Antiproliferative and cytotoxic properties of bevacizumab on different ocular cells

AIM: To evaluate the antiproliferative and cytotoxic properties of bevacizumab, a monoclonal antibody against vascular endothelial growth factor (VEGF), on human retinal pigment epithelium (ARPE19) cells, rat retinal ganglion cells (RGC5), and pig choroidal endothelial cells (CEC). METHODS: Monolayer cultures of ARPE19, RGC5, and CEC were used. Bevacizumab (0.008-2.5 mg/ml), diluted in culture medium, was added to cells that were growing on cell culture dishes. Cellular proliferative activity was monitored by 5'-bromo-2'-deoxyuridine (BrdU) incorporation into cellular DNA and the morphology assessed microscopically. For cytotoxicity assays ARPE19, RGC5, and CEC cells were grown to confluence and then cultured in a serum depleted medium to ensure a static milieu. The MTT test was performed after 1 day. The "Live/Dead" viability/cytotoxicity assay was performed and analysed by fluorescence microscopy after 6, 12, 18, 24, 30, 36, and 48 hours of incubation. Expression of VEGF, VEGF receptors (VEGFR1 and VEGFR2) and von Willebrand factor was analysed by immunohistochemistry. RESULTS: No cytotoxicity of bevacizumab on RGC5, CEC, and ARPE19 cells could be observed after 1 day. However, after 2 days at a bevacizumab concentration of 2.5 mg/ml a moderate decrease in ARPE19 cell numbers and cell viability was observed. Bevacizumab caused a dose dependent suppression of DNA synthesis in CEC as a result of a moderate antiproliferative activity (maximum reduction 36.8%). No relevant antiproliferative effect of bevacizumab on RGC5 and ARPE19 cells could be observed when used at a concentration of 0.8 mg/ml or lower. CEC and ARPE 19 cells stained positively for VEGF, VEGFR1, and VEGFR2. More than 95% of the CEC were positive for von Willebrand factor. CONCLUSIONS: These experimental findings support the safety of intravitreal bevacizumab when used at the currently applied concentration of about 0.25 mg/ml. Bevacizumab exerts a moderate growth inhibition on CEC when used in concentrations of at least 0.025 mg/ml. However, at higher doses (2.5 mg/ml) bevacizumab may be harmful to the retinal pigment epithelium.     

Toxicity testing of the VEGF inhibitors bevacizumab,ranibizumab and pegaptanib in rats both with and without prior retinal ganglion cell damage

Purpose: To evaluate the effects of intravitreally introduced vascular endothelial growth factor (VEGF) inhibitors in rat eyes with healthy retinal ganglion cells (RGC) and into others with N‐methyl‐D‐aspartate (NMDA)‐induced RGC damage. Methods: Bevacizumab, ranibizumab and pegaptanib were intravitreally injected each at two different concentrations. Respective vehicles of the three substances served as controls. In a different group, additionally a rat anti‐VEGF antibody was injected after NMDA treatment. Retrogradely labelled RGC were counted on retinal wholemounts 1 week or 2 months after intravitreal introduction of the VEGF inhibitors. Electron microscopy (EM) was performed on normal rat eyes 2 months after introduction of the VEGF inhibitors. Results: RGC counts in healthy rat eyes were essentially unchanged from those of the control animals after the administration of both low and high concentrations of bevacizumab, ranibizumab or pegaptanib. Compared to the other two substances, however, high doses of pegaptanib and its respective vehicle significantly decreased RGC after 1 week and led to a marked increase of mitochondrial swelling in EM. In eyes with NMDA‐induced RGC damage, no changes of RGC numbers were detected after rat anti‐VEGF antibody or bevacizumab, ranibizumab and pegaptanib at both tested concentrations. Conclusions: Even at higher doses, bevacizumab and ranibizumab showed no toxic effects on RGC in vivo in either untreated rats or in the NMDA‐induced RGC damage model. Also a rat anti‐VEGF antibody showed no adverse effects after NMDA. Anti‐VEGF therapy therefore appears safe even for eyes with additional excitotoxic RGC damage. Potential harm from the pegaptanib carrier solution at very high local concentrations cannot be excluded.     

