Preclinical trials in autosomal dominant AD: implementation of the DIAN-TU trial.

The Dominantly Inherited Alzheimer's Network Trials Unit (DIAN-TU) was formed to direct the design and management of interventional therapeutic trials of international DIAN and autosomal dominant Alzheimer's disease (ADAD) participants. The goal of the DIAN-TU is to implement safe trials that have the highest likelihood of success while advancing scientific understanding of these diseases and clinical effects of proposed therapies. The DIAN-TU has launched a trial design that leverages the existing infrastructure of the ongoing DIAN observational study, takes advantage of a variety of drug targets, incorporates the latest results of biomarker and cognitive data collected during the observational study, and implements biomarkers measuring Alzheimer's disease (AD) biological processes to improve the efficiency of trial design. The DIAN-TU trial design is unique due to the sophisticated design of multiple drugs, multiple pharmaceutical partners, academics servings as sponsor, geographic distribution of a rare population and intensive safety and biomarker assessments. The implementation of the operational aspects such as home health research delivery, safety magnetic resonance imagings (MRIs) at remote locations, monitoring clinical and cognitive measures, and regulatory management involving multiple pharmaceutical sponsors of the complex DIAN-TU trial are described.

Photosensitization of retinal pigment epithelium by protoporphyrin IX.

BACKGROUND: Clinical evidence of injury to the retinal pigment epithelium is an important feature of age-related macular degeneration, but the mechanism of this injury is unknown. Blue-light-dependent activation of the blood-borne photosensitizer protoporphyrin IX is known to produce free radicals which may damage cells and tissues. This study was undertaken to determine the effect of blue light and protoporphyrin IX on retinal pigment epithelial cells in vitro. METHODS: Third-passage porcine retinal pigment epithelial cells were plated in six-well culture plates at 100,000 cells/well and grown to confluence. Retinal pigment epithelial cells were then incubated in culture media with and without 35 micrograms/dl protoporphyrin IX and exposed to low intensity (118 microW/cm2) blue, blue-free, or full-spectrum white light in an irradiating incubator for 16 h on/8 h off cycles for 7 days. Some of the wells were shielded from light (dark controls). Retinal pigment epithelial cells were examined by light microscopy and were trypsinized and counted after 7 days. RESULTS: White light with and without protoporphyrin IX and protoporphyrin IX in dark conditions did not decrease the retinal pigment epithelial cell count significantly. Blue light alone and blue light with protoporphyrin IX decreased the cell count by 22 +/- 4% and 35 +/- 3% compared to the controls, respectively. CONCLUSION: Blue wavelength light without exogenous protoporphyrin IX has a cytotoxic effect on confluent cultures of retinal pigment epithelium, suggesting that endogenous photosensitizers may be present in retinal pigment epithelial cells. Protoporphyrin IX has an additive cytotoxic effect in the presence of blue light, suggesting that this photosensitizer is capable of mediating blue-light-induced retinal pigment epithelial damage. Since protoporphyrin IX is present in blood and tissue fluids, and the retina is chronically exposed to light, protoporphyrin IX-mediated free radical formation may occur in vivo and may play a role in retinal pigment epithelial changes that occur early in the pathogenesis of age-related macular degeneration.

Effect of mitomycin-C on human retinal pigment epithelium in culture.

