Good Things Come in Threes: Genetically Engineered Neural Stem Cells Mitigate Chronic CNS Autoimmunity
Authors: David Pleasure
Journal: Molecular Therapy (2016)

Patients with relapsing-remitting multiple sclerosis (MS) are now customarily treated with immunomodulatory drugs. These treatments are often successful in reducing the incidence of new acute inflammatory demyelinative plaques but are far less capable of preventing disseminated central nervous system (CNS) axon loss and the resulting progression of nonremitting clinical neurological deficits. This dichotomous response to these immunomodulatory therapies suggests that there is some fundamental pathophysiological difference in MS between the acute plaques and the disseminated axon loss.1Mahad DH Trapp BD Lassmann H Pathological mechanisms in progressive multiple sclerosis.Lancet Neurol. 2015; 14: 183-193Abstract Full Text Full Text PDF PubMed Scopus (712) Google Scholar As reported in this issue of Molecular Therapy,2Li X Zhang Y Yan Y Ciric B Ma C-G Gran B et al.Neural stem cells engineered to express three therapeutic factors mediate recovery from chronic stage CNS autoimmunity.Mol Ther. 2016; 24: 1456-1469Abstract Full Text Full Text PDF PubMed Scopus (38) Google Scholar Li et al. employed a popular chronic MS model, experimental autoimmune encephalomyelitis (EAE), induced in C57BL/6 mice by immunization against myelin oligodendrocyte glycoprotein peptide 35–55 (MOG peptide),3Soulika AM Lee E McCauley E Miers L Bannerman P Pleasure D Initiation and progression of axonopathy in experimental autoimmune encephalomyelitis.J Neurosci. 2009; 29: 14965-14979Crossref PubMed Scopus (109) Google Scholar to develop a therapy based on genetically engineered, bone marrow–derived neural stem cells (bNSCs) that might prove effective against MS progression. MOG peptide EAE in C57BL/6 mice is characterized by the subacute onset of multifocal inflammatory demyelinative CNS lesions and accompanying weakness, followed by long-lasting, nonremitting weakness, progressive CNS axon loss, and persistently poor CNS remyelination.3Soulika AM Lee E McCauley E Miers L Bannerman P Pleasure D Initiation and progression of axonopathy in experimental autoimmune encephalomyelitis.J Neurosci. 2009; 29: 14965-14979Crossref PubMed Scopus (109) Google Scholar In the new study, Li et al. transduced murine bNSCs with a construct that expresses a “cocktail” comprising the anti-inflammatory cytokine interleukin-10 (IL-10), the neurotrophic factor NT-3, and the soluble LINGO antagonist, LINGO-1-Fc, following exposure of the cells to tetracycline. LINGO-1 antagonists are able to promote remyelination in the CNS through stimulating differentiation of oligodendrocyte precursor cells. When these engineered bNSCs were administered intravenously during the subacute phase of MOG peptide EAE, they trafficked efficiently to the CNS, and the weakness of the mice was substantially diminished. Administration of the engineered bNSCs later in the course of this illness diminished the severity of long-term weakness and was accompanied by (i) a shift in spinal cord microglial polarization toward M2 (alternative activation), (ii) improved spinal cord axonal transport, as evidenced by a diminution in axonal accumulation of amyloid precursor protein, and (iii) enhanced spinal cord myelin content. Microglial polarization toward M2 has been linked to reduced inflammation and the restoration of homeostasis. Li et al. observed that the infused bNSCs did not give rise to long-lasting neurons or oligodendroglia but instead served as an effective vehicle for delivery of the three therapeutic factors (IL-10, NT-3, and LINGO-1-Fc) to the CNS of the MOG peptide EAE mice. Li and colleagues thus demonstrated that bNSCs can substitute for CNS-derived NSCs as a CNS delivery system for immunomodulatory, trophic, and pro-regenerative factors, substantially increasing the potential feasibility of this approach for treatment of patients with primary or secondary progressive MS. In additional experiments, the authors showed that omission from the bNSCs of one or more of the three therapeutic molecules sharply diminished their therapeutic efficacy. Certainly the most exciting finding by Li et al. is that intravenous administration of their triple-transduced bNSCs on day 60 post-immunization with MOG peptide, a time point long after most systemic inflammatory cells have been cleared from the CNS in MOG peptide EAE,3Soulika AM Lee E McCauley E Miers L Bannerman P Pleasure D Initiation and progression of axonopathy in experimental autoimmune encephalomyelitis.J Neurosci. 