Evaluation of Factors Related to Late Recurrence - Later than 10 Years after the Initial Treatment - in Primary Breast Cancer

Tumors can recur years after treatment, and breast cancer is especially noted for long periods of dormancy. The status of the cancer during this period is poorly understood. As a model to study mechanisms of dormancy, we used murine D2.0R mammary carcinoma cells, which are poorly metastatic but form occasional metastases in liver and other organs after long latency. Highly metastatic D2A1 cells provided a positive, metastatic control. Our goals were to learn how the cell lines differ in survival kinetics in a secondary site and to seek evidence for the source of D2.0R dormancy. In spontaneous metastasis assays from mammary fat pad injections, we found evidence for dormancy because of a persistence of large numbers of solitary cells in the liver. To quantify the fate of cells after arrival in liver, experimental metastasis assays were used. To permit identification of cells that had not divided, cells were labeled before injection with fluorescent nanospheres, which were diluted to undetectable levels by cell division. Cancer cells were injected i.v. to target them to the liver and coinjected with reference microspheres to monitor cell survival. Dormancy was defined as retention of nanosphere fluorescence in vivo, as well as negative staining for the proliferation marker Ki67. A large proportion of D2.0R cells persisted as solitary dormant cells. No metastases formed, but viable cells could be recovered from the liver 11 weeks after injection. Large numbers of solitary, dormant, Ki67-negative D2A1 cells were also detected against a background of progressively growing metastases. Thus, this study identified a possible contributor to tumor dormancy: solitary, dormant cells that persist in tissue. If such cells are present in patients, they could contribute to tumor recurrence and would not be susceptible to current therapeutic strategies targeting proliferating cells.