A common theme runs through all the recent advances in endothelial keratoplasty (EK), be it Descemet’s stripping automated endothelial keratoplasty (DSAEK), Descemet’s membrane endothelial keratoplasty (DMEK), or nanothin DSAEK (NT-DSAEK). The less tissue transplanted, the faster the recovery, the less risk of rejection, and the better the postoperative visual recovery. (See “Background,” below.) However, when less tissue is used to carry the corneal endothelial cells (CEC), the more technically challenging the procedure becomes, resulting in the slower adoption of superior methods. The good news: The development of injected cultured CECs may be the ideal endothelial cell transplant to replace primary CEC failure.

Here, we explain the benefits, technique, mechanism, safety and efficacy of injected cultured CECs.

Benefits

Injected cultured CECs remove the need for a carrier when reestablishing a healthy endothelial cell count, and restore a high CEC density without altering the normal corneal curvature. This allows for optimal postop visual outcomes.

Additionally, the lack of a CEC carrier eliminates the risk of graft detachment and decreases the risk of rejection. Further, this therapy is not technically challenging, which lowers the barrier for widespread adoption. In early clinical studies, injected CECs have been used to replenish endothelial cells in Fuchs’ endothelial corneal dystrophy, pseudophakic bullous keratopathy, pseudoexfoliation syndrome keratopathy and corneal graft failure.⁶

Technique

Donor cell preparation consists of isolating CECs with multiple steps of cultivation to ensure high-quality cell characteristics. In brief, the CECs are isolated by stripping Descemet’s membrane, followed by enzyme treatment to remove the collagen matrix. These cells undergo analysis to confirm their biological characteristics and to verify criteria for surgical use.

The cells are stored in media that maintain the cells’ biochemical characteristics. Prior to injection, the cell suspension is supplemented with a Rho-associated protein kinase (ROCK) inhibitor.

The recipient eye is prepared by mechanically removing abnormal extracellular material and degenerated CECs on Descemet’s membrane in the central 8 mm diameter area of the cornea. This is performed with a silicone cannula through a 1.6 mm incision at the corneal limbus under local anesthesia. A total of 1x10⁶ CECs are injected with a syringe into the anterior chamber. The patient is then required to lay face-down for three hours to encourage CECs to settle on the inner cornea, and allow for cell adhesion. Postoperative care includes topical steroids and prophylactic antibiotics.⁶

Mechanism

Face-down positioning promotes the migration of CECs toward the inner corneal surface. Adhesion of the CECs is promoted by ROCK inhibition through the suppression of the Rho/ROCK/MLC (myosin light chain)-signaling cascade. Without the ROCK inhibitor, the dissociated cells contract due to actin cytoskeleton activation, which leads to a significant decrease in the cell surface area. Further, when actin contracts, this leads to a decrease in focal adhesion complexes that are necessary for cell adhesion. Therefore, ROCK inhibition promotes cell adhesion with a dual approach of preventing cell contraction by relaxing actin and by increasing focal adhesion complexes that promote CEC adhesion to the inner corneal surface.⁷

Safety

In 2018, Kinoshita et al. published the first clinical study on the safety of cultured CECs. The researchers evaluated the potential for injected CECs to cause IOP elevation secondary to cells migrating through the trabecular meshwork. They reported no perioperative IOP elevation or any abnormal collection of CECs, with the exception of one patient at 8 months follow-up. This elevation was attributed to steroid-induced glaucoma, due to the timing of the increased pressure and no trabecular meshwork abnormalities.

In the 5-year follow-up period, no other IOP elevations were observed. Additionally, this long-term observational follow-up showed no allogenic immune reaction or anterior uveitis development in any of the eyes undergoing CEC injection therapy.⁸

Questions regarding the potential for CECs to pass through the trabecular meshwork into systemic circulation through the episcleral venous drainage was investigated. This was addressed with early animal studies that tracked CECs by fluorescein-labeled tracing and polymerase chain reaction, which did not find evidence of spread to the multiple organ systems evaluated.⁷ Since 40% to 50% of injected CECs adhere to the cornea, it is thought the remaining cells are removed by the host immune system or through autophagy.

