OSTEOLYSIS IN CANCER: A ROLE FOR THE GENETIC MANIPULATION OF RANK EXPRESSION2

Abstract

Tumour cells induce osteolysis by producing multiple cytokines that indirectly activate osteoclasts; this process is dependent upon surface expression of a protein known as “receptor activator of nuclear factor kB ligand (RANK-L)” on osteoblasts (OB), and subsequent osteoclast (OC) interaction via surface expressed RANK. Harnessing this RANK-RANK-L interaction has potential for reducing cancer osteolysis. The aim of this study is to prevent tumour-induced osteolysis by ablating osteoclast activation.

A monocyte cell line (RAW 264.7) was grown in vitro in the presence of RANK-L and recombinant mouse macrophage colony stimulating factor (rmM-CSF) to produce osteoclasts. Tumour-associated cytokines IL-1a, TNF-a, and IL-6, and the regulatory cytokine osteoprotegerin (OPG) were added to assess osteoclast cell number (cytospin analysis with TRAP staining) and function (resorption pit number on dentine slices). Short interfering sequence of RNA directed towards RANK receptor (RANK RNAi) was used to assess the effect of abrogating RANK-RANK-L signaling in this pathway.

Tumour-associated cytokines failed to significantly alter OC cell number or function in the model tested. When TNF-a, IL-1a, and IL-6 were added together, the effect on OC function was variable, without a clear trend towards OC activation. The addition of the cytokine OPG revealed a trend towards reducing OC function, but this did not reach statistical significance. RANK RNAi also revealed a trend towards reducing OC function in the presence and absence of tumour-associated cytokines.

Tumour associated cytokines failed to enhance OC function using the monocyte cell line RAW 264.7. Both OPG and RANK RNAi revealed a trend towards reducing OC function, although further testing is required to confirm this observation. Future direction with include analysis of fresh bone marrow-derived OC, which may be more appropriate for this model.