Demyelination the loss of myelin that insulates and protects axons is a common pathological feature of many human neurodegenerative diseases including Multiple sclerosis (MS) and spinal cord injury. Chronic demyelination can ultimately lead to axonal loss, culminating in extensive disability through defects in neurologic function. The demyelination that defines many human neurodegenerative pathologies is progressive over time; however, studies indicate that myelin repair can occur during the course of some of diseases as well as animal models . While it is generally thought that endogenous oligodendrocyte precursor cells (OPCs) are largely responsible for spontaneous remyelination, it is unclear why these cells are only able to transiently induce myelin repair in the presence of ongoing disease. Along these lines, surgical transplantation of ES-derived human OPCs into sites of demyelination are directly capable of inducing remyelination of the damaged axons and/or modifying the local environment to stimulate and support remyelination by endogenous OPCs. Therefore, understanding the mechanisms by which engrafted huOPCs respond to the microenvironmental niche and survive is of critical importance to ensure the long-term survival and function of these cells for sustaining functional recovery. To this end, we have determined that the cytokine IFN-g promotes apoptosis (death) of human ES-derived OPCs and this is dependent, in part, through induction of the chemokine CXCL10. Importantly, we have insight into the molecular mechanisms by which IFN-g/CXCL10 mediates huOPC apoptosis. Moreover, we have made the novel observation that signaling through the chemokine receptor CXCR2 protects huOPCs from IFN-g/CXCL10-mediated death. Therefore, these findings highlight a previously unrecognized mechanism by which OPCs may protect themselves from death during chronic demyelinating diseases. The experiments outlined in this research proposal will further define the mechanisms associated with IFN-g/CXCL10 death as well as protection following CXCR2 signaling. The discoveries revealed from these experiments will potentially be clinically relevant as it may be possible to enhance survival of engrafted huOPCs as well as endogenous OPCs and thus increase the remyelination potential of these cells.
Statement of Benefit to California:
With over 37 million people, California is the most populated state in the US - and continues to grow. Combined with an increasing aged community, the medical demands placed on the health care workers will also continue to accelerate at a dramatic rate. As such, there will an increase in the number of neurologic diseases/injuries in which loss of myelin i.e. demyelination will be a common pathologic feature. Moreover, many of these demyelinating diseases will be chronic in which a patient may have symptoms associated with demyelination for a life-time. Therefore, interventional cell replacement therapies in which a remyelinating cell can be surgically engrafted into a human with a demyelinating disease represents a significant step forward for treatment of these devastating diseases. Indeed, the FDA has recently approved the use of oligodendrocyte progenitor cells (OPCs) derived from human embryonic stem (ES) cells for treatment of demyelination associated with spinal cord injury. A critical feature will be to understand how to sustain the long-term survival of the engrafted cells to insure functional recovery of the patient. This proposal will identify mechanisms associated with survival of OPCs derived from ES cells. Therefore, the long-term gain to California and its citizens that is derived from these experiments is i) the potential for increased health benefits derived from understanding how to sustain engrafted cell survival and improving the overall quality of life for these individuals; ii) increased productivity as a result of patients with demyelinating disease returning to work-force, iii)diminished economic burden on health care industry to care for patients with demyelinating diseases; and iv) employment for scientists/health care workers.
Human oligodendrocyte precursor cells (hOPCs) are capable of mediating remyelination in human demyelinating diseases such as multiple sclerosis (MS), making them an attractive population for cellular therapies. However, several obstacles to clinical translation exist including overcoming the inflammatory environment commonly encountered in demyelinating diseases. The applicant has data suggesting that hOPCs are susceptible to interferon gamma (IFN-g)-induced apoptosis mediated by the secretion of the chemokine CXCL10, and that this IFN-g/CXCL10-induced cell death can be blocked by signaling of the CXCR2 chemokine receptor. In this application, the principal investigator (PI) proposes two aims to further explore the mechanism by which IFN-g, CXCL10, and CXCR2 mediate hOPC cell survival. In Aim 1, human embryonic stem cell (hESC)- and human induced pluripotent stem cell (hiPSC)- derived OPCs will be used in gene silencing experiments to define the signaling pathways responsible for IFN-g/CXCL10-mediated apoptosis and CXCR2-mediated protection from apoptosis. In Aim 2, the PI will use murine models to demonstrate the in vivo relevance of these findings and interrogate the in vivo importance of CXCR2 signaling in protecting transplanted OPCs from apoptosis.
Significance and Innovation:
- Survival of engrafted OPCs in demyelinating diseases is an important problem. Improving the lives of individuals with demyelinating diseases like MS would have enormous impact.
- While the applicant concentrates on an important pathway, the relationship between elucidating the mechanism of cell survival in this model and successful treatment of demyelinating diseases is not clear. Loss of OPCs is an ongoing process in diseases like MS, and there is evidence for remyelination during the course of the disease. Thus, OPC replacement alone is not likely to be curative.
- Reviewers suggested that understanding how to prevent OPC loss through the promotion of protective mechanisms would have greater significance than would the proposed studies. In this regard, a proposal that utilized iPS cell-derived OPCs from unaffected and MS patients to study survival pathways would have greater potential impact.
- While there is some novelty in Aim 2, this project is not particularly innovative, as it relies on a fairly standard set of techniques and approaches.
- The comparison of hESC- versus iPSC-derived OPCs is interesting and may have impact on future treatment strategies.
Feasibility and Experimental Design:
- Although the preliminary data (PD) support the underlying hypothesis and suggest in vivo relevance, there were some discrepancies between statements in the text and what was demonstrated in the provided figures. Reviewers voiced concerns that the magnitude of the data may have been over-interpreted.
- Despite evidence in the PD that additional pathways might contribute to hOPC apoptosis, the applicant does not address these pathways in the experiments, provide rationale for focusing on the described pathway at the exclusion of the others, nor discuss what this might imply for the study of OPC apoptosis in vivo.
- Several aspects of the experimental design are inadequately described in the proposal. Examples include insufficient discussion of controlling for non-specific effects during gene silencing, inadequate descriptions of techniques for measuring cell survival and remyelination, and limited description of the in vivo experiments proposed in Aim 2 (see next bullet point for details).
- Although the proposed in vivo experiments are feasible in theory, there is no clear rationale provided for the choice of in vivo models in relation to human disease. Moreover, the experimental plan lacked the details necessary to convince reviewers that the applicant is prepared to undertake these labor intensive, complex experiments.
- The protocol for inducing good quality hOPCs from pluripotent stem cells is not described, and PD demonstrating the ability of the research team to generate this cell type is not included.
- The proposed research is designed to give clear, albeit incremental, results. Alternate plans are adequately discussed.
- Although human disease models are not used, this project could yield novel data regarding how CXCR2 regulates OPC survival.
- The specific aims and the research proposal have achievable milestones and should be feasible in less than the proposed 3-year timeline.
Principal Investigator (PI) and Research Team:
- The PI has a substantial track record of success in neuroimmunology that supports the present project and has an excellent publication record.
- The research team is appropriately qualified to conduct the proposed research, although a more detailed description their ability to generate hOPCs from iPSC would have been desirable. The collaboration with the co-investigator (Co-I) is integral to the success of the project.
- The salary breakdown for the Co-I is not well described or justified.
Responsiveness to RFA:
- The proposed research adequately and appropriately addresses the goals and objectives of the RFA. It focuses on basic mechanisms by which hOPCs might be affected by cytokines and chemokines known to be present in disease models where OPC transplantation represents a potential therapeutic strategy.