Photo Activated Chromophore for Keratitis-Corneal Cross-linking
Corneal collagen cross-linking is a technique that uses riboflavin (Vitamin B2) and UV-A light (~365-370nm) to treat corneal ectasia or progressive keratoconus via stiffening of the cornea. Photo Activated Chromophore for Keratitis-Corneal Cross-linking (PACK-CXL) is an emerging application of the same technique to treat infectious keratitis. Infectious keratitis has been conventionally treated with antibiotics and antifungals. PACK-CXL offers a potential alternative as a primary or adjunctive treatment to topical antibiotics, which may become increasingly important in the context of escalating antimicrobial resistance. Clinically, in cases of microbial keratitis with corneal melt that are refractory to conventional antibiotic therapy, effective treatment may limit the need for emergent keratoplasty, which is associated with higher rates of infection and rejection. Additionally, since minor traumatic corneal ulcers are one of the leading causes of blindness in developing countries, PACK-CXL may also become a less expensive alternative to antibiotics and antifungals.
PACK-CXL is presumed to work via three main mechanisms. Firstly, the planar ring structure of the chromophore intercalates between the bases of DNA and RNA, which causes lesions in the chromosomal strands and thereby limits microbial replication. Secondly there is a release of reactive oxygen species that directly damages the pathogen cell walls. Lastly activated chromophores cross-link the corneal stroma via covalent bonds. This changes the tertiary structure of the fibers altering the sites of collagenases, rendering them more resistant to enzymatic degradation.
The first clinical usage of UV-A and riboflavin treatment for melting corneal ulcers was in 2000. Four patients with non-infectious corneal ulcers that had failed conventional treatment and required either a conjunctival flap or keratoplasty. PACK-CXL was given as an alternative treatment. Three patients had stoppage of their corneal melting. The fourth patient required a keratoplasty when the ulcer was still present three weeks post-treatment.
In 2008, five patients with keratitis of confirmed infectious origin was treated with PACK-CXL. Three patients with mycobacterial keratitis and two with fungal keratitis were refractory to topical treatments. Each patient was treated with PACK-CXL and afterwards appropriate antimicrobial and antifungal treatment was continued. Four patients had halting of the melting process. The fifth patient, who had a contact lens-induced fungal keratitis, continued to have corneal melting due to an immune reaction and underwent a keratoplasty.
Currently, PACK-CXL does not have FDA approval for the treatment of infectious keratitis. However, studies have revealed some clear contraindications:
- History or suspicion of herpes simplex keratitis, because latent herpes can be reactivated by UV radiation
- Severe keratitis with deeper infiltrates, because the standard CXL protocol produces an effective treatment depth limited to 250-300 μm
Most studies have used the protocol established by a study in 2003 known as the Dresden or standard protocol. In PACK-CXL, as in standard corneal cross-linking, the central 8-10 mm of the epithelium is removed. The patient is treated with 0.1% riboflavin drops at frequent intervals for 30 minutes. The patient is then positioned with UVA light at 365 nm with the desired irradiance of 3 mW/ cm2 applied at approximately 5 cm from the cornea for 30 min. Modifications of the standard protocol have involved changing the UVA treatment time. One study altered the UVA treatment to 30 mW/cm2 for 3 minutes and demonstrated the effectiveness of this protocol. A current ongoing study is testing the effectiveness of 3 mW/cm2 for 30 minutes or 9 mW/cm2 for 10 minutes, 18 mW/cm2 for 5 minutes, 30 mW/cm2 for 3 minutes.
Another study utilized a transepithelial approach, as there were already epithelial defects in the ulcerated regions, thereby modifying the Dresden protocol to not further disturb the epithelium. In this study of moderate to severe keratitis, adjunct PACK-CXL did not result in a reduction in the median size of the stromal infiltrate or the epithelial defects at day 7 or day 30 post-treatment when compared to the standard of care using topical antibiotics.
The consensus regarding the efficacy of PACK-CXL in treating microbial keratitis has been mixed. A meta-analysis of twenty-five studies that included 210 eyes of 209 patients of which 175 eyes underwent PACK-CXL concluded that it may have “some acceptable evidence” in the management of bacterial keratitis. Evidence regarding fungal and protozoan infection was weaker. However, additional randomized, prospective, controlled trials are required to judge its efficacy. Investigations to date have been limited by retrospective or uncontrolled study design, demographic or other differences between study groups, unclear inclusion and exclusion criteria, enrollment underpowered to achieve statistical significance, lack of masking, lack of well-defined dependent variables such as infiltrate depth and ulcerative area, failure to standardize operative and postoperative regimens and rescue regimens and need for robust interpretation of the data. Some observations common to the cumulative case series were (1) that smaller ulcers showed higher success rate of adjuvant PACK-CXL as determined by resolution of epithelial defect and stromal infiltrate; (2) eyes with hypopyon had a worse prognosis; (3) deeper stromal involvement was less amenable to successful treatment; (4) PACK-CXL contraindicated in eyes with herpes simplex keratitis.
