Enzymatic Manipulation of the Vitreous Cavity
POLLY A. QUIRAM, MD, PhD
During the past decade, significant progress has been made toward understanding the role of the vitreous in retinal disease. It is known that traction at the vitreoretinal interface contributes to the retinal pathology seen in proliferative diabetic retinopathy, proliferative vitreoretinopathy, macular puckers, diabetic macular edema, vitreomacular traction syndrome, macular holes, and retinal detachments.1
Surgical management of these disorders often targets separating the posterior hyaloid from the internal limiting membrane (ILM), thereby creating a posterior vitreous detachment (PVD). Mechanical separation of the vitreoretinal interface with vitrectomy may not remove all of the vitreous or completely separate the vitreoretinal junction, as cortical vitreous fibrils are left behind on the ILM.2 Incomplete removal of the vitreous may result in surgical failure.3 Fortunately, with a better understanding of vitreous anatomy and biochemistry, enzymatic agents have been used to cleave the vitreoretinal junction and chemically manipulate the vitreous.3
The term "pharmacologic vitreodynamics" has been coined to describe the use of enzymatic agents to alter the vitreous cavity. This term details the complex mechanical and biochemical changes that occur following PVD formation, which includes alterations in molecular flux.4 Current clinical trials are investigating the efficacy of these agents as surgical adjuncts and alternatives to surgery. This article will review the current understanding of these vitreous-altering enzymatics and summarize ongoing clinical investigations.
|Polly A Quiram, MD, PhD, is a vitreoretinal surgeon at Vitreoretinal Surgery PA in Minneapolis and an associate professor at the University of Minnesota. She reports no financial interest in any product mentioned in this article. Dr. Quiram can be reached via e-mail email@example.com.|
IDEAL PHARMACOLOGIC AGENTS: PLASMIN AND MICROPLASMIN
Several authors have proposed substances, enzymatic and nonenzymatic, to cleave the vitreoretinal juncture or liquefy the central vitreous as an adjunct to vitreoretinal surgery or as a method to resolve vitreous traction.5-9 Whereas previously studied agents have been shown to be toxic to the retina,10-12 enzymatic manipulation can be performed with agents with a more favorable safety profile such as hyaluronidase, plasmin, and microplasmin.13-15 While hyaluronidase (Vitrase) has been shown to cause liquefaction of the vitreous without induction of a PVD,9 microplasmin and plasmin cause vitreous liquefaction and induction of a PVD without damage to the retina.16,17
Hyaluronidase is a highly purified ovine enzyme that primarily digests the proteoglycan hyaluronan, which comprises a large component of the vitreous body.14,18 It was originally studied in patients with dense vitreous hemorrhage to promote vitreous liquefaction and resolve vitreous hemorrhage, but phase 3 clinical trials concluded that hyaluronidase is slightly more effective than placebo.18
Although hyaluronidase has been shown to decrease vitreous macromolecule size, suggesting a role for vitreous liquefaction, it cannot induce a PVD.19,20 Effective enzymatic manipulation of the vitreous cavity requires vitreous liquefaction and complete vitreoretinal interface separation (PVD) without damaging the retina. Both plasmin and microplasmin safely eliminate vitreous traction and result in an internal limiting membrane clean of cortical vitreous as shown in animal and human studies.8,9
Plasmin is an 88-kD nonspecific protease capable of hydrolyzing the glycoproteins laminin and fibronectin, which bridge collagen fibers between the posterior vitreous cortex and the ILM (Figure 1).2 Gandorfer and associates reported that a single injection of plasmin enzyme can cleave the vitreoretinal junction without causing morphological changes to the retina in postmortem porcine eyes. Autologous plasmin enzyme (APE), which is isolated from a patient's serum, has been extensively studied in humans for treatment of macular holes, diabetic retinopathy, and congenital x-linked retinoschisis.16,21,22 In these eyes, plasmin enzyme facilitates the release of the posterior vitreous from the retina. Plasmin has been show to be safe at concentrations 10 times (4 U) the amount needed for PVD formation (0.4 U) without ERG changes or morphologic changes to the ILM and retina.23
Figure 1. Structure of plasmin enzyme an 88-kD nonspecific protease capable of hydrolyzing the glycoproteins laminin and fibronectin. The active domain is highlighted in red. Microplasmin consists of the active domain minus the "kringle" domains.
