Gisela Velez

Humanized anti-interleukin-2 (IL-2) receptor alpha therapy: long-term results in uveitis patients and preliminary safety and activity data for establishing parameters for subcutaneous administration

Therapy for severe uveitis is frequently long-term immunosuppression using systemic corticosteroids and cytotoxic agents, but side effects make long-term therapy difficult. A long-term (>4 year) Phase I/II single armed interventional study using intravenous anti-IL-2 receptor alpha treatments (daclizumab) and a short-term Phase II study evaluating the use of a subcutaneous daclizumab formulation were conducted. Patients were tapered off their systemic immunosuppressive therapy and received daclizumab infusions or subcutaneous injections at intervals varying from 2 to 6 weeks. In the long-term study, seven of ten enrolled patients were tapered from their original immunosuppressive medications and maintained exclusively on repeated daclizumab infusions for control of their uveitis for over 4 years. No patient was permanently removed from therapy for an adverse event ascribed to the medication. The use of 6-week infusion intervals led to recurrence of uveitis, while 2- to 4-week intervals did not. Only one patient developed measurable anti-daclizumab antibodies but this disappeared when subcutaneous therapy was begun. In the short-term study, four of the five patients receiving the subcutaneous formulation met the study endpoints for success within the first 12 weeks. All five were successful by 26 weeks. These studies provide preliminary evidence that regularly administered long-term daclizumab therapy can be given in lieu of standard immunosuppression for years to treat severe uveitis and that subcutaneously administered daclizumab appeared to be a clinically viable treatment strategy. These studies suggest that anti-IL-2 receptor blockade could be useful in the treatment of Th1-mediated autoimmune conditions.

A Double-masked, Randomized Study to Investigate the Safety and Efficacy of Daclizumab to Treat the Ocular Complications Related to Behcet's Disease

Purpose: To investigate the safety and efficacy of daclizumab (Zenapax, humanized anti-Tac, HAT) in controlling the ocular manifestations of Behçets disease. Design: Randomized, placebo-controlled, double-masked clinical trial. Participants: Seventeen participants with Behçets disease experiencing at least two prior ocular attacks and requiring treatment with immunosuppressive agents for the ocular complications of Behçets disease. Methods: Participants received either intravenous placebo or daclizumab (1 mg/kg) infusions every two weeks for six weeks, then every four weeks while continuing their standard immunosuppressive regimens. If clinically indicated, tapering of the standard immunosuppressive medications was allowed after six months of study enrollment. Complete ocular and physical examinations and an adverse event assessment were performed at baseline and prior to each study infusion. Main Outcome Measures: Primary safety endpoints were the development of a life-threatening complication or a severe opportunistic infection. Primary efficacy outcomes were the number of ocular attacks and an assessment of systemic immunosuppressive medications required during the study, including the ability to taper concomitant immunosuppressive therapy. Results: Nine participants randomized to daclizumab and eight to placebo were followed monthly. Follow-up ranged from one to 34 months, with a median follow-up of 15 months. Two participants randomized to daclizumab discontinued study therapy prior to the end of the study for personal reasons. No participant experienced a safety endpoint, and visual acuity remained stable in all participants during the course of the study. Ten participants (six daclizumab, four placebo) experienced ocular attacks requiring therapy. The median ocular attack rate during the study was greater in the daclizumab arm than the placebo arm (median 1.27 vs. 0.17 attacks/year, respectively). Participants in the placebo arm also experienced a greater reduction in the immunosuppressive medication score compared to participants receiving daclizumab (median –4.0 vs. –1.0, respectively). Conclusions: The observed results in the placebo group demonstrate that careful follow-up and treatment with standard combination immunosuppressive therapy can be effective for the management of the ocular complications of Behçets disease. In our small study, there was no suggestion that daclizumab was beneficial in comparison with placebo. However, the low observed attack rate limited our ability to make a definitive treatment group comparison.

