For nearly 20 years, neovascular age-related macular degeneration (AMD) has been getting all the attention owing to the introduction of intravitreally administered anti-vascular endothelial growth factor (anti-VEGF) drugs. These agents have successfully stopped disease progression and saved vision in many affected patients.

However, focus has been shifting to the overwhelmingly large population of patients who have the dry, or non-exudative, form of AMD, the insidious type that does not involve the growth of abnormal blood vessels but erodes vision over time. This point was made by Dr Rishi P. Singh, president of Cleveland Clinic Martin North and South hospitals in Stuart, Florida, United States, and professor of ophthalmology at Cleveland Clinic Lerner College of Medicine in Ohio, US.

The end stage of dry AMD is known as geographic atrophy (GA). At this stage of the disease, the activities of daily living—reading, driving, facial recognition and so on—are severely affected, leading to overall serious deterioration of the patient’s condition and quality of life.

 have provided evidence that intravitreal injection of certain inhibitors can cause regression or slowing of vision loss over time in people with GA. “This is how clinicians are currently attempting to treat GA,” said Dr Singh. Findings from these studies have shown advancements in the ability to prevent progression of GA by 15–25%. In the case of GA outside the fovea, that ability increased to 35% — “a good step in the right direction,” he stated.

Dr Singh pointed out that previously, physicians had not looked at GA as closely as they could have, mainly because treatments are only now on the horizon. Optical coherence tomography (OCT) is the main imaging modality, and the emphasis in these images is on identifying exudative AMD and its characteristic fluid or haemorrhages during clinical evaluations. “GA requires reappraisal of these diagnostic techniques to ascertain that the patients with the most need are identified for treatment through the clinical trials,” Dr Singh said.

Dr Singh explained that in routine clinical practice, a few factors come into play, and he acknowledged the need to do better when evaluating and treating patients with neovascular AMD. For example, the number of treatments prescribed is substantially lower in clinical practice than in clinical trials. Additionally, fluorescein angiography is not carried out routinely in clinical practice as it is in clinical trials, although this test may not always be necessary in the clinical setting.

In studies of GA, numerous tests are performed, such as microperimetry and low-luminance vision testing, that are also not routinely performed in clinics; Dr Singh noted that the added benefits in the clinical setting are questionable. In clinical trials, these tests quantify the benefit of the drugs under evaluation for treating GA.

“If I want a very precise measurement of the clinical effect of a drug, tools such as low-luminance vision and microperimetry are invaluable to determine the treatment effect over time,” Dr Singh explained. Although these specific tests are not used often by ophthalmologists, fundus autofluorescence and OCT are used to determine the size and spread of the GA as patients are followed.

Regarding treatments, the effects of the drugs being evaluated are individualised to each patient. “Each patient dictates the clinical suspicions based on how much the disease has progressed and [whether] further intervention is needed,” Dr Singh said.

He noted that he is very pragmatic in these cases when ordering testing. “I use an evidence-based treatment approach, in that if I order a particular test or additional imaging, it is going to help me make useful decisions about a particular patient’s treatment plan,” he explained. This more thoughtful approach will ease the burden in the clinic.

There are several ongoing clinical trials for GA candidates, all of which attack the complement cascade, C3 and C5, both of which are in the alternative complement cascade. Pegcetacoplan (Empaveli, Apellis Pharmaceuticals), a targeted C3 therapy, was recently fast-tracked by the US Food and Drug Administration (FDA) based on the results of three studies: the OAKS, DERBY and FILLY trials.

The drug successfully slowed the progression of GA. “Pegcetacoplan showed significant benefit in its Phase 3 trial and it has a relatively good safety profile. [However], some patients converted to neovascular AMD as a result of treatment with pegcetacoplan, which we need to learn more about,” Dr Singh observed.

Avacincaptad pegol (Zimura, Iveric Bio), a C5 inhibitor, is now being evaluated in a Phase 3 study. The results of the GATHER1 study were similar to those of the OAKS and DERBYstudies. GATHER2,4 which compared monthly avacincaptad treatment with sham treatment, will report results during autumn 2022. “These findings will determine how we will use the drug in clinical practice,” Dr Singh stated.

Dr Singh believes the future of these therapies is very bright, especially concerning the approximately 50% of patients who require the assistance of carers to travel to appointments and other activities. Considering this patient burden, a reduction in disease progression would be a major step forward for patients and carers.

Data from the current studies clearly indicate that there is a great deal of hope for beneficial future therapies for this patient population. Dr Singh recommended that patients under the care of a retina specialist or ophthalmologist should determine whether they are candidates for treatment with these drugs when they become clinically available.

Patients should recognise the importance of family history and lifestyle in the development of GA, both of which are strong predictors of disease development. Smoking, a diet high in saturated fat and not eating fish or green leafy vegetables are key elements in GA development. A change in habits is a good step towards possibly preventing the disease.