A major issue has been a large difference in the defined inclusion and exclusion criteria of the studies that have been done with PACK-CXL. For each study listed below, the distinction between the type of pathogen and the depth of the infiltrate is emphasized since it may have implications on the resulting data.
Makdoumi K, et al.
The strength of this prospective non-randomized study is that PACK-CXL was used as a primary treatment for bacterial keratitis, instead of an adjunct treatment with standard therapy. The weakness of this study was the lack of grading of the ulcers treated, although it was noted that the majority of the ulcers were not severe.
- Enrolled 16 eyes of 16 patients
- Included patients that were ≥ 18 years and had suspected bacterial keratitis
- Excluded patients that had prior antibiotic treatment for the current episode of keratitis, suspicion of a non-bacterial keratitis, pachymetry values below 400 μm, stromal infiltrate in close proximity to limbus, pregnancy, breast-feeding, participation in any ophthalmological study in which the follow-up had not been completed or suspicion that the patient might not be able to complete the follow-up after treatment
- The primary end point was defined as epithelialization with non-progressive infiltration at two consecutive visits. All patients achieved the primary end point
- Mean time from treatment to epithelial healing of the cornea was 7.1 days (range 1-14 days, median 5.5)
- Visual acuity at the latest visit
- Increased in 7 patients
- Unchanged in 8 patients
- Decreased in 1 patient
- Corneal epithelization without intervention was achieved in 15 patients
- In one patient with negative culture and a past-medical history of glaucoma, CRVO, rubeosis received amniotic membrane transplant 14 days post-treatment due to advanced corneal edema and an epithelial defect prior to keratitis.
- Antibiotic administration was necessary in two cases.
- The first patient had history of diabetes mellitus, proliferative retinopathy with bilateral panretinal laser photocoagulation. The primary ulcer healed but secondary ulcer with an alternate localization had appeared and was treated with antibiotics.
- The second patient initially had deep corneal infiltrate reaching Descemet’s membrane with 1 mm hypopyon and marked anterior uveitis. Day 2 of post-treatment showed an intact corneal epithelium and a complete regression of the hypopyon. Anti-inflammatory therapy was initiated with the addition of an antibiotic 12 days post-PACK-CXL due to a recurrent secondary small epithelial defect and persistence of anterior uveitis.
- The secondary end point was to register any side-effects or complications of the treatment. None was observed.
Said DG, et al.
The strength of the study is that it is a prospective randomized clinical trial of advanced cases of microbial keratitis with melting that compares patients treated with standard PACK-CXL protocol within 48 hours of assessment followed by standard medical treatment with patients in the control group received medical treatment only. The weakness is that the mean size of the ulcer was larger in the PACK-CXL group (5.62±1.88 x 6.22±1.98 mm; p = 0.004) than in the control group (3.97±2.5 x 4.22±2.18 mm; p = 0.007) post-randomization
- Enrolled 40 eyes of 40 patients (21 in the PACK-CXL group and 19 in the control)
- Inclusion criteria ≥ 18 years and had an infective corneal ulcer with a possible bacterial, fungal, Acanthamoeba, or mixed origin with evident corneal melting
- Exclusion criteria excluded <18 years, corneal ulceration in proximity (1 mm) to the corneal limbus, underlying autoimmune disease, history of herpetic eye disease, corneal thickness less than 400 mm with epithelium, or pregnancy or nursing
- No difference in average time till healing. 39.76±18.22 days in the PACK-CXL group and 46.05±27.44 days in the control group (p = 0.68).
- No difference in after treatment corrected distance visual acuity. 1.64±0.62 logMAR in the PACK-CXL group and 1.67±0.48 logMAR in the control group (p = 0.68).
- Three patients in the control group had corneal perforation and one patient from the control group experienced infection recurrence. No patients in the PACK-CXL group had either perforation or recurrence. Because of overlapping 95% CI of perforation within the PACK-CXL and the control groups no significant conclusions can be drawn about the effect of PACK-CXL on the reduction or prevention of perforation.