New developments in sustained release drug delivery for the treatment of intraocular disease

Over the past 20 years, drug development has led to numerous medications with the potential to treat intraocular disease. These medications usually require systemic or frequent topical administration, which can limit the types of medications that can be used and the diseases that can be treated effectively. Topical administration of medications can successfully treat a number of ocular diseases such as glaucoma, inflammation of the anterior segment or ocular surface, and external diseases of the eye. Nevertheless, topical delivery often fails to provide therapeutic levels in the vitreous cavity or posterior segment, and therefore is inadequate in the treatment of vitreoretinal diseases. Drugs can often be delivered to the posterior segment by injection via the pars plana, as is the case, for example, with ganciclovir, foscarnet, and cidofovir. However, depending on the rate of clearance from the vitreous of a particular medication, large boluses and frequent administrations may be required to ensure therapeutic levels over an extended period of time. Frequent injections in clinical practice may not be practical for chronic diseases that can sometimes require multiple weekly administrations over months or years. In addition, multiple intraocular injections can lead to an increased likelihood of complications, such as vitreous haemorrhage, retinal detachment, and endophthalmitis. The blood-ocular barrier is the main obstacle in the treatment of intraocular disease with systemic medications. Poor penetration of many drugs into the eye not only limits the number of medications available for the treatment of ocular diseases, but also limits the extent to which those available can be used without incurring serious systemic side effects. Medications, such as prednisone, cytotoxic agents for the treatment of intraocular inflammation, and antivirals for the treatment of cytomegalovirus (CMV) retinitis often cause severe side effects at the dosages needed to achieve their desired therapeutic effect. This is particularly problematic …

Growth Factors Outside the PDGF Family Drive Experimental PVR

PURPOSE Proliferative vitreoretinopathy (PVR) is a recurring and problematic disease for which there is no pharmacologic treatment. Platelet-derived growth factor (PDGF) in the vitreous is associated with experimental and clinical PVR. Furthermore, PDGF receptors (PDGFRs) are present and activated in epiretinal membranes of patient donors, and they are essential for experimental PVR. These observations suggest that PVR arises at least in part from PDGF/PDGFR-driven events. The goal of this study was to determine whether PDGFs were a potential therapeutic target for PVR. METHODS Experimental PVR was induced in rabbits by injecting fibroblasts. Vitreous specimens were collected from experimental rabbits or from patients undergoing vitrectomy to repair retinal detachment. A neutralizing PDGF antibody and a PDGF Trap were tested for their ability to prevent experimental PVR. Activation of PDGFR was monitored by antiphosphotyrosine Western blot analysis of immunoprecipitated PDGFRs. Contraction of collagen gels was monitored in vitro. RESULTS Neutralizing vitreal PDGFs did not effectively attenuate PVR, even though the reagents used potently blocked PDGF-dependent activation of the PDGF alpha receptor (PDGFRalpha). Vitreal growth factors outside the PDGF family modestly activated PDGFRalpha and appeared to do so without engaging the ligand-binding domain of PDGFRalpha. This indirect route to activate PDGFRalpha had profound functional consequences. It promoted the contraction of collagen gels and appeared sufficient to drive experimental PVR. CONCLUSIONS Although PDGF appears to be a poor therapeutic target, PDGFRalpha is particularly attractive because it can be activated by a much larger spectrum of vitreal growth factors than previously appreciated.