Dr Singh is hoping that future versions of drugs for GA could involve sustained drug delivery, eliminating the need for monthly or every-other-month treatments, and combination therapies.

Dr Singh serves as a consultant to Allergan, Bausch + Lomb, Genentech, Regeneron Pharmaceuticals, Novartis, AsclepiX Therapeutics, Gyroscope Therapeutics and Apellis Pharmaceuticals.

1. Liao DS, Grossi FV, El Mehdi D, et al. Complement C3 inhibitor pegcetacoplan for geographic atrophy secondary to age-related macular degeneration: a randomized phase 2 trial. 

2. Apellis announces top-line results from phase 3 DERBY and OAKS studies in geographic atrophy (GA) and plans to submit NDA to FDA in the first half of 2022. Press release. Apellis Pharmaceuticals, Inc. 9 Sep 2021. Accessed 4 Oct 2022. 

investors.apellis.com/news-releases/news-release-details/apellis-announces-top-line-results-phase-3-derby-and-oaks

3. Jaffe GJ, Westby K, Csaky KG, et al. C5 inhibitor avacincaptad pegol for geographic atrophy due to age-related macular degeneration: a randomized pivotal phase 2/3 trial. 

4. Iveric Bio completes patient enrollment of GATHER2 pivotal clinical trial of Zimura ahead of schedule. Press release. Iveric bio, Inc. 26 Jul 2021. Accessed 4 Oct 2022. 

investors.ivericbio.com/news-releases/news-release-details/iveric-bio-completes-patient-enrollment-gather2-pivotal-clinical

Articles

Giving patients access to new treatments and information on changing their lifestyle habits could result in major steps towards minimising or even preventing geographic atrophy.

Spotlight on GA: A bright future with potential treatments

Patient expectations for cataract surgery are very high, especially when it comes to advanced-technology implants. To meet those expectations, we need accurate preoperative measurements as a basis for IOL calculations, and we cannot have accurate measurements without a stable tear film.

Objective tests such as tear osmolarity and matrix metalloproteinase 9 (MMP-9) offer efficient ways to screen the tear film and, in turn, may act as predictors of whether preoperative measurements will be accurate. Studies show that cataract patients with abnormal tear film osmolarity have highly variable keratometric readings (up to nearly 3.5 D, taken 2 weeks apart).

With treatment and stabilisation of the tear film, more accurate measurements are captured, reducing residual refractive error.

 Here is a look at how I screen my cataract patients, aggressively treat them before surgery and initiate long-term therapy postoperatively for chronic dry eye disease.

Given the prevalence of dry eye disease in the cataract age group, we have to look at every cataract patient as a dry eye consultation. This is true even when patients are asymptomatic or do not recognise mild symptoms of dry eye disease.

Patients might have intermittent blurred vision that they blame on the cataracts or fatigue, when in fact the problem is tear film instability. If we do not treat the condition before surgery, dry eye symptoms often worsen from the stress of surgery, at which point patients blame their surgeon for “giving them dry eye”.

Objective tests make dry eye screening accurate and efficient. I use an osmolarity test (TearLab Osmolarity System, TearLab Corporation), in which a score above 308 mOsm/L and/or an interocular difference of 8 mOsm/L or higher indicates an abnormal tear film. When the two eyes exhibit different numbers, the diagnosis and treatment decisions are based on the higher number.

In addition to osmolarity testing, I also perform meibography (TearScience LipiView, Johnson & Johnson Vision) to show patients their missing or architecturally abnormal gland structures. Another point-of-care test that our technicians perform is an MMP-9 test for inflammation (InflammaDry, Quidel Corporation). If this test shows a stripe ranging from faint pink to bright red, inflammation is present. We need to treat the inflammation before cataract surgery.

Finally, I educate patients about dry eye and their diagnostic test results. It is so important to make believers of them and engage them in their own treatment process before surgery.

I give patients a card with their osmolarity number and a scale on the back with the colour-coded interpretation of their number. I explain to them, “We have to tune your ocular surface to get the best results from cataract surgery,” and typically they are highly motivated to adhere to their customised treatment for the next few weeks.

When patients are ready for cataract surgery, they do not want to wait months for their dry eye treatment to work, but they do not mind waiting a few weeks to ensure a better outcome. I treat the tear film aggressively and schedule patients to return this for their measurements in about 3 weeks, depending on disease severity. I explain that they must be adherent, or I will not be able to acquire reliable data for surgery.

I start patients on a topical corticosteroid for 2 weeks to quell the inflammation (either Alcon’s Eysuvis or loteprednol if there are insurance coverage issues for Eysuvis). Patients start oral omega supplements, sometimes starting with a liquid formula before transitioning to capsules long term.

If meibography shows impacted meibomian glands, I perform thermal pulsation (TearScience LipiFlow, Johnson & Johnson Vision) to evacuate the inspissated meibum and allow the ingested omega fatty acids to filter through the cleaned meibomian gland orifices onto the ocular surface. If patients have blepharitis, I recommend microblepharoexfoliation of the lid margins (BlephEx, Alcon) to debulk the debris, break up the biofilm, and minimise the risk of infection and endophthalmitis.

