PS1317 VITAMIN D3 INHIBITS GROWTH OF HODGKIN CELL LINES AND IMPROVES SENSITIVITY TO CHEMOTHERAPY AT PHYSIOLOGICAL DOSES IN VITRO AND IN VIVO

Background: Vitamin D (calcitriol) is essential to normal cell function and has known anti‐cancer activity, including anti‐proliferative effects 1 . Recently, our group reported vitamin D deficiency leads to reduced progression‐free and overall survival in a large cohort of prospectively treated first‐line Hodgkin Lymphoma (HL) patients 2 . Vitamin D supplementation may be synergistic with other cancer therapies 1 although an effect in HL has not been described. Aims: (1) To investigate the in vitro effect of calcitriol on HL cell line growth; alone and in combination with chemotherapy.(2) To examine the in vivo effect of calcitriol in combination with the commonly used chemotherapy drug doxorubicin on tumour growth. Methods: In our in vitro study, we incubated HL cell lines (L428, L1236 and KMH2) 3–5 with calcitriol at varying concentrations (10 –12 M – 10 –8 M). We defined a physiological dose of calcitriol (10 –10  M) prior to analysis 6 . Next, we combined chemotherapy agents (doxorubicin, etoposide, gemcitabine and vinorelbine) with calcitriol to determine dose response curves. Luminescence was measured at 24 hour intervals. We calculated cellular proliferation as relative luminescence units, expressed as a percentage of baseline. For the in vivo study, 8‐week old NSG mice were bilaterally, subcutaneously injected with 5 x10 6 KMH2 cells into each flank and tumour growth was measured by caliper twice weekly. Vitamin D treatment 10,000IU/kg [Colecalciferol] was given subcutaneously twice weekly. Doxorubicin was administered on day 10 (1.5 mg/kg intra‐peritoneal injection) in the chemotherapy treated groups. Established tumours (>50mm 3 ) were compared in 4 groups; controls (n = 6), vitamin D monotherapy (n = 8), doxorubicin monotherapy (n = 8), and combination vitamin D and doxorubicin therapy (n = 8). Results: In our in vitro study, we found that calcitriol is toxic to HL cell lines at a dose of 10 –8 M, 2‐log higher than our defined physiological dose (10 –10 M). For the L428 and KMH2 cell lines, culture with 10 –9 M calcitriol (1‐log above physiological range) also reduced the rate of cellular proliferation although did not inhibit growth completely. Physiological dose calcitriol did not inhibit HL cell proliferation. The in vitro combination of physiological calcitriol with doxorubicin and etoposide did inhibit growth of multiple cell lines at the lower end of dose response curves. In our animal model we demonstrate a markedly reduced rate of tumour growth when vitamin D was administered in combination with doxorubicin (Figure 1). Summary/Conclusion: We report for the first time in HL the anti‐proliferative effect of vitamin D in vitro and in vivo . The physiologically active form of vitamin D (calcitriol) was effective in our cell ine model, and the dietary for of vitamin D (colecaliferol) was effective in our in vivo model. We note that high dose (10 –8 M) calcitriol is cytotoxic to HL cell lines, and that at physiological levels calcitriol in combination with low‐dose chemotherapy has additive anti‐proliferative effects. In addition, we found a marked additive anti‐proliferative effect of the combination of doxorubicin with vitamin D in our KMH2‐NSG mouse model of HL. Our findings suggest vitamin D to be an important factor in the growth of HL tumours. Future studies could explore the mechanism of this effect by examining vitamin D receptor (VDR) activity in vivo and the immune‐modulation effects of supplemental vitamin D therapy (e.g. VDR+ T‐cells) on the microenvironment.