Pathogenesis of bone disease



One of the most important clinical features in multiple myeloma is the development of osteolytic bone disease. The bone destruction in MM is mediated by normal osteoclasts activated by myeloma cells.

Bone omeostasis is regulated by a balance between osteoprotegerin, RANKL, and DKK1.

RANKL increases  osteoclast activity.
DKK1  suppresses osteoblast activity.

Osteoprotegerin is a secreted member of the TNF receptor superfamily which is produced by osteoblasts. It acts as soluble decoy receptor for RANKL, and it blocks its effects. Thus, it inhibits the differentiation and activation of osteoclasts. Serum OPG levels are lower in patients with MM than in healthy individuals. TNF-kappaB-knockout mice have osteopetrotic bones, while mice lacking osteoprotegerin have severe osteoporosis.

RANK (= Receptor activator of nuclear factor-kappaB) is a receptor on osteoclast precursors that induces the formation of osteoclasts by signaling through the NF-kB and Jun N-terminal kinase pathways. RANK ligand (RANKL) is a membrane-bound protein produced by osteoblasts and marrow stromal cells, and it is subsequently cleaved into a soluble form (sRANKL). RANKL binds to RANK on osteoclast precursors and it stimulates their differentiation and activity. RANKL is a member of TNFs.

Myeloma cells express RANKL and induce RANKL expression in bone marrow stromal cells. Myeloma cells also inhibit production and induce degradation of OPG. These effects result in an increased RANKL-to-OPG ratio, that promotes the formation and activation of osteoclasts. Patients with myeloma bone disease have inappropriately low serum and bone marrow levels of OPG.

Osteolytic bone disease in multiple myeloma is due to DKK1 (Dickkopft-related protein 1), an inhibitor of osteoblast function. DKK1 is a soluble inhibitor of the canonical Wnt signaling secreted by MM cells. Inhibition of Wnt/β-catenin/T-cell factor (TCF) signaling induces proliferation of mesenchymal stem cells and inhibits their differentiation into osteoblasts.



Osteoclasts in the bone marrow of a patient with multiple myeloma (courtesy of Dr. Michael G. Bayerl - Hematopathology, Penn State Hershey Medical Center):






Osteoprotegerin is bound, internalized, and degraded by multiple myeloma cells.
Blood. 2002 Oct 15;100(8):3002-7.
Standal T, Seidel C, Hjertner Ø, Plesner T, Sanderson RD, Waage A, Borset M, Sundan A.
This study shows that MM cells can bind, internalize, and degrade OPG, thereby providing a possible explanation for the lower levels of OPG in the BM of patients with MM.

Recombinant osteoprotegerin decreases tumor burden and increases survival in a murine model of multiple myeloma.
Cancer Res. 2003 Jan 15;63(2):287-9.
Vanderkerken K, De Leenheer E, Shipman C, Asosingh K, Willems A, Van Camp B, Croucher P.
In this study, recombinant osteoprotegerin decreased serum paraprotein and tumor burden, and it increased survival in mice injected with 5T33MM cells. Osteoclast number was decreased. This study provide a proof of concept that the inhibition of the interaction between RANKL and RANK with OPG not only inhibits the development of myeloma bone disease, but it also decreases tumor growth and prolongs survival.

Bisphosphonates and osteoprotegerin as inhibitors of myeloma bone disease.
Cancer. 2003 Feb 1;97(3 Suppl):818-24.
Croucher PI, Shipman CM, Van Camp B, Vanderkerken K.
This study assesses the ability of recombinant osteoprotegerin (Fc.OPG) and bisphosphonates to inhibit the development of bone disease in the 5T2MM murine model of MM. Both bisphosphonates and Fc.OPG prevented the development of osteolytic bone lesions in 5T2MM mice.

A phase I study of AMGN-0007, a recombinant osteoprotegerin construct, in patients with multiple myeloma or breast carcinoma related bone metastases.
Cancer. 2003 Feb 1;97(3 Suppl):887-92.
Body JJ, Greipp P, Coleman RE, Facon T, Geurs F, Fermand JP, Harousseau JL, Lipton A, Mariette X, Williams CD, Nakanishi A, Holloway D, Martin SW, Dunstan CR, Bekker PJ.
This is a randomized, double-blind study that evaluated safety and effect on bone resorption of AMGN-0007 in 28 patients with multiple myeloma and 26 patients with breast carcinoma and lytic bone lesions. AMGN-0007 is a recombinant OPG, developed as a potential therapeutic agent in the treatment of bone disease. Patients were randomized to receive a single dose of either AMGN-0007 SC or pamidronate 90 mg IV. AMGN-0007 suppressed bone resorption as indicated by a sustained, and profound decrease of urinary urinary N-telopeptide of collagen (NTX), surrogate marker of bone resorption. Changes were comparable to those observed with pamidronate.