With the tear film stabilised, as evidenced by an osmolarity score below 308 mOsm/L and an absent red line on the MMP-9 test, I am ready to take measurements, calculate the implant powers and proceed with surgery.

Post-op: Transition to long-term treatments

Ocular surface disease is chronic; patients will need to use customised therapies long term. I explain to patients that to get the most from their new implants, they need to keep their tear film healthy. Postoperatively, patients continue with their omegas, a heated moisture mask, lid scrubs and an annual thermal and evacuation treatment if warranted.

For patients with meibomian gland dysfunction, I often perform intense pulsed-light therapy (Lumenis OptiLight or ESW Vision’s E-Eye). This four-session treatment is scheduled every 2–3 weeks and can then be repeated as a single treatment semi-annually for maintenance. Most patients perceive the benefits, with decreased lid margin hyperaemia and increased ocular comfort, after the sessions are complete.

Additionally, for many patients’ long-term care, I prescribe ciclosporin (Allergan’s Restasis or Sun Pharmaceutical Industries’ Cequa), or lifitegrast ophthalmic solution (Xiidra, Novartis Pharmaceuticals Corporation) 5% eye drops, or varenicline solution (Tyrvaya, Oyster Point Pharma) nasal spray 0.03%. Each patient’s treatment needs must be customised and carefully monitored.

When we stabilise the tear film, refractive surprises become less frequent. Moreover, patients do not suddenly experience dry eye symptoms postoperatively and blame their surgeon.

For the process to work, we have to ensure adherence by engaging patients in their surgical journey. I find this empowers them. They like tracking their osmolarity number. In addition, by starting a dry eye care routine leading up to surgery, patients are well prepared to continue their maintenance therapy after surgery.

Dr Matossianspecialises in refractive cataract surgery and dry eye disease.Sheis the founder of Matossian Eye Associates, located in Pennsylvania and New Jersey, United States.

1. Epitropoulos AT, Matossian C, Berdy GJ, et al. Effect of tear osmolarity on repeatability of keratometry for cataract surgery planning. 

2. Matossian C. Impact of thermal pulsation treatment on astigmatism management and outcomes in meibomian gland dysfunction patients undergoing cataract surgery. 

Correcting abnormal tear film osmolarity before cataract surgery enables more accurate measurements to be captured, reducing residual refractive error.

Cataract predictability: Screen and treat unstable tear film in weeks

The prevalence of myopia is increasing worldwide: in 2050, half of the world’s population is expected to be myopic, with the estimated prevalence highest for Asian regions.

 The proportion of the population with the condition is also increasing in Europe,

Uncorrected myopia does not only cause far objects to appear blurry; with increasing severity of the disorder, the risk for serious eye diseases increases. For example, retinal detachment, choroidal neovascularisation and glaucoma are linked to severe myopia and can cause irreversible blindness.

Myopia is caused by a mismatch of refractive power and axial length of the eye, so that the focal point of the eye lies in front of the retina, creating a blurry image. This mismatch usually develops (and increases) due to excessive axial growth of the eyeball during childhood. Over recent years, several therapies to inhibit myopia progression at this critical age have been developed.

The long-established therapy: Low-dose atropine

The most popular therapy, especially in Asia, is the daily application of atropine eye drops ranging from 0.01% to 0.05%.

 Owing to the higher numbers of myopic children in Asian countries, most of the randomised and placebo-controlled studies of low-dose atropine were performed with Asian children, with only a handful of studies including European children.

The key message concerning atropine therapy is that the higher the dose, the more myopia progression is inhibited. After 3 years of daily application of 0.05%, 0.025% or 0.01% atropine eye drops, children showed mean myopia progressions of −0.73 D, −1.31 D and −1.60 D, respectively. Accordingly, mean axial length increases of 0.50 mm, 0.74 mm and 0.89 mm were observed after 3 years.

Atropine eye drops at 0.01% may not show beneficial effects in every case, so increasing the dose should be considered.8 However, the higher the dose, the stronger the possible side effects. Therefore, one needs to carefully evaluate the highest tolerable dose for each individual case.

 Spectacle lenses with defocus incorporated multiple segments (DIMS) technology. The defocusing segments are only visible in the shadow.

In the Netherlands, drops of 0.5% and 1% are applied to children with progressive myopia. This shows great success in terms of inhibiting myopia progression but also causes strong side effects, such as disturbances of accommodation or photosensitivity, which need to be tackled with photochromic multifocal spectacles.

Apart from pharmacological intervention with atropine, there are also optical therapies with specially designed contact or spectacle lenses. These are usually based on the theory of myopic defocus, in which focus in front of the retina is created in order to stop/inhibit further axial length growth. This is realised by adding a second image shell focused in front of the retina to the primary image shell focused on the fovea, or by ‘bending’ the image shell so that it lies centrally on the fovea but in front of the retina in the periphery.