Microplasmin (ThromboGenics Ltd., Dublin, Ireland) is a recombinant protein containing 1 active protease site of human plasmin, but it lacks many of the remaining "kringle" domains.24 It functions as a direct-acting thrombolytic agent, and it is a much smaller molecule (29 kD) than human plasmin (88 kD). It has been proposed that the smaller size of microplasmin enables the molecule to penetrate epiretinal tissue more effectively than plasmin.25
Microplasmin alters the vitreous cavity by vitreous liquefaction and cleavage of the vitreoretinal interface with a single intravitreal injection of 62 μg or 125 μg.16 Microplasmin works rapidly with induction of PVD and vitreous liquefaction within 30 minutes.25,26 Since microplasmin is a recombinant protein, it can be prepared and manufactured in large quantities and the labor-intensive preparation that is needed for the production of plasmin enzyme can be avoided.
CLINICAL TRIALS: MICROPLASMIN AS A SURGICAL ADJUNCT
MIVI III is a phase 2b randomized, placebo-controlled, double-masked, parallel-group, dose-ranging, US-based study to determine the efficacy of microplasmin as a surgical adjunct in vitrectomy surgery. This study enrolled patients with vitreomacular traction or macular holes in whom vitrectomy was indicated but had no evidence of a complete PVD. Exclusion criteria included patient with evidence of complete macular PVD on biomicroscopy, B-scan, or OCT. Patients were randomized to sham or microplasmin injection (25, 75, or 125 μg dose). Injection was performed 7 days before planned pars plana vitrectomy surgery. The primary endpoint was the proportion of patients achieving total PVD at the 7-day visit. A secondary endpoint included the number of patients avoiding surgical intervention. If vitrectomy was performed, presence of PVD was determined at the time of surgery. The surgery was cancelled if the patient showed significant visual improvement, release of vitreous traction, or closure of the macular hole (Figures 2 and 3).
Figure 2. MIVI III Stage 1 Macular Hole. (A) Optical coherence tomography demonstrating evidence of a macular hole with visual acuity of 20/63. (B) OCT image 1 week after intravitreal injection of microplasmin. Macular hole is closed and surgery is cancelled. Visual acuity of 20/63. (C) OCT image 6 months after intravitreal injection of microplasmin. Macular hole is closed and visual acuity improved to 20/25.
Figure 3. MIVI III Stage 2 Macular Hole. (A) OCT demonstrating evidence of a macular hole with visual acuity of 20/63. (B) OCT image 1 week after intravitreal injection of microplasmin. Macular hole is closed and surgery is cancelled. Visual acuity of 20/50. (C) OCT image 1 month after intravitreal injection of microplasmin. Macular hole is closed and visual acuity improved to 20/25.
Of the 125 patients enrolled, 55% had macular holes and 38% had vitreomacular traction. Induction of a PVD occurred in 21% of all patients receiving microplasmin injection. A PVD was identified in 30% of patients receiving the highest microplasmin dose (125 μg). Surgery was avoided in 27% of patients with VMT and 35% of patients with macular holes (125 μg dose). These results showed that microplasmin is well tolerated, is effective as a surgical adjunct, achieves PVD prior to vitrectomy, can achieve VMT resolution and macular hole closure without the need for vitrectomy, and has optimal dose-response effect at 125 μg.
CLINICAL TRIALS: MICROPLASMIN TO AVOID SURGICAL INTERVENTION
The European MIVI IIT (Traction) was a randomized, sham-injection controlled, double-masked, ascending-dose, dose-ranging trial of microplasmin for nonsurgical PVD induction for treatment of vitreomacular traction. Inclusion criteria was evidence of VMT by ultrasound and OCT with the presence of macular thickening >250 μm. Common conditions included VMT, macular holes, or tractional diabetic macular edema. Visual acuity had to be 20/40 or worse in the study eye and 20/400 or better in the fellow eye.