Fluorescein Angiographic Findings In Primary Intraocular Lymphoma

Purpose Primary intraocular lymphoma (PIOL), also known as primary central nervous system lymphoma, is a rare yet blinding and fatal disease. Often presenting with ocular involvement, it can masquerade as posterior or intermediate uveitis, thus delaying diagnosis. A noninvasive ancillary test such as fluorescein angiography could be helpful in raising the level of suspicion in the diagnosis of this disease. Methods Results of fluorescein angiography (FA) and clinical characteristics of 17 patients (31 eyes) who presented to the National Eye Institute with the diagnosis of PIOL (confirmed by histopathologic analysis) were reviewed. Results The most common angiographic characteristics included disturbances at the level of the retinal pigment epithelium (RPE), such as granularity (19 eyes [61%]), blockage (17 eyes [55%]), and late staining (14 eyes [45%]). These changes are well correlated to histopathologic findings of lymphoma cells located between the RPE and Bruchs membrane. Perivascular staining or leakage and cystoid macular edema were rare. Other less common findings included pigment epithelial detachments and punctate hyperfluorescent lesions. Clinical characteristics found in eyes for which results of FA were available included vitreitis (29 eyes [94%]), subretinal infiltrates (19 eyes [61%]), and anterior chamber cells (10 eyes [32%]). In some cases, clinical examination did not correlate with FA findings. Conclusions Although PIOL may present with a normal angiographic phenotype, extensive RPE changes demonstrated by FA, combined with the absence of perivascular staining or leakage and macular edema, may be associated with and distinctive of PIOL.

N-Acetylcysteine Suppresses Retinal Detachment in an Experimental Model of Proliferative Vitreoretinopathy

Proliferative vitreoretinopathy (PVR) is a complication that develops in 5% to 10% of patients who undergo surgery to correct a detached retina. The only treatment option for PVR is surgical intervention, which has a limited success rate that diminishes in patients with recurring PVR. Our recent studies revealed that antioxidants prevented intracellular signaling events that were essential for experimental PVR. The purpose of this study was to test whether N-acetyl-cysteine (NAC), an antioxidant used in a variety of clinical settings, was capable of protecting rabbits from PVR. Vitreous-driven activation of PDGFRalpha and cellular responses intrinsic to PVR (contraction of collagen gels and cell proliferation) were blocked by concentrations of NAC that were well below the maximum tolerated dose. Furthermore, intravitreal injection of NAC effectively protected rabbits from developing retinal detachment, which is the sight-robbing phase of PVR. Finally, these observations with an animal model appear relevant to clinical PVR because NAC prevented human PVR vitreous-induced contraction of primary RPE cells derived from a human PVR membrane. Our observations demonstrate that antioxidants significantly inhibited experimental PVR, and suggest that antioxidants have the potential to function as a PVR prophylactic in patients undergoing retinal surgery to repair a detached retina.

Pathological Signaling via Platelet-Derived Growth Factor Receptor Involves Chronic Activation of Akt and Suppression of p53

ABSTRACT In contrast to direct activation of platelet-derived growth factor (PDGF) receptor α (PDGFRα) via PDGF, indirect activation via growth factors outside the PDGF family failed to induce dimerization, internalization, and degradation of PDGFRα. Chronically activated, monomeric PDGFRα induced prolonged activation of Akt and suppressed the level of p53. These events were sufficient to promote both cellular responses (proliferation, survival, and contraction) that are intrinsic to proliferative vitreoretinopathy (PVR) and induce the disease itself. This signature signaling pathway appeared to extend beyond PVR since deregulating PDGFRα in ways that promote solid tumors also resulted in chronic activation of Akt and a decline in the level of p53.

Expression of PDGFRα Is a Determinant of the PVR Potential of ARPE19 Cells

PURPOSE Previous studies indicate that the expression of platelet-derived growth factor (PDGF) receptor α (PDGFRα) dramatically increases the ability of fibroblasts to induce experimental proliferative vitreoretinopathy (PVR). The purpose of this study was to determine whether PDGFRα contributed to the PVR potential of retinal pigment epithelial (RPE) cells, one of the most abundant cell types in PVR membranes. METHODS PDGFRα expression in human ARPE19 cells was increased or decreased by stably expressing the PDGFRα cDNA or short hairpin (sh) RNA directed at PDGFRα, respectively. The level of PDGFRα expression in the resulting panel of cell lines was either barely detectable (KD), standard (similar to the level of primary RPE cells), or overexpressed approximately 80-fold. Western blot analysis was used to assess the level of p53 and the activation state of PDGFRα and Akt. The following cellular responses were monitored: proliferation, apoptosis, and contraction. The PVR potential of cells was tested in a rabbit model of PVR in which cells were coinjected with platelet-rich plasma into the vitreous. RESULTS Comparison of KD and overexpressing cells indicated that high-level expression of PDGFRα dramatically augmented signaling events, cellular responses, and the PVR potential of ARPE19 cells. However, all these outcomes were also significantly increased, albeit not as robustly, by PDGFRα expression to the level typically present in RPE cells. CONCLUSIONS Even though RPE cells express substantially less PDGFRα than fibroblasts, it significantly boosts PVR-related signaling events, cellular responses, and the PVR potential of ARPE19 cells. These studies suggest that inhibiting activation, signaling, or both by PDGFRα has the potential to prevent the development of PVR.