In addition to the central zone with normal correction, multifocal contact lenses have a second focus with positive refractive power, which creates a second image shell in front of the retina. Here we can see an addition-dependent effect: higher addition leads to less myopia progression and axial length growth.

After 3 years of contact lens wear with high addition (+2.50 D) or medium addition (+1.50 D), children showed mean myopia progression of −0.60 D and −0.89 D, respectively, compared with −1.05 D in the group with monofocal contact lenses. Mean axial length growth was 0.42 mm, 0.58 mm and 0.66 mm, respectively, for the groups with high addition, medium addition and monofocal contact lenses.

Orthokeratology lenses follow the same optical principle: by flattening central corneal radii to reduce corneal refractive power and thus correct myopia, the mid-peripheral corneal radii are steepened. Therefore, the refractive power mid-peripheral cornea increases, and a second image shell is created in front of the retina in the midperiphery.

Orthokeratology lenses are worn at night; during sleep, the cornea is modulated so that daytime vision is possible without any optical correction. Unlike refractive surgery, this process is reversible. A detailed review concluded that orthokeratology lenses are effective in inhibiting myopia progression,

 but high compliance is needed to not only inhibit myopia progression but also achieve the desired effect of myopia correction during the day.

Other soft contact lenses specifically designed to inhibit myopia progression showed a duration-dependent effect. The treatment effect correlated positively with daily wearing time: the longer the daily wearing time, the less myopia progression in terms of increasing refractive error.

All types of contact lenses place special demands in terms of handling the lenses, hygiene and a care routine, which must be considered when contact lenses are recommended for children.

The newcomer: Spectacle lenses with multisegments

Like contact lenses, specially designed spectacle lenses offer optical correction and therapy of myopia at the same time. However, spectacle lenses have additional advantages over contact lenses: they do not require special handling and they can be worn all day, every day, which has shown a positive effect in terms of myopia progression.

Although spectacle lenses and contact lenses both use the principle of myopic defocus, its implementation is fundamentally different. Because spectacle lenses have a higher vertex distance than contact lenses, they offer the possibility to implement the defocus, which should primarily act on the periphery of the retina, outside the central zone of the lens.

Therefore, with spectacle lenses, when looking straight ahead through the defocus-free central zone, the defocus acts only in the periphery, not on the fovea, and the patient has clear vision. The situation is different with contact lenses: because they sit directly on the cornea, the defocus is always present.

Most promising in terms of inhibition of myopia progression and tolerability appear to be spectacle lenses with numerous tiny lenses, with positive refractive power that are incorporated in the mid-periphery of the front surface and thus create a second image shell in front of the peripheral retina. The tiny additional lenses are hardly visible; looking at someone wearing spectacles with these, one cannot tell that they are special lenses with a therapeutic function.

After 2 years wearing spectacle lenses with highly aspherical (HAL) and slightly aspherical lenslets (SAL), children experienced average myopia progression of −0.66 D and −1.04 D, respectively, and average axial length growth of 0.34 mm and 0.51 mm, which, again, shows the addition-dependent effect of optical therapies on inhibiting myopia progression.

Children wearing spectacle lenses with defocus incorporated multiple segments (DIMS) technology (see 

) experienced an average myopia progression after 3 years of −0.52 D and an average axial length growth of about 0.31 mm.

 Owing to the different implementation of myopic defocus, these spectacle lenses provide better visual acuity and contrast sensitivity in primary gaze than multifocal contact lenses.

Example axial length growth of patient's right (blue) and left (yellow) eye in comparison with the underlying age-dependent physiological axial length growth. Tolerance range includes measurement inaccuracies. 

Abbreviations: f, female; m, male. (Images courtesy of Dr Hakan Kaymak)

All myopia therapies should be carried out over many years because the critical phase of axial eye growth extends from childhood until adolescence.

 Control examination should take place on a regular basis, at least every year. In addition to the standard determination of refraction and, if necessary, adjustment of the optical correction, it is essential to measure axial length at each examination with high accuracy.

Axial length growth is a more sensitive means of assessing myopia progression than looking at the actual refraction change. By measuring axial length, myopia progression can be detected at an earlier stage, even before it would be reflected in a change in refractive state (see 

As the eye grows from birth to adulthood, not all axial length growth detected can be attributed to progressive myopia: only axial length growth that is excessive over the age-dependent physiological growth of the eye leads to myopia. As a specific goal for any myopia therapy, it is proposed to focus on the underlying physiological axial length growth versus the axial length growth in excess thereof that brings about the progressive myopia.

Dr Kaymak is head surgeon with a focus on retina, macula and vitreous body at Breyer, Kaymak & Klabe, Duesseldorf, which is one of the largest private ophthalmology practices in Germany. His personal interest is myopia therapy. Currently, he receives consulting fees from Hoya Lens Deutschland GmbH.

