TY - JOUR
T1 - Combined DNA flow cytometry and sorting with k-ras2 mutation spectrum analysis and the prognosis of human sporadic colorectal cancer
AU - Geido, Elio
AU - Sciutto, Andrea
AU - Rubagotti, Alessandra
AU - Oliani, Cristina
AU - Monaco, Roberto
AU - Risio, Mauro
AU - Giaretti, Walter
PY - 2002
Y1 - 2002
N2 - Background: Activation of the k-ras2 pathways and chromosomal instability leading to aneuploidy in human sporadic colorectal cancer (sCRC) is essential to the tumor cell ability to survive, grow, and metastatize. Methods: The study included 135 patients with sCRC who were followed up for a median of 72 months. Multiple fresh-frozen fragments obtained from superficial and invasive areas of the tumors were mixed and used to detect the degree of DNA aneuploidy (DNA index [DI]) and S-phase fraction by two scatter signals and 4,6-diamidino-2- phenylindole-2-hydrocloride (DAPI) fluorescence flow cytometry (FCM). PCR amplification and k-ras2 mutation spectrum analysis were performed using enriched epithelial nuclei after sorting DNA aneuploid nuclei and DNA diploid nuclei from which tissue-infiltrating lymphocytes were absent. Results: DNA aneuploidy was detected in 98 (73%) and k-ras2 mutations in 54 cases (40%). Univariate analyses of overall survival with both Dukes' A to D or B to C series of cases showed that DNA multiple aneuploidy, k-ras2 mutations, older age, and distal site, but not increased S-phase fraction, were predictive of worse outcome. Multivariate Cox models strongly indicated that k-ras2 mutations, but neither single nor multiple DNA aneuploidy, were an independent prognostic factor in both series of patients. In particular, with B and C Dukes' stage patients (n = 110), the relative risk (RR) of death was above 2.5 with k-ras2 mutations and above 3 with the G→C/T transversions. Conclusion: Combined FCM and k-ras2 analysis may be used to predict long-term increased risk of death in sCRC patients.
AB - Background: Activation of the k-ras2 pathways and chromosomal instability leading to aneuploidy in human sporadic colorectal cancer (sCRC) is essential to the tumor cell ability to survive, grow, and metastatize. Methods: The study included 135 patients with sCRC who were followed up for a median of 72 months. Multiple fresh-frozen fragments obtained from superficial and invasive areas of the tumors were mixed and used to detect the degree of DNA aneuploidy (DNA index [DI]) and S-phase fraction by two scatter signals and 4,6-diamidino-2- phenylindole-2-hydrocloride (DAPI) fluorescence flow cytometry (FCM). PCR amplification and k-ras2 mutation spectrum analysis were performed using enriched epithelial nuclei after sorting DNA aneuploid nuclei and DNA diploid nuclei from which tissue-infiltrating lymphocytes were absent. Results: DNA aneuploidy was detected in 98 (73%) and k-ras2 mutations in 54 cases (40%). Univariate analyses of overall survival with both Dukes' A to D or B to C series of cases showed that DNA multiple aneuploidy, k-ras2 mutations, older age, and distal site, but not increased S-phase fraction, were predictive of worse outcome. Multivariate Cox models strongly indicated that k-ras2 mutations, but neither single nor multiple DNA aneuploidy, were an independent prognostic factor in both series of patients. In particular, with B and C Dukes' stage patients (n = 110), the relative risk (RR) of death was above 2.5 with k-ras2 mutations and above 3 with the G→C/T transversions. Conclusion: Combined FCM and k-ras2 analysis may be used to predict long-term increased risk of death in sCRC patients.
KW - Aneuploidy
KW - Chromosome instability
KW - Oncogenes
KW - Tumor progression
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U2 - 10.1002/cyto.10109
DO - 10.1002/cyto.10109
M3 - Article
C2 - 12210601
AN - SCOPUS:0037103892
VL - 50
SP - 216
EP - 224
JO - Cytometry Part B - Clinical Cytometry
JF - Cytometry Part B - Clinical Cytometry
SN - 1552-4949
IS - 4
ER -