Advancing mesenchymal stem/stromal cells-based therapies for neurologic disease

Advancing mesenchymal stem/stromal cells-based therapies for neurologic disease

The past decade has seen a dramatic expansion in the develop‐ment and implementation of experimental cellular therapies in human patients. Mesenchymal stem/stromal cells (MSCs) are at the forefront of this effort, with over 650 registered clinical trials that employ MSCs as the principle therapeutic agent (www.clinical‐trials.gov). MSCs represent a specialized type of stromal cells fi rst identi fi ed in bone marrow based on their capacity to di ff erentiate into skeletal cell types (adipocytes, chondrocytes, and osteoblasts) and support hematopoiesis. MSCs or MSC‐like cells are now be‐lieved to reside in most tissues as perivascular cells or pericytes, with isolates from bone marrow, adipose tissue and umbilical cord being the most widely studied. In addition to their stem/progenitor properties, MSCs have also been shown to possess a broad range of e ff ector functions including angiogenic, anti‐in fl ammatory and im‐muno‐modulatory activities that are associated, in large part, with secretion of paracrine acting proteins and exosomes/micro‐vesicles.erefore, while originally developed for treating skeletal and he‐matologic diseases, MSC‐based therapies now target a diverse array of inflammatory, ischemic, and auto‐immune diseases, with the most promising results coming from clinical trials in patients with steroid resistant graversus host disease (Squillaro et al., 2016).

Shifting paradigms: Studies conducted in the late 1990’s including work from my own laboratory, which showed MSCs injected directly into the CNS of newborn mice acquired characteristics of neural cells (Kopen et al., 1999), suggested that adult stem cells possess broader than expected plasticity.is spurred many labs to identify culture conditions that promoted neural commitment of MSCs for use in cell replacement strategies to treat neurologic diseases. Cell replacement strategies remain a viable option as evidenced by a re‐cent study showing that embryonic stem cell‐derived basal forebrain cholinergic neurons transplanted into two di ff erent mouse models of Alzheimer’s disease functionally integrated into the endogenous basal forebrain cholinergic projection system resulting in improved learning and memory performance (Yue et al., 2015). However, the lack of de fi nitive evidence showing that MSCs can be reprogrammed to generate electrically excitable neurons suggests their use in this regard may be limited. Nevertheless, the realization that chronic inflammation and pathogenic immune responses are prominent features of many neurological disorders has provided a new path forward to exploit the anti‐in fl ammatory and immuno‐suppressive properties of MSC as a therapeutic option for these diseases. To date MSC‐based clinical trials have been evaluated in patients afflicted with cerebral ataxia, amyotrophic lateral sclerosis, multiple sclero‐sis, multiple systems atrophy, Parkinson’s disease, and Alzheimer’s disease , and the number of these clinical trials is exceeded only by those directed at skeletal‐related diseases (Squillaro et al., 2016).

Non-human primates as a model system for translations studies: Translational studies conducted in large animal models can be extremely valuable for advancing human clinical trials due to their similarities to human anatomical structure, host immune respons‐es, and disease pathophysiology. Over the past decade our laborato‐ry conducted a series of pre‐clinical studies in non‐human primates to evaluate the engraftment kinetics, anatomical distribution and transplant immunology of MSCs following direct intracranial injection, with the goal of exploiting the cells to treat neurologic sequelae associated with lysosomal storage diseases. For example, we demonstrated that stereotactic‐guided injection of unmatched MSCs from a male donor into the caudate putamen of female, infant macaques was safe and well‐tolerated based on physical ex‐aminations, body weight measures, and serological testing (Isakova et al., 2007). Using a battery of age‐ and species‐appropriate tests we further showed that MSC administration yielded no adverse e ff ects on animal cognition, fi ne and course motor function, behav‐ior, or neural development up to 6 months post‐transplant.ese fi ndings were signi fi cant since animals were monitored throughout their fi rst year of life during which social behavior, motor skills and cognitive abilities are rapidly developing. Analysis of brain tissue further revealed that overall MSC engraftment levels were 17.8‐fold higher (P＜ 0.05) in infantvs. young adult transplant recipients with a maximal observed di ff erence of 180‐fold.is result was also highly favorable in that patients with storage diseases that involve neurodegeneration develop neurologic complications at an early age and therefore early intervention is critical toward retarding dis‐ease progression. In subsequent studies we showed that intracranial administration of allogeneic but not autologous MSCs induced a weak allograft response that involved expansion of NK, B and T cell subsets in peripheral blood, and that its magnitude was depen‐dent upon the degree of MHC mismatch between the MSC donor and transplant recipient. This finding was further substantiated by the fact that secondary challenge with allogeneic donor MSCs induced allo‐antibody production and elevated levels of CD3−CD16+HLADR+myeloid dendritic cells, which play a major role in peripheral tolerance (Isakova et al., 2010, 2014).

