Corneal collagen crosslinking (CXL), initially described by Spoerl and colleagues,¹ has extended far beyond its original indication of progressive keratoconus. Some clinicians might wonder how this technique can be so versatile and used in so many different ways. The versatility comes from the fact that crosslinking is one of the most basic elements of corneal architecture. Just as an architect can conceptualize different elements of a building using similar means and techniques, it is only now that we are recognizing the possibilities that the technique of crosslinking has for acting on corneal tissue.

Research in the field of CXL is highly dynamic; techniques, concepts, and indications are constantly evolving. This article attempts to provide a comprehensive outlook on the current state of the art of CXL technology in 2015. Given the speed of development, an outlook article written 12 months from now for 2016 would (hopefully) read quite differently.

CXL FOR ECTASIA

Ectasia of various origins (keratoconus, pellucid marginal degeneration, and post-LASIK ectasia) remains the main indication for CXL. Epithelium (epi)-off techniques using no more than 9 mW/cm² seem to be efficient for these indications.^2-4 The year 2014 has again seen vigorous debate on refinements of the technique, intended to either accelerate CXL or reduce postoperative pain and inflammation.

Accelerated CXL. Intensities of up to 30 mW/cm² are already in clinical use to accelerate the procedure; however, there is still doubt regarding the biomechanical effectiveness of CXL at these high intensities. Our research group showed⁵ that the biomechanical effect of CXL decreases distinctly at irradiations higher than 9 mW/cm.² The Bunsen-Roscoe law of reciprocity states that, “a photochemical reaction will stay constant if the total energy is constant: a shortened irradiation time at higher irradiance should lead to the same increase in biomechanical stiffness as a longer irradiation time at lower irradiance.”⁵ Apparently, this law of photochemistry cannot be applied to the biomechanical effect of CXL in a living organism (Figure 1).

Most published clinical evidence pertains to 3 mW/cm² for 30 minutes (the original Dresden protocol) or 9 mW/cm² for 10 minutes. For intensities higher than 9 mW/cm², publications are sparse and lack documented stabilization of progression.⁶

Epi-on techniques to reduce postoperative pain and inflammation. The success of epi-on techniques depends on both riboflavin and oxygen availability (see Why Do High Intensities and Epi-On Techniques Fail in Stabilizing Corneal Biomechanics?). Various riboflavin solutions have been tested for the ability to strengthen corneal biomechanics in the laboratory; yet some commercially available solutions fail to work under either experimental or clinical conditions.^7,8 Likewise, some modifications of epi-on solutions that seem to induce a satisfying effect in the laboratory results seem promising; however, clinical evidence has not been reported yet⁹

CXL PLUS

CXL in combination with refractive laser surgery procedures, also referred to as CXL Plus, may be applied to improve visual quality in keratoconus by depth-limited tissue regularization (the Athens protocol);¹⁰ to prevent ectasia after LASIK (LASIK Xtra); or to treat low myopia as a standalone procedure. A few warnings:

• Although the Athens protocol can improve visual quality and increase distance BCVA, clinicians should proceed with caution, as long-term data (more than 5 years) on biomechanical stability is sparse.
• LASIK Xtra has been proposed to reduce the risk of iatrogenic ectasia; however, given that ectasia after LASIK has become an extremely rare event, proving that the combination of rapid CXL and LASIK is effective would require a large study with more than 10,000 eyes. In other words, with LASIK Xtra, the surgeon is opting to conduct an additional procedure after LASIK that has not been proven to be effective but that can increase the risk of unpredictable changes in corneal curvature.
• The flattening effect of CXL in normal corneas has not yet been well defined. If a normal cornea behaves even remotely similarly to a keratoconic cornea, then we have not yet learned to control the process of flattening precisely enough to allow refractive corrections of myopia.

PACK-CXL FOR INFECTIOUS KERATITIS

Infectious keratitis and related corneal ulcers are a major cause of global blindness, with several million people affected every year in developing countries.¹¹ In view of the alarming 2014 report of the World Health Organization (WHO) regarding increasing resistance to antibiotics in medicine, alternative therapies to antibiotic therapy would be highly appreciated. The use of photoactivated chromophore for infectious keratitis CXL (PACK-CXL)¹¹ might represent such an approach.

