Lens regeneration in children

BRIEF COMMUNICATIONS ARISING Improving outcomes in congenital cataract ARISING FROM

H. Lin et al. Nature 531, 323–328 (2016); doi:10.1038/nature17181

Lens regeneration after cataract surgery in infants is a clinical phenomenon with which paediatric ophthalmologists battle. Lin et al.1 reported a novel surgical technique that aimed to convert this postoperative complication into an alternative therapy for children aged under 24 months. However, the early outcomes reported in the experimental group fall short of outcomes achievable through conventional treatment in this population. As the authors offer no comment or explanation for this discrepancy, this new approach cannot be considered effective or safe for affected children. There is a Reply to this Comment by Liu, Y. et al. Nature 556, https://dx.doi.org/10.1038/nature26150 (2018). Regeneration of residual lens cells following surgical removal of congenital cataract can result in re-opacification, which needs to be treated by further intraocular surgery, necessitating repeated general anaesthetics during a sensitive period of neurodevelopment. The adaptation of this regenerative process by Lin et al.1, in which a novel surgical method of cataract removal preserves endogenous lens cells, enabling functional lens regeneration, may eventually lead to the development of treatments for degenerative disease. However, issues relating to adult cataract (an age-related degenerative process) and congenital and infantile cataract are conflated in their report. Cataract is virtually universal in older age, making cataract surgery one of the most common surgical procedures, with enviably excellent visual outcomes. By contrast, infantile cataract is uncommon, affecting 3–15 per 10,000 children worldwide and, by definition, present from birth or early infancy2. We now understand that mutations within the genes responsible for the production or orchestration of the lens epithelial progenitor/stem cells (LECs) are responsible for the majority of bilateral congenital or infantile cataract, even in cases where there is no family history3. Thus the treatment approach adapted by Lin et al.1, which relies on regeneration of lens stem cells without addressing the persisting underlying genetic defect, cannot be definitive. Children treated using this technique may require further surgery but Lin et al. do not acknowledge this in the article. The rationale for the trial reported by Lin et al.1 is the need to address adverse outcomes associated with the use of artificial intraocular lenses, which are implanted in some children to replace the focusing power of the removed cataractous lens4. However, surgery with intraocular lens implantation was not the ‘control’ standard approach evaluated within the report. Equivalence in vision outcomes between their intervention and control groups was reported, but these should also have been assessed against the extant benchmark. Outcomes in infantile cataract have improved substantially over the past few decades, largely owing to the application of basic neuroscientific understanding of sensitive and critical periods in visual neurodevelopment. Cataractrelated childhood visual impairment is largely due to bilateral stimulus deprivation amblyopia: the failure to restore a normal trajectory of visual neurodevelopment during a brief and finite window of opportunity. This critical window closes at some point during the first six months of life. Thus, younger age at surgery is the strongest predictor of better visual outcome and, in cataract present from birth, the window for intervention is conventionally considered to be the first six to eight weeks of life. Hence whole-population newborn screening programmes exist in many countries to ensure early diagnosis of congenital cataract and prompt referral for specialist treatment. In settings where such programmes do not yet exist, late diagnosis and treatment, resulting in irreversible amblyopia, drives poor visual outcomes. We have previously, on behalf of the British Isles Congenital Cataract Interest Group, reported outcomes within a contemporaneous, nationally