Are intravitreal bevacizumab and ranibizumab effective in a rat model of choroidal neovascularization?

Background  Vascular endothelial growth factor (VEGF) is an important stimulator of choroidal neovascularization (CNV). Bevacizumab (Avastin), ranibizumab (Lucentis) and pegaptanib sodium (Macugen) are anti-VEGF medications that have been used in the treatment of CNV. The purpose of our study is to evaluate the efficacy and safety of intravitreal injections of bevacizumab, ranibizumab and pegaptanib sodium in the treatment of CNV in a rat model. Methods  Multiple CNV lesions were induced by laser photocoagulation of the retina in Brown-Norway rats. After 3 weeks, 17 rats were divided into three groups and received intravitreal injections of bevacizumab, ranibizumab or pegaptanib sodium in different dosages. The lesions were evaluated by fluorescein angiography 1, 7, 14, and 28 days later to assess the efficacy of these medications. Results  Different doses of bevacizumab did not show any effect on stopping the leakage on fluorescein angiography on days 1, 7, 14, and 28. Ranibizumab and pegaptanib sodium did not stop the leakage of CNV either. No angiographic or histopathologic toxicity was observed. Conclusions  These three anti-VEGF agents did not show any therapeutic effect on stopping CNV leakage in rats. Previous experiments with ranibizumab in monkeys resulted in a significant decrease in leakage of CNV. The difference may be due to the fact that both ranibizumab and bevacizumab are humanized and species-specific. There are several studies evaluating the effect of bevacizumab in non-primates. Since bevacizumab is humanized, the results of studies on non-primates may not be similar to humans and non-human primates. Grant Support: Leir Foundation The authors have full control of the primary data and will provide it to Graefe’s Archive for Clinical and Experimental Ophthalmology at their request. Neither author has any conflict of interest, financial or otherwise, to report.     

Preclinical aspects of anti-VEGF agents for the treatment of wet AMD: ranibizumab and bevacizumab

Three anti-vascular endothelial growth factor (VEGF) therapies are currently used for the treatment of patients with wet age-related macular degeneration (AMD): pegaptanib, ranibizumab, and bevacizumab. Ranibizumab is an antibody fragment approved for the treatment of wet AMD. Bevacizumab is a full-length antibody registered for use in oncology but unlicensed for wet AMD. However, it is used off-label worldwide not only for wet AMD but also for various other ocular diseases associated with macular edema and abnormal vessel growth. We consider aspects of ranibizumab and bevacizumab in relation to their molecular characteristics, in vitro and in vivo properties, and preclinical safety data. Before 2009, most studies described the short-term toxicity of bevacizumab in multiple cell types of the eye. Since 2009, an increasing number of studies have compared the properties of ranibizumab and bevacizumab and investigated their impact on retinal cell functioning. Compared with bevacizumab, ranibizumab neutralizes VEGF better at low concentrations, maintains efficacy for longer, and has a higher retinal penetration and potency. Studies in animals demonstrate ranibizumab to be better localized to the injected eye, whereas bevacizumab appears to have a greater effect in the fellow eye. In humans, a localized and systemic effect has been reported for both molecules. In conclusion, overlapping yet distinct pharmacological properties of ranibizumab and bevacizumab indicate that safety or efficacy data from one cannot be extrapolated to the other.     

Comparison of choroidal and retinal endothelial cells: characteristics and response to VEGF isoforms and anti-VEGF treatments

