Long-term safety of human retinal progenitor cell transplantation in retinitis pigmentosa patients

Abstract

Background: Retinitis pigmentosa is a common genetic disease that causes retinal degeneration and blindness for which there is currently no curable treatment available. Vision preservation was observed in retinitis pigmentosa animal models after retinal stem cell transplantation. However, long-term safety studies and visual assessment have not been thoroughly tested in retinitis pigmentosa patients.

Methods: In our pre-clinical study, purified human fetal-derived retinal progenitor cells (RPCs) were transplanted into the diseased retina of Royal College of Surgeons (RCS) rats, a model of retinal degeneration. Based on these results, we conducted a phase I clinical trial to establish the safety and tolerability of transplantation of RPCs in eight patients with advanced retinitis pigmentosa. Patients were studied for 24 months.

Results: After RPC transplantation in RCS rats, we observed moderate recovery of vision and maintenance of the outer nuclear layer thickness. Most importantly, we did not find tumor formation or immune rejection. In the retinis pigmentosa patients given RPC injections, we also did not observe immunological rejection or tumorigenesis when immunosuppressive agents were not administered. We observed a significant improvement in visual acuity (P < 0.05) in five patients and an increase in retinal sensitivity of pupillary responses in three of the eight patients between 2 and 6 months after the transplant, but this improvement did not appear by 12 months.

Conclusion: Our study for the first time confirmed the long-term safety and feasibility of vision repair by stem cell therapy in patients blinded by retinitis pigmentosa.

Trial registration: WHO Trial Registration, ChiCTR-TNRC-08000193 . Retrospectively registered on 5 December 2008.

Keywords: Cell transplantation; Progenitor cell; Retina; Retinitis pigmentosa; Visual improvement.

Fig. 1

Characterization of retinal progenitor cells (RPCs). a Undifferentiated RPCs were stained with early eye field markers, including paired box protein 6 (PAX6) and SRY (sex determining region Y)-box 2 (SOX2), and for the immature neural cell marker Nestin. RPCs were stained negative for glial fibrillary acidic protein (GFAP). b Flow cytometry profiles of RPCs for subpopulations expressing PAX6, SOX2, Nestin, and GFAP, respectively. c The graph shows gene expression of retinal progenitor and mature retinal markers (fold increase) in hRPCs compared with human embryonic stem cells (hESCs) assayed by real-time quantitative polymerase chain reaction (RT-qPCR). d Normal female (46 XX) karyotype of the clinical RPCs. e Photoreceptor differentiation by retinoic acid (RA) treatment in vitro. RA(+) groups showed RPCs were positive for the mature photoreceptor markers recoverin and rhodopsin and negative for Ki67 after RA inducing. RA(-) groups are the controls

Fig. 2

Transplantation of retinal progenitor cells (RPCs) repopulated the RCS rat outer nuclear layer and increased electroretinal function. a Distribution of the transplanted cells (arrows) in the subretinal space at 6 weeks after xenotransplantation indicated by DiI staining. Horizontal cellular migration could be visualized. b RPCs stained with anti-human mitochondria were seen in the outer nuclear layer (ONL), some cells were double-labeled recoverin. In addition, the ganglion cell layer (GCL) and inner nuclear layer (INL) were also marked in the host retina. b’ Enlarged area reflecting the differentiation of transplanted cells. c,c’ RPCs were double-labeled with anti-human mitochondria and rhodopsin. d Mean and standard deviation measurements of the ONL thickness in the RPC-grafted area were significantly higher compared to the control group (P < 0.05). e Representative electroretinography (ERG) (5 dB flash under scotopic conditions) recorded at 3 and 6 weeks after cell transplantation. f Mean b-wave amplitude peaks were significantly higher in transplanted animals compared to phosphate-buffered saline (PBS) controls at 3 and 6 weeks after cell transplantation (P < 0.05). Scale bars = 50 μm (b,c), 10 μm (b’,c’)

Fig. 3

Representative retinal appearance and morphologic features before and after cell transplantation for patient 6. Retinal structures were imaged by optical coherence tomography (OCT) before and after surgery. a OCT imaging with horizontal scanning along the superior temporal macular area before surgery. Notice the characteristic of local photoreceptor atrophy. b On postoperative day 7, a mass of transplanted cells (arrows) is most evident as dense medium reflectivity, which is located between the degenerated photoreceptor layer and retinal pigment epithelium layer. c Transplanted cells are still present although the thickness has been noticeably reduced (arrows) 1 month after surgery

Fig. 4

Retinal morphological changes after RPC transplantation in patient 6. a–d Baseline images, a’–d’ 12-month follow-up, and a”–d” 24-month follow-up. a Color fundus photographs, b fluorescein angiograms, c autofluorescence imaging in the macular area, d foveal optical coherence tomography (OCT), and e horizontal OCT scanning along the injection site. a’,a” No retinal hemorrhage or edema occurred after RPC transplantation. b’,b” The characteristics of fluorescent leakage did not change after transplantation. c’,c” No obvious autofluorescence destruction in the macular area after RPC transplantation, except for a minimal area of hypo-autofluorescence (arrow). This disrupted RPE layer corresponded to the injection site. d’,d” Foveal depression was maintained pre- and postoperatively, indicating that no macular edema occurred. e The injection site before surgery (box and arrow indicate the direction of OCT scanning). e’,e” Signs of the injected cells could not be observed at 12 and 24 months post-implant and, in this patient, retinal scarring was evident with local retinal thickening

Fig. 5

Best corrected visual acuity outcomes following RPC transplantation. The mean best corrected visual acuity (BCVA) in the treated eyes significantly improved, albeit only slightly, at 2, 3, and 6 months compared to the baseline measurements (*P = 0.029, 0.013, and 0.019, respectively; n = 8). MAR minimum angle of resolution