INTRA-ARTERIAL IMMUNOSELECTED CD34+ STEM CELLS FOR ACUTE ISCHEMIC STROKE BANERJEE, S., ET. AL. STEM CELLS TRANSLATIONAL MEDICINE Presentation by Airan Jansen Program Administrator CIRM Bridges to Stem Cell Research California Polytechnic University, Pomona California State University, Los Angeles

STROKE The loss of brain function due to a reduction or loss of blood supply to the brain Two types of stroke Ischemic-blood clot blocks the blood flow in an artery within the brain Hemorrhagic-Blood vessel bursts within the brain Stroke is the 4th leading cause of death in the US (~140,000 Americans die from stroke each year) ~800,000 people in the US suffer a stroke each year Stroke is the leading cause of adult disability High Blood pressure, high cholesterol and smoking are major risk factors for stroke

ISCHEMIC STROKE Occurs when an artery that supplies oxygen to the brain becomes blocked (usually by blood clots) 85% of strokes are ischemic Current Treatments for Ischemic Stroke include: Tissue plasminogen activator (tPA)- works to dissolve the clot and restore blood flow. This treatment should be administered within 3 hours of stroke. Endovascular procedures- a catheter is sent to the blocked site in order to remove the clot. Patients must meet certain criteria to be eligible for this treatment (image courtesy of Covidien)

ISCHEMIC STROKE There remains a need for treatments that can reduce tissue injury and progression and enhance functional recovery in patients with stroke Stem cells offer the promise of a novel reparative strategy for ischemic stroke victims

MESENCHYMAL STEM CELLS OR MARROW STROMAL CELLS (MSCS) MSCS are located in the adult Bone Marrow (BM) MSCS have the ability to migrate from the BM through the blood- brain barrier and enter the forebrain and cerebellum MSCS have been shown to differentiate into nerve cells (astrocytes and neurons) after injection into the neonatal mouse brain (Kopen, et. al., Proc Natl Acad Sci, 1999) MSCS can be divided into CD34+ and CD34- subsets and previous studies have shown CD34+, but not CD34- cells, can be used to treat stroke in rodents (13)

EVIDENCE THAT CD34+ CELLS CAN BE USED FOR ACUTE STROKE THERAPY 1.Increased mobilization of CD34+ cells into the blood after a stroke positively correlates with clinical recovery (Paczkowska, E., et. al., Eur J Haematol, 2005). 2.Trials of therapy with CD34+ cells in rodent models of stroke have shown functional recovery and a reduced size of the damaged area (infarct) by increasing production of nerve cells due to stimulation of blood vessel formation (angiogenesis) (Taguchi, A., et.al., J Clin Invest, 2004) Compared with CD34 – cell transplantation (D), increased evidence of activated vasculature was observed in animals receiving CD34 + cells (E), as detected with mouse specific anti-CD13 antibody (Taguchi, A., et.al., J Clin Invest, 2004).

EVIDENCE THAT CD34+ CELLS CAN BE USED FOR ACUTE STROKE THERAPY (CONTINUED) 3. Recent trial of intra-arterial bone marrow stem cell treatment of acute stroke showed a positive correlation between the number of CD34+ cells and functional outcome at 1 month. M Moniche F et al. Stroke. 2012;43: Clinical outcomes. A, Modified Rankin Scale at 6 months Modified Rankin Scale

THREE MAIN ROUTES OF CENTRAL NERVOUS SYSTEM DELIVERY OF CD34+ CELLS 1.Intravenous 1.PRO: Would not cause further complications in stroke patients 2.CON: Most of the transplanted cells would be absorbed by non-target organs (i.e. lungs, spleen, liver) before they reach the brain. 2.Direct surgical cranial implantation 1.PRO: Most efficient method of delivery 2.CON: potential for increased cerebral inflammation and edema as well as problems associated with anesthesia in unstable patients 3.Intra-arterial 1.PRO: Directed therapy without the risks of major surgery

AIM OF STUDY To establish safety and feasibility of autologous CD34+ selected stem/progenitor cell therapy- delivered into the middle cerebral artery (via the femoral artery in the leg) in patients with acute (<7days) ischemic stroke. /

PATIENT INCLUSION CRITERIA

PATIENT EXCLUSION CRITERIA

COHORT SELECTION PACS: Partial (brain) anterior circulation stroke TACS: Total (brain) anterior circulation stroke

OVERVIEW OF EXPERIMENTAL PROCEDURES 1. Bone marrow aspiration from stroke patient to obtain CD34+ cells using lidocaine anesthesia only. Aspiration is done <7 days after stroke has occured

OVERVIEW OF EXPERIMENTAL PROCEDURES CONTINUED 2. Isolation of CD34+ cells using CD34+ antibody-coated magnetic beads 3. CD34+ cell infusion via femoral (leg) artery puncture to deliver cells to middle cerebral artery.

COMPARATIVE STROKE SCALES National Institutes of Health Stroke Scale Modified Rankin Score Scale ScoreStroke Severity 0No symptoms 1No Significant disability. Able to carry out usual activities, despite some symptoms. 2Slight disability. Able to look after own affairs without assistance, but unable to carry out all previous activities. 3Moderate disability. Requires some help, but able to walk unassisted. 4Moderately severe disability. Unable to attend to own bodily needs without assistance, and unable to walk unassisted. 5Severe disability. Requires nursing care and attention, bedridden, incontinent. 6Dead.

CLINICAL OUTCOMES Figure 2. National Institutes of Health Stroke Scale scores on admission and at days Figure 3. Modified Rankin Scores on admission and at days 0-180

PERCENTAGE CHANGE OF INFARCT VOLUME

SERIAL BRAIN IMAGING FROM BASELINE TO DAY 180 IN PATIENTS 1 AND 2 Patient 1Patient 2

SERIAL BRAIN IMAGING FROM BASELINE TO DAY 180 IN PATIENTS 3, 4 AND 5 Patient 3Patient 4Patient 5

DISCUSSION AND FUTURE DIRECTION All patients showed improvements in clinical scores and reduction of lesion volume within 6 months. Previously, patients with TACS have a poor prognosis (96% of such patients die or become dependent at 6 month follow up (Bamford J., et. al. Lancet 1991) Of the 5 patients studied 4 were of the TACS subtype In this study, 3 of the 4 TACS patients were independent at the final follow up visit. Future studies will need to further investigate the following: Treatment in patients older than 80 years of age Ideal timing of transplantation relative to time of stroke Ideal dose of stem cells to be delivered Trials with more patients and control groups will also need to be studied

CONCLUSION This is the 1 st study in humans that demonstrates the safe and feasible treatment of patients with acute severe ischemic stroke using direct intra-arterial infusion of autologous CD34+ selected cells. ALL patients improved up to 6 months of follow-up