1. Holden BA, Fricke TR, Wilson DA, et al. Global prevalence of myopia and high myopia and temporal trends from 2000 through 2050. 

2. Williams KM, Bertelsen G, Cumberland P, et al. Increasing prevalence of myopia in Europe and the impact of education.

3. Williams KM, Verhoeven VJM, Cumberland P, et al. Prevalence of refractive error in Europe: the European Eye Epidemiology (E(3)) Consortium. 

4. Saw S-M, Gazzard G, Shih-Yen EC, Chua W-H. Myopia and associated pathological complications. 

5. Flitcroft DI. The complex interactions of retinal, optical and environmental factors in myopia aetiology.

6. Wong TY, Ferreira A, Hughes R, et al. Epidemiology and disease burden of pathologic myopia and myopic choroidal neovascularization: an evidence-based systematic review. 

7. Yam JC, Li FF, Zhang X, et al. Two-year clinical trial of the low-concentration atropine for myopia progression (LAMP) study: Phase 2 report. 

8. Kaymak H, Graff B, Schaeffel F, et al. A retrospective analysis of the therapeutic effects of 0.01% atropine on axial length growth in children in a real-life clinical setting. 

9. Sacchi M, Serafino M, Villani E, et al. Efficacy of atropine 0.01% for the treatment of childhood myopia in European patients. 

10. Joachimsen L, Böhringer D, Gross NJ, et al. A pilot study on the efficacy and safety of 0.01% atropine in German schoolchildren with progressive myopia.

11. Yam JC, Zhang XJ, Zhang Y, et al. Three-year clinical trial of low-concentration atropine for myopia progression (LAMP) study: Continued versus washout: Phase 3 report. 

12. Polling JR, Tan E, Driessen S, et al. A 3-year follow-up study of atropine treatment for progressive myopia in Europeans. 

13. Walline JJ, Walker MK, Mutti DO, et al. Effect of high add power, medium add power, or single-vision contact lenses on myopia progression in children: The BLINK randomized clinical trial. 

14. Cho P, Tan Q. Myopia and orthokeratology for myopia control. 

15. Lam CSY, Tang WC, Tse DY-Y, et al. Defocus incorporated soft contact (DISC) lens slows myopia progression in Hong Kong Chinese schoolchildren: a 2-year randomised clinical trial. 

16. Bao J, Huang Y, Li X, et al. Spectacle lenses with aspherical lenslets for myopia control vs single-vision spectacle lenses: a randomized clinical trial. 

17. Kaymak H, Neller K, Schütz S, et al. Vision tests on spectacle lenses and contact lenses for optical myopia correction: a pilot study. 

18. Lam CS, Tang WC, Lee PH, et al. Myopia control effect of defocus incorporated multiple segments (DIMS) spectacle lens in Chinese children: results of a 3-year follow-up study. 

19. Truckenbrod C, Meigen C, Brandt M, et al. Longitudinal analysis of axial length growth in a German cohort of healthy children and adolescents. 

20. Kaymak H, Graff B, Neller K, et al. Myopietherapie und Prophylaxe mit “Defocus Incorporated Multiple Segments”-Brillengläsern. 

21. Chamberlain P, La Lazon de Jara P, Arumugam B, Bullimore MA. Axial length targets for myopia control. 

Axial length growth is a sensitive means of assessing myopia progression in children. Drops and special lenses are options for inhibiting excessive axial length growth.

Controlling the growing problem of myopia in children

Low-grade inflammation seems to be associated with baseline disease in inherited retinal dystrophies (IRDs), according to Dr Christine N. Kay, a surgeon at Vitreoretinal Associates, in Gainesville, Florida, United States. It can be argued that patients with IRDs may be predisposed to inflammation. Adeno-associated virus (AAV)-based gene therapy in these patients may trigger severe inflammatory responses that can lead to permanent visual loss in some.

Examples of diseases include zonular instability in cataract surgeries resulting in IOL dislocations in retinitis pigmentosa (RP), presence of cells in the anterior vitreous, cystoid macular oedema in RP and retinal antibodies in serum samples from patients with RP.

A murine study found that inflammation preceded retinal degeneration in 

 Dr Kay also cited Phase 1/2 trials of gene therapy for 

-associated Leber congenital amaurosis, choroideremia and 

-associated X-linked RP, in which ocular inflammation was dose dependent, with patients who received higher doses at higher risk for vitritis, retinitis and choroiditis.

“At … best … gene therapies treat disease and restore/rescue function,” Dr Kay explained. “At worst … [they introduce] a new antigen and create a new problem.”

Among the numerous factors that stimulate inflammation are delivery mode; vector/capsid manufacturing; promoters and transgene expression; surgical considerations; host characteristics; and dose (the most clinically relevant inflammation occurs with doses of 1 x 1011 vg/eyeor higher).