Efficacy

Recently, long-term observational data have been published that provide support to the efficacy of injected cultured CECs. In the initial clinical cohort, the inclusion criteria required no CECs on specular microscopy, a central corneal thickness (CCT) greater than 630 µm, the presence of epithelial edema and a best-corrected visual acuity (BCVA) worse than 20/40 (Snellen).⁶

In 2020, Numa et al. reported that at 5 years, 90.9% (10/11) of eyes maintained a CEC count of more than 500 cells/mm². Of these, 80% (8/10) had a CEC count of greater than 1000 cells/mm². An improvement in the CEC hexagonality and a decrease in cell variability was observed over time, which indicates the cells continued to stabilize over the 5-year period. In these same patients, the CCT was less than 590 µm in 90.9% (10/11) of eyes. The 5-year vision outcomes were a BCVA of 20/40 maintained in 90.9% (10/11) of eyes. Of these, 80% (8/10) had a BCVA of 20/25 or better, with 60% (6/10) better than 20/20. These initial results are equivalent with previously reported outcomes for DSAEK and DMEK.⁸

A Promising Solution

The latest long-term data is promising, showing that injected cultivated CECs have comparable clinical outcomes to traditional EK. Another advantage of cultured CECs is that a single donor cornea can provide enough high-quality CECs for multiple recipients. Early clinical studies show the injection of cultivated CECs holds promise to be the ideal treatment for endothelial failure that can restore proper corneal function, has low complications and is easy to implement. It is currently undergoing further FDA clinic trials. CP

References:

1. EBAA Eye Banking Statistical Report. Eye Bank Association of America. 2019;1 ed.
2. Melles GRJ, Ong TS, Ververs B, van der Wees J. Descemet membrane endothelial keratoplasty (DMEK). Cornea. 2006;25(8):987-990.
3. Melles GRJ, Wijdh RHJ, Nieuwendaal CP. A technique to excise the descemet membrane from a recipient cornea (descemetorhexis). Cornea. 2004;23(3):286-288.
4. Cheung AY, Hou JH, Bedard P, et al. Technique for Preparing Ultrathin and Nanothin Descemet Stripping Automated Endothelial Keratoplasty Tissue. Cornea. 2018;37(5):661-666.
5. Busin M., Patel, AK, Scorcia, V, Ponzin, D. Microkeratome-assisted preparation of ultrathin grafts for descemet stripping automated endothelial keratoplasty. Invest Ophthalmol Vis Sci. 2012;53(1):521-4.
6. Kinoshita S, Koizumi N, Ueno M, et al. Injection of cultured cells with a ROCK inhibitor for bullous keratopathy. N Engl J Med. 2018;378(11):995-1003.
7. Okumura N, Sakamoto Y, Fujii K, et al. Rho kinase inhibitor enables cell-based therapy for corneal endothelial dysfunction. Sci Rep. 2016;6:1-11.
8. Numa K, Imai K, Ueno M, et al. Five-Year Follow-Up of First Eleven Cases Undergoing Injection of Cultured Corneal Endothelial Cells for Corneal Endothelial Failure. Ophthalmology. 2020 Sep 5;S0161-6420(20)30853-8.ophtha.2020.09.002

 

DR. DE LA PRESA is an ophthalmic surgeon specializing in cornea, cataract and ocular surface reconstruction. He has earned numerous honors and awards for his research, including the NASA Earth & Space Health Innovation Award, AMA Healthier Nation Innovation Award, ASCRS Resident Excellence Award, and the Harry Friedman Research Award. He is completing a fellowship in cornea and external disease at the Cincinnati Eye Institute & the University of Cincinnati, and will be starting practice at Grene Vision Group in Wichita, KS. 

DR. HOLLAND is the director of Cornea Services at Cincinnati Eye Institute and professor of Clinical Ophthalmology at the University of Cincinnati. He has published extensively in both basic and clinical research, and is the author of over 250 articles in peer-reviewed journals. He is the former President of ASCRS and the Cornea Society. He was awarded the Binkhorst Medal by ASCRS in 2008, the Castroviejo Award by the Cornea Society in 2013, and the Life Achievement Honor Award by the AAO in 2012. He is the co-editor of Cornea, and is known internationally for his expertise in ocular surface reconstruction.