Neovascular eye diseases such as wet age-related macular degeneration and proliferative diabetic retinopathy are two of the most common causes of irreversible visual loss. Although mediated by vascular endothelial growth factor (VEGF), the mechanisms of these diseases are not fully understood. Molecular inhibitors of VEGF including pegaptanib, ranibizumab and bevacizumab are used as treatments for these diseases. However, there have been very few direct comparisons between these agents, and as dose and treatment regimes differ their relative efficacies are hard to determine. In vitro comparisons tend to use cells from different sites or species, which show heterogeneity in their responses. The aim of this study was to compare the characteristics of primary cultures of isolated human choroidal endothelial cells (hCEC) and retinal endothelial cells (hREC), and their proliferation responses to stimulation with VEGF 121 and 165, and to compare the anti-proliferative effects of these three drugs. hCEC and hREC were positive for the cell markers VEGFR1, VEGFR2, CD31, CD34 and von Willebrand's factor (vWF), with greater expression of CD34 on the hREC compared to hCEC. Contrary to previous assumptions VEGF isoforms 121 and 165 were found to be equally potent in stimulating endothelial cell proliferation. However, hREC exhibited higher proliferation with either VEGF isoform compared to hCEC. The anti-VEGF treatments ranibizumab and bevacizumab were effective in decreasing proliferation of hCEC induced by the two VEGF isoforms, individually and in combination, with ranibizumab being moderately more effective, particularly in hREC. Pegaptanib was effective in controlling the proliferation of hCEC stimulated by VEGF 165, but was ineffective against the stimulatory effect of VEGF 121. There were found to be significant differences in microvascular endothelial cells from the retina and choroid, both in the expression of cell markers and their behaviour in response to growth factors and currently available anti-VEGF agents, highlighting the importance of targeting treatments to specific intraocular vascular beds and/or diseases.     

Comparison of subconjunctivally injected bevacizumab, ranibizumab, and pegaptanib for inhibition of corneal neovascularization in a rat model

AIM:To compare the efficacies of subconjunctival bevacizumab, ranibizumab, and pegaptanib sodium injections for the inhibition of corneal neovascularization in an experimental rat model. METHODS:Sixteen corneas of 16 rats were chemically cauterized and randomized into four groups:bevacizumab group that treated with 0.05mL/1.25mg bevacizumab, ranibizumab group that treated with 0.05mL/0.5mg ranibizumab, pegaptanib group that treated with 0.05mL/0.15mg pegaptanib sodium, and control group that treated with 0.05mL saline solution. Digital photographs of the corneas were taken and analyzed using an image analysis software program. All corneas were excised and examined histologically on the 15^th day. RESULTS: Each treatment group had significantly less neovascularized corneal areas and fewer blood vessels than the control group (all P<0.05). In addition, bevacizumab group had significantly less neovascularized corneal areas and fewer blood vessels than ranibizumab and pegaptanib groups (both P<0.05). However, there was no significant difference between the ranibizumab and pegaptanib groups regarding percentage of neovascularized corneal areas and number of blood vessels (both P>0.05). CONCLUSION:Subconjunctival bevacizumab, ranibizumab, and pegaptanib sodium were effective with no corneal epitheliopathy for inhibiting corneal neovascularization after corneal burn in rats. Bevacizumab was more effective than ranibizumab and pegaptanib sodium.     

Antipermeability and antiproliferative effects of standard and frozen bevacizumab on choroidal endothelial cells

BACKGROUND: Bevacizumab is an antiangiogenic compound developed to target tumour vessels. Its off-label use in ophthalmology requires in vitro testing on ocular cells. AIM: To quantify the antipermeability and antiproliferative effects of bevacizumab on cultured choroidal endothelial cells (CECs). It was examined whether deep-freezing of bevacizumab attenuates its antiangiogenic activity. METHODS: Porcine CECs were cultured in permeable insert systems. Permeability of the cell monolayers was quantified by a fluorescent isothiocyanate-dextran assay after treatment with vascular endothelial growth factor (VEGF; 20-100 ng/ml) alone and in combination with bevacizumab (0.1-1 mg/ml). Proliferation of the CECs was tested using a "wound scratch" assay. The experiments were repeated with bevacizumab after freezing at -20 degrees C for 5 days. RESULTS: Bevacizumab significantly reduced VEGF-induced permeability in a dose-dependant manner. A molar ratio of 2.6:1 of bevacizumab to VEGF was required for complete blocking of VEGF-induced rise in permeability. CEC proliferation was significantly blocked by bevacizumab (0.5 mg/ml). Thawed bevacizumab after deep freezing showed a moderate, but not statistically significant loss in activity. CONCLUSION: Bevacizumab significantly reduces VEGF-induced permeability and proliferation of CECs. Freezing and thawing of bevacizumab will affect its biological activity.     