Evaluation and prognostic value of serum osteoprotegerin in multiple myeloma.
Br J Haematol. 2005 Jun;129(5):706-7.
Depil S, Mathiot C, Leleu X, Moreau AS, Faucompré JL, Hennache B, Bauters F, Bataille R, Facon T.
This study of serum levels of OPG in 140 patients with newly diagnosed MM found that OPG >2.4 ng/mL is an adverse independent prognostic factor for overall survival.



Human myeloma cells stimulate the receptor activator of nuclear factor-kappa B ligand (RANKL) in T lymphocytes: a potential role in multiple myeloma bone disease.
Blood. 2002 Dec 15;100(13):4615-21.
Giuliani N, Colla S, Sala R, Moroni M, Lazzaretti M, La Monica S, Bonomini S, Hojden M, Sammarelli G, Barillè S, Bataille R, Rizzoli V.
T cells may regulate bone resorption through cross-talk between RANKL, and interferon gamma (IFN-gamma), which suppresses osteoclastogenesis. This study found that human myeloma cell lines (HMCLs) increased the expression and secretion of RANKL in activated T lymphocytes, and that the release of IFN-gamma by T lymphocytes was reduced in presence of MM cells. RANKL mRNA was up-regulated in T lymphocytes in the BM of MM patients. This suggests that T cells are probably involved in MM-induced osteolysis through overexpression of RANKL.

RANK-Fc: a therapeutic antagonist for RANK-L in myeloma.
Cancer. 2003 Feb 1;97(3 Suppl):802-12.
Sordillo EM, Pearse RN.
RANK-Fc is a recombinant RANKL antagonist formed by fusing the extracellular domain of RANK to the Fc portion of human IgG1. In vitro, RANK-Fc suppresses the formation of osteoclasts in cocultures of MM with bone marrow and osteoblast/stromal cells. This study showed the ability of RANK-Fc to block the development of MM-induced bone disease in vivo. Administration of the RNAAKL antagonist RANK-Fc not only limited the osteoclastogenesis, but it also caused a reduction in tumor burden and serum paraprotein in SCID-hu-MM mice.

RANK ligand and osteoprotegerin in myeloma bone disease.
Blood. 2003 Mar 15;101(6):2094-8.
Sezer O, Heider U, Zavrski I, Kühne CA, Hofbauer LC.



The role of the Wnt-signaling antagonist DKK1 in the development of osteolytic lesions in multiple myeloma.
N Engl J Med. 2003 Dec 25;349(26):2483-94.
Tian E, Zhan F, Walker R, Rasmussen E, Ma Y, Barlogie B, Shaughnessy JD Jr.
In this study, purified plasma cells from the bone marrow of 36 patients with newly diagnosed MM were studied to identify molecules responsible for the osteolytic process. Dickkopf 1 (DKK1) was one of the genes differently expressed between MM patients with and those without focal bone lesions. A recombinant DKK1 protein was found to  inhibit the differentiation of osteoblast precursor cells in vitro.

Antibody-based inhibition of DKK1 suppresses tumor-induced bone resorption and multiple myeloma growth in vivo.
Blood. 2007 Mar 1;109(5):2106-11.
Yaccoby S, Ling W, Zhan F, Walker R, Barlogie B, Shaughnessy JD Jr.
This study tested the effect of anti-DKK1 therapy on bone metabolism and tumor growth in SCID-rab mice. The bone mineral density (BMD) of implanted myelomatous bone in mice treated with DKK1-neutralizing antibodies increased. Histology showed an increased numbers of osteoblasts and reduced number of osteoclasts. At the same time, the MM growth was inhibited.

Dkk1-induced inhibition of Wnt signaling in osteoblast differentiation is an underlying mechanism of bone loss in multiple myeloma.
Bone. 2008 Apr;42(4):669-80.
Qiang YW, Barlogie B, Rudikoff S, Shaughnessy JD Jr.
This study showed that Dkk1 promotes osteolysis by abrogating osteoblast differentiation mediated by bone morphogenic protein 2 (BMP-2). The autocrine Wnt signaling in osteoblasts is necessary to induce BMP-2-mediated differentiation of the pre-osteoblast cells. Wnt signaling alone does not induce this differentiation.

