Consequences of Intermittent PTH Treatment on Myeloma Growth, Bone Disease and Molecular Profiling of Whole Myelomatous Bone

Abstract 4063

Intermittent PTH (iPTH) treatment is bone anabolic and has been successfully used for treatment of osteoporosis. The aims of the study were to investigate the effects of iPTH-induced bone formation on myeloma progression and unravel molecular mechanisms associated with these effects, using the SCID-rab and SCID-hu models. Using our reported procedure (Xin et al., BJH 2007), we established a novel myeloma cell line, Hg, capable of sequential passaging in experimental models. Hg cells have similar gene expression profiling (GEP) as the original patient's plasma cells, are classified in the MMSET subtype and express DKK1. SCID-hu or SCID-rab (8-10 hosts/group in each model) mice engrafted with Hg myeloma cells were subcutaneously treated with saline or iPTH (80 ug/kg/day) for 4 weeks. Overall, whereas BMD of the myelomatous bones in saline-treated hosts was similarly reduced in SCID-rab and SCID-hu mice by 14±5%, it was increased by 10±2% in iPTH-treated hosts (p<0.002 saline vs. iPTH-treated hosts). Histologic and histomorphometric analyses revealed increased bone formation parameters and no effect on number of osteoclasts. The bone anabolic effect of iPTH was associated with reduced myeloma growth by >50% (p<0.03) assessed by measurement of human immunoglobulin level in mice sera and histologically. Treatment with iPTH also increased BMD and attenuated myeloma growth in SCID-rab mice engrafted with myeloma cells from 10 patients. We found by qRT-PCR that type-1 PTH receptor was not expressed by myeloma cells and that PTH had no direct effect on in vitro growth of myeloma cells indicating that PTH anti-myeloma effect is indirectly mediated through modulation of the BM microenvironment. To shed light on molecular mechanisms associated with iPTH effects, human GEP and qRT-PCR validation of selected genes were preformed on whole myelomatous human bones from Hg-bearing SCID-hu mice treated with saline or iPTH for 4 weeks (5 hosts/group). Mice were sacrificed 2 hours after the last injection. Treatment with iPTH upregulated 343 genes and downregulated 410 genes ('2 folds, p<0.05) in myelomatous bones. There was a remarkable alteration in expression of genes associated with cAMP (e.g. RGS1/2 upregulation) and Wnt (e.g. LRP4 upregulation; DKK1 downregulation) signaling, upregulation of growth factors and receptors involved in bone remodeling (e.g. FGFR1/2, FDGFA, FDGFRA, TGFB, TGFBR1), and increased expression of osteoblast markers (e.g. osteocalcin, RUNX2). Interestingly, although iPTH induced upregulation of RANKL there was a reduction in expression of osteoclastic genes (e.g. ACP5/TRAP, NFATC1), probably due to increased osteoblast numbers, downregulation of inflammatory genes (e.g. AIF1) and upregulation of anti-inflammatory genes (e.g. TNFAIP6, CXCL14). Moreover, iPTH reduced expression of typical myeloma associated genes (e.g. CD38, WHSC1, IRF4) and had no effect on expression of myeloma growth factors such as IL6 and IGF1. Expression of certain documented anti-myeloma factors was upregulated by iPTH (e.g. decorin). We conclude that iPTH-induced bone formation in myelomatous bones is mediated by activation of multiple signaling pathways involved in osteoblastogenesis, and attenuated bone resorption and myeloma growth through mechanisms involving increased production of anti-myeloma factors by osteoblasts and minimizing inflammatory conditions induced by myeloma. Treatment with iPTH may be a promising approach for myeloma bone disease and tumor progression.