Testing of prostate cancer biopsy specimens from metastatic sites for p53 protein accumulation and gene mutation may provide useful prognostic information and could influence the recommended course of treatment.

Human tobacco-related cancers exhibit a high frequency of G to T transversions in the mutation hot spot region of the p53 tumor suppressor gene, possibly the result of specific mutagens in tobacco smoke, most notably benzo [a] An increasing body of molecular epidemiological evidence suggests that regions ofthe p53 gene may be a selective target of environmental carcinogens etiologically associated with specific human cancers (4-19). In this context, a high frequency of G to T transversions in the mutation hot spot regions of the p53 gene (2, 3) have been identified in human hepatocellular carcinomas and strongly correlated with the mutagenic effects of dietary aflatoxin B1 exposurf (4,(13)(14)(15).Similarly, UV-B-specific dipyrimidine G to T transitions in the p53 gene have been characterized in human squamous cell carcinomas of the skin (16,17). Unique mutations in p53 have likewise been reported in radon-associated lung cancers (18) and in human epithelial cells exposed to nickel(II) in vitro (19), providing evidence for an important molecular target for the genotoxic effects of these agents. Of particular clinical and epidemiological interest are the observations of a high frequency of G to T transversions among p53 mutations in tobacco-related human neoplasias, including small-cell and non-small-cell lung cancers (5-8), esophageal carcinomas (10-12), and squamous cell carcinomas of the head and neck (9). In several ofthese studies, the presence of p53 mutations in tumors was strongly correlated with lifetime cigarette consumption (7) or a history of heavy smoking (9).It has been suggested (4-12) that the frequent G to T transversion in the p53 gene in tobacco-related cancers may be associated with specific mutagens in tobacco smoke, most notably activated benzo [a] We have previously investigated the role of p53 alterations in the development and progression of the two-stage mouse skin tumorigenesis model induced by 7,12-dimethylbenz- (24) following hematoxylin/eosin staining. Primary explant cultures of skin tumors were prepared as described (22). Papillomas harvested for analyses were subjected to 35-38 weeks of B[a]P treatment, while carcinomas analyzed were subjected to 50 weeks of carcinogen exposure. In the complete carcinogenesis protocol with DMBA, papillomas and carcinomas used in molecular analyses were obtained from mice subjected to 34-52 weeks of carcinogen exposure. The two-stage carcinogenesis protocol using DMBA as initiator has been described (22

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Cyclins and cyclin-dependent kinases (Cdks) are central to regulation of the cell cycle. Their abnormal expression may cause loss of cell-cycle control and result in autonomous cell growth, a critical feature of neoplasias. In this study, using immunoblotting, we analyzed the protein levels of several G1/S cyclins (cyclins D1, D2, D3, A, and E) and their respective Cdks (Cdk 2, 4, and 6) in 17 mouse squamous cell carcinomas (SCCs) and 18 mouse skin tumor cell lines. Overexpression of these cell cycle-related genes was frequent in tumors and cell lines. Of special interest was the fact that a group of cell lines that became more aggressive after animal passaging expressed more cyclins D2 and D3 than their respective parental lines did. In addition, SCCs had higher cyclin D3 expression levels than papillomas, and metastases had higher levels than the respective primary tumors, indicating that overexpression of cyclin D3 may be associated with increased aggressiveness of mouse SCC. Interestingly, overexpression of cyclin E was seen in most SCCs induced by a complete carcinogenesis protocol with benzo[a]pyrene (B(a)P) and only in a few SCCs induced by a two-stage carcinogenesis protocol using 7,12-dimethylbenz[a]anthracene as initiator. In contrast, more of the latter tumors overexpressed cyclin D1 and D2 than those induced by B(a)P. Thus, it is possible that different components of the cell-cycle machinery are involved in proliferative dysfunctions that take place during tumor development with different carcinogenesis protocols. Taken together, these results indicate that overexpression of G1 cyclins and their related Cdks is a significant molecular abnormality that could be involved in the process of tumor progression.

Treatment of B6C3F1 mice with concentrations of 62.5-625 p.p.m. 1,3-butadiene by inhalation for up to 2 years causes a significantly increased incidence of Harderian gland (HG) neoplasms over untreated controls (Melnick,R., Huff,J., Chou,B.J. and Miller,R.A. Cancer Res., 50, 6592-6599, 1990). Since a specific K-ras mutation (codon 13 GGC-->CGC) had previously been described in lung and liver tumors from 1,3-butadiene-treated B6C3F1 mice, we analyzed 23 adenomas and six adenocarcinomas of the HG from mice exposed to 1,3-butadiene for this mutation and mutations in the H-ras gene. We also examined ras activation in 16 spontaneously occurring HG adenomas and one adenocarcinoma. DNA samples were prepared from paraffin-embedded tissues and analyzed by PCR followed by direct sequencing methods. Only one 1,3-butadiene-induced HG tumor contained the K-ras codon 13 mutation previously detected in lung and liver tumors. However, 16/29 HG tumors from the treated B6C3F1 mice contained H-ras codon 61 mutations. The mutations detected were: 12 CAA-->CGA transitions, two CAA-->CTA and two CAA-->AAA transversions. Eleven of 17 spontaneous HG tumors contained mutations in H-ras codon 61: five CAA-->CGA transitions, two CAA-->CTA transversions and four CAA-->AAA transversions. While the spectrum of ras mutations did not differ between the spontaneously occurring and chemically induced tumors, these data indicate that activation of H-ras contributes to the process of HG tumorigenesis in both groups of these neoplasms.