Nerve myelination is an essential process in the formation and function of the central nervous system (CNS) and peripheral nervous system (PNS) compartments. The myelin sheath around axons is necessary for the proper conduction of electric impulses along nerves. Loss of myelin occurs in a number of diseases, among which are Multiple Sclerosis (MS) affecting the CNS, Guillain-Barre Syndrome, CIDP and others (see Abramsky and Ovadia, 1997; Trojaborg, 1998, Hartung et al, 1998). While of various etiologies, such as infectious pathogens or auto-immune attacks, demyelinating diseases all cause loss of neurological functions and may lead to paralysis and death. While present therapeutical agents reduce inflammatory attacks in MS and retards disease progression, there is a need to develop therapies that could lead to remyelination and recovery of neurological functions (Abramsky and Ovadia, 1997, Pohlau et al, 1998).
The synthesis of myelin is a function of specialized glial cells: the oligodendrocytes in the CNS and the myelinating Schwann cells in the PNS. These two cell types in their fully differentiated state may be called myelinating cells. Myelin is a lipid membrane structure containing a number of different proteins. Myelin basic proteins (MBP) represent the major components (30%) of CNS and also of PNS myelin proteins. Expression of the MBP and other genes encoding the various myelin proteins (e.g. P0, PMP-22, MAG in PNS, PLP, MOG in CNS), is turned on during the terminal differentiation of oligodendrocytes and myelinating Schwann cells. The origin of these cells is in the embryonal neural crest (Fraser, 1991) from which they migrate, and undergo a differentiation that proceeds in a number of steps. Schwann cell (SC) development appears to involve three main steps: 1) the generation of precursors (pSC) from migrating cells; 2) the proliferation and transition to embryonic SC (eSC) expressing the S1100 protein; 3) the postnatal terminal differentiation of part of the eSC population into myelinating SC that express MBP and other myelin proteins (Kioussi and Gruss, 1996). The cells migrating from the neural crest give rise not only to pSC but also to sensory and sympathic neurons, to smooth muscle cells and to cells which reach the skin and hair follicles and become pigmented melanocytes. The fate of the neural crest cells is affected by various inducing factors: differentiation to glial cells, to neurons and to muscles is promoted by Neuregulins such as glial growth factor (GGF), by BMP2/4 and by TGF-β respectively (Anderson, 1997). The differentiation to melanocytes may be promoted by growth factors such as bFGF or PDGF or SDF (Stocker et al., 1991; Anderson, 1997).
The ultimate differentiation of Schwann and oligodendrocyte progenitors into actively myelinating cells and myelination itself seems to depend on signals generated by the interaction between neuronal axons and the glial cells (Lemke and Chao, 1988; Trapp et al, 1988). When axon-Schwann cell contact is interrupted, as after nerve damage, the cells reverse to a non-myelinating state and expression of myelin protein genes is lost (Jessen and Mirsky, 1991). To be able to stimulate myelination or remyelination, after neural diseases or trauma, it would be extremely important to identify factors that are able to induce the synthesis of myelin.
Injury to CNS induced by acute insults including trauma, hypoxia and ischemia can affect both neurons and white matter. Although most attention has been paid to processes leading to neuronal death, increasing evidence suggests that damage to oligodendrocytes, which myelinate axons, is also a specific component of CNS injury. Thus oligodendrocyte pathology was demonstrated at very early phase after brain ischemia (3 hours) in rats, suggesting that these cells are even more vulnerable to excitotoxic events than neuronal cells (Pantoni et al. 1996). One potential candidate mediating cell death is the marked elevation of glutamate concentration that accompanies many acute CNS injuries (Lipton et al. 1994). Indeed, beside neurons even oligodendrocytes were found to express functional glutamate receptors belonging to the AMPA/kainate subtype. Moreover oligodendrocytes display high vulnerability to glutamate application (McDonald et al. 1998).