老年性黄斑变性治疗方法试验比较的启示

老年性黄斑变性(AMD)治疗方法试验比较(CATT)结果显示,倍伐单抗(商品名Avastin)和雷珠单抗按照相同的给药计划给药,临床观察12个月,患者视力效果等同.雷珠单抗按需求给药组在视力上与每一个月给药组等同.倍伐单抗是一个对抗血管内皮生长因子(VEGF)的全长单克隆抗体(MAb),能够对抗VEGF所有有生物活性的亚型,阻止VEGF和受体结合;雷珠单抗是把倍伐单抗和抗原结合部的片段分离出来,在临床上完成了4期Ⅰ/Ⅱ观察的制剂.二者在价格上悬殊巨大,倍伐单抗50美元/支,而雷珠单抗2000美元/支.在如此价格差距下,如果倍伐单抗能够改善视力和黄斑解剖,其临床应用的性价比会优于雷珠单抗.CATT研究结果支持全球继续使用玻璃体腔入路的给药途径使用倍伐单抗作为对雷珠单抗的一种廉价选择.

Abstract：

Comparison of age-related macular degeneration treatments trials(CATT)reported the results of the first year:At 1 year,bevacizumab and ranibizumab had equivalent effects on visual acuity when administered according to the same schedule.Ranibizumab given as needed with monthly evaluation had effects on vision that were equivalent to those of ranibizumab administered monthly.Bevacizumab is a monoclonal antibody(MAb)and full-length anti-VEGF antibody that successfully prevents binding of all the biologically active isoforms of VEGF to the receptor.Ranibizumab is a isolated antigen-binding fragment (Fab)from bevacizumab using recombinant genetic techniques.Ranibizumab has been clinically evaluated in 4 Phase Ⅰ/Ⅱ studies.The cost of Avastin is ＄50 per dose compared with ＄ 2000 per dose for Ranibizumab.If Avastin could improve vision and macular anatomy,then it would not only be clinically superior to Ranibizumab.The CATT data support the continued global use of intravitreal bevacizumab as an effective.low-cost alternative to ranibizumab,particularly for patients paying all costs out of pocket.     

Bevacizumab modulates epithelial‐to‐mesenchymal transition in the retinal pigment epithelial cells via connective tissue growth factor up‐regulation

Purpose: To investigate the effect of bevacizumab treatment on connective tissue growth factor (CTGF) expression and the induction of epithelial‐to‐mesenchymal transition in ARPE‐19 cells and human donor retinal pigment epithelium (HRPE) cells in vitro. Methods: We quantitated the protein and gene expression level of CTGF by ELISA. The effect of Fc–Fc receptor (Fc–FcR) interaction on CTGF expression was evaluated by CD64 siRNA silencing. Expression of epithelial‐to‐mesenchymal transition markers, alpha‐smooth muscle actin (α‐SMA) and zona occludens protein (ZO‐1) was evaluated by Western blot. Cell migration and collagen gel contraction assay were examined by light microscopy, and collagen production was measured by ELISA. Results: Bevacizumab stimulation increased CTGF expression in ARPE‐19 and HRPE cells in a dose‐dependent manner. CD64 gene silencing inhibited the effect of bevacizumab‐induced CTGF up‐regulation. Bevacizumab increased the expression of α‐SMA and decreased the expression of ZO‐1 in ARPE‐19 cells. Bevacizumab also caused the release of type‐1 collagen and increased cell migration and contraction of collagen. Conclusions: Bevacizumab exerts pro‐fibrotic effects on human RPE cells at clinical doses by up‐regulation of CTGF expression via an Fc–FcR interaction. This effect of bevacizumab may be one of the underlying mechanisms involved in age‐related macular degeneration therapy or intravitreal bevacizumab‐associated complications.     