2009; 29: 14965-14979Crossref PubMed Scopus (109) Google Scholar was still effective in substantially diminishing the severity of long-term neurological disability. Why were IL-10, NT-3, and LINGO-1-Fc reasonable choices for improving the prognosis for recovery in this chronic MS model? The primary beneficial effect of augmenting CNS IL-10 content may have been to suppress long-term activation of deleterious microglial and astroglial innate immune responses.4Mayo L Pires Da Cunha A Madi A Beynon V Yang Z Alvarez JI et al.IL-10-dependent Tr1 cells attenuate astrocyte activation and ameliorate chronic central nervous system inflammation.Brain. 2016; 139: 1939-1957Crossref PubMed Scopus (67) Google Scholar,5Hucke S Herold M Liebmann M et al.The farnesoid-X-receptor in myeloid cells controls CNS autoimmunity in an IL-10-dependent fashion.Acta Neuropathol. 2016; (e-pub ahead of print 6 July 2016)PubMed Google Scholar NT-3, in addition to exerting immunomodulatory effects, augments oligodendroglial myelin protein synthesis and remyelination, at least in part via a post-transcriptional mechanism.6Coelho RP Yuelling LM Fuss B Sato-Bigbee C Neurotrophin-3 targets the translational initiation machinery in oligodendrocytes.Glia. 2009; 57: 1754-1764Crossref PubMed Scopus (26) Google Scholar,7Yang J Yan Y Xia Y Kang T Li X Ciric B et al.Neurotrophin 3 transduction augments remyelinating and immunomodulatory capacity of neural stem cells.Mol Ther. 2014; 22: 440-450Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar Finally, inhibiting LINGO-1, by administration of either monoclonal antibodies8Mi S Miller RH Tang W Lee X Hu B Wu W et al.Promotion of central nervous system remyelination by induced differentiation of oligodendrocyte precursor cells.Ann Neurol. 2009; 65: 304-315Crossref PubMed Scopus (254) Google Scholar or NSCs transduced to express the soluble LINGO-1 antagonist LINGO-1-Fc,9Li X Zhang Y Yan Y Ciric B Ma CG Chin J et al.LINGO-1-Fc-transduced neural stem cells are effective therapy for chronic stage experimental autoimmune encephalomyelitis.Mol Neurobiol. 2016; (e-pub ahead of print 6 June 2016)Google Scholar enhances both axonal survival and remyelination in MOG peptide EAE. Li and colleagues' study has some limitations. The conclusion that infusion of the engineered bNSCs rescued axons and enhanced remyelination when administered during the chronic phase of MOG peptide EAE was based on light microscopic analysis of myelin basic protein and toluidine blue–stained spinal cord histological sections at a single time point. By this means, they documented increased myelin basic protein immunoreactivity, greater numbers of myelinated axons, and diminished myelinated axon g-ratios (indicative of thicker myelin sheaths) in mice that had received bNSCs engineered to express all three therapeutic molecules in comparison to mice given bNSCs engineered to express only one or two of the three therapeutic molecules. However, those results do not conclusively discriminate between protection of existing myelin and enhanced remyelination of demyelinated axons. Transmission electron microscopic studies of the mice at multiple time points would be necessary to confirm that their engineered bNSCs elicited axonal rescue and remyelination by documenting increased numbers of unmyelinated and high g-ratio (i.e., remyelinating) axons. This same research group had reported such a transmission electron microscopic study in an earlier publication.7Yang J Yan Y Xia Y Kang T Li X Ciric B et al.Neurotrophin 3 transduction