PURPOSE: To determine the effect of mitomycin-C on confluent and non-confluent human retinal pigment epithelium (RPE) in tissue culture. METHODS: The effect of mitomycin-C on confluent RPE was determined by treating first passage confluent cells with 0.01, 0.1, 1, 10, 100 or 1000 micromolar (microM) mitomycin-C for 1, 3, or 7 days. The cell viability after treatment was determined by using an esterase stain. The effect of mitomycin-C on proliferating RPE was determined by incubating non-confluent cells with the above concentrations of mitomycin-C for 20 min, 1 hour or 24 hours. RESULTS: Mitomycin-C can be toxic to a confluent RPE monolayer, and the LD50 is 421, 28.8 or 0.0632 microM when cells are continually exposed to mitomycin-C for 1, 3 or 7 days, respectively. Exposure to mitomycin-C at concentrations > or = 10 microM for 20-60 min significantly inhibits proliferation of non-confluent RPE. A 24 hour exposure of RPE to 1 microM mitomycin-C markedly inhibits proliferation of non-confluent RPE with minimal toxicity to confluent RPE. CONCLUSIONS: Since exposure of human RPE to mitomycin-C for 24 hours can inhibit cell proliferation at concentrations which are well-tolerated by confluent RPE, mitomycin-C may be a suitable agent for inhibiting RPE proliferation in vivo.

Retinal pigment epithelial debridement as a model for the pathogenesis and treatment of macular degeneration.

PURPOSE: To determine the effects of the absence of the retinal pigment epithelium on the choriocapillaris and outer retina by performing retinal pigment epithelial cell debridement with mitomycin C to inhibit cell proliferation pharmacologically in the porcine eye. METHODS: A pars plana vitrectomy was performed in 12 eyes, and two neurosensory retinal detachments per eye were created by injecting 10(-3) mg/ml mitomycin C and 0.25% edetic acid into the subretinal space. Twenty minutes later, the retinal pigment epithelium was debrided, and the retina was reattached with a fluid-gas exchange. RESULTS: Bruch's membrane was devoid of native retinal pigment epithelium, and the choriocapillaris was patent immediately after debridement. No proliferation of the retinal pigment epithelium occurred 1 week after debridement, and choriocapillaris atrophy was present beneath areas of Bruch's membrane that were devoid of retinal pigment epithelium. Four weeks postsurgery, choriocapillaris atrophy persisted in all debrided blebs, although unpigmented retinal pigment epithelium repopulated portions of Bruch's membrane in one of three blebs. Outer retinal atrophy was present in areas of Bruch's membrane with no retinal pigment epithelium and no choriocapillaris 4 weeks postsurgery. The choriocapillaris was patent in areas of mitomycin C injection without debridement. CONCLUSION: Absence of the retinal pigment epithelium leads to atrophy of the choriocapillaris within 1 week after surgery. This finding provides an animal model to study transplantation of retinal pigment epithelium onto bare patches of Bruch's membrane in age-related macular degeneration and other diseases and provides insight into the pathogenesis of nonexudative age-related macular degeneration.

Débridement of the pig retinal pigment epithelium in vivo.

OBJECTIVE: To study the morphologic effects of surgical débridement of the retinal pigment epithelium (RPE) in an animal model. METHODS: A pars plana vitrectomy was performed in the domestic pig, and a neurosensory retinal detachment was created by injecting the calcium-chelating agent edetic acid (commonly referred to as ethylenediaminetetraacetic acid or EDTA) into the subretinal space through a retinotomy. Twenty minutes later, the RPE was débrided by gently brushing Bruch's membrane with a soft-tip silicone catheter. Dissociated RPE was aspirated from the subretinal space, and the retina was reattached with a fluid-gas exchange. RESULTS: Light microscopic analysis confirmed that Bruch's membrane was devoid of native RPE and the choriocapillaris was morphologically intact immediately after débridement. Photoreceptor outer segments were disrupted and foreshortened immediately after RPE débridement. One to 4 weeks later, a layer of hypopigmented RPE covered most of the previously débrided areas of Bruch's membrane. The choriocapillaris was intact in areas of Bruch's membrane that were repopulated by hypopigmented RPE, and remained intact 12 weeks after débridement. Some regions of Bruch's membrane near the retinotomy remained devoid of RPE for more than 4 weeks after débridement. The choriocapillaris was atrophic and there was extensive disruption of the outer retinal layers in these areas. CONCLUSIONS: The RPE healed in most areas after surgical débridement of the RPE in the experimental animal. Atrophy of the choriocapillaris was present in areas of poor RPE healing near the retinotomy.