Evaluation of the Response to Ranibizumab Therapy following Bevacizumab Treatment Failure in Eyes with Diabetic Macular Edema

Background/Aims

Bevacizumab and ranibizumab are routinely used to treat diabetic macular edema (DME). We aim to evaluate the usefulness of switching to ranibizumab therapy following bevacizumab treatment failure in eyes with DME.

Methods

We performed a retrospective analysis of a consecutive group of patients with DME who received ranibizumab injections following the failure of bevacizumab injections. The injections were delivered following a pro re nata protocol every 4–6 weeks. The data collected included demographics, systemic and ophthalmic findings, as well as the central subfield thickness according to spectral-domain OCT.

Results

Eight eyes (5 patients) were included in the study. The median number of bevacizumab injections prior to the switch to ranibizumab was 4, and the median number of ranibizumab injections during the study was 2. The mean follow-up period was 541 ± 258 days. The mean central retinal thickness (CRT) (±SEM) was 539 ± 75 μm before the initiation of bevacizumab treatment, and 524 ± 43 μm after the last bevacizumab injection (p = 0.7). It reduced to 325 ± 26 μm following the ranibizumab injections (p = 0.0063). The best-corrected visual acuity (BCVA) improved in 4 eyes and remained stable in 4 eyes following the ranibizumab injections.

Conclusion

A ranibizumab therapy was effective in reducing the CRT in eyes that failed bevacizumab therapy. A BCVA improvement can also occur in these eyes. Switching between anti-vascular endothelial growth factor compounds may be beneficial in eyes with DME.Key Words: Ranibizumab, Bevacizumab, Diabetic macular edema, Treatment failure     

Vascular endothelial growth factor plasma levels before and after treatment of neovascular age‐related macular degeneration with bevacizumab or ranibizumab

Purpose:  To evaluate the changes of vascular endothelial growth factor (VEGF) plasma levels after intravitreal injections of ranibizumab or bevacizumab in patients with exudative age‐related macular degeneration (AMD). Methods:  Forty‐three patients with exudative AMD and 19 age‐ and sex‐matched control patients without chorioretinal diseases were studied. Nineteen patients were treated with intravitreal ranibizumab 0.5 mg, 24 with intravitreal bevacizumab 1.25 mg. Blood samples were collected just before the first injection, and 28 days after three initial consecutive injections performed in 4‐weekly intervals (loading dose). Concentration of VEGF in the plasma was measured by ELISA. Results:  At baseline, the median VEGF concentrations in controls were 180.97 pg/ml, in the bevacizumab group 189.72 pg/ml and in the ranibizumab group 191.36 pg/ml. VEGF plasma concentrations in patients with wet AMD were comparable to controls (p = 0.225). Twenty‐eight days after the third injection, a significant reduction of 42% in the median VEGF plasma levels was found in bevacizumab‐treated patients (109.97 pg/ml; p = 0.0002) but not in ranibizumab‐treated patients (189.97 pg/ml; p = 0.198) where a reduction of 0.7% in the median value was found. Conclusions:  Intravitreal bevacizumab significantly reduced VEGF plasma levels until 28 days after intravitreal injection in patients with exudative AMD. Ranibizumab did not achieve a significant plasma VEGF reduction at the same time‐point. These findings alert to the potential systemic safety differences between the two drugs after intravitreal administration.     

Twelve-month safety of intravitreal injections of bevacizumab (Avastin®): results of the Pan-American Collaborative Retina Study Group (PACORES)

