Laura Baqué Vidal, PhD
Project Manager
Department of Clinical Sciences, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden
Gynecology and Reproductive Medicine, Karolinska Universitetssjukhuset, Stockholm, Sweden
Disclaimer/Conflict of interest statement: none.
Brightfield images of (A) human pluripotent stem cell-derived retinal pigment epithelial cells at day 60 of differentiation, and depigmented human pluripotent stem cell-derived retinal pigment epithelial cells (B) at day 60+3 of differentiation, right before freezing.
What?
The retinal pigment epithelial (RPE) cells are specialized cells that lie on the back of the eye. Precisely, they are between the Bruch membrane (a membrane that physically separates the eye to the choroid or blood vessels) and the retina (multilayer of neurons that transduces light into electrical signal in the brain through the optic nerve). RPE cells are not neurons, instead, they are epithelial cells that play a fundamental role in maintaining the homeostasis of the retina: visual cycle recycling, phagocytosis of old photoreceptor fragments, secretion of growth factors, transport of nutrients and water, capture of not-transduced light (1).
The RPEs have a very characteristic phenotype when they are mature: they have a cobblestone morphology (like a hexagonal shape) making a kind of mosaic with pigmentation inside and bright tight junctions on their connections with each other (Figure A). The pigmentation observed in the cells is a sign of maturation and it is present for the granules of melanin that are present in these cells to absorb the leftover light that the retina doesn’t use to “see”.
When cells are that mature they are not proliferative, but they are senescent, therefore they cannot be cryopreserved. For several reasons, to do a cell product for multiple patients, one needs to be able to store the cell product until patient delivery. In Figure B, the mature RPEs have been replated (harvested and plated again) for them to regain proliferative capacity and, therefore, to be able to be frozen and stored (2).
Why?
Age-related macular degeneration (AMD) is a prevalent progressive blindness disease in the elderly worldwide. Epidemiologists project a growing number of patients of over 280 million by the end of 2040 (3). There is no cure or even effective treatment for the cause of the disease, which is not fully understood yet (4). The native RPEs of these patients start degenerating and dying. When these cells cannot maintain homeostasis, the retina also degenerates and discontinues to process the light. Therefore, a black hole of vision is created where the degeneration has started -in the macula, explaining the name of the disease. We think that cell therapy is the best shot for these patients to stop the disease's progression or even, in the early stages, stop it from starting the loss of vision (black hole). For a comprehensive summary of how gene and cell therapy could help on this disease refer to here.
Who?
Some different academic groups and companies are pursuing the strategy of making a stem-cell-based RPE cell products. Currently, though, there is none already available on the market. Most of them are in preclinical stage and few are in phase I/IIa.
The only medicines that are available for these patients are now two types that aim to slow down the progression of the disease. One group is the anti-VEGF intravitreal injections to block the outgrowth of capillary vessels into the eye (5) and another group are anti-complement treatments also by intravitreal injection like pegcetacoplan or avacincaptad pegol (6,7).
When?
The interest on transplanting RPE into AMD patients started in the early 90, when fetal material was used to transplant young and healthy RPEs (8) that were able to slightly rescue some visual acuity. After, and because of the ethically concerning issue around aborted material as well as limited source, some primary RPE cell lines were used to transplant into patients. Those fail to functionally rescue vision and to identify as mature RPEs. Finally, since the new century with the discovery of the stem cell potential, the efforts have been on developing protocols to get healthy, mature and pure RPEs for stem-cell-based RPE transplantation.
Where?
The leading countries developing stem-cell-based RPE cell products that are or were on clinical trials are USA, Japan, Israel, UK, China and Republic of Korea. In some other countries, developments are still on the preclinical stage: Sweden, Spain, India, Finland, Singapore.
How?
