FMS Kinase Inhibitors: Current Status and Future Prospects.

FMS, first discovered as the oncogene responsible for Feline McDonough Sarcoma, is a type III receptor tyrosine kinase that binds to the macrophage or monocyte colony-stimulating factor (M-CSF or CSF-1). Signal transduction through that binding results in survival, proliferation, and differentiation of monocyte/macrophage lineage. Overexpression of CSF-1 and/or FMS has been implicated in a number of disease states such as the growth of metastasis of certain types of cancer, in promoting osteoclast proliferation in bone osteolysis, and many inflammatory disorders. Inhibition of CSF-1 and/or FMS may help treat these pathological conditions. This article reviews FMS gene, FMS kinase, CSF-1, IL-34, and their roles in bone osteolysis, cancer biology, and inflammation. Monoclonal antibodies, FMS crystal structure, and small molecule FMS kinase inhibitors of different chemical scaffolds are also reviewed.

E2a/Pbx1 oncogene inhibits terminal differentiation but not myeloid potential of pro-T cells.

E2a/Pbx1 is a fusion oncoprotein resulting from the t(1;19) translocation found in human pre-B acute lymphocytic leukemia and in a small number of acute T-lymphoid and myeloid leukemias. It was previously suggested that E2a/Pbx1 could cooperate with normal or oncogenic signaling pathways to immortalize myeloid and lymphoid progenitor cells. To address this question, we introduced the receptor of the macrophage-colony-stimulating factor (M-CSF-R) in pro-T cells immortalized by a conditional, estradiol-dependent, E2a/Pbx1-protein, and continuously proliferating in response to stem cell factor and interleukin-7. We asked whether M-CSF-R would be functional in an early T progenitor cell and influence the fate of E2a/Pbx1-immortalized cells. E2a-Pbx1 immortalized pro-T cells could proliferate and shifted from lymphoid to myeloid lineage after signaling through exogenously expressed M-CSF-R, irrespective of the presence of estradiol. However, terminal macrophage differentiation of the cells was obtained only when estradiol was withdrawn from cultures. This demonstrated that M-CSF-R is functional for proliferation and differentiation signaling in a T-lymphoid progenitor cell, which, in addition, unveiled myeloid potential of pro-T progenitors. Moreover, the block of differentiation induced by the E2a/Pbx1 oncogene could be modulated by hematopoietic cytokines such as M-CSF, suggesting plasticity of leukemic progenitor cells. Finally, additional experiments suggested that PU.1 and eight twenty-one transcriptional regulators might be implicated in the mechanisms of oncogenesis by E2a/Pbx1.

Recent advances in colony stimulating factor-1 receptor/c-FMS as an emerging target for various therapeutic implications.

Colony stimulating factor-1 (CSF-1) is one of the most common proinflammatory cytokine responsible for various inflammatory disorders. It has a remarkable role in the development and progression of osteoarthritis, cancer and other autoimmune disease conditions. The CSF-1 acts by binding to the receptor, called colony stimulating factor-1 receptor (CSF-1R) also known as c-FMS resulting in the cascade of signalling pathway causing cell proliferation and differentiation. Interleukin-34 (IL-34), recently identified as another ligand for CSF-IR, is a cytokine protein. Both, CSF-1 and IL-34, although two distinct cytokines, follow the similar signalling pathway on binding to the same receptor, CSF-1R. Like CSF-1, IL-34 promotes the differentiation and survival of monocyte, macrophages and osteoclasts. This CSF-1R/c-FMS is over expressed in many cancers and on tumour associated macrophages, consequently, have been exploited as a drug target for promising treatment for cancer and inflammatory diseases. Some CSF-1R/c-FMS inhibitors such as ABT-869, Imatinib, AG013736, JNJ-40346527, PLX3397, DCC-3014 and Ki20227 have been successfully used in these disease conditions. Many c-FMS inhibitors have been the candidates of clinical trials, but suffer from some side effects like cardiotoxicity, vomiting, swollen eyes, diarrhoea, etc. If selectivity of cFMS inhibition is achieved successfully, side effects can be overruled and this approach may become a novel therapy for treatment of various therapeutic interventions. Thus, successful targeting of c-FMS may result in multifunctional therapy. With this background of information, the present review focuses on the recent developments in the area of CSF-1R/c-FMS inhibitors with emphasis on crystal structure, mechanism of action and various therapeutic implications in which c-FMS plays a pivotal role. The review on structure activity relationship of various compounds acting as the inhibitors of c-FMS which gives the selection criteria for the development of novel molecules is also being presented.

