Pamidronate causes apoptosis of plasma cells in vivo in patients with multiple myeloma Sharon Gordon, 1 Miep H. Helfrich, 1 Hamdi I. A. Sati, 2 Michael Greaves, 1 Stuart H. Ralston, 1 Dominic J. Culligan, 2 Richard L. Soutar 3 and Michael J. Rogers 1 1 Department of Medicine and Therapeutics, and 2 Department of Haematology, University of Aberdeen Medical School, Aberdeen, and 3 Haematology Department, Western Infirmary, Glasgow, UK Received 5 February 2002; accepted for publication 18 April 2002 Summary. Anti-resorptive bisphosphonates, such as pamidronate, are an effective treatment for osteolytic dis- ease and hypercalcaemia in patients with multiple myel- oma, but have also been shown to cause apoptosis of myeloma cell lines in vitro. In this study, we found that a single infusion of pamidronate, in 16 newly diagnosed patients with multiple myeloma, caused a marked increase in apoptosis of plasma cells in vivo in 10 patients and a minimal increase in four patients (P <0®05). The nitrogen- containing bisphosphonates pamidronate and zoledronic acid also induced apoptosis of authentic, human bone marrow-derived plasma cells in vitro. Apoptosis of plasma cells in vitro was probably caused by inhibition of the mevalonate pathway and loss of prenylated small GTPases, as even low concentrations (‡ 1 lmol/l) of zoledronic acid caused accumulation of unprenylated Rap1A in cultures of bone marrow mononuclear cells in vitro. GGTI-298, a spe- cific inhibitor of geranylgeranyl transferase I, also induced apoptosis in human plasma cells in vitro, suggesting that geranylgeranylated proteins play a role in signalling path- ways that prevent plasma cell death. Our results suggest that pamidronate may have direct and/or indirect anti- tumour effects in patients with multiple myeloma, which has important implications for the further development of the more potent nitrogen-containing bisphosphonates, such as zoledronic acid, in the treatment of myeloma. Keywords: bisphosphonate, myeloma, apoptosis, pamidro- nate, prenylation. Multiple myeloma (MM) is a plasma cell malignancy associated with high morbidity and short survival. The disease is characterized by the clonal expansion of malig- nant plasma cells within the bone marrow. Factors secreted by myeloma cells and/or other cells in the bone marrow microenvironment stimulate osteoclastic bone resorption, leading to characteristic osteolytic lesions, bone pain, fracture, generalized osteoporosis and hypercalcaemia. The global incidence of MM is increasing but there has been no substantial improvement in prognosis. New and more effective approaches to the treatment of MM are therefore essential. Bisphosphonates, such as pamidronate (PAM) and clodr- onate (CLO), are an important class of agents that inhibit osteoclast-mediated bone resorption and are widely used in the treatment of cancer-induced osteolytic disease (Russell et al, 1999; Kanis & McCloskey, 2000; Lipton, 2000). Both PAM and CLO have been shown in large, controlled clinical trials to reduce tumour-related skeletal morbidity and have become accepted treatment for the lytic bone disease and hypercalcaemia associated with MM (Lahtinen et al, 1992; Berenson et al, 1996, 1998; McCloskey et al, 1998; Kanis & McCloskey, 2000). Zoledronic acid (ZOL), which is a more potent antiresorptive bisphosphonate than PAM, has recently been reported to be at least as effective as PAM in the treatment of tumour-induced hypercalcaemia (Berenson et al, 2001a,b; Major et al, 2001). As well as effectively treating lytic bone disease, PAM therapy has also been associated with increased survival or reduced bone marrow plasmacytosis in some patients with myeloma (Berenson et al, 1998; Dhodapkar et al, 1998), suggesting that these compounds may have anti-tumour effects in addition to their ability to inhibit bone resorption. We and others have found that relatively high concentrations (100 lmol/l) of PAM, ZOL and other nitrogen-containing bisphosphonates are cytotoxic, cause cell-cycle arrest and induce apoptosis in human myeloma cell lines in vitro (Shipman et al, 1997, 1998; Aparicio et al, 1998; Derenne et al, 1999; Tassone Correspondence: Dr M. J. Rogers, Bone Research Group, Depart- ment of Medicine and Therapeutics, University of Aberdeen Medical School, Polwarth Building, Foresterhill, Aberdeen AB25 2ZD, UK. E-mail: m.j.rogers@abdn.ac.uk British Journal of Haematology, 2002, 119, 475–483 Ó 2002 Blackwell Publishing Ltd 475