Hello, my name is Neeraj Bajatia. I'm Chief of Neurocritical Care in the Program in Trauma and Professor and Vice Chair in the Department of Neurology at the University of Maryland School of Medicine. I want to thank the organizers and the Congress for inviting me to talk to you today about stem cell therapy in ischemic stroke. Just to start off with a little bit of epidemiology of stroke, it's always important to understand that the incidence of the disease that we're studying, stroke is the second leading cause of death and the third leading cause of death and disability combined worldwide. Interestingly enough, the overall burden of disease when you compare 1990 to 2019 has seen an increase in the burden of stroke. When we think about stroke therapy in recovery, the vast majority of ischemic stroke patients are not eligible for reperfusion therapy such as intravenous thrombolytics or intra-arterial thrombectomy and so the focus is really on neuroprotection and neurorepair. They still remain very important in the management of acute stroke patients. Recovery largely depends on the size of the lesion, the internal milieu of the brain injury itself and the age and comorbid status of the patient and really can be simplified and some would say maybe oversimplified into three broad time points. Time 1 or T1 is really this first time point or epoch where we think of in the acute, hyper-acute setting where you see rapid changes in blood flow and edema, pro-inflammatory mechanisms. T2 represents this kind of bluish area here where you start to see spontaneous behavioral recovery or neurologic recovery. Reparative processes are starting to become initiated, reaching peak levels and suggesting that this may be a golden period for initiating exogenous restorative therapies. And T3, so-called time 3 in the reddish area over here, really thinking about it months later is when you start to see spontaneous behavioral gains that have started to reach a plateau and the responsiveness to many restorative interventions may start to plateau as well. When we think about the rationale for stem cell therapy therefore and in the context of the overall recovery pattern after an ischemic stroke, stem cell therapy works by a process involved integration into the host brain and provide neuroprotection via down regulation of inflammatory immune responses, inhibition of apoptosis in the transplanted host, increase in endogenous repair via vascular regeneration, and induction of host and brain plasticity and migration of endogenous stem cells. So really, when you think about the regenerative therapy in the form of stem cells, whether it be neural stem cells, hematopoietic stem cells, and mesenchymal stem cells, they can prevent the stroke-related tissue damage and promote repair of damaged tissue and enhance functional recovery. Transplanted stem cells have been shown to migrate to areas of ischemia. While there is the potential for differentiation into neurons and neurogenesis, they also have been shown to induce endogenous neural stem cell migration, activate microglia to inhibit inflammatory pathways, and induce angiogenesis. So really, these are the multiple mechanisms by which stem cell therapy may provide benefit to an injured brain. So if we were to look at stem cell-based approach for stroke and stroke therapy, really, you kind of break it down into four different categories, the source of the stem cells, the route of administration, timing of administration, and outcome measurement itself. So in terms of the source of stem cells, we can think of those stem cells that are so-called pluripotential that can differentiate into phenotypes from all three embryonic layers and the risk of oncogenesis mutation. Neural stem cells are the type of multipotent stem cells that can differentiate into neurons or astrocytes. Various forms of neural stem cells located in dentate gyrus and subventricular zone can differentiate in neural stem cells from pluripotential stem cells, fetal, that is the fetal type, can transdifferentiate from somatic cells, i.e. skin, fibroblast, or urine or blood cells. This alleviates the ethical dilemma of stem cells itself. Mesenchymal stem cells derived from adipose or bone marrow tissue have been shown to be promising for regulating microglial phenotypes to reduce inflammatory response after ischemia and stimulate the regenerative processes that are prone to accumulation and aggression in other organs, particularly the lungs itself. Enhancing stem cell for transplantation means to improve stem cell delivery and function. This can be done via multiple different mechanisms as is illustrated in this cartoon. You can have hypoxic ischemic preconditioning, which enhances the function of the stem cells. Enhance expression of trophic factors such as GDNF or FGF1. Pre-treatment that can enhance the function once it's transplanted, as well as short-term exposure to oxidative stress. These stem cells can be pre-treated to improve homing, to improve access to the blood-brain barrier, and improve survival in the target lesion by any one of these mechanisms that's described in this slide. The next important thing to think about