Using knowledge gained from these studies, we then set out to test the e fficacy of MSCs in a non‐human primate model of early onset Krabbe disease (Baskin et al., 1998). Krabbe disease is one of over 50 types of lysosomal storage diseases in humans and is caused by a de fi ciency in galactosylceramidase (GALC) activity.e dis‐ease is characterized by loss of myelin‐forming oligodendrocytes and progressive demyelination resulting in severely impaired motor function. Disease symptoms including marked irritability, spastici‐ty, and seizures appear within 3–6 months of age in human infants and the disease is often fatal by the second year of life with few e ff ective treatment options. Neuro‐in fl ammation that manifests as robust astrogliosis, microglial activation, and macrophage recruit‐mentis also now recognized as a critical aspect of the pathophys‐iology of this disease (Potter and Petryniak, 2016). Consequently, MSC administration may disrupt the feed forward loop of microg‐lia activation, oligodendrocyte death, demyelination, and inflam‐mation characteristic of this disease.

To assess this we treated an infant rhesus macaque that exhibit‐ed symptoms consistent with severe early onset Krabbe disease in humans including a noticeable tremor, weak posture and impaired ambulation (Isakova et al., 2016). This diagnosis was confirmed by lack of GALC activity in the animal’s peripheral blood cells. At seven weeks of age the animal was administeredviadirect intracranial injec‐tionpartially matched MSCs from a healthy donor, which expressed a similar repertoire of Mamu A1 and Mamu E alleles in order to mini‐mize the risk of allograreaction. While we still observed a transient increase in peripheral blood lymphocyte counts several weeks post MSC administration, we were unable to detect evidence of allo‐anti‐body production in the transplant recipient aer secondary antigen challenge, suggesting that the donor cells were weakly immunogenic.

The infant also exhibited extremely low conduction velocities (CVs) for the tibial, ulnar and median nerves consistent with a diagnosis of severe Krabbe disease. At one month post MSC ad‐ministration CVs for the tibial nerve, which controls hind limb movements and posture, increased by 3.7‐fold, and this change was consistent with a measurable improvement in coordination, leg and arm resistance, and ambulation based on physical examinations. CV values for the ulnar nerve also increased by 1.5‐fold over base‐line and remained at or above pre‐treatment levels for the duration of the study. These changes were accompanied by a marked de‐crease in distal latencies measured for the tibial and ulnar nerves at 30 days post MSC administration.erefore, despite the severity of its symptoms, the afflicted infant responded rather rapidly to MSC administration based as evidenced by improved nerve conduction velocities, motor control, ambulation and learning. While this cases study did not assess e ff ects on neuro‐in fl ammation, outcomes are consistent with previous reports indicating that MSCs exert trophic e ff ects that improve survival of myelin‐producing oligodendrocytes when transplanted in a rodent model of toxicity‐induced demyelin‐ation (Jaramillo‐Merchan et al., 2013) and that MSC conditioned media reduces functional de fi cits in a mouse model of experimen‐tal autoimmune encephalitis by promoting maturation of oligo‐dendroglial progenitors toward mature myelin producing cells (Bai et al., 2012). These positive outcomes provide a basis for further testing using this large animal model, which provides a means to rigorously pursue mechanistic‐based studies that can inform clini‐cal trials to improve e ffi cacy in human patients.

Refinements for moving forward: A growing number of early stage clinical trials have demonstrated the safety of MSC‐based therapies in humans, and completed trials to date suggest that MSCs may be e ffi cacious in treating a range of neurologic disorders (Squillaro et al., 2016). However, while MSC‐based therapies have shown a clear bene fi t in some patient populations, other trails have yielded suboptimal results or failed to meet their primary e fficacy endpoints. Developing efficacious MSC‐based therapies is criti‐cally dependent on a number of key factors, such as selection of the appropriate patient population, adequate rigor in trial design, development of appropriate dosing strategies, and selection of rigorous and well‐de fi ned metrics to assess patient outcomes. How‐ever, choice of the appropriate human MSC donor population and the manufacturing scheme employed to generate clinical cell doses is also important but oen overlooked despite the fact that human MSC populations exhibit signi fi cant donor‐to‐donor heterogeneity.erefore, there remains a critical need to develop metrics to assess the relative potency of clinical grade MSC preparations so they can be carefully matched to the appropriate patient populations. As a step toward this goal our laboratory recently described a Clinical Indica‐tions Prediction (CLIP) scale that predicts the therapeutic ef fi cacy of di ff erent human MSC isolates for a given disease indication based onTWIST1expression levels.is scale arose from our studies showing that stem/progenitor and e ff ector functions of MSCs are coordinate‐ly regulated byTWIST1, and that one could predictably alter these properties in MSCs by manipulating expressed levels of this protein (Boregowda et al., 2016). While continued validation of the CLIP scale is needed, the scale itself is advantageous over other potency assays as it predicts di ff erences in growth, survival, stem/progentior, and effector functions of MSCs rather than just a single function, and can easily be correlated to quanti fi able functional assays. For ex‐ample, we reported thatTWIST1levels are positively correlated with CFU‐F activity.erefore, use of a standard CFU‐F assay provides a simple means to orient a given MSC preparation on the CLIP scale. Importantly, the scale can be expanded to incorporate additional metrics as the role ofTWIST1in MSCs is further explored.