Bamdad S, et al.
The strength of this study is that it is a prospective randomized clinical trial focusing only on moderate bacterial corneal ulcers. Patients were treated with PACK-CXL and then with standard medical therapy or standard therapy only. The weakness of this study includes the lack of details about the bacteria identified in cultures and not measuring the final visual acuity post-treatment.
- Enrolled 32 eyes of 32 patients (16 in the PACK-CXL group and 16 in the control group
- Grading of ulcers was defined as the following
- Grade 1 (mild): nonaxial, <2 mm in size, involved the superficial 1/3 of the cornea with mild anterior chamber reaction
- Grade 2 (moderate ulcers): 2 to 6 mm in size, involved the superficial 2/3 of the cornea, and had 4+ anterior chamber reactions
- Grade 3 (severe): >6 mm in size, extending to the inner 1/3 of the cornea or with severe hypopyon
- Exclusion of patients with corneal perforation, corneal descemetocele, collagen vascular disease, or immunocompromising diseases or those requiring emergency keratoplasty.
- Day 1 post-treatment: no difference in the size of epithelial defects (p = 0.16), area of infiltrates (p = 0.89) or in the grade of ulcers (p = 0.99) in the PACK-CXL when compared to the control group.
- Day 7 post-treatment: smaller epithelial defects (p = 0.001) and smaller area of infiltrates (p = 0.001) but no difference in the grade of ulcers (p = 0.56) in the PACK CXL group when compared to the control group.
- Day 14 post-treatment: smaller epithelial defects (p = 0.001), smaller area of infiltrates (p = 0.001) and significant differences in the grade of the ulcers (p = 0.001) in the PACK CXL group when compared to the control group.
Alio JL, et al.
This is a meta-study that identified 12 unique articles (including Makdoumi et. al) that used PACK-CXL for the treatment of infectious keratitis. The strength of a meta-study is that it analyzes the overall effectiveness of PACK-CXL in multiple studies. The weakness of this analysis was that none of the included studies had control groups.
- A search was performed in Medline for studies from January 2000 to January 2013 that described treatment of infectious keratitis with PACK-CXL. All papers were case reports or case series with level 3 evidence according to Cochrane systemic metanalysis protocol. Articles were individually reviewed to identify original clinical data and bibliographies were searched to manually add additional articles in peer-reviewed journals.
- Exclusion of in vitro, animal and non-English studies
- Primary outcome was defined as healing of the corneal ulcer with no progressive infiltration, block of corneal melting and secondary endpoint of recovery of visual acuity
- 104 eyes were analyzed, which included
- Bacterial infection in 58 eyes (57%)
- Gram-positive bacteria in 44 eyes (43%), with 4 Mycobacterium (3.6%)
- Gram-negative bacteria in 14 eyes (13%)
- Fungal infection in 14 eyes (13%)
- Acanthamoeba infection in 7 eyes (7%)
- Culture negative or not performed in 26 eyes (25%)
- Bacterial infection in 58 eyes (57%)
- Treatment was effective in blocking corneal melting overall at 85% (95%; CI 0.77 to 0.91). Treatment was effective in the following order: in Gram-negative bacteria (13/14; 92%), in Gram-positive bacteria (37/44; 84%), Acanthamoeba (5/7; 71%) and lastly fungus (8/13; 61%).
- Gram-positive Mycobacterium required corneal transplants for all patients
- The mean time of reepithelization post PACK-CXL treatment was 20.7±28.1 days (3 to 145 days). The mean reepithelization days was:
- 11.3 (3 to 45) for Gram-positive bacterial keratitis
- 12.6 (4 to 30) for Gram-negative bacterial keratitis
- 53.2 (3 to 120) for fungal keratitis
- 66.6 (8 to 145) for Acanthamoeba keratitis
- 19.25 for culture negative keratitis
- Few complications were noted after PACK-CXL treatment, namely:
- An eye with Candida keratitis developed corneal edema
- An eye culture-negative infection developed a severe dendritic lesion.