The immune response varies according to the mode of administration (subretinal versus intravitreal). Most trials today evaluate vectors injected subretinally, whereas suprachoroidal injections are used less frequently.

“Intravitreal delivery … is [generally] more inflammatory than subretinal delivery and can lead to [increases in] aqueous vector levels, systemic biodistribution to spleen/lymph nodes, [and] systemic dissemination from … anterior chamber [outflow],” Dr Kay said. This delivery, in turn, can trigger more “neutralising antibody responses, which can interfere with effective transgene expression and lead to greater humoral responses and … reduced therapeutic efficacy.”

Regarding the vector capsid serotype, baseline immunity can be species-dependent in animal testing. Dose-dependent increases in binding and neutralising antibodies after AAV injection and in increased serum levels of binding antibodies appear to be correlated with inflammation. However, injection dose is the most important factor affecting increased serum antibody level.

In terms of the roles that vector design and manufacturing play in inflammation, we need to know more about the effects of the production platform (triple transfection in HEK293 cells and herpes- and baculovirus-based systems), of the empty capsid ratio (which correlates with increased transduction efficiency) and of process residuals and contaminants. (Manufacturing of AAV vectors produces three types of capsid—empty, partial and full; the ratio pinpoints the number of empty capsids.) Nevertheless, as previously noted, it is vector dose that correlates most closely with the inflammatory response.

As another study reports, strong correlation has been observed between AAV toxicity and 

 Ubiquitous promoters (retinal pigment epithelial promoters) were more toxic than photoreceptor-specific promoters; the reason may be that ubiquitous promoters drive higher levels of transgene expression in more cell types.

Surgical delivery of AAV also plays a role in inflammation. Pro-inflammatory factors of surgery include variability in bleb size, location and dilution with intraocular irrigation (BSS); surgery-related adverse events (particularly haemorrhage); operative duration; and vitreous egress of the vector.

Finally, host (recipient) characteristics include patient age, pigmentation (of the iris), immune system hyperactivity, human leukocyte antigen type and neutralising antibodies. AAV can activate innate pattern recognition receptors that lead to the release of inflammatory cytokines and type I interferons.

A delayed (adaptive) immune response occurs days or weeks after innate immunity. A cellular response can result in increased T cells and a humoral response in increased B cells, plasma cells and antibodies.

In the battle between AAV and the immune system, these are the important points to remember:

Although inflammation is par for the course in ocular gene therapy, ophthalmologists rarely discuss it;

Inflammation occurs with subretinal as well as intravitreal injections;

Inflammation may be subtle (subretinal deposits seen on an OCT) and may be correlated with loss of therapeutic efficacy;

Inflammation is typically dose-dependent, can manifest in multiple ways, and can occur and recur in adults;

Preventing inflammation should be the goal because once it develops, treatment can be difficult and require long-term management and extensive interventions;

Optimisation of capsid design for low empty capsid ratio, a clean manufacturing profile and promoter choice is required;

Intelligent and target-specific choice of immunosuppressant management, including general agreement on corticosteroid use, is key;

Alternative steroid-sparing immunosuppression can be used if a specified therapy requires long-term management; and

The use of a periocular steroid injection seems to have become routine in subretinal trials, but carries potential accompanying risks of cataract and glaucoma.

Because “our primary responsibility as physicians is to do no harm … we need to acknowledge inflammation in AAV-based gene therapy … discuss it, and learn … to control it,” Dr Kay concluded.

This article was adapted from Dr Christine Kay’s presentation at the 25th annual meeting of the American Society of Gene & Cell Therapy in May 2022. Dr Kay is an investigator for and consultant to numerous companies involved in retinal gene therapy research.

1. Blank T, Goldmann T, Koch M, et al. Early microglia activation precedes photoreceptor degeneration in mouse model of CNGB1-linked retinitis pigmentosa. 

2. Bainbridge JWB, Mehat MS, Sundaram V, et al. Long-term effect of gene therapy on Leber’s congenital amaurosis

3. Xue K, Jolly JK, Barnard AR, et al. Beneficial effects on vision in patients undergoing retinal gene therapy for choroideremia. 

4. Cehaic-Kapetanovic J, Xue K, Martinez-Fernandez de la Camara C, et al. Initial results from a first-in-human gene therapy trial on X-linked retinitis pigmentosa caused by mutation in RPGR. 

5. Bucher K, Rodríguez-Bocanegra E, Dauletbekov D, et al. Immune responses to retinal gene therapy using adeno-associated viral vectors—implications for treatment success and safety. 

6. Timmers AM, Newmark JA, Turunen HT, et al. Ocular inflammatory response to intravitreal injection of adeno-associated virus vector: relative contribution of genome and capsid. 

-regulatory sequences are correlated with ocular toxicity. 

Inflammation is par for the course in ocular gene therapy; preventing it should be the goal because once it develops, treatment can be difficult.