augments remyelinating and immunomodulatory capacity of neural stem cells.Mol Ther. 2014; 22: 440-450Abstract Full Text Full Text PDF PubMed Scopus (52) Google Scholar However, the bNSCs employed in that earlier publication had been engineered to express only NT-3; furthermore, they were intravenously infused only during the subacute and not the chronic phase of MOG peptide EAE. Another limitation of the present study is that MOG peptide EAE, although manifesting most severely in the spinal cord, gives rise to inflammatory demyelinative lesions and axon loss in all regions of the CNS; these are likely to contribute to the clinical neurological deficits. Therefore, the effects of the engineered bNSCs on forebrain, brainstem, and cerebellum of the MOG peptide EAE mice deserve scrutiny. Moreover, because lymph node–like meningeal immune cell aggregates appear to play an important role in the progression of MS,10Howell OW Reeves CA Nicholas R Carassiti D Radotra B Gentleman SM et al.Meningeal inflammation is widespread and linked to cortical pathology in multiple sclerosis.Brain. 2011; 134: 2755-2771Crossref PubMed Scopus (539) Google Scholar it would be particularly interesting to evaluate the effects of the engineered bNSC infusions on the accumulation and properties of immune cell populations within the meninges of the MOG peptide EAE mice. Finally, because of recent clinical observations that therapies directed against B cells can slow worsening of unremitting neurological deficits in patients with secondary and primary progressive MS, it would be useful to examine the effects of the engineered bNSC infusions on the B-cell lineage in the periphery, the meninges, and the CNS. Li and colleagues were not the first to explore a multipronged therapy for chronic MOG peptide EAE. For example, Basso et al.11Basso AS Frenkel D Quintana FJ Costa-Pinto FA Petrovic-Stojkovic S Puckett L et al.Reversal of axonal loss and disability in a mouse model of progressive multiple sclerosis.J Clin Invest. 2008; 118: 1532-1543Crossref PubMed Scopus (184) Google Scholar were motivated to investigate data indicating roles for oxidant injury and impaired CNS glutamate homeostasis in damaging axons and the oligodendroglial lineage in EAE and MS. They found that intraperitoneal administration of the antioxidant fullerene conjugated to an NMDA receptor antagonist diminished axon loss and demyelination in a variant of the chronic MOG peptide model in which substitution of NOD for C57BL/6 mice better simulated the clinical transition of relapsing-remitting to secondary progressive EAE. The results of this study suggest that engineering bNSCs to express an antioxidant and/or an ionotropic glutamate receptor antagonist in addition to IL-10, NT-3, and LINGO-1-Fc might further improve therapeutic efficacy. How successful has the rodent EAE model at predicting therapeutic responses in human MS? The immunomodulatory drugs glatiramer acetate and natalizumab, first validated in EAE, have since proven efficacious in patients with MS. However, therapy with γ-interferon or inhibitors of tumor necrosis factor-α, both of which diminish EAE severity, instead increased the severity of MS.12Steinman L Zamvil SS Virtues and pitfalls of EAE for the development of therapies for multiple sclerosis.Trends Immunol. 2005; 26: 565-571Abstract Full Text Full Text PDF PubMed Scopus (198) Google Scholar Therefore, we cannot be sure that delivery to the CNS of bNSCs expressing the particular molecules chosen by Li et al. will slow MS progression. Also, although the authors convincingly demonstrated the efficacy of intravenously infused bNSCs as a murine CNS delivery system, how effectively will they deliver therapeutic molecules to the larger brains of patients with MS? These questions might best be addressed by advancing from murine MOG peptide EAE to studies of MOG peptide EAE in nonhuman primates.13‘t Hart BA van Kooyk Y Geurts JJ Gran B The primate autoimmune encephalomyelitis model; a bridge between mouse and man.Ann Clin Trans Neurol. 2015; 2: 581-593Crossref PubMed Google Scholar