representative cohort of children undergoing surgery in the British Isles for congenital and infantile cataract in the first two years of life (IoLunder2 study)4. These outcomes are comparable to those found in other contemporary reports5 and are more than twofold better than those reported by Lin et al. in either their experimental or control groups. Indeed, the mean acuity achieved in their trial is the threshold for legal definition of blindness, an outcome that would lead most ophthalmologists, and probably most parents, to question the value of this new proposed intervention. Effective treatment for congenital cataract requires, alongside surgery, post-operative management of the loss of refractive (focusing) power through removal of the natural lens. Failure to appropriately manage this results in dense amblyopia. As the method described in Lin et al. involves an 8-month post-operative period of lens regeneration with consequent partially obscured and poorly focused vision, the inevitable resultant amblyopia may explain the poor visual outcomes. The authors offer no other explanation, and do not describe how the rapidly changing and highly defocused refractive state (moving through 18 diopter units of refractive error in 8 months) was managed. Had the report adhered to the international reporting standard of CONSORT6 it might be have been possible to assess the quality (internal validity) and generalizability (external validity) of the trial. For example, it is necessary to know: whether the control and intervention groups were equivalent with respect to baseline clinical characteristics (particularly age at surgery); how randomization was undertaken; the power calculation and the primary outcome and secondary outcomes on which this was based; how clustering by surgeon was addressed; and how clustering and correlation of outcome data were addressed in the analysis, given that both eyes of each subject were treated and analysed. The authors have described their study as a phase 1 trial, but the aim of such an investigation is to assess adverse outcomes of treatment, such as uncorrected high or irregular refractive outcome, which will drive the visual system towards amblyopia and resultant visual impairment. As the paper stands, it is not possible to agree with its principal conclusion that it provides evidence supporting the superiority of the novel treatment. A tempered report, clearly articulating the limitations of the approach with respect to outcome and permanency of effect, would have avoided giving the false impression that the new approach can be expected to supercede current treatment practices. A.L.S. and J.R. are supported by the National Institute for Health Research (NIHR) Biomedical Research Centres at Moorfields Eye Hospital NHS Foundation Trust and UCL Institute of Ophthalmology, and at the UCL Institute of Child Health/Great Ormond Street Hospital for children. A.L.S. is supported by fellowships from the Ulverscroft Vision Research Group and the Academy of Medical Sciences. Ameenat Lola Solebo1,2,3,4, Christopher J. Hammond5 & Jugnoo S. Rahi1,2,3,4 1 Lifecourse Epidemiology and Biostatistics Section, Population, Policy and Practice Programme, Institute of Child Health, University College London, London, UK. email: [email protected] 2 Great Ormond Street Hospital/Institute of Child Heath NIHR Biomedical Research Centre, London, UK. 3 Moorfields Eye Hospital and Institute of Ophthalmology NIHR Biomedical Research Centre, London, UK. 5 A P R I L 2 0 1 8 | VO L 5 5 6 | NAT U R E | E 1

© 2018 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.

BRIEF COMMUNICATIONS ARISING 4 Ulverscroft Vision Research Group, University College London Institute of Child Health, London, UK. 5 King’s College Hospital London, Academic Section of Ophthalmology, School of Life Course Sciences, Faculty of Life Sciences and Medicine, St Thomas’s Hospital Campus, London, UK.

Received 13 April 2016; accepted 23 January 2018. 1. 2. 3.

Lin, H. et al. Lens regeneration using endogenous stem cells with gain of visual function. Nature 531, 323–328 (2016). Solebo, A. L. in Congenital Cataract: A Concise Guide to Diagnosis and Management (eds Lloyd, I. C. et al.) (Springer, 2016). Gillespie, R. L. et al. Personalized diagnosis and management of congenital cataract by next-generation sequencing. Ophthalmology 121, 2124–2137 (2014).

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5. 6.

Solebo, A. L., Russell-Eggitt, I., Cumberland, P. M. & Rahi, J. S. Risks and outcomes associated with primary intraocular lens implantation in children under 2 years of age: the IoLunder2 cohort study. Br. J. Ophthalmol. 99, 1471–1476 (2015). Ye, H. H., Deng, D. M., Qian, Y. Y., Lin, Z. & Chen, W. R. Long-term visual outcome of dense bilateral congenital cataract. Chin. Med. J. (Engl.) 120, 1494–1497 (2007). Schulz, K. F., Altman, D. G. & Moher, D. CONSORT 2010 statement: updated guidelines for reporting parallel group randomised trials. PLoS Med. 7, e1000251 (2010).