is the route of administration. We think about intravenous, intra-arterial, intracerebral, intrathecal, intraperitoneal, or intranasal approaches. Each has their advantage and disadvantage. With intracerebral approaches, the advantages may be, while it may be effective, it allows for precise implantation. It's been studied widely in both experimental and clinical models. It is, of course, very invasive and may have poor cell availability. Intra-arterial approaches are slightly less invasive, that is, maybe slightly safer than the intracerebral approaches, but there's still potential for microemboli. Many of the studies in both in animals and humans have, however, used this approach. Intravenous approaches are, again, less invasive than intracerebral approaches, but actually have a disadvantage of poor cell and fragment. It's really not well studied, both in animals and in patients in clinical trials. In terms of intranasal, this is an attractive feature of administering stem cells because it can bypass the blood-brain barrier, but has a disadvantage for poor cell and fragment, and has not been widely studied to date in clinical and experimental models. As with many things that have to do with therapies for stroke, timing of administration is extremely important. When we talk about acute timing, within days of the stroke, it has been shown in preclinical studies to result in improved functional scoring outcomes at 21 days post-stroke. Certain delivery mechanisms allow for the opportunity of multiple doses to be delivered over time. Subacute delivery, in many studies, the literature will wait at least seven days, may be of benefit to wait until after the initial reperfusion injury has passed, as that substrate may not be ideal for migration or integration of transplanted stem cells. And then, of course, in the chronic phase, which occurs months after, the approach there and the strategy there is to really optimize the recovery pattern as we see that as a so-called, quote-unquote, fixed deficit or one that's plateaued, as we showed in the previous slides. When we think of outcome measurement, it can be broken down into three broad categories, those that are cognitive or neuropsychological assessments, global functional assessments, and then more specific motor recovery scores or specific deficit scoring changes. And if we go back to our initial slide timeline of what happens after a stroke, in the acute setting, perhaps there's a greater focus on those global functional measurements that may have an impact with therapy at the acute setting. And then as you work your way out in terms of days and weeks and months, you start to see that the overall emphasis on recovery may be more specialized, may be more related to the actual deficits that occur in the patient that had the stroke itself. With that as a way of background, understanding where the field of stem cell therapy is within stroke and what are the factors that we must consider when we're devising or thinking about new interventions related to stem cell therapy in stroke, we can review the Pisces Phase 1 study. This study utilized an immortalized human neural stem cell line from which a drug product was developed for allogenic therapy. In preclinical testing, it showed a dose-dependent improvement in sensor motor function in rats implanted with CTX-DP4 weeks after MCA occlusion. In this Phase 1 study, it was an open-label, single-site dose escalation study. And the treatment population was men aged 60 or older with a stable disability. And by stable disability, they were looking for those individuals who had a NIH Stroke Scale 6 or greater and a modified Rankin score between 2 and 4, between 6 and 60 months after stroke. So this is really in the subacute phase or the chronic phase, I'm sorry, the chronic phase of stroke recovery that we described in the previous slides. In a sequential manner, individuals were implanted with a single dose of either 2 million and then subsequently 5 million, 10 million, or 20 million cells by a stereotactic ipsilateral putaminal injection, so an intracerebral method. Clinical and brain imaging data were collected over the next two years and the primary endpoint here, of course, was safety as this was the first in human study of a stem cell transplantation in a stroke population. So adverse events and neurological changes were also looked at very carefully. So when we look at the results from this study, 11 individuals were recruited with a mean age of 69 with a range of 60 to 82 years old. As you can see here, approximately 3 per group were administered the stem cells with the last group of 20 million cells being administered in two subjects. The time from stroke onset to implantation ranged from 12 months out to 51 months with a mean time from stroke was about 29 months. Three of the individuals had subcortical infarcts only and seven had right hemispheric infarcts. As you can see in this topographical display of all the different infarct patterns across all the patients together, kind of co-registered upon each other. You can see the risk factors for this stroke are detailed here in Table 1 as well, as well as their baseline