Two cohorts of 15 patients received 75 μg or 125 μg of microplasmin or sham with a 4:1 randomization of microplasmin vs sham. Follow-up data were collected for 6 months. Of the 30 patients enrolled, 16 (53%) had VMT 6 (20%) had macular holes, and 8 (27%) had tractional diabetic macular edema. Intravitreal injection of microplasmin was well tolerated with no adverse events including retinal tears, retinal detachments, or endophthalmitis reported.
Results showed complete VMT resolution in 40% of patients, with the best results (50% resolution) seen with the higher dose of microplasmin (125 μg) (Figure 4). Closure of macular holes without vitrectomy occurred in 50% of eyes (Figure 5). A greater than 3-line improvement of visual acuity occurred in 20% of patients. The MIVI 2T trial results show that microplasmin relieves VMT in 40% of patients with an abnormal vitreoretinal interface.
Figure 4. Results of MIVI 2T Trial: Vitreomacular traction (VMT). (A) OCT demonstrating evidence of VMT with visual acuity of 20/100. (B) OCT image 1 week after intravitreal injection of microplasmin. Vitreoretinal traction is released and visual acuity improved to 20/63. (C) OCT image 3 months after intravitreal injection of microplasmin. No evidence of VMT is present and visual acuity is 20/16.
Figure 5. Results of MIVI 2T Trial: Stage 2 Macular Hole. (A) OCT demonstrating evidence of a stage 2 macular hole. (B) OCT image 3 days after intravitreal injection of microplasmin. Macular hole is closed with improvement of visual acuity. (C) Resolving subretinal fluid 28 days after injection of microplasmin. Visual acuity has improved 11 letters from baseline.
MIVI-TRUST (Microplasmin for Vitreous Injection — Tractional Release withoUt Surgical Treatment) is the US version of the MIVI IIT European trial. It is a randomized, placebo-injection controlled, double-masked, phase 3 trial. This study is designed to demonstrate the safety and efficacy of intravitreal injection of microplasmin for the nonsurgical treatment of focal vitreomacular traction. Endpoints include the proportion of patients with nonsurgical resolution of focal vitreomacular traction, closure of macular holes, or subjects with total PVD at day 28.
Inclusion criteria include the presence of focal vitreomacular traction that is related to decreased visual function with symptoms such as metamorphopsia, decreased visual acuity, or other visual complaint. BCVA must be 20/25 or worse in the study eye and BCVA of 20/800 or better in the nonstudy eye. Exclusion criteria include PDR, exudative macular degeneration, retinal vein occlusion, vitreous hemorrhage, high myopia, macular hole >400μm, history of retinal detachment in the study eye, or vitrectomy in the study eye. The follow-up period is 6 months and the patient allocation to microplasmin or sham injection is 2:1.
Interestingly, this is the first FDA-approved study to use OCT data to determine a primary endpoint. This study is currently enrolling patients.
FUTURE DIRECTIONS AND IMPLICATIONS OF "CHEMICAL VITRECTOMY"
Evidence has shown that enzymatic manipulation of the vitreous (chemical vitrectomy) can potentially relieve vitreoretinal traction, alter the influx of molecules, and alter oxygen levels in the vitreous cavity. For example, diabetic macular edema and diabetic retinopathy have the potential to be treated with microplasmin. A complete PVD is a strong negative risk factor for the progression of diabetic retinopathy27 and a complete posterior vitreous separation may allow for resolution of diabetic macular edema.16
Multiple growth factors, including vascular endothelial growth factor (VEGF), have been implicated in the pathogenesis of diabetic macular edema and neovascularization.28-30 The author and associates have previously shown that the creation of a microplasmin-assisted PVD increases vitreal oxygen and increases the rate of oxygen exchange within the vitreous cavity in animal models.19 It has been reported that induction of a PVD may alter the flux of molecules in the vitreous cavity and influence the concentration of vitreal growth factors.4,31 Data from cat models suggest that microplasmin-assisted PVD leads to decreased levels of vitreal VEGF.32 Microplasmin-induced PVD may increase vitreous oxygenation, thereby decreasing retinal ischemia and altering growth factor production.
It is well known that stabilization of proliferative diabetic retinopathy and diabetic macular edema often occurs following vitrectomy, possibly due to increased oxygenation, removal of the vitreous scaffold and an increase in molecular flux.33-38 It is not known if "chemical vitrectomy" has the same effect, but microplasmin may be able to delay the progression of diabetic retinopathy and diabetic macular edema by inducing a prophylactic PVD. Prospective clinical trials will be needed to address this theory.
