The Role of Vitamin A in the Progression of Dry AMD

Aziz A. Khanifar, MD
Vitreoretinal Surgery Fellow
Weill Cornell Medical College
New York NY

Srilaxmi Bearelly, MD, MHS
Assistant Professor of Ophthalmology
The Duke Center for Macular Diseases
Duke Eye Center
Durham, NC

An important pathological characteristic of age-related macular degeneration (AMD) is the accumulation of lipofuscin in the retinal pigment epithelium (RPE). Lipofuscin may interfere with normal RPE cellular function and result in RPE death with associated loss of overlying photoreceptors. A major molecular component of lipofuscin is N-retinylidene-N-retinylethanolamine (A2E), which is derived from retinoid byproducts.¹ These retinoid byproducts result from vitamin A metabolism which occurs with the phototransduction cascade. Fundus autofluorescence (FAF) images allow monitoring of lipofuscin in vivo in patients with dry AMD and are utilized increasingly as outcome measures in clinical trials for geographic atrophy. Vitamin A uptake by the RPE has been hypothesized as a therapeutic target in the management of AMD. It has been proposed that reducing vitamin A entry into the visual cycle may reduce the production of retinoid byproducts and lead to a decrease in accumulation of A2E, thereby reducing toxicity to the RPE. Understanding the role of vitamin A in AMD requires an understanding of lipofuscin composition, toxicity, and imaging.

What is lipofuscin?
In post-mitotic RPE cells, autofluorescent storage bodies known as lipofuscin granules accumulate with age and disease. These granules, which are mainly the byproducts of phagocytosed photoreceptor outer segments, accumulate as a normal process of aging in healthy humans. However, the rate and amount of lipofuscin accumulation is much greater with retinal degenerations such as AMD, Stargardt's disease, and Best disease. Although not all intracellular modulators of lipofuscin accumulation are known, the roles of ABCA4, ELOVL4, and RPE65 genes have been documented.^2-5 For example, Radu et al reported a statistically significant increase in lipofuscin granules in the RPE of ABCA4 knockout mice that were a fed a diet high in vitamin A.⁵ Patients with mutations in the ABCA4 gene typically express autosomal recessive Stargardt's disease. This is even more pronounced with the autosomal dominant model for Stargardt's disease involving a mutation in ELOVL4 gene.³ In both these models, there is a markedly increased formation of lipofuscin and the retinoid byproduct A2E, followed by RPE atrophy. Maiti et al reduced lipofuscin production in a murine model by blocking RPE65, a critical enzyme for the retinoid cycle.⁴

How does lipofuscin affect the RPE?
Increased lipofuscin granule accumulation accelerates RPE death by several proposed mechanisms. A key molecular component of lipofuscin is A2E, which is formed by the combination of all-trans-retinal and phosphatidylethanolamine in a 2:1 ratio. A2E causes a dose-dependent increase in plasma membrane permeability through phospholipid bilayer solubilization and membrane bleb formation.¹ There is also evidence that A2E is phototoxic. Singlet oxygen generation following blue light absorption leads to formation of A2E-epoxides at double bond locations. These epoxides then directly damage the RPE DNA through fragmentation.¹ In addition, recent in vitro studies have implicated A2E in complement activation,⁶ as well as alterations in cholesterol metabolism.⁷

How can lipofuscin be imaged in vivo?
Delori and colleagues, using both in vivo and ex vivo spectrophotometric analysis, demonstrated that the dominant fluorophore with FAF is RPE lipofuscin.⁸ They showed that optimal excitation occurs at 510 nm with peak emission occurring at 630 nm, and reported FAF patterns in healthy patients as well as those with retinal diseases.⁸ Multiple animal experiments have demonstrated the vitamin A-dependent nature of FAF.^9-11 Both Robison and Katz showed a reduction in FAF corresponding to a diet with low retinoid content.^9,10 Radu and colleagues reported decreased lipofuscin and FAF with pharmacologic reduction of serum retinol,¹¹ and increased RPE lipofuscin pigments with vitamin A supplementation in a mouse model.⁵

Several other investigators have examined the clinical implications of increased autofluorescence found at the borders of geographic atrophy and its relationship to progression of atrophy in patients with dry AMD.^12-17 As a result, FAF is utilized increasingly for clinical trials evaluating new therapies for dry AMD.

Are treatments targeting the vitamin A pathway being conducted?
Currently, fenretinide is being evaluated as treatment for dry AMD in a phase II clinical trial (clinicaltrials.gov identifier NCT00429936) by Sirion Therapeutics, Inc (San Diego, CA). Fenretinide competes with retinol to bind retinol binding protein (RBP), and promotes clearance of retinol and RBP. Therefore, fenretinide theoretically limits the amount of retinol entering the RPE and the visual cycle. Investigators conducting the study propose this will lead to a decrease in retinoid precursors available for the formation of A2E. Preclinical data demonstrates a dose-dependent reduction in A2E.¹⁸ Side effects of retinol-reducing therapy may include delays in dark adaptation. Marmor and colleagues reported a significant dose-dependent delay in dark adaptation and rod electroretinogram responses in two children receiving fenretinide for the treatment of neuroblastoma. They also report a return to normal function with cessation of fenretinide.¹⁹

Conclusion
Vitamin A may play a role in the pathogenesis of AMD via formation of A2E and lipofuscin, which are implicated in RPE and photoreceptor cell death. The exact mechanism leading to the accumulation of retinoid byproducts and A2E in AMD is not known. However, clinicians are increasingly utilizing FAF to monitor lipofuscin accumulation and progression of AMD, and researchers are investigating means of reducing retinol entry into the visual cycle as a way of decreasing lipofuscin-mediated cell death.

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