Resistance to the Antineoplastic Agent Gallium Nitrate Results in Marked Alterations in Intracellular Iron and Gallium Trafficking: Identification of Novel Intermediates

Abstract

Gallium (Ga) shows significant antitumor activity by markedly interfering with iron (Fe) metabolism, and ⁶⁷Ga is used as a radio-imaging agent for cancer detection. Therefore, the mechanisms involved in ⁶⁷Ga uptake, metabolism, and resistance are critical to understand. The development of tumor lines that are gallium-resistant suggests ⁶⁷Ga uptake may be different in these cells, providing an opportunity for understanding intracellular ⁶⁷Ga and ⁵⁹Fe transport and gallium resistance. In this study, gallium-resistant cells were used to assess ⁶⁷Ga and ⁵⁹Fe uptake using native polyacrylamide gel electrophoresis autoradiography. In contrast to the common view that ⁶⁷Ga and ⁵⁹Fe use the same uptake pathways, we show that the trafficking of these two metal ions is different in cells either resistant (R) or sensitive (S) to gallium. Indeed, in contrast to ⁵⁹Fe, little ⁶⁷Ga is incorporated into ferritin, with most present as a labile ⁶⁷Ga pool. We also report unique changes in ⁶⁷Ga and ⁵⁹Fe trafficking between R and S cells. In particular, in R cells, there was a distinct transferrin-transferrin receptor 1-hemochromatosis protein (HFE) complex (band B) not observed in S cells. Furthermore, because HFE regulates iron and gallium uptake, the two Tf-TfR1-HFE complexes in R cells may be involved in reduced ⁶⁷Ga and ⁵⁹Fe uptake compared with S cells. In S cells, a novel iron-binding intermediate (band D) was identified that was not present in R cells and may be a “sensitivity factor” to gallium. In contrast to the general view that ⁶⁷Ga and ⁵⁹Fe use the same or similar uptake pathways, we show that their distribution and trafficking is markedly different in R and S cells.