Download (direct link):
The biologically active form of M-CSF is a homodimer. These homodimers can exist as integral cell surface proteins, or may be released from their producer cell by proteolytic cleavage, thus yielding the soluble cytokine.
The M-CSF receptor is a single chain, heavily glycosylated, polypeptide of molecular mass 150 kDa. Its intracellular domain displays tyrosine kinase activity which is capable of autophosphorylation, as well as phosphorylating additional cytoplasmic polypeptides.
GRANULOCYTE-MACROPHAGE COLONY STIMULATING FACTOR (GM-CSF)
GM-CSF is also known as CSF-a or pluripoietin-a. It is a 127 amino acid, single chain, glycosylated polypeptide, exhibiting a molecular mass in the region of 22 kDa. Its 3-D structure exhibits a bundle of four a-helices and a two-stranded antiparallel b-sheet (Figure 6.2). It thus resembles the other CSFs. GM-CSF is produced by various cells (Table 6.3) and studies have indicated that its biological activities include:
• Proliferation/differentiation factor of haemopoietic progenitor cells, particularly those yielding neutrophils (a variety of granulocyte) and macrophages, but also eosinophils, erythrocytes and megakaryocytes. In vivo studies also demonstrate this cytokine’s ability to promote haemopoiesis.
• Activation of mature haemopoietic cells, resulting in:
- enhanced phagocytic activity;
- enhanced microbiocidal activity;
- augmented anti-tumour activity;
- enhanced leukocyte chemotaxis.
The intact GM-CSF receptor is a heterodimer, consisting of a low-affinity a-chain and a b-chain, which also forms part of the IL-3 and IL-5 receptors (the b-chain alone does not bind GM-CSF). The a-chain is a 80 kDa glycoprotein and exhibits only a short intracellular domain. The larger b-chain (130 kDa) displays a significant intracellular domain. Signal transduction involves the (tyrosine) phosphorylation of a number of cytoplasmic proteins (Figure 6.3).
The multiplicity of activities attributed to GM-CSF (particularly by in vitro studies), along with the redundancy exhibited by this cytokine, renders difficult appraisal of its most significant physiological role (this is, of course, also true in the case of many other cytokines). Studies
Figure 6.2. 3-D structure of GM-CSF. Photo from Walter et al. (1992), by courtesy of the Protein Data Bank: http://www.pdb.org/
designed to elucidate its exact role, by generation of mutant mice lacking GM-CSF, have yielded surprising results. These mice were found to be healthy and fertile, and were capable of sustaining normal steady-state haemopoiesis. In fact, the only physiological abnormality revealed was associated with the lungs, where a major build-up of surfactant was noted (surfactant is a mixture of various phospholipids and proteins, which appears to function mainly to reduce surface tension at the alveolar air-liquid interface). Although GM-CSF can undoubtedly influence haemopoiesis when administered experimentally, the above finding casts serious doubt on the physiological significance of such activity. It also illustrates the power of the technique by which knock-out mice, lacking a specific gene, can be generated, and shows how this technique can help delineate the true physiological role of that gene’s product.
HAEMOPOIETIC GROWTH FACTORS 261
^_________J Cell cytoplasm
Figure 6.3. The GM-CSF receptor. Ligand binding appears to promote the phosphorylation of various cytoplasmic polypeptide substrates (at least in part via an associated JAK 2 kinase), leading to signal transduction
CLINICAL APPLICATION OF CSFs
As haemopoietic growth factors serve to stimulate the production of mature blood cells, their clinical application in diseases characterized by sub-optimal production of specific blood cell types was obvious. Several CSF preparations have gained regulatory approval, or are currently being assessed in clinical trials (Table 6.4). G-CSF and GM-CSF have proved useful in the treatment of neutropenia. All three CSF types are (or are likely to be) useful in the treatment of infectious diseases, some forms of cancer and the management of bone marrow transplants (Table 6.4), as they stimulate the differentiation/activation of white blood cell types most affected by such conditions.
Table 6.4. Colony-stimulating factors approved for medical use or in clinical trials
Product Status Indication Company
Nepogen (filgrastim; Approved Neutropenia caused by Amgen Inc.
Bone marrow transplants
Leukine (sargramostim; Approved Autologous bone marrow Immunex