Evidence for tau expression in cells of monocyte lineage and its in vitro phosphorylation by v-fms kinase.

The v-fms encoded kinase, which is known to associate with cytoskeletal elements, was shown to phosphorylate histiocytic proteins that had physical and immunological attributes in common with tau. Tau is a microtubule associated protein that is aberrantly phosphorylated in Alzheimer's disease of the brain. Since c-fms expression is normally exclusive to cells of monocyte lineage, these cells were examined for expression of tau. Heat stable tau was identified in cultured peripheral blood monocytes. This represents the first molecular characterization of tau in cells of non-neural origin.

Recognition of activated CSF-1 receptor in breast carcinomas by a tyrosine 723 phosphospecific antibody.

The macrophage colony stimulating factor receptor (CSF-1R), the product of the c-fms proto-oncogene, plays an important role in regulating the normal proliferation and differentiation of macrophages and trophoblasts. However, the abnormal expression of CSF-1R transcripts and protein by human breast carcinomas has been shown to correlate with advanced stage and poor prognosis. Ligand activated CSF-1R dimers transphosphorylate several tyrosines in their cytoplasmic domains which provide recognition sites for various effector proteins in multiple signal transduction pathways. In cells transformed by the c-fms oncogene, one of the major CSF-1R phosphotyrosines, pTyr723 is important for phenotypic expression of anchorage-independent growth and metastasis. In order to investigate the relationship between receptor activation/phosphorylation and cellular phenotypes in vitro and in vivo, we prepared a CSF-1R phosphorylation-state specific antibody raised against a specific phosphopeptide of CSF-1R, which included phosphorylated tyrosine 723. On immunoblots of lysates from cells expressing CSF-1R, this antibody recognizes phosphorylated CSF-IR in CSF-1 stimulated cells but not in unstimulated cells. As an immunohistochemical reagent, this antibody stained 52% of invasive human breast tumors (72% of CSF-1R positive cases) in a sample of 114 cases and 38% of carcinoma in situ. This data represents the first direct evidence of in vivo phosphorylation of CSF-1R in human breast carcinomas.

SGK1, a potential regulator of c-fms related breast cancer aggressiveness.

The aggressive behavior of breast cancer cells can at times be modulated by hormonal mechanisms. Exposure to glucocorticoids (GC) has been shown to stimulate the invasiveness, motility and adhesiveness of breast cancer cells containing the glucocorticoid receptor. This is largely explained by GC-associated overexpression of the c-fms proto-oncogene, which encodes the receptor for the colony stimulating factor-1 (CSF-1). Our objective is to investigate additional GC-associated genetic alterations that could modulate c-fms related malignant behavior in breast cancer cells. A microarray technique using an oligonucleotide array representing 16,700 known expressed human genes was used to analyze the gene expression profile of breast cancer cells exposed to dexamethasone (Dex) or vehicle. Results were confirmed by western blot analysis. Six genes were found to be consistently differentially overexpressed in the Dex-exposed cells compared to control. We focused on serum-glucose kinase 1 (SGK1), a serine-threonine kinase known to be involved in intracellular signal transduction pathways and induced by GC and serum. An adhesion assay was performed on extracellular matrix after exposing the breast cancer cells to Dex, CSF-1 or to Dex or CSF-1 plus LY294002, a functional inhibitor of SGK1 action. Exposure to LY294002 significantly decreased both CSF-1 and Dex-induced adhesiveness to the level of control cells. SGK1 may act as a downstream intracellular regulator of c-fms, particularly of c-fms-induced adhesiveness of breast cancer cells after exposure to GC or CSF-1. This finding may have implications for potential therapeutic interventions aimed at decreasing the aggressiveness of breast cancer cells.

Autocrine inhibition of the c-fms proto-oncogene reduces breast cancer bone metastasis assessed with in vivo dual-modality imaging.

