Role of Rab4A in the enhancement of cancer cell invasion induced by plasma samples collected during radiotherapy in patients with triple-negative breast cancer.

Abstract

The relapse rate for early-stage triple-negative breast cancer (TNBC) is significantly higher than that of other breast cancer subtypes. To reduce the relapse rate, we evaluated the relevance of targeting Rab4A to inhibit the increase in cancer cell invasion induced by radiotherapy (RT). The ability of cancer cells to invade into adjacent tissues correlates with the proteolytic activity of membrane type 1 matrix metalloproteinase (MT1-MMP) on their surface, whose turnover is regulated by Rab4A. When RAB4A was downregulated using shRNA in the TNBC cells D2A1 and MDA-MB-231, a significant reduction in the proteolytic activity of MT1-MMP and the invasion capacity of these TNBC cells were measured. Plasma samples from six early-stage TNBC patients were collected before RT and after the fourth radiation dose. Plasma samples collected during RT from three of these patients increased the invasiveness of TNBC cells, compared to plasma collected before RT. Of them, two experienced a recurrence of their cancer, while the third patient had not yet completed the 5-year follow-up. The downregulation of RAB4A inhibited the RT-induced increase in cancer cell invasion. In contrast, in the three patients who did not relapse after 5-year follow-up, plasma collected during RT did not significantly increase the invasiveness of the MDA-MB-231 cells, compared to their plasma collected before RT. The decrease in RAB4A expression did not result in a significant difference in the invasiveness of cancer cells when incubated with plasma collected before or during RT from these patients. Metastatic development was observed when D2A1 cells were preincubated before i.v. injection in Balb/c mice with plasma collected during RT from a patient who experienced early cancer relapse; this was also inhibited by downregulating RAB4A. In summary, our preliminary results generate the hypothesis that RT may increase cancer cell invasion and metastasis formation, which is associated with the Rab4A-mediated turnover of MT1-MMP on the cancer cell surface.