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Randomized Controlled Trial
. 2014 Dec;16(12):1720-32.
doi: 10.1016/j.jcyt.2014.07.011. Epub 2014 Sep 18.

Bilateral Administration of Autologous CD133+ Cells in Ambulatory Patients With Refractory Critical Limb Ischemia: Lessons Learned From a Pilot Randomized, Double-Blind, Placebo-Controlled Trial

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Free PMC article
Randomized Controlled Trial

Bilateral Administration of Autologous CD133+ Cells in Ambulatory Patients With Refractory Critical Limb Ischemia: Lessons Learned From a Pilot Randomized, Double-Blind, Placebo-Controlled Trial

Amish N Raval et al. Cytotherapy. .
Free PMC article

Abstract

Background aims: CD133+ cells confer angiogenic potential and may be beneficial for the treatment of critical limb ischemia (CLI). However, patient selection, blinding methods and end points for clinical trials are challenging. We hypothesized that bilateral intramuscular administration of cytokine-mobilized CD133+ cells in ambulatory patients with refractory CLI would be feasible and safe.

Methods: In this double-blind, randomized sham-controlled trial, subjects received subcutaneous injections of granulocyte colony-stimulating factor (10 μg/kg per day) for 5 days, followed by leukapheresis, and intramuscular administration of 50-400 million sorted CD133+ cells delivered into both legs. Control subjects received normal saline injections, sham leukapheresis and intramuscular injection of placebo buffered solution. Subjects were followed for 1 year. An aliquot of CD133+ cells was collected from each subject to test for genes associated with cell senescence.

Results: Seventy subjects were screened, of whom 10 were eligible. Subject enrollment was suspended because of a high rate of mobilization failure in subjects randomly assigned to treatment. Of 10 subjects enrolled (7 randomly assigned to treatment, 3 randomly assigned to control), there were no differences in serious adverse events at 12 months, and blinding was preserved. There were non-significant trends toward improved amputation-free survival, 6-minute walk distance, walking impairment questionnaire and quality of life in subjects randomly assigned to treatment. Successful CD133+ mobilizers expressed fewer senescence-associated genes compared with poor mobilizers.

Conclusions: Bilateral administration of autologous CD133+ cells in ambulatory CLI subjects was safe, and blinding was preserved. However, poor mobilization efficiency combined with high CD133+ senescence suggests futility in this approach.

Keywords: angiogenesis; critical limb ischemia; peripheral artery disease; stem cell.

Figures

Figure 1
Figure 1
Intramuscular Injection Protocol. Left panel: Schematic and actual injection locations mapped to 10 major muscle groups in the leg. Middle panel: Posterior injection locations. Right panel: Injection procedure.
Figure 2
Figure 2
Trial design. G-CSF: granulocyte colony stimulating factor, IM: intramuscular IV: intravenous, GFR: glomerular filtration rate, ABI: ankle-brachial index.
Figure 3
Figure 3
Baseline to 12 month change in 6 minute walk distance by intention to treat and actual treatment allocation in surviving subjects. (*paired, t-test)
Figure 4
Figure 4
Baseline to 12 month change in Walking Impairment Questionnaire (WIQ) Score by intention to treat and actual treatment allocation in surviving subjects. (*paired t-test)
Figure 5
Figure 5
Baseline to 12 month change in SF36 categories by actual treatment allocation.
Figure 6
Figure 6
Gene array assay for senescence associated genes. 6A. Compared to normal healthy donor CD133+ cells, CD133+ cell obtained from subjects randomized to the treatment arm (n=6) had 2 fold or greater expression in several senescence markers indicated above with horizontal p value cut off of 0.1. 6B. Subjects treated with CD133+ compared to mobilization failure subjects showed >2 fold upregulation in 4 of 5 genes expressing senescence markers and > 2 fold downregulation of 1 of 5 genes expressing an anti-senescence marker. Red vertical lines indicate 2-fold change in gene expression while the black horizontal line indicates a p value of 0.1. CLI: critical limb ischemia

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