Biomaterials-enabled cornea regeneration in patients at high risk for rejection of donor tissue transplantation

Abstract

The severe worldwide shortage of donor organs, and severe pathologies placing patients at high risk for rejecting conventional cornea transplantation, have left many corneal blind patients untreated. Following successful pre-clinical evaluation in mini-pigs, we tested a biomaterials-enabled pro-regeneration strategy to restore corneal integrity in an open-label observational study of six patients. Cell-free corneal implants comprising recombinant human collagen and phosphorylcholine were grafted by anterior lamellar keratoplasty into corneas of unilaterally blind patients diagnosed at high-risk for rejecting donor allografts. They were followed-up for a mean of 24 months. Patients with acute disease (ulceration) were relieved of pain and discomfort within 1-2 weeks post-operation. Patients with scarred or ulcerated corneas from severe infection showed better vision improvement, followed by corneas with burns. Corneas with immune or degenerative conditions transplanted for symptom relief only showed no vision improvement overall. However, grafting promoted nerve regeneration as observed by improved touch sensitivity to near normal levels in all patients tested, even for those with little/no sensitivity before treatment. Overall, three out of six patients showed significant vision improvement. Others were sufficiently stabilized to allow follow-on surgery to restore vision. Grafting outcomes in mini-pig corneas were superior to those in human subjects, emphasizing that animal models are only predictive for patients with non-severely pathological corneas; however, for establishing parameters such as stable corneal tissue and nerve regeneration, our pig model is satisfactory. While further testing is merited, we have nevertheless shown that cell-free implants are potentially safe, efficacious options for treating high-risk patients.

Fig. 1

Regenerated corneas of Göttingen mini-pigs at 12-months post-grafting with RHCIII-MPC compared to healthy, unoperated control corneas. a control cornea versus RHCIII-MPC implanted cornea both showing comparable optical clarity. Serial block face-scanning electron microscopy (SBF-SEM) of single sections show that the epithelium is multilayered with comparable morphology including a layer of basal cells. Underlying the epithelium are stromal keratocytes arranged in lamellae. Scale bars, 50 µm. 3D reconstructions of the corneas show that both regenerated neo-cornea and healthy control comprise stromas with keratocytes arranged in highly ordered lamellae. b Light transmission profile of regenerated neo-corneas compared to healthy contralateral corneas. c Backscatter profile of regenerated neo-corneas compared to healthy contralateral corneas

Fig. 2

Patient corneas before and after grafting with RHCIII-MPC implants at last follow-up. Patients are divided into three groups based on their pre-operative diagnoses: infection (herpes simplex viral and fungal keratitis), burns (alkali and thermal) and other (failed graft and post-stroke neurotrophic keratitis). Post-operation, regenerated neocorneas from Patients 1 and 2 are mostly clear. In Patients 3 and 4, where stem cell deficiency is present, some superficial vessels concurrent with conjunctival invasion are seen. Patient 5 has a mostly clear cornea encircled by blood vessels but has invaded in one quadrant, while Patient 6’s cornea remains hazy. Patient 2 has an unrelated nasal pterygium

Fig. 3

Regenerated patient corneas. a In vivo confocal images of the regenerated cornea from Patient 1 at 24 months post-operation, showing the regenerated epithelium, regenerating nerve (arrowhead) and stroma. The unoperated endothelium remains intact. b Changes in corneal touch sensitivity before and after RHCIII-MPC implantation as measured by Cochet-Bonnet aesthesiometry. The average pressure required to elicit a blink response from corneas before surgery, after implantation, and in comparison to the normal, healthy corneas. Touch sensitivity is inversely related to the pressure needed to elicit a blink response from the patients. Note: *p < 0.05 compared to unoperated contralateral eyes (Kruskal–Wallis test with Dunn’s correction for multiple comparisons)