It may be possible to use PACK-CXL in the treatment of bacterial, fungal, and mixed corneal ulcers; a proof-of-principle study was conducted by our group in 2008.¹³ This technique seems to work efficiently in early and more superficial ulcers¹⁴ and as an adjuvant or last-resort treatment in large and deep ulcers.¹⁵ In the distant future, PACK-CXL may also be an option for treating Acanthamoeba keratitis. However, it has not been shown to be effective yet, and more laboratory and clinical research is needed.^16,17

At the time of this writing, a large prospective, randomized multicenter trial was scheduled to commence in December 2014 to focus on PACK-CXL as a first-line treatment modality in previously untreated corneal infiltrates and small ulcers. PACK-CXL alone will be compared with antimicrobial treatment, the current standard of care.

ELUCIDATING CXL MOLECULAR MECHANISMS

In the past, CXL technology was mostly driven by an interesting trial-and-error approach, by which new clinical techniques were sometimes introduced with little to no experimental, laboratory, or clinical data supporting the claim. An ideal way to better understand which genes and proteins play a role in crosslinking would be to work in transgenic organisms, in which a single gene out of many thousands can be selectively overexpressed (enhanced) or knocked out.

The mouse would be an ideal animal for that purpose because thousands of transgenic strains exist already. There is an obstacle with using a mouse model, however: Whereas epi-off CXL is possible without too many obstacles (Figure 2), analyzing the biomechanical response of the mouse cornea has failed so far due to technical limitations—the mouse cornea is only 1 mm in diameter. After several years of building customized setups and experimenting, we succeeded in measuring stress-strain measurements in the mouse eye (Kling et al, in preparation for publication). Our mouse model might open new alleys for the identification of yet unknown factors and players in this fascinating field.

Farhad Hafezi, MD, PhD, is Medical Director of The ELZA Institute, Zurich, Switzerland; Professor of Ophthalmology, University of Geneva, Switzerland; and Clinical Professor of Ophthalmology, University of Southern California, Los Angeles. Dr. Hafezi states that he is a named co-inventor of PCT/CH 2012/000090 application (ultraviolet light source). He may be reached at e-mail: farhad@hafezi.ch.

1. Spoerl E, Huhle M, Kasper M, Seiler T. Increased rigidity of the cornea caused by intrastromal cross-linking. Ophthalmologe. 1997;94(12):902-906.
2. O’Brart DP, Kwong TQ, Patel P, et al. Long-term follow-up of riboflavin/ultraviolet A (370 nm) corneal collagen cross-linking to halt the progression of keratoconus. Br J Ophthalmol. 2013;97(4):433-437.
3. Richoz O, Mavrakanas N, Pajic B, Hafezi F. Corneal collagen cross-linking for ectasia after LASIK and photorefractive keratectomy: Long-term results. Ophthalmology. 2013;120(7):1354-1359.
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10. Kanellopoulos AJ, Binder PS. Collagen cross-linking (CCL) with sequential topography-guided PRK: A temporizing alternative for keratoconus to penetrating keratoplasty. Cornea. 2007;26(7):891-895.
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13. Iseli HP, Thiel MA, Hafezi F, et al. Ultraviolet A/riboflavin corneal cross-linking for infectious keratitis associated with corneal melts. Cornea. 2008;27(5):590-594.
14. Price MO, Tenkman LR, Schrier A, et al. Photoactivated riboflavin treatment of infectious keratitis using collagen crosslinking technology. J Refract Surg. 2012;28(10):706-713.
15. 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;121(7):1377-1382.
16. Berra M, Galperin G, Boscaro G, et al. Treatment of acanthamoeba keratitis by corneal cross-linking. Cornea. 2013;32(2):174-178.
17. Richoz O, Gatzioufas Z, Hafezi F. Corneal collagen cross-linking for the treatment of acanthamoeba keratitis. Cornea. 2013;32(10):189.