The clinical production challenges of stem-cell-based RPE products are the general ones for any Advanced Therapy Medical Products (ATMP) and other specific ones for these cells. Generally, all ATMP products face the adaptation towards a Good Manufacturing Practice (GMP) environment which involves the selection of raw materials that are certified to be used as GMP, selecting a starting material that also qualifies as GMP with all quality controls around viruses and transmissible spongiform encephalopathies (TSE) in place, as well as identify (or build) a clean room GMP facility that can host your production activities. More on the specific sides of the manufacturing of stem-cell-based RPE cells is the scale-up: depending on your process, but, usually, these cells grow on 2D, therefore a lot of surface is needed, and therefore, multiple flasks or multilayered flasks are needed. A second concern is purity. Some labs have established protocols that yield > 98% pure RPEs, whereas others use cell sorting procedures to select the RPEs from other cellular impurities, which implies a more difficult process as well as expensive. Another issue is the injection of these RPEs into the subretinal space, two alternatives are coexisting at the moment: 1) using a cell suspension, which makes the process a little easier, since it’s only cells and a solvent to be injected through a cannula, and 2) transplanting the cells in a sheet, which implies difficulties on the surface in which the cells are plated, the manual culture of this process (usually involved cutting and culture in transwells) and the device that one may need to inject which would need to be customized. Overall, the production of these cells are, in comparison with other ATMP, on the easier side, since the differentiation protocol is quite straightforward, no major purity problems appear, there is no virus transduction in the process to clean up and there are quite many identity and functionality assays widely accepted in the community to set as quality controls. As a first-of-its-kind product, regulatory authorities face the challenge of reviewing all applications and understanding the uniqueness of this investigational product, which, sometimes, takes more effort than expected and might delay the product development programme.
Did you know that…
RPE cells have granules of melanin, the same substance found in the skin, that protects us from UV light and is responsible of the color of our hair. That makes it easier to work with these cells because when cultured in any clear plastic or glass they can be easily spotted for their dark brown color. They usually have a pattern of melanin when they are plated in 2D, like a mosaic and when they are collected in a tube, one can see a dispersion of brown cells in the solution -especially if it’s a transparent solvent.
References
- Strauss O. The retinal pigment epithelium in visual function. Physiol Rev. 2005;85(3):845–81.
- Baqué-Vidal L, Main H, Petrus-Reurer S, Lederer AR, Beri NE, Bär F, et al. Clinically compliant cryopreservation of differentiated retinal pigment epithelial cells. Cytotherapy. 2024;000:1–11.
- Jonas JB, Cheung CMG, Panda-Jonas S. Updates on the epidemiology of age-related macular degeneration. Asia-Pacific Journal of Ophthalmology. 2017;6(6):493–7.
- Ambati J, Ambati BK, Yoo SH, Ianchulev S, Adamis AP. Age-related macular degeneration: Etiology, pathogenesis, and therapeutic strategies. Surv Ophthalmol. 2003;48(3):257–93.
- Gehrs KM, Anderson DH, Johnson L V., Hageman GS. Age-related macular degeneration - Emerging pathogenetic and therapeutic concepts. Ann Med. 2006;38(7):450–71.
- Pavluk L, Kaya M. Apellis.com. 2023 [cited 2024 Feb 17]. FDA Approves SYFOVRETM (pegcetacoplan injection) as the First and Only Treatment for Geographic Atrophy (GA), a Leading Cause of Blindness. Available from: https://investors.apellis.com/news-releases/news-release-details/fda-approves-syfovretm-pegcetacoplan-injection-first-and-only
- Communications & Investor Relations. AstellasPharma. 2023. Iveric Bio Receives U.S. FDA Approval for IZERVAYTM (avacincaptad pegol intravitreal solution), a New Treatment for Geographic Atrophy .
- Algvere P V., Berglin L, Gouras P, Sheng Y. Transplantation of fetal retinal pigment epithelium in age-related macular degeneration with subfoveal neovascularization. Graefe’s Archive for Clinical and Experimental Ophthalmology. 1994;232(12):707–16.
#ScientificSpotlight