Breast cancer cells preferentially home to the bone microenvironment, which provides a unique niche with a network of multiple bidirectional communications between host and tumor, promoting survival and growth of bone metastases. In the bone microenvironment, the c-fms proto-oncogene that encodes for the CSF-1 receptor, along with CSF-1, serves as one critical cytokine/receptor pair, functioning in paracrine and autocrine fashion. Previous studies concentrated on the effect of inhibition of host (mouse) c-fms on bone metastasis, with resulting decrease in osteolysis and bone metastases as a paracrine effect. In this report, we assessed the role of c-fms inhibition within the tumor cells (autocrine effect) in the early establishment of breast cancer cells in bone and the effects of this early c-fms inhibition on subsequent bone metastases and destruction. This study exploited a multidisciplinary approach by employing two non-invasive, in vivo imaging methods to assess the progression of bone metastases and bone destruction, in addition to ex vivo analyses using RT-PCR and histopathology. Using a mouse model of bone homing human breast cancer cells, we showed that an early one-time application of anti-human c-fms antibody delayed growth of bone metastases and bone destruction for at least 31 days as quantitatively measured by bioluminescence imaging and computed tomography, compared to controls. Thus, neutralizing human c-fms in the breast cancer cell alone decreases extent of subsequent bone metastasis formation and osteolysis. Furthermore, we are the first to show that anti-c-fms antibodies can impact early establishment of breast cancer cells in bone.

Tumor necrosis factor-alpha increases circulating osteoclast precursor numbers by promoting their proliferation and differentiation in the bone marrow through up-regulation of c-Fms expression.

Osteoclasts are essential cells for bone erosion in inflammatory arthritis and are derived from cells in the myeloid lineage. Recently, we reported that tumor necrosis factor-alpha (TNFalpha) increases the blood osteoclast precursor (OCP) numbers in arthritic patients and animals, which are reduced by anti-TNF therapy, implying that circulating OCPs may have an important role in the pathogenesis of erosive arthritis. The aim of this study is to investigate the mechanism by which TNFalpha induces this increase in OCP frequency. We found that TNFalpha stimulated cell division and conversion of CD11b+/Gr-1-/lo/c-Fms- to CD11b+/Gr-1-/lo/c-Fms+ cells, which was not blocked by neutralizing macrophage colony-stimulating factor (M-CSF) antibody. Ex vivo analysis of monocytes demonstrated the following: (i) blood CD11b+/Gr-1-/lo but not CD11b-/Gr-1- cells give rise to osteoclasts when they were cultured with receptor activator NF-kappaB ligand and M-CSF; and (ii) TNF-transgenic mice have a significant increase in blood CD11b+/Gr-1-/lo cells and bone marrow proliferating CD11b+/Gr-1-/lo cells. Administration of TNFalpha to wild type mice induced bone marrow CD11b+/Gr-1-/lo cell proliferation, which was associated with an increase in CD11b+/Gr-1-/lo OCPs in the circulation. Thus, TNFalpha directly stimulates bone marrow OCP genesis by enhancing c-Fms expression. This results in progenitor cell proliferation and differentiation in response to M-CSF, leading to an enlargement of the marrow OCP pool. Increased marrow OCPs subsequently egress to the circulation, forming a basis for elevated OCP frequency. Therefore, the first step of TNF-induced osteoclastogenesis is at the level of OCP genesis in the bone marrow, which represents another layer of regulation to control erosive disease.

c-fms is present in primary tumours as well as in their metastases in bone marrow.

The expression of c-fms oncoprotein in different primary tumours as well as in their metastases in bone marrow, was shown. All the samples were fixed and processed by the acetone, methyl benzoate, xylene procedure (AMeX), which was suitable for studying oncoprotein expression not only in primary tumours but also in bone marrow (BM) biopsies. Among the patients suffering from acute myeloid leukaemia (AMeL), positive c-fms cells were found in 55% cases. On the contrary, patients with lymphocytic cell disorders have not had detectable c-fms oncogene product in BM biopsies.c-fms oncoprotein was also detected in some primary tumour specimens (lung carcinoma, cervical carcinoma, gastric carcinoma, breast carcinoma and melanoma) and their metastases in BM, while it was not present in normal uterine tissue. There was a positive correlation between c-fms oncoprotein expression in primary and metastatic tumours. Our results showed that c-fms product is confined, not only to some normal, but also to the variety of malignant cells of different origin.