- 16 eyes (15%) required deep or lamellar keratoplasty
- 7/44 (16%) of Gram-positive bacterial keratitis
- 1/14 (7%) of Gram-negative bacterial keratitis
- 2/7 (28%) of Acanthamoebal keratitis
- 5/13 (38%) of fungal keratitis
- 1/26 (4%) of culture negative keratitis
Prajna NV, et al14
Published in Ophthalmology in February, 2020, the authors of this article performed a randomized controlled clinical trial that randomized patients "with moderate vision loss from a smear-positive fungal ulcer" to either topical natamycin alone, topical natamycin + CXL, topical amphotericin alone, or topical amphotericin + CXL. They found that, regardless of antifungal used, there was no benefit to CXL. Patients who underwent CXL were no more likely to re-epithelialize by 3 weeks or 3 months, they had no improvement in infiltrate or scar size, and they actually had worse final visual acuity.
- Conjunctival injection, cells, flare in the anterior chamber and increased hypopyon
- Failure to control microbial keratitis, including corneal perforation
- Exacerbation of herpes simplex keratitis
- Potential corneal endothelial cell loss and corneal edema
- Price MO, Price FW. Corneal cross-linking in the treatment of corneal ulcers. Curr Opin Ophthalmol. 2016. doi:10.1097/ICU.0000000000000248
- Schnitzler E, Spörl E, Seiler T. [Irradiation of cornea with ultraviolet light and riboflavin administration as a new treatment for erosive corneal processes, preliminary results in four patients]. Klin Monbl Augenheilkd. 2000. doi:10.1055/s-2000-10344
- Iseli HP, Thiel MA, Hafezi F, Kampmeier J, Seiler T. Ultraviolet a/riboflavin corneal cross-linking for infectious keratitis associated with corneal melts. Cornea. 2008. doi:10.1097/ICO.0b013e318169d698
- Wollensak G, Spoerl E, Seiler T. Riboflavin/ultraviolet-A-induced collagen crosslinking for the treatment of keratoconus. Am J Ophthalmol. 2003. doi:10.1016/S0002-9394(02)02220-1
- Knyazer B, Krakauer Y, Baumfeld Y, Lifshitz T, Kling S, Hafezi F. Accelerated Corneal Cross-Linking With Photoactivated Chromophore for Moderate Therapy-Resistant Infectious Keratitis. Cornea. 2018;37(4):528-531. doi:10.1097/ICO.0000000000001498
- Hafezi F. Swiss PACK-CXL Multicenter Trial for the Treatment of Infectious Keratitis. https://clinicaltrials.gov/ct2/show/NCT02717871. Published 2016.
- Kasetsuwan N, Reinprayoon U, Satitpitakul V. Photoactivated Chromophore for Moderate to Severe Infectious Keratitis as an Adjunct Therapy: A Randomized Controlled Trial. Am J Ophthalmol. 2016. doi:10.1016/j.ajo.2016.02.030
- Papaioannou L, Miligkos M, Papathanassiou M. Corneal Collagen Cross-Linking for Infectious Keratitis: A Systematic Review and Meta-Analysis. Cornea. 2016. doi:10.1097/ICO.0000000000000644
- Makdoumi K, Mortensen J, Sorkhabi O, Malmvall BE, Crafoord S. UVA-riboflavin photochemical therapy of bacterial keratitis: A pilot study. Graefe’s Arch Clin Exp Ophthalmol. 2012. doi:10.1007/s00417-011-1754-1
- Said DG, Elalfy MS, Gatzioufas Z, et al. Collagen cross-linking with photoactivated riboflavin (PACK-CXL) for the treatment of advanced infectious keratitis with corneal melting. Ophthalmology. 2014. doi:10.1016/j.ophtha.2014.01.011
- Mittal R, Garg P. Re: Said et al.: Collagen cross-linking with photoactivated riboflavin (PACK-CXL) for the treatment of advanced infectious keratitis with corneal melting (Ophthalmology 2014;121:1377-82). Ophthalmology. 2014. doi:10.1016/j.ophtha.2014.06.043
- Bamdad S, Malekhosseini H, Khosravi A. Ultraviolet A/riboflavin collagen cross-linking for treatment of moderate bacterial corneal ulcers. Cornea. 2015. doi:10.1097/ICO.0000000000000375
- Alio JL, Abbouda A, Valle DD, del Castillo JMB, Fernandez JAG. Corneal cross linking and infectious keratitis: A systematic review with a meta-analysis of reported cases. J Ophthalmic Inflamm Infect. 2013. doi:10.1186/1869-5760-3-47
14. Prajna NV, Radhakrishnan N, Lalitha P, et al. A Randomized Clinical Trial Evaluating the Effect of Adjuvant Cross-Linking on Outcomes in Fungal Keratitis. Ophthalmology. Feb 2020. Vol 127, Number 2, pp 159-166.