Serum concentrations of DKK-1 correlate with the extent of bone disease in patients with multiple myeloma.
Eur J Haematol. 2008 Jun;80(6):490-4.
Kaiser M, Mieth M, Liebisch P, Oberländer R, Rademacher J, Jakob C, Kleeberg L, Fleissner C, Braendle E, Peters M, Stover D, Sezer O, Heider U.
This study compared serum levels of DKK-1 in 184 untreated MM patients to those of 33 MGUS patients. Serum DKK-1 was elevated in MM as compared with MGUS (mean 11,963 pg/mL vs. 1,993 pg/mL. Serum DKK-1 levels correlated with the Durie and Salmon stage. MM patients without lytic lesions in x-rays had significantly lower DKK-1 levels than patients with lytic bone lesions. Serum DKK-1 correlated with the number of bone lesions.

Myeloma-derived Dickkopf-1 disrupts Wnt-regulated osteoprotegerin and RANKL production by osteoblasts: a potential mechanism underlying osteolytic bone lesions in multiple myeloma.
Blood. 2008 Jul 1;112(1):196-207.
Qiang YW, Chen Y, Stephens O, Brown N, Chen B, Epstein J, Barlogie B, Shaughnessy JD Jr.
MM cell-derived DKK1 disrupts Wnt3a-regulated OPG and RANKL expression in osteoblasts. Wnt3a-induced OPG expression is reduced in osteoblasts cocultured with DKK1-expressing MM cell lines or primary MM cells. DKK1 induces osteolysis by inhibiting Wnt-regulated differentiation of osteoblasts, and by promoting osteoclastogenesis after increase of the RANKL/OPG ratio.



Enhanced production of osteopontin in multiple myeloma: clinical and pathogenic implications.
Br J Haematol. 2003 Oct;123(2):263-70.
Saeki Y, Mima T, Ishii T, Ogata A, Kobayashi H, Ohshima S, Ishida T, Tabunoki Y, Kitayama H, Mizuki M, Katada Y, Asaoku H, Kitano M, Nishimoto N, Yoshizaki K, Maeda M, Kon S, Kinoshita N, Uede T, Kawase I.
This study examined the production of osteopontin (OPN) in MM cells and plasma OPN levels in 30 MM patients, 21 MGUS patients, and 30 healthy volunteers. OPN was produced in large amounts in MM cell lines and BM cells from MM patients, whereas no OPN was produced by BM cells from patients with MGUS. Plasma OPN levels of MM patients were significantly higher than those of MGUS patients and healthy volunteers. OPN levels correlated with progression and osteolysis of MM. Therefore, MM cells produce OPN, which may contribute to osteoclastic bone resorption.

Myeloma cells suppress bone formation by secreting a soluble Wnt inhibitor, sFRP-2.
Blood. 2005 Nov 1;106(9):3160-5.
Oshima T, Abe M, Asano J, Hara T, Kitazoe K, Sekimoto E, Tanaka Y, Shibata H, Hashimoto T, Ozaki S, Kido S, Inoue D, Matsumoto T.
This study showed that MM cells in vitro produce a soluble Wnt inhibitor, secreted Frizzled-related protein 2 (sFRP-2), but not other Wnt inhibitors including sFRP-1, sFRP-3, and dickkopf 1 (DKK-1) at the protein level. Most MM cells from patients with advanced osteolytic lesions also expressed sFRP-2. Secretion of sFRP-2 inhibits bone formation by  inhibiting the canonical Wnt pathway.

Wnt3a signaling within bone inhibits multiple myeloma bone disease and tumor growth.
Blood. 2008 Jul 15;112(2):374-82.
Qiang YW, Shaughnessy JD Jr, Yaccoby S.
These authors studied the effect of Wnt3a on bone disease and growth of MM cells. Wnt3a activated canonical signaling in MM cell lines, but it had no effect on MM cell growth in vitro. Expression of Wnt3a in MM cells conferred no growth advantage in vitro. SCID-hu mice with MM treated with recombinant Wnt3a had increased bone formation and decreased MM growth.

Activin A promotes multiple myeloma-induced osteolysis and is a promising target for myeloma bone disease.
Proc Natl Acad Sci U S A. 2010 Mar 16;107(11):5124-9.
Vallet S, Mukherjee S, Vaghela N, Hideshima T, Fulciniti M, Pozzi S, Santo L, Cirstea D, Patel K, Sohani AR, Guimaraes A, Xie W, Chauhan D, Schoonmaker JA, Attar E, Churchill M, Weller E, Munshi N, Seehra JS, Weissleder R, Anderson KC, Scadden DT, Raje N.



Giampaolo Talamo, M.D.