Causes of ocular inflammation linked to AAV gene therapy

This is the fourth in a series of articles that summarises an afternoon of presentations, plus a debate, on the topic of corneal ectasia that was held as part of the recent 29th Cologne Adventsymposium, an annual meeting organised by Laserforum e.V., in Cologne, Germany. Chaired by Drs Omid Kermani and Georg Gerten, the presentations each considered the condition from a different perspective and are available 

Part 1, published in the June 2022 issue of Ophthalmology Times Europe®, 

considered the contribution of eye rubbing to the development of keratoconus

. Part 2, published in the July/August 2022 issue, looked at 

developments in diagnostics and classification methods for ectatic corneal diseases

. Part 3, published in the September 2022 issue, covered 

. In this final instalment of the series, we look at recent developments in corneal cross-linking. This is based on the presentation that Prof. Farhad Hafezi gave during the event.

Reviewed by Dr Omid Kermani and Prof. Farhad Hafezi.

 Comparison of corneal stiffening achieved with different epi-off CXL irradiation protocols: blue is the Dresden protocol (30 minutes at 3 mW/cm2); red is 10 minutes at 9 mW/cm

; green is 5 minutes at 18 mW/cm2; purple is control with no cross-linking.

 (Image courtesy of Prof. Hafezi, adapted from Hammer et al. 2014

Prof. Hafezi began his presentation by explaining that there are three essential components for every corneal cross-linking (CXL) procedure: UV light, riboflavin and oxygen. When CXL was first developed in the early 2000s, it involved corneal epithelial cell debridement (making it an “epi-off” procedure) to enable riboflavin to be applied to the cornea and saturate the stroma.

This was followed by the delivery of UV, total dose (fluence) 5.4 J/cm

-intensity light for 30 minutes. This is known as the Dresden protocol.

One of the basic laws of photochemistry is the Bunsen–Roscoe law of reciprocity. If all reagents are in excess, then the amount of reaction that occurs depends solely on fluence—not the speed at which the light energy is delivered. Under these conditions, doubling light intensity and halving the irradiation time should have the same effect.

But would this principle hold true in CXL? Prof. Hafezi’s group tested the biomechanics of several different accelerated CXL protocols (

) and found a 10-minute protocol that, although it induces significantly less corneal stiffening than the Dresden protocol, is enough to stabilise a keratoconus in most cases.

Comparison of stiffening results from different epi-off cross-linking protocols. 

(Image adapted from Abrishamchi et al. 2021

Prof. Hafezi explained that “the reason that increasing acceleration of the cross-linking reaction results in decreasing stiffening effects is that the speed of oxygen diffusion into the stroma is the rate-limiting factor in cross-linking. If the cross-linking reaction is too fast, the oxygen cannot diffuse in fast enough and efficacy is lost, unless intensity is increased to compensate.” If there is no oxygen in the cornea, no stiffening occurs at all.

Until recently, Prof. Hafezi always reverted to the 30-minute Dresden protocol for particularly aggressive forms of keratoconus, such as in a young child or an ectasia after LASIK. However, he has now developed and published an accelerated epi-off protocol that compensates for the acceleration-related loss of stiffness by increasing fluence (

). This new protocol achieves the same stiffness as the Dresden protocol while saving the surgeon 20 minutes of time.

“The holy grail for cross-linking would be a truly effective epi-on procedure,” Prof. Hafezi said. Leaving the epithelium intact would mean faster healing, less haze and an even lower infection risk. “The quest for good, efficient epi-on has been going on for more than a decade. What do we need to make it work?”

If the epithelium remains in place, then the UV irradiation needs to be increased because the epithelium blocks around 15 to 20% of the light. This is relatively easy: increase the fluence from 5.4 to 7.2 J/cm

The next element is riboflavin. Riboflavin is a relatively large molecule, so transporting it through the tight junctions of the epithelium is not easy. One method is iontophoresis, which uses an electrical current to encourage transport.

The third factor is oxygen diffusion. The presence of the epithelium forms yet another barrier to oxygen diffusion into the cornea. It has been proposed that pulsing the light on and off might be effective: the idea is that oxygen is consumed when the light is on, so the oxygen can re-diffuse during the light’s off period.

The next question was: what happens with a slow procedure? Irradiating for 1 hour, instead of 30 minutes, with half the intensity, produces very efficient epi-on cross-linking.

 Prof. Hafezi said, “This was a proof of principle, and as a scientist I am happy—but as a clinician I don’t want to spend an hour in the operating room.”

One approach to increasing the amount of oxygen in the cornea during CXL, goggles that are fed a continuous stream of 100% oxygen, was proposed 2–3 years ago. However, Prof. Hafezi was not keen: “This makes sense. It should work. I don’t like it. Why? Because it is additional technology and I want to simplify the procedure, not make it more complicated and more expensive.”

“It is not necessary to have 100% oxygen surrounding the eye; what is needed is enough oxygen in the stroma for the cross-linking reaction to proceed effectively. What was needed was an intelligent combination of everything we have learned so far.”

