Treatment with gallium nitrate: evidence for interference with iron metabolism in vivo.

Gallium, when bound to transferrin, has been previously shown to cause tumor cell cytotoxicity by preventing cellular uptake of transferrin bound iron in vitro. Patients treated with constant infusion gallium nitrate for carcinoma show a rise in serum iron within 6 hr of the start of treatment. Serum iron returns to baseline by 24 hr post-infusion. Atomic analysis of iron and gallium content of Sephadex G-150 fractions of treatment sera indicate that about an equimolar amount of gallium and iron are associated with transferrin. These gallium and iron concentrations result in inhibition of transferrin mediated iron uptake in vitro, and in vivo allow for > 90% saturation of transferrin with metal. All seven patients who completed two courses of gallium therapy exhibited hypochromic microcytic anemia (mean fall in hemoglobin 3.5 grams %). Evidence for red cell iron depletion was confirmed by an increase (mean 3.3-fold) in zinc protoporphyrin levels. Since transferrin receptor increases on gallium treated iron requiring cells in vitro, we assessed cell surface transferrin receptor on peripheral blood lymphocytes by measuring fluorescent transferrin receptor antibody binding. A population of highly transferrin receptor positive cells peaks at 48 hr into the infusion. DNA analysis as well as double staining indicate the majority of transferrin receptor positive cells are unstimulated B lymphocytes. These studies provide the first documentation that constant infusion gallium treatment results in significant interference with iron metabolism and evidence for tissue iron depletion in vivo. These changes may correlate with therapeutic effects of gallium such as tumor response.

Effects of transferrin-indium on cellular proliferation of a human leukemia cell line.

In previous studies, we have demonstrated that transferrin-gallium inhibits cellular proliferation by a mechanism whereby cellular iron utilization is impaired. Since indium, a similar class 3A metal, has not been well studied, we examined its effects on cellular iron uptake and cellular proliferation. In these studies, we provide evidence that indium, when bound to transferrin, has a 50-fold higher effect on inhibition of cellular proliferation than indium added as indium salt. Cells exposed to relatively low concentrations of transferrin-indium exhibit markedly increased transferrin receptor expression but, as with transferrin-gallium, these cells incorporate an inappropriately low amount of iron, suggesting that there is a defect in the release of internalized iron from transferrin. In further studies, we utilize a monoclonal antibody against transferrin receptor that inhibits transferrin-mediated iron uptake. This antibody exhibits a dose-related inhibition of cellular proliferation, and when both transferrin-indium and monoclonal antibody are added to media, there is a more than additive effect on inhibition of cellular proliferation.

Transferrin synthesis by small cell lung cancer cells acts as an autocrine regulator of cellular proliferation.

Since transferrin is required for cellular proliferation, we investigated transferrin synthesis by a small cell lung cancer line (NCI-H510) that survives in serum-free media without added transferrin. Immunoassays for human transferrin demonstrated that these cells contained immunoreactive human transferrin. Immunofluorescence studies showed that the protein is expressed on the surface of cells, presumably bound to transferrin receptor. Media conditioned by NCI-H510 cells support proliferation of human leukemic cells that would not survive in media lacking transferrin. [35S]Methionine incorporation documented transferrin synthesis by NCI-H510 cells as well as three other small cell lines. Transferrin synthesis by NCI-H510 cells increased more than 10-fold when cells entered active phases of the cell cycle, and this increase was seen before large increases in transferrin-receptor expression. Further experiments examining the effects of agents that affect iron metabolism show that the addition of transferrin-iron or hemin to the media is associated with a more rapid initial rate of proliferation and lower rates of transferrin synthesis than control cells. Gallium salts, which inhibit iron uptake, inhibited proliferation of these cells. If the cells recovered from this effect, transferrin synthesis remained greatly increased compared to control. We conclude that transferrin synthesis by these malignant cells is ultimately related to an iron requirement for cellular proliferation. It appears that this synthesized transferrin acts as part of an important autocrine mechanism permitting proliferation of these cells, and perhaps permitting tumor cell growth in vivo in areas not well vascularized.

The place of threaded pin fixation in the treatment of slipped upper femoral epiphysis.

Pin fixation in the treatment of slipped upper femoral epiphysis was evaluated in 60 patients admitted to the authors' hospital in Melbourne between 1970 and 1978. Forty-three cases were reviewed at an average of eight years following initial treatment. For chronic slips, in situ fixation with pins prevented further slip and promoted growth plate closure. The complication rate was low. Best results were achieved with two or three threaded pins placed into the posteroinferior segment of the femoral head to avoid avascular necrosis. Early upper femoral osteotomy was not required, as considerable bone remodeling occurred even after growth plate closure. In contrast, avascular necrosis was common following treatment of acute, severe slips, even with gentle internal rotation of the leg to reduce the acute component of the slip and pinning.