Background Vascular endothelial growth factor (VEGF) plays an important role in many diseases of the posterior pole that are characterized by macular edema and/or intraocular neovascularization. Recently anti-VEGF agents such as ranibizumab and pegaptanib sodium have been shown to be beneficial in the treatment of choroidal neovascularization (CNV) secondary to age-related macular degeneration (ARMD). However in most parts of the world, both pegaptanib sodium and ranibizumab are not readily available. Bevacizumab, a humanized recombinant monoclonal IgG antibody that binds and inhibits all VEGF isoforms, has been proposed as an alternative treatment option. Methods A total of 1,265 consecutive patients were injected with bevacizumab for diseases such as proliferative diabetic retinopathy, diabetic macular edema, retinal vein occlusions, and CNV of several etiologies including ARMD at eight Latin American institutions from 1 September 2005 to 31 January 2006. Of these 1,265, 92 were excluded because they were injected once and lost to follow-up. The remaining 1,173 patients constitute the subjects of this retrospective, multicenter, open label, uncontrolled interventional case series that reports the cumulative systemic and ocular adverse events following intravitreal bevacizumab during 12 months of follow-up. Patients were examined at baseline and then monthly. If the patients were unable to attend the 12-month visit, a telephone interview was conducted to assess for possible systemic complications. Results A total of 4,303 intravitreal injections of bevacizumab on 1,310 eyes was reported. All 1,173 patients were accounted for at the 12-month visit. Systemic adverse events were reported in 18 (1.5%) patients. These included seven (0.59%) cases of an acute elevation of systemic blood pressure, six (0.5%) cerebrovascular accidents, five (0.4%) myocardial infarctions, two (0.17%) iliac artery aneurysms, two (0.17%) toe amputations and five (0.4%) deaths. Ocular complications included seven (0.16%) bacterial endophthalmitis, seven (0.16%) tractional retinal detachments, four (0.09%) uveitis, and a case (0.02%) each of rhegmatogenous retinal detachment and vitreous hemorrhage. Conclusion Despite the limited follow-up, repeated intravitreal injections of either 1.25 mg or 2.5 mg of bevacizumab appears to be safe and well tolerated during the 1st year. Presented in part at the combined Club Jules Gonin-Retina Society Meeting (October 2006, Cape Town, South Africa). For a complete listing of participating members of PACORES see the Appendix. The authors have no financial interests in the subject matter presented.     

Induction of vascular endothelial growth factor receptor expression in human umbilical vein endothelial cells after repeated bevacizumab treatment in vitro

AIM: To investigate the mechanism underlying the loss of responsiveness to anti-vascular endothelial growth factor (VEGF) treatment after repeated injections for choroidal neovascularization, VEGF and VEGF receptor (VEGFR) expressions were evaluated following repeated bevacizumab treatments in hypoxic human umbilical vein endothelial cells (HUVECs) in vitro. METHODS: HUVECs were incubated under hypoxic conditions in two media of different bevacizumab concentrations (1.0 or 2.5 mg/mL) for 17h, and then in a new medium without bevacizumab for 7h. This procedure was repeated twice more. A culture with an identical volume of excipients served as the control. Cytotoxicity and cell proliferation were assessed using 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide and Ki-67 assays, respectively. Levels of VEGF and VEGFR were assessed using enzyme-linked immunosorbent assay and Western blot respectively. RESULTS: Cytotoxic effects were not reported for either bevacizumab concentration. Cell proliferation was not reduced after anti-VEGF treatments. VEGF level after single treatment was significantly higher than that of the control and after repeated treatments. Phosphorylated VEGFR-2 expression increased significantly after single and repeated bevacizumab treatments compared with the control. The 1.0 mg/mL bevacizumab induced significantly higher expressions of VEGFR-2 than the 2.5 mg/mL in single and repeated treatment groups. CONCLUSION: Bevacizumab treatment of HUVECs elevated VEGFR expression in both single and repeated treatments, indicating a mechanism for the reduced efficacy of anti-VEGF therapy in ocular neovascular disorders.     