modified Rankin score. The majority of the patients were 3 or 4 with a couple of patients who had a modified Rankin of 2 or at least one patient on a modified Rankin of 2 at the time of transplantation. This being a safety study, really a first-in-human trial, the primary endpoint here was looking at safety. And while there were four SAEs reported, none of them were symptomatic. And you can see the different SAEs that were reported here. While there was a couple believed to be related to the procedure itself, such as an extradural hematoma or subdural hematoma, which were asymptomatic, they did not seem to impact on the disease course for each of the patients that had those complications. Many of the complications occurred more than six months after surgery did not seem to be related at all to the procedure itself, and were rather related to pre-stroke risks that were inherent to these patients. Interestingly, though this study was not designed to look at outcome, they did find a trend in improvement across all outcome measures, perhaps except the modified Rankin scale. And this may be due to the insensitivity of the modified Rankin scale to look for fine changes in recovery patterns. And what you see here in this inset, the graph, is an improvement in the NIH Stroke Scale score over a period of 24 months across all subjects. You can see that the NIH Stroke Scale score was relatively stable at the time of surgery when the implantations occurred. And overall, as a trend, you can see that the Stroke Scale scores did improve over the subsequent two years. This is similar to the trend that was seen for other endpoints, most notably for motor recovery patterns. In fact, when looking at the motor recovery patterns, they seem to have the most robust change in response across all patterns that were observed. The results from the PISCES-1 study really greatly informed the PISCES-2 study, which is a single-arm, open-label study, focusing on adults aged greater than 40, so broadening the age category from 60 to 40, also including women, not just men. In this study, the emphasis was looking at motor recovery, so they recruited those individuals who had significant upper limb motor deficits two to 13 months after ischemic stroke, so in the subacute to chronic stroke phase. 20 million cells were injected via stereotactic injection to the putamen ipsilateral to the cerebral infarct. And the primary outcome was an improvement by two or more points on the research arm test subtest two at three months after implantation. As you can see here, they assessed about 40 subjects for eligibility. 18 were excluded for various reasons, and there were 23 treated. However, two were lost to follow-up, and one passed away, which was unrelated to the study, or unrelated to the intervention itself. So they have a completion of 20 subjects with 12-month follow-up. Overall, the treatment was well-tolerated. There were no significant adverse events that were related to the study intervention itself. The primary endpoint was reaching 15% of the subjects by 12 months. The improvement is greatly dependent on the baseline motor pairs. As you can see in the graph here on the top right, those who had essential plesia in the arm did not demonstrate an improvement in their function, whereas those who had less severe deficits, such as a motor score or limb score of 2, had the greatest improvement in motor deficits as a result of the intervention. This data provided evidence for targeting a population for intervention, that is, a population that may benefit most or be enriched by the injection of stem cells. And as you can see in the table here, you can see the further breakout in the response time of the percent of improvement by two points did seem to really require the full 12 months of observation period for there to be a dramatic improvement seen in the motor deficits. And again, like in the Phase I study, the overall benefit or improvement in the rank and scale score was also seen, though not as robustly, as the motor score. So with that said, as a summary, when you think about stem cell therapy for stroke, and this was a graphic I pulled from clinicaltrials.gov just the preceding week, we're seeing that there's an increasing number of clinical trials that are mostly in the exploratory phase of development worldwide, many centered in China as well as three ongoing trials certainly in the United States at present and a couple in Europe and the Middle East. Many are showing that they're able to pass through the safety hurdle, but identifying the optimal target still remains yet to be seen. And this will also determine what the optimal regimen will be, both in terms of the timing, dose, and targeted measure outcome. So when we think about stem cell and stroke, we're really at the point where we're starting to take the basic science to the transitional level. We're trying to optimize transplant regimen to identify where we can do this the most safely and with the greatest benefit. Thank you for your time and I would look forward to any questions that we may have.

stem cell therapy

ischemic stroke

neuroprotection

functional recovery

clinical trials