WAF1/Cip1 gene polymorphism and expression in carcinomas of the breast, ovary and endometrium

WAF1/Cip1 is a recently described downstream effector of the tumor suppressor gene, p53. WAF1/Cip1 codes for a 21 kilodalton protein (p21) which is a universal inhibitor of cyclins and is thus critical in cell cycle control. To understand if mutations of WAF1/Cip1 occur in cancer, we screened 56 cases of invasive breast carcinoma, 35 cases of ductal carcinoma in situ (DCIS) of the breast, 56 ovarian carcinomas, 43 endometrial carcinomas, and 30 borderline tumors of the ovary for mutations in the second exon of WAF1/Cip1 (90% of the open reading frame). Single strand conformation polymorphism (SSCP) analysis demonstrated an altered mobility pattern for exon 2 in 12 invasive breast cancers (15%), 5 ductal carcinomas in situ of the breast (14%), 7 endometrial carcinomas (16%), 15 invasive ovarian carcinomas (17%) and in none of the ovarian tumors of low malignant potential. Overall, 16% (39 of 243) of the cases had an altered codon 31. Each case with altered SSCP, analyzed by DNA sequencing and/or restriction analysis, showed the same alteration of codon 31, a C to A transition encoding a change in amino acid sequence from serine to argininc. DNA from the blood of 21 normal individuals showed the same alteration in WAF1/Cip1 in 4 cases (19%). Furthermore, paired normal tissue was available for 3 of 20 breast carcinomas with the codon 31 alteration. Normal DNA from all three cases showed the same alteration of codon 31 as found in the tumor tissue. These results indicate that the codon 31 alteration is a polymorphism. p21 expression, identified by immunohistochemistry, was found to vary in a pattern that depended both on the presence or absence of the codon 31 polymorphism and on the tissue type. In breast CIS, we found higher staining in tumor nuclei as compared to normal nuclei (p=0.0016). There was no difference in staining in the other tumor cases. In breast and ovarian carcinomas there was no correlation between p21 protein expression and the presence or absence of the alteration at the 31st codon. However, in endometrial carcinomas, an increased percentage of nuclei immunopositive for p21 was associated (p=0.027) with the presence of arginine at codon 31. These data suggest that WAF1/Cip1 may play a role in breast CIS, and furthermore, altered p21 may have a different function than the normal p21 in endometrial cancers.