There is now a published epi-on protocol that consists of normal air (no oxygen boost), iontophoresis to push riboflavin into the cornea, pulsed light, gentle acceleration, high fluence, and there are 3-year follow-up clinical data showing that it is effective.

The next question was, could epi-on CXL be undertaken without iontophoresis? Prof. Hafezi’s group has been assessing the ability of penetration enhancers to get riboflavin to pass through the epithelial cell tight junctions, spending almost a year at the Swiss Federal Institute of Technology using multiphoton microscopy to assess the extent of riboflavin penetration, in order to find a penetration enhancer combination that achieves a stromal riboflavin concentration that is similar to that achieved with epi-off CXL.

This means there is now a gently accelerated high-fluence epi-on protocol, which takes approximately 30 minutes in total (including 15 minutes of irradiation), that can be performed at the slit lamp. This protocol is currently undergoing clinical evaluation.

Prof. Hafezi reminded the audience that the original Dresden protocol required a minimum stromal thickness of 400 µm in order to protect the corneal endothelium from being exposed to potentially damaging levels of UV irradiation. Historically, surgeons have artificially increased the corneal thickness prior to treatment, either by swelling the cornea with hypo-osmolaric riboflavin

 or through the use of a riboflavin-soaked contact lens.

However, these methods have disadvantages. The amount of swelling achieved by the hypoosmolar riboflavin is unpredictable, and, because contact lenses form an additional barrier to oxygen diffusion, their use reduces the stiffening efficacy by around 30%.

What Prof. Hafezi proposes now is what he describes as individualised cross-linking. Previously, surgeons adapted the cornea to the CXL protocol; now the technique can be adapted to each patient’s cornea. His research group modelled the interaction of oxygen, UV light, riboflavin and the stroma, and made an algorithm to predict the extent of cross-linking based on these parameters.

Using this, they developed the “sub400” protocol, which uses measurements of the thinnest point of the cornea to calculate the optimal duration of a 3 mW/cm

 intensity irradiation (via a simple look-up table) in an otherwise standard epi-off CXL protocol that leaves the same un-cross-linked stromal safety margin as the Dresden protocol to avoid harming the endothelium.

Clinical results have been very encouraging, with around 80% of corneas having successful prevention of ectasia progression after 1 year, even in corneas as thin as ~200 µm that would otherwise have gone on to require keratoplasty.

This opens up a possibility for a new indication for CXL, keratoglobus. Prof. Hafezi has published a case report of sub400 CXL being used to successfully halt disease progression.

 The C-eye portable cross-linking device in use. 

Describing his aims, Prof. Hafezi said, “We developed the sub400 protocol and epi-on for one reason: we want to democratise access to cross-linking, moving away from an expensive infrastructure toward equipment we all share, which is the slit lamp, and this is why we developed the C-eye device” (EMAGine AG, Zug, Switzerland; 

According to Prof. Hafezi, the device is “small but high-tech. It can treat at the slit lamp with more than 11 different protocols—for keratoconus, for refractive, for keratitis—so if you ask me how often I go back to the laying position, I do, but only for special cases such as patients that are not compliant or who require general anaesthesia.” 

 Surgeon’s view of the cornea through C-eye during slit lamp cross-linking. 

What about riboflavin distribution when treating at the slit lamp: is it affected by gravity? Prof. Hafezi said, “The short answer is yes but it takes an hour before the first changes become apparent, which is more than enough time to perform cross-linking”.

With regards to fixation, it is much better with an upright patient who is comfortable and has a fixation light on the other eye so they know where to look. However, is the Dresden protocol, which takes half an hour, achievable?

“We don’t need to [perform this],” explained Prof. Hafezi. “Most cross-linking procedures take around 10–12 minutes; PACK [photo-activated chromophore for keratitis] cross-linking for keratitis takes around 6 minutes, and that’s easily feasible, especially if you give the patient the surgeon’s chair so they are really comfortable and spend 30 seconds at the beginning to make sure that the height is well adjusted.”

He added: “The last question is the most important: can we leave the operating room for the procedure room or even the examination room? Yes, we can. With epi-on there is no question at all: the epithelium is on. With epi-off, you do not only stiffen the cornea but also sterilise it, so the answer is yes.”

The trend of the past 4 or 5 years has been for office-based procedures. Intravitreal injections

 have been shown to be safe performed in the procedure room, or even the examination room, with no increased risk for endophthalmitis, so cross-linking in the procedure room should be possible.

Prof. Hafezi concluded his presentation by reiterating that cross-linking at the slit lamp broadens access to the procedure.

 It is suitable for all indications—epi-on, epi-off, keratoconus, PACK cross-linking—and is available to patients with a variety of special needs.

Corneal cross-linking (CXL) is improving all the time, although all protocols retain the basic requirements for oxygen, riboflavin and UV light. Recent developments mean that CXL at the slit lamp is feasible, which will improve access to the procedure.

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