Immortalization and transformation of human mammary epithelial cells by a tumor-derived Myc mutant.

The Myc transcription factor is commonly dysregulated in many human cancers, including breast carcinomas. However, the precise role of Myc in the initiation and maintenance of malignancy is unclear. In this study we compared the ability of wild-type Myc (wt Myc) or Myc phosphorylation deficient mutants (T58A, S62A or T58A/S62A) to immortalize and transform human mammary epithelial cells (HMECs). All Myc constructs promoted cellular immortalization. As previously reported in other cells, the Myc T58A mutant tempered apoptotic responses and increased Myc protein stability in HMEC cells. More importantly, we now show that HMECs overexpressing the Myc T58A mutant acquire a unique cellular phenotype characterized by cell aggregation, detachment from the substrate and growth in liquid suspension. Coincident with these changes, the cells become anchorage-independent for growth in agarose. Previous studies have shown that wt Myc can collaborate with hTERT in inducing HMEC anchorage-independent growth. We have verified this observation and further shown that Myc T58A was a stronger facilitator of such co-transformation. Thus, our findings indicate that differences in Myc protein phosphorylation modulate its biological activity in human breast epithelial cells and specifically that the T58A mutation can facilitate both cellular immortalization and transformation. Finally, we used the isogenic cell lines generated in this study to identify a subset of genes whose expression is greatly altered during the transition from the immortal to the anchorage-independent states.

Dihydroartemisinin is cytotoxic to papillomavirus-expressing epithelial cells in vitro and in vivo.

Nearly all cervical cancers are etiologically attributable to human papillomavirus (HPV) infection and pharmaceutical treatments targeting HPV-infected cells would be of great medical benefit. Because many neoplastic cells (including cervical cancer cells) overexpress the transferrin receptor to increase their iron uptake, we hypothesized that iron-dependent, antimalarial drugs such as artemisinin might prove useful in treating HPV-infected or transformed cells. We tested three different artemisinin compounds and found that dihydroartemisinin (DHA) and artesunate displayed strong cytotoxic effects on HPV-immortalized and transformed cervical cells in vitro with little effect on normal cervical epithelial cells. DHA-induced cell death involved activation of the mitochondrial caspase pathway with resultant apoptosis. Apoptosis was p53 independent and was not the consequence of drug-induced reductions in viral oncogene expression. Due to its selective cytotoxicity, hydrophobicity, and known ability to penetrate epithelial surfaces, we postulated that DHA might be useful for the topical treatment of mucosal papillomavirus lesions. To test this hypothesis, we applied DHA to the oral mucosa of dogs that had been challenged with the canine oral papillomavirus. Although applied only intermittently, DHA strongly inhibited viral-induced tumor formation. Interestingly, the DHA-treated, tumor-negative dogs developed antibodies against the viral L1 capsid protein, suggesting that DHA had inhibited tumor growth but not early rounds of papillomavirus replication. These findings indicate that DHA and other artemisinin derivatives may be useful for the topical treatment of epithelial papillomavirus lesions, including those that have progressed to the neoplastic state.