Plasmin Is the Major Protease Responsible for Processing PDGF-C in the Vitreous of Patients with Proliferative Vitreoretinopathy

PURPOSE Proliferative vitreoretinopathy (PVR) is the primary cause of failure of retinal reattachment surgery. Growth factors such as platelet-derived growth factor (PDGF) are strongly associated with PVR. Of the five PDGF family members, PDGF-C predominates in the vitreous of experimental and clinical PVR. PDGF-C is secreted as a latent protein that requires proteolytic processing for activation. Although tissue plasminogen activator (tPA) is primarily responsible for processing PDGF-C in cultured cells, it constitutes a minority of the processing activity in the vitreous of experimental animals and in patients with PVR. Identifying the major PDGF-C processing protease was the purpose of this study. METHODS The presence of serum proteins in the vitreous was detected by Coomassie blue staining and Western blotting. PDGF-C processing activity was detected in an in vitro processing assay using either native or recombinant PDGF-C as the substrate. Plasmin activity was blocked using alpha(2)-plasmin inhibitor. Phosphorylation of the PDGF receptor (PDGFR) was monitored by antiphosphotyrosine Western blotting. Vitreous specimens were collected from experimental rabbits or from patients undergoing vitrectomy to repair retinal detachment or for other reasons. RESULTS A number of prominent serum proteins (albumin and IgG) were detected in the vitreous of all patients undergoing retinal surgery. The level of these proteins markedly increased in the vitreous of rabbits as they developed PVR. These observations suggested that serum-borne proteases are also likely to be present in the vitreous. Indeed, plasmin (a protease capable of processing PDGF-C) was present in the vitreous from PVR rabbits and retinal surgery patients. Plasmin was dramatically more effective than tPA in processing PDGF-C in an in vitro assay. Blocking plasmin activity eliminated most of the processing activity in the vitreous of patients and rabbits with PVR. CONCLUSIONS Plasmin was the major PDGF-C processing protease in the vitreous of PVR rabbits and patients undergoing retinal surgery. Blocking plasmin prevented the generation of active PDGF-C, which is the major PDGF isoform relevant for PVR. These observations are the first report of an in vivo protease responsible for processing PDGF-C. In addition, plasmin was identified as a novel therapeutic target for patients with PVR.

High-dose intravitreal ganciclovir and foscarnet for cytomegalovirus retinitis

PURPOSE To describe the chronic use of high doses of intravitreal ganciclovir, in combination with foscarnet, for the treatment of cytomegalovirus retinitis. METHODS A 31-year-old man with human immunodeficiency virus (HIV) infection and unilateral active cytomegalovirus retinitis was treated with escalating intravitreal injections of ganciclovir (up to 3.0 mg twice a week) in combination with foscarnet (up to 2.4 mg twice a week) over the course of approximately 1 year. RESULTS Complete regression of the retinitis was obtained with high doses of intravitreal ganciclovir and foscarnet. Visual acuity in the affected eye remained 20/20 throughout the course of therapy. No ganciclovir retinal toxicity was identified. CONCLUSION High doses of intravitreal ganciclovir in combination with foscarnet can be well tolerated and may be required to successfully control cytomegalovirus retinitis in some patients.