Conclusion: The number and scope of MSC‐based clinical trials continues to show robust expansion both with respect to treatment of neurologic diseases as well as many other maladies. However, as more advanced phase trials that employ MSCs are completed their outcomes will dictate the long‐term viability of the fi eld. To ensure their success, it is essential to pursue development of potency as‐says that serve as reliable predictors ofin vivoe ffi cacy and that are also acceptable to the appropriate regulatory authorities who over‐see trial approval.e di fficulty in developing such assays to date clearly indicates the need for continued translational research using the most appropriate animal models available. Indeed, robust col‐laborative e ff orts that engage basic scientists, disease model experts and clinicians is needed in order to develop MSC‐based therapies to their full potential.

Donald G. Phinney＊

＊Correspondence to: Donald G. Phinney, Ph.D., dphinney@scripps.edu.

Accepted: 2017-01-11

orcid: 0000-0002-8688-2619 (Donald G. Phinney)

Bai L, Lennon DP, Caplan AI, DeChant A, Hecker J, Kranso J, Zaremba A, Miller RH (2012) Hepatocyte growth factor mediates mesenchymal stem cell‐induced recov‐ery in multiple sclerosis models. Nat Neurosci 15:862‐870.

Baskin GB, Ratterree M, Davison BB, Falkenstein KP, Clarke MR, England JD, Vanier MT, Luzi P, Ra fi MA, Wenger DA (1998) Genetic galactocerebrosidase deficiency (globoid cell leukodystrophy, Krabbe disease) in rhesus monkeys (Macaca mulatta). Lab Anim Sci 48:476‐482.

Boregowda SV, Krishnappa V, Haga CL, Ortiz LA, Phinney DG (2016) A clinical indications prediction scale based on TWIST1 for human mesenchymal stem cells. EBioMedicine 4:62‐73.

Isakova IA, Baker KC, Dufour J, Phinney DG (2016) Mesenchymal stem cells yield transient improvements in motor function in an infant rhesus macaque with severe early‐onset Krabbe disease. Stem Cells Transl Med DOI:10.5966/ sctm.2015‐0317.

Isakova IA, Dufour J, Lanclos C, Bruhn J, Phinney DG (2010) Cell‐dose‐dependent increases in circulating levels of immune e ff ector cells in rhesus macaques follow‐ing intracranial injection of allogeneic MSCs. Exp Hematol 38:957‐967.e951.

Isakova IA, Baker K, DuTreil M, Dufour J, Gaupp D, Phinney DG (2007) Age‐ and dose‐related e ff ects on MSC engrament levels and anatomical distribution in the central nervous systems of nonhuman primates: identi fi cation of novel MSC subpopulations that respond to guidance cues in brain. Stem Cells 25:3261‐3270.

Isakova IA, Lanclos C, Bruhn J, Kuroda MJ, Baker KC, Krishnappa V, Phinney DG (2014) Allo‐reactivity of mesenchymal stem cells in rhesus macaques is dose and haplotype dependent and limits durable cell engrament in vivo. PLoS One 9:e87238.

Jaramillo‐Merchan J, Jones J, Ivorra JL, Pastor D, Viso‐Leon MC, Armengol JA, Molto MD, Geijo‐Barrientos E, Martinez S (2013) Mesenchymal stromal‐cell transplants induce oligodendrocyte progenitor migration and remyelination in a chronic demyelination model. Cell Death Dis 4:e779.

Kopen GC, Prockop DJ, Phinney DG (1999) Marrow stromal cells migrate through‐out forebrain and cerebellum, and they di ff erentiate into astrocytes aer injection into neonatal mouse brains. Proc Natl Acad Sci U S A 96:10711‐10716.

Potter GB, Petryniak MA (2016) Neuroimmune mechanisms in Krabbe’s disease. J Neurosci Res 94:1341‐1348.

Squillaro T, Peluso G, Galderisi U (2016) Clinical trials with mesenchymal stem cells: an update. Cell Transplant 25:829‐848.

Yue W, Li Y, Zhang T, Jiang M, Qian Y, Zhang M, Sheng N, Feng S, Tang K, Yu X, Shu Y, Yue C, Jing N (2015) ESC‐derived basal forebrain cholinergic neurons ameliorate the cognitive symptoms associated with Alzheimer’s disease in mouse models. Stem Cell Rep 5:776‐790.

10.4103/1673-5374.198978

How to cite this article:Phinney DG (2017) Advancing mesenchymal stem/stromal cells-based therapies for neurologic disease. Neural Regen Res 12(1):60-61.

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