The ability to induce vitreous liquefaction and a complete posterior vitreous detachment with a single intravitreal injection of microplasmin has potentially significant implications for management of multiple vitreoretinopathies. If used as a surgical adjunct, it has the potential to facilitate more complete removal of the vitreous gel, decrease surgical time, and reduce intraoperative complications. As prophylactic therapy, its uses may include treatment for vitreomacular traction syndrome, proliferative diabetic retinopathy, macular holes and pretreatment before pneumatic retinopexy to decrease the incidence of new retinal breaks. We look forward to the promising results of ongoing clinical trials. RP
1. Sebag J. Pharmacologic vitreolysis. Retina. 1998;18:1-3.
2. Gandorfer A, Ulbig M, Kampik A. Plasmin-assisted vitrectomy eliminates cortical vitreous remnants. Eye. 2002;16:95-97.
3. Trese MT Enzymatic vitreous surgery. Semin Opthalmol. 2000;15:116-121
4. Goldenberg D, Trese M. Pharmocalogic vitreodynamics and molecular flux. Dev Ophthal. 2009;44:31-36.
5. Hermel M, Schrage NF. Efficacy of plasmin enzymes and chondroitinase ABC in creating posterior vitreous separation in the pig: a masked, placebo-controlled in vivo study. Grafes Arch Clin Exp Ophthalmol. 2007;245:399-406.
6. Sebag J. Molecular biology of pharmacologic vitreolysis. Trans Am Ophthalmol Soc. 2005,103:473-494.
7. Tezel TH, Del Priore LV, Kaplan HJ. Posterior vitreous detachment with Dispase. Retina. 1998;8:7-15.
8. Staubach F, Nober V, Janknecht P. Enzyme-assisted vitrectomy in enucleated pig eyes: a comparison of hyaluronidase, chondroitinase, and plasmin. Curr Eye Res. 2004;29:261-268.
9. Wang ZL, Zhang X, Xun X, et al. PVD Following plasmin but not hyaluronidase: implications for combination pharmacologic vitreolysis therapy. Retina. 2005;25:35-43.
10. Zhu D, Chen H, Xu X. Effects of intravitreal dispase on vitreoretinal interface in rabbits. Curr Eye Res. 2006;31:935-946.
11. Jorge R, Oyamaguchi EK, Cardillo JA, Gobbi A. Intravitreal injection of dispase causes retinal hemorrhages in rabbit and human eyes. Curr Eye Res. 2003;26:107-112.
12. Hagemann GS, Russell SR. Chondroitinate-mediated disinsertion of the primate vitreous body. Invest Ophthalmol Vis Sci. 1994;35(suppl):28.
13. Sakuma T, Tanaka M, Mitzota A, Inoue J. Safety of in vivo pharmacologic vitreolysis with recombinant microplasmin in rabbit eyes. Invest Ophthalmol Vis Sci. 2005;46:3295-3299.
14. Kuppermann BD, Thomas EL, De Smet MD, Grillone LR. Safety results of two phase III trials of an intravitreous injection of highly purified ovine hyaluronidase (Vitrase®) for the management of vitreous hemorrhage. Am J Ophthalmol. 2005;140:585-597.
15. Liotta LA, Goldfarb RH, Brundage R, Siegal GP. Effect of plasminogen activator (urokinase), plasmin, and thrombin on glycoprotein and collagenous components of basement membrane. Cancer Res. 1981;41:4629-4636.
16. Williams JG, Trese MT, Williams GA, Hartzer M. Autologous plasmin enzyme in the surgical management of diabetic retinopathy. Ophthalmology 2001;108:1902-1905.
17. Gandorfer A, Rohlender M, Sethi C et al. Posterior vitreous detachment induced by microplasmin.Invest Opthalmol Vis Sci 2004, 45:641-647.
18. Kuppermann BD, Thomas EL, De Smet MD, Grillone LR. Pooled efficacy results from two multinational randomized controlled clinical trials of a single intravitreous injection of highly purified ovine hyaluronidase (Vitrase®) for the management of vitreous hemorrhage. Am J Ophthalmol 2005;140:573-584.