Choroidal endothelial cells transmigrate across the retinal pigment epithelium but do not proliferate in response to soluble vascular endothelial growth factor

The purpose of this study was to investigate the effects of soluble VEGF on human choroidal endothelial cell (CEC) transmigration across an RPE monolayer as it relates to choroidal neovascularization in AMD. In coculture assays, ARPE-19 (ARPE) was plated on the undersides of Transwell inserts having 0.4 microm pores. Primary human CECs were then plated into the insert. CECs in the Transwell inserts were counted after 72 hr of growth. CEC proliferation was also measured after culturing CECs in ARPE-CEC coculture-conditioned media or in media with exogenous VEGF121 and/or VEGF165 added. Transmigration assays were performed on Transwells with 8.0 microm pores: green-labelled CECs were plated in Transwell inserts with or without red-labelled ARPE plated on the undersides of the insert. In some transmigration assays, ARPE was plated into the wells to provide a chemotactic gradient for CEC transmigration. After 72 hr CECs were plated, green cells were counted either within the well media as CECs that transmigrated the epithelial monolayer, or on the underside of the insert as CECs that transmigrated the Transwell insert to but not beyond the ARPE monolayer. A neutralizing antibody to VEGF was added to the wells of Transwells at the time the CECs were plated in the insert and transmigrated CECs were counted. VEGF protein was measured in the conditioned media of ARPE and CEC coculture and in transmigration assays. Compared to control, CEC proliferation significantly increased when CECs were cultured in coculture conditioned media (p=0.001) or in coculture assays (p<0.001). However, there was no effect on CEC proliferation when VEGF121, VEGF165, or both were added to solo CECs. Antibody to VEGF did not reduce the proliferative effects of coculture conditioned media on CEC. ARPE plated in the well significantly increased CEC transmigration (p<0.001) compared to transmigration assays without ARPE in the well. VEGF protein measured in the well media of transmigration assays having ARPE within the well was significantly greater than in the assays without ARPE within the well (p<0.004). Exogenous neutralizing antibody to VEGF significantly reduced transmigration, and this effect was dose-dependent. VEGF provides a chemotactic gradient for human CECs to transmigrate across a monolayer of ARPE. Neutralization of VEGF in the media partially reduces transmigration. Whereas soluble VEGF does not increase proliferation of solo CECs, coculture conditioned media enhances proliferation, suggesting that growth factors other than VEGF cause CEC proliferation. These findings may have relevance to the transformation of occult CNV into CNV within the neurosensory retina in AMD.     

从贝伐单抗应用的窘境议医疗管理政策的瓶颈

贝伐单抗（Avastin）和雷珠单抗（Lucentis）是同一公司研发的有效的抗血管内皮生长因子制剂,前者被用于治疗肿瘤,后者被用于治疗湿性年龄相关性黄斑变性（AMD）,但前者价格低于后者数十倍。近年的研究表明,两者在治疗AMD等新生血管性眼病的效果与安全性相似,眼科医师试图用其治疗新生血管性眼病,但有＂超适应证用药＂的困惑及风险。本文介绍了国外眼科界对＂超适应证用药＂的观点、国内执业医师法的相关规定及世界银行建议中国基本药物的仿制药战略。期望人们认识到,医师与患者在医疗过程中有着共同的立场、愿望及风险。     

Bevacizumab(Avastin)抑制角膜新生血管的应用新进展

诱发角膜新生血管的因素涉及各种生长因子。研究表明:在角膜新生血管中广泛存在的血管内皮生长因子(vascular endothelial growth factor,VEGF)起着主要作用。一种可行的治疗角膜新生血管策略是:通过特异性的中和抗VEGF抗体竞争性的结合VEGF,从而抑制VEGF活性。近年来,利用抗VEGF治疗策略,抑制脉络膜新生血管获得了很好的效果。靶向VEGF治疗药物的疗效和安全性已经证明。因此我们设想,局部应用新的抗VEGF药物,如贝伐单抗、兰尼单抗等可以有效地抑制角膜新生血管,恢复角膜透明和视力。     

Pegaptanib and ranibizumab for neovascular age-related macular degeneration: a systematic review