Neuregulins such as GGF, which act on embryonic Schwann cell precursors, are also survival, growth and maturation factors for postnatal oligodendrocytes and Schwann cells in damaged nerves, and GGF is one of the mitogenic factors provided by axonal contact (Topliko et al, 1996). Recombinant hGGF2 could enhance remyelination upon prolonged administration in a murine model for Multiple Sclerosis (Cannella et al, 1998) or in crushed peripheral nerve (Chen et al, 1998). Another cytokine that is induced in Schwann cells by axonal contact is the Ciliary neurotrophic factor CNTF (Lee et al, 1995). CNTF, as well as leukemia inhibitory factor (LIF), was shown to promote survival of oligodendrocytes from optic nerve cultured in vitro with bFGF or PDGF, and to increase the number of MBP expressing oligodendrocytes in these cultures (Mayer et al, 1994). However, when added to glial precursor cells, CNTF and LIF appear rather to favor astrocyte differentiation and induce expression of the astrocyte GFAP marker, while on oligodendrocytes it would have mainly a survival action with little effect on the level of MBP gene expression (Kahn and De Vellis, 1994, Bonni et al, 1997). Nevertheless, combinations of CNTF with brain-derived neurotrophic factor BNDF improve recovery of an injured peripheral sciatic nerve (Ho et al, 1998).
CNTF and LIF are cytokines acting through a common receptor system which comprises the LIF receptor (LIFR) and the gp130 chain, the latter being also part of the Interleukin-6 (IL-6) receptor complex (Ip et al, 1992). CNTF and LIF are, therefore, part of the IL-6 family of cytokines. In the case of CNTF and LIF, signal transduction operates through dimerization of LIFR with gp130, whereas in the case of IL-6 the signal is generated by the dimerization of two gp130 chains (Murakami et al, 1993). In order to bind gp130, IL-6 makes a complex with an IL-6 Receptor chain, which exists on certain cells as a gp80 transmembrane protein but whose soluble form can also function as an IL-6 agonist when provided from outside the cell (Taga et al, 1989, Novick et al, 1992). By fusing the entire coding regions of the cDNAs encoding the soluble IL-6 receptor (sIL-6R) and IL-6, a recombinant IL6RIL6 chimera can be produced in CHO cells (Chebath et al, 1997, WO99/02552). This IL6RIL6 chimera has enhanced IL-6-type biological activities and it binds with a much higher efficiency to the gp130 chain in vitro than does the mixture of IL-6 with sIL-6R (Kollet et al, 1999).
A review of the effects of IL-6 on cells of the central and peripheral nervous system indicates that the cytokine may have protective effects on neuronal cells as well as participate in inflammatory neuro-degenerative processes (Gadient and Otten, 1997, Mendel et al, 1998). On glial cells, CNTF and LIF were much more active than IL-6 to stimulate astrocyte differentiation and there was no effect on myelin protein producing cells (Kahn and De Vellis. 1994). IL-6 was found to prevent glutamate-induced cell death in hippocampal (Yamada et al. 1994) as well as in striatal (Toulmond et al. 1992) neurons. The IL-6 mechanism of neuroprotection against toxicity elicited by NMDA, the selective agonist for NMDA subtype of glutamate receptors, is still unknown. In fact IL-6 was found to enhance the NMDA-mediated intracellular calcium elevation. In transgenic mice expressing higher levels of both IL-6 and soluble IL-6R (sIL6-R), an accelerated nerve regeneration was observed following injury of the hypoglossal nerve as shown by retrograde labeling of the hypoglossal nuclei in the brain (Hirota et al, 1996). In that work, the addition of IL-6 and sIL-6R to cultures of dorsal root ganglia (DRG) cells showed increased neurite extension in neurons, but no effect on myelinating cells was reported.
In the light of the data presented above, CNTF, LIF or a mixture of IL-6 and sIL-6R have not been shown to induce the terminal differentiation of glial cells into myelinating cells. However, as outlined above, stimulation of myelinating cells differentiation would be of great benefit for patients suffering from demyelinating or neurodegenerative diseases.
Citation of any document herein is not intended as an admission that such document is pertinent prior art, or considered material to the patentability of any claim of the present application. Any statement as to content or a date of any document is based on the information available to applicants at the time of filing and does not constitute an admission as to the correctness of such statement.