assessed reproducibility of both the numerical ratios and the tumor 18q LOH status. Results: Sample quality issues prevented analysis of 13 tumors (10%). The frequency of 18q LOH was 64%. The probability of agreement on the same tumor evaluated twice was 92% (Cohen's kappa 0.83). Intra-class correlation coefficients for individual markers were consistently over 0.90. Tissue specimens must have high (>85%) tumor cell content for reproducibility. Differences among patients affected reproducibility much more than effects of laboratory and sample type (DNA vs. histologic section). Conclusions: This study validates reproducibility of the 18q LOH assay for both clinical categorization of status and numerical ratios if samples are high quality and appropriate for scalpel microdissection. [Table: see text]

Abstract Aberrant crypt foci (ACF) are the earliest visible neoplastic lesions in the colorectum. The natural history of these lesions and their role in the adenoma-carcinoma sequence are unknown. We studied ACF in a subset of patients enrolled in the Adenoma Prevention with Celecoxib (APC) Trial to determine whether the presence of ACF in the rectum correlated with that of adenomas in the entire large intestine. Participants were randomized to placebo (n=17), celecoxib 200 mg bid (n=15), or celecoxib 400 mg bid (n=13). Magnification chromoendoscopy (MCE) was performed to identify and biopsy ACF within the rectum at baseline and after 8-12 months of medication use. A total of 670 ACF were identified in 45 patients, and 70 of these were examined histologically. All of the ACF examined were non-dysplastic. Analysis showed no differences in ACF number between baseline and post-treatment studies (8.3±1.0 vs. 6.3±1.1) and no modulation by treatment (celecoxib vs placebo; p=0.77). Immunohistochemical studies showed that ACF contained cells exhibiting increased proliferation, but lacked other features of neoplasia such as increased Cox-2 expression and microvascular density, nuclear localization of β-catenin, or decreased expression of the tumor suppressors SMAD4, estrogen receptor (ERα) or MGMT. Of the secondary markers assessed at baseline, only SMAD4 expression correlated with post-treatment adenoma recurrence. Taken together, these data showed that non-dysplastic ACF are not accurate surrogate endpoint biomarkers of colorectal adenoma risk or NSAID chemoprevention response.

Abstract Aberrant crypt foci (ACF) are the earliest visible neoplastic lesions in the colorectum. The natural history of these lesions and their role in the adenoma-carcinoma sequence are unknown. We studied ACF in a subset of patients randomized to placebo (n = 17), celecoxib (200 mg twice daily; n = 15), or celecoxib (400 mg twice daily; n = 13) in the Adenoma Prevention with Celecoxib (APC) trial. Magnification chromoendoscopy was done to identify, count, and biopsy ACF within the rectum at baseline and after 8 to 12 months of treatment. A total of 655 ACF were identified in 45 patients. We examined 70 of these ACF histologically, and all 70 were nondysplastic. Cohort characteristics and APC trial treatment results for substudy patients were similar to those of the overall APC trial. There was no significant modulation of ACF by celecoxib (versus placebo; P = 0.77). Immunohistochemical comparison of ACF with adjacent normal mucosa showed that ACF had an increased proliferative index as determined by Ki-67 (P &lt; 0.0001), but lacked other features of neoplasia such as increased cyclooxygenase-2 expression and microvessel density, nuclear localization of β-catenin, or decreased expression of the tumor suppressors SMAD4, Estrogen Receptor α, or MGMT. Only baseline SMAD4 expression in ACF correlated with posttreatment adenoma recurrence (independent of treatment arm; P = 0.01). The presence or number of nondysplastic ACF did not correlate with a higher risk of synchronous advanced or recurrent adenomas. Our overall results indicated that nondysplastic ACF were not accurate surrogate endpoint biomarkers of recurrent colorectal adenomas in the APC trial.