AIMS: To assess the clinical effectiveness of pegaptanib sodium and ranibizumab for neovascular age-related macular degeneration (AMD). METHODS: A systematic review of randomised controlled trials (RCTs) identified through searching 12 electronic databases, bibliographies and consultation with experts and manufacturers. RCTs were eligible if they assessed the effects of pegaptanib or ranibizumab with best supportive care, sham injection or photodynamic therapy (PDT) on patients with subfoveal choroidal neovascularisation associated with wet AMD and examined outcomes including visual acuity and adverse events. RESULTS: Three RCTs of ranibizumab (MARINA, ANCHOR, FOCUS) and two of pegaptanib (VISION study) met the inclusion criteria. The RCTs included patients with different lesion types. The studies showed statistically significant benefit on different measures of visual acuity for patients receiving pegaptanib, ranibizumab or ranibizumab with PDT compared to control (sham injection, PDT or sham injection with PDT) after 12 months. These differences appeared to be clinically significant. Although adverse events were common among those receiving pegaptanib or ranibizumab, they were considered mild to moderate in nature. Meta-analysis of ranibizumab trials and indirect comparison of the two drugs were not possible due to differences in the study populations' lesion types. However, results from the RCTs of ranibizumab tended to show a greater effect on visual acuity than results from the RCT of pegaptanib. CONCLUSIONS: Pegaptanib and ranibizumab appear to slow or stop the progression of neovascular AMD. Uncertainty remains over the relative benefits of pegaptanib compared with ranibizumab and other unlicensed drugs (eg, Avastin), due to the nature of the evidence. Head-to-head RCTs and economic evaluations comparing these alternatives are needed.     

Bevacizumab (Avastin) eye drops inhibit corneal neovascularization

Background To analyze the ability of bevacizumab (Avastin) eye drops to inhibit corneal neovascularization. Design: interventional case series involving five patients (age: 42 ± 14 years). Methods Patients with aggressive corneal neovascularisation not responding to conventional therapy were treated with bevacizumab (Avastin) eye drops (5x/day; 5 mg/ml) for 0.5 to 6 months (mean: 3.6 ± 2; four patients with limbal stem cell deficiency [three due to chemical burns and one inherited] and one after perforating keratoplasty). Results Bevacizumab eye drops were well tolerated without obvious corneal side-effects. All five patients showed a reduction in the neovascularized area (decrease 48 ± 28%; 13–75%). Conclusions Bevacizumab eye drops seem to inhibit corneal neovascularization without obvious corneal epithelial side-effects.     

Anti-VEGF therapy: comparison of current and future agents

With the identification of vascular endothelial growth factor (VEGF) and the confirmation of its pathophysiologic link to retinal and choroidal angiogenesis, numerous agents have been designed to inhibit its activity. It is noteworthy that anatomic and visual benefits have been associated with the use of anti-VEGF agents such as pegaptanib (Macugen) and to a greater extent, ranibizumab (Lucentis) and bevacizumab (Avastin), particularly in the management of neovascular age-related macular degeneration (AMD). Clinical trials and case series have confirmed the utility of these agents. However, shortcomings of the current drugs such as short half-life, intraocular dosing, limited effectiveness in some patients, and potential systemic side effects continue to drive the development of new agents. In this article, we review current anti-VEGF therapies and discuss future developments.     

“Balancing risk in ophthalmic prescribing: assessing the safety of anti-VEGF therapies and the risks associated with unlicensed medicines”

Vascular endothelial growth factor (VEGF) inhibitor medications such as ranibizumab, pegaptanib and bevacizumab are in use for the treatment of neovascular age-related macular degeneration (AMD) and other retinal conditions, although only ranibizumab and pegaptanib are approved for these conditions. In contrast, bevacizumab was developed for the intravenous systemic treatment of colorectal cancer and is not formulated for intravitreal use, but is commonly used off-label in ophthalmology. European Union legislation permits the use of drugs outside the terms of their licence (‘off-label’) only under certain circumstances, such as during clinical trials, compassionate/named patient use in the absence of a licensed alternative, emergency scenarios (e.g., pandemics) or at the discretion of a treating physician. In such cases, patients should be fully informed regarding their treatment and any potential risks involved. Off-label drug use can be an important tool to provide patients with treatment in cases of unmet medical need. However, the use of an unlicensed medicinal product, when a suitable licensed alternative is available, puts prescribing physicians at risk of liability if safety issues arise. Emerging clinical evidence suggests safety differences exist between ranibizumab and bevacizumab.