19. Quiram PA, Leverenz VA, Baker RM, Dang L. Microplasmin-induced posterior vitreous detachment affects vitreous oxygen levels. Retina. 2007;27:1090-1096.
20. Hikichi T, Kado M, Yoshida A. Intravitreal injection of hyaluronidase cannot induce posterior vitreous detachment in the rabbit. Retina 2000;20:195-198
21. Trese MT, Williams GA, Hartzer MK. A new approach to stage 3 macular holes. Ophthalmology. 2000; 107:1607-1611.
22. Wu WC, Drenser KA, Capone A, Williams GA, Trese MT. Plasmin enzyme-assisted vitreoretinal surgery in congenital X-linked retinoschisis: surgical techniques based on a new classification system. Retina. 2007;27:1079-1085.
23. Wang F, Wang Z, Sun X, Wang F. Safety and efficacy of dispase and plasmin in pharmacologic vitreolysis. Invest Ophthalmol Vis Sci 2004;45:3286-3290.
24. Nagai N, Demarsin E, Van Hoef B, et al. Recombinant human microplasmin: production and potential therapeutic properties. Thromb Haemost. 2003;1:307-313.
25. Asami T, Terasaki H, Kachi S, et al. Ultrastructure of internal limiting membrane removed during plasmin-assisted vitrectomy from eyes with diabetic macular edema. Ophthalmology. 2004;111:231-237.
26. Sebag J, Ansari RR, Suh KI. Pharmacologic vitreolysis with microplasmin increases vitreous diffusion coefficients. Graefes Arch Clin Exp Ophthalmol. 2007;245:576-580.
27. Ono R, Kakehashi A, Yamagami H, et al. Prospective assessment of proliferative diabetic retinopathy with observations of posterior vitreous detachment. Int Ophthalmol. 2005;26:15-19.
28. Paques M, Massin P, Gaudric A. Growth factors and diabetic retinopathy. Diabetes Metabolism. 1997;23:125-130.
29. Patel JI, Tombran-Tink J, Hykin PG, Gregor ZJ. Vitreous and aqueous concentrations of proangiogenic, antiangiogenic factors and other cytokines in diabetic retinopathy patients with macular edema: implications for structural differences in macular profiles. Exp Eye Res. 2006;82:798-806.
30. Boulton M, Gregor Z, McLeod D, et al. Intravitreal growth factors in proliferative diabetic retinopathy: correlation with neovascular activity and glycaemic management. Br J Ophthalmol. 1997;81:228-233.
31. Stefánsson E, Loftsson T. The Stokes-Einstein equation and the physiological effects of vitreous surgery. Acta Ophthalmol Scand. 2006;84:718-719.
32. Quiram PA, Leverenz V, Baker R, Loan D. Enzymatic induction of a posterior vitreous detachment alters molecular vitreodynamics in animal models. [ARVO Abstract]. Invest Ophthalmol Vis Sci. 2007; Abstract nr 83.
33. Holekamp NM, Shui YB, Beebe DC, Vitrectomy surgery increases oxygen exposure to the lens: a possible mechanism for nuclear cataract formation. Am J Ophthalmol. 2005;139:302-310.
34. Stefansson E, Landers MB How does vitrectomy affect diabetic macular edema? Am J Ophthalmol. 2006;141:984.
35. Grigorian R, Bhagat N, Lanzetta P, et al Pars plana vitrectomy for refractory diabetic macular edema. Semin Ophthalmol. 2003;18:116-120.
36. Recchia FM, Ruby AJ, Carvalho, et al. Pars plana vitrectomy with removal of the internal limiting membrane in the treatment of persistent diabetic macular edema. Am J Ophthalmol. 2005;139:447-454.
37. Gotzaridis IV, Lit IS, D'Amico DJ. Progress in vitreoretinal surgery for proliferative diabetic retinopathy. Semin Ophthalmol. 2001;16:31-40.
38. Rosenblatt BJ, Shah GK, Sharma S, et al Pars plana vitrectomy with internal limiting membranectomy for refractory diabetic macular edema without a taut posterior hyaloid. Graefes Arch Clin Exp Ophthalmol. 2005;243:20-25.
Retinal Physician, Issue: July 2009