September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Production of clinical-grade patient-specific iPSCs and transplantable photoreceptor precursor cells for the treatment of retinal degenerative blindness.
Author Affiliations & Notes
  • Budd A Tucker
    Ophthalmology, Wynn Institute for Vision Research, Iowa City, Iowa, United States
  • Luke A Wiley
    Ophthalmology, Wynn Institute for Vision Research, Iowa City, Iowa, United States
  • Erin R Burnight
    Ophthalmology, Wynn Institute for Vision Research, Iowa City, Iowa, United States
  • Kristan S Worthington
    Ophthalmology, Wynn Institute for Vision Research, Iowa City, Iowa, United States
  • Cathryn M Cranston
    Ophthalmology, Wynn Institute for Vision Research, Iowa City, Iowa, United States
  • Kristin R Anfinson
    Ophthalmology, Wynn Institute for Vision Research, Iowa City, Iowa, United States
  • Robert F Mullins
    Ophthalmology, Wynn Institute for Vision Research, Iowa City, Iowa, United States
  • Edwin M Stone
    Ophthalmology, Wynn Institute for Vision Research, Iowa City, Iowa, United States
  • Footnotes
    Commercial Relationships   Budd Tucker, None; Luke Wiley, None; Erin Burnight, None; Kristan Worthington, None; Cathryn Cranston, None; Kristin Anfinson, None; Robert Mullins, None; Edwin Stone, None
  • Footnotes
    Support  Wynn Institute Endowment for Vision Research, Stephen A. Wynn Professorship for Regenerative Ophthalmology, NIH RO1-EY024605, NIH RO1-EY024588
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 6064. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to Subscribers Only
      Sign In or Create an Account ×
    • Get Citation

      Budd A Tucker, Luke A Wiley, Erin R Burnight, Kristan S Worthington, Cathryn M Cranston, Kristin R Anfinson, Robert F Mullins, Edwin M Stone; Production of clinical-grade patient-specific iPSCs and transplantable photoreceptor precursor cells for the treatment of retinal degenerative blindness.. Invest. Ophthalmol. Vis. Sci. 2016;57(12):6064.

      Download citation file:


      © ARVO (1962-2015); The Authors (2016-present)

      ×
  • Supplements
Abstract

Purpose : Stem cell-derived photoreceptor precursor cells have the potential to restore vision to patients with extensive vision loss from retinal degenerative diseases. Creating these cells from the patients for whom they are intended has the advantage of perfect immunologic matching. The purpose of this study was to develop methods for manufacturing transplantable retinal cells in a low-cost, non-profit environment, under current good manufacturing practices (cGMP).

Methods : Skin biopsies were obtained from legally blind adult patients (n=35) with inherited retinal degeneration and fibroblast lines were established under ISO class 5 cGMP conditions. For seven of the patients with the poorest vision (light perception or worse), patient-specific induced pluripotent stem cells (iPSCs) were generated via transient expression of the OCT4, SOX2, KLF4, and c-MYC. Karyotyping and TaqMan Scorecard analysis were used for clonal validation. Retinal photoreceptor precursor cells were derived using a stepwise cGMP-compliant 3D differentiation protocol. Immunocytochemical analysis and confocal microscopy were used to monitor retinal development. Bolus injections of differentiated cells (1-5x106) were transplanted into immune compromised mice to assess safety.

Results : We succeeded in establishing iPSC lines from all 7 blind individuals ranging in age from 32-86 years. Thirty days post-differentiation, polarized neural epithelium containing PAX6-, SOX2-, and OTX2-positive retinal progenitor cells were seen. After 60 days, PAX6- and OTX2-positive cells were found to segregate into distinct lamina corresponding to presumptive RPE and photoreceptor precursor cells. After 90 days of differentiation, post-mitotic photoreceptor precursor cells positive for CRX, NRL and NR2E3 were observed. Four weeks after transplantation into immune compromised animals revealed no evidence of abnormal proliferation or tumor formation.

Conclusions : cGMP-compliant protocols were developed for skin biopsy and dermal fibroblast isolation, iPSC generation and retinal specific differentiation. Clinical-grade retinal cells were successfully generated from all 7 patients attempted, including 4 patients over the age of 70 years. Post-mitotic photoreceptor precursor cells retain their retinal phenotype and do not form tumors following transplantation in immune compromised recipients.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×