Author Contributions All authors contributed equally to all aspects of authorship. Competing Financial Interests Declared none. doi:10.1038/nature26148

Lens regeneration in children ARISING FROM

H. Lin et al. Nature 531, 323–328 (2016); doi:10.1038/nature17181

Congenital cataracts are the primary cause of treatable childhood blindness worldwide, affecting about four infants per 10,000 live births1. Current surgical techniques have helped thousands of patients, but the limitations of these techniques2–4 led Lin et al.5 to report an alternative approach that they claim leads to a regenerated lens with refractive capacity. We have concerns regarding features of the presented data and the conclusions reached in the Article. This has implications for our patients who ask for such surgical intervention. There is a Reply to this Comment by Liu, Y. et al. Nature 556, http://dx.doi.org/10.1038/ nature26150 (2018). Embryologically, the lens of the eye is derived from the surface ectoderm6,7 and shares many properties of ectodermal tissue. Lens cortex continues to be produced throughout life, and lens size continues to increase owing to progressive elongation of slowly dividing lens epithelial cells (LECs) into fibre cells that lose their nuclei8–13. LECs are capable of unwanted proliferation in aged eyes; ophthalmologists observe this routinely after cataract extraction, when LECs proliferate and form semi-transparent scar-like tissue such as Soemmerring (Sömmering) rings and lens pearls14. Lens transparency is maintained as the result of highly organized fibre cell packing with extracellular spaces that are narrower than the wavelength of light. The authors claim that if they perform a cataract extraction in a way that reduces the injury to the lens epithelium, the normal natural epithelial healing response will lead to a new biconvex, transparent lens with flexibility (accommodative power). However, the authors concede in their Reply to this BCA that with their intervention it is unreasonable to expect a completely normal regenerated lens. Without a completely normal lens, including clarity, normal shape, and normal size, one would predict that normal visual function would not develop and children’s visual development would suffer from amblyopia. The lack of clarity and normal structure of the regenerated lentoids is highlighted for example in figure 3b of Lin et al.5, where the image of a rabbit lens seven weeks after surgery shows an optically irregularly shaped lens, not the biconvex shape of a normal lens6. In addition, figure 3c and f of ref. 5 shows images of rabbit lenses with posterior subcapsular cataracts in the visual axis that would decrease vision. Furthermore, histopathology was not presented in the rabbit regenerated lenses at maximum axial length, and thus extended data figure 7a of Lin et al.5 is insufficient to assess the quality and normality of lens regeneration. To demonstrate a normal reconstituted lens, it is necessary to provide sagittal lens sections and lens measurements that encompass the nodal point of the eye. The lens imperfections

demonstrated in figure 4a of ref. 5 are multiple, not single including the visual axis, and represent more than just the loss of LECs and scarring at the site of capsulorhexis. Such changes are by definition cataractous. The images of postoperative human eyes in figure 5f of ref. 5 appear to show hazy (that is, cataractous) lenses. More importantly, extended data figure 8e of ref. 5 shows no regenerated lens six months after lens surgery. Instead, a single light reflex is shown, which is likely to be from the lens capsule alone with no lens cortex and no evidence of a clear, biconvex crystalline lens that would be required for normal vision (compare the single lens capsule slit lens reflex of extended data figure 8e of ref. 5 with the normal lens reflex in figure 3b). In the clinical setting, a clear view by indirect ophthalmoscopy (as shown in extended data figure 8c, d of ref. 5) in no way signifies that the lens casts a clear and focused image on the retina. It is easy for indirect ophthalmoscopy to get clear views through lenses with legally blinding cataracts. There is no information about the authors’ management of the children’s refractive status. Importantly, there remains the distinct possibility that the proposed surgical technique may cause inferior visual outcomes. Comparing visual acuity measures in studies of children can be inherently difficult because of different clinical characteristics and the use of different measurement techniques. Nevertheless, the visual outcomes reported in both the experimental and conventionally treated groups were unacceptably poor. Six months after surgery, the mean visual acuity was equivalent to 20/200 (6/60), whereas other investigators have reported that 60% of children with bilateral infantile cataracts achieve an acuity of 20/40 or better after standard ophthalmic care, and almost all achieve better acuity than the mean visual acuity reported by Lin et al.5. We would welcome a graphic, detailed distribution of the visual acuity results from the authors’ patients (rather than only means or summarized data). Follow-up data covering more than one year represents another important clinical parameter. Finally, as the authors state that they did not exclude all genetic causes of congenital cataracts, and that they did not perform any gene editing or manipulation of the lens epithelial cells, we question why the proliferating LECs, presumably with genetic variants causing the original congenital cataracts, did not recapitulate the original lens opacity. One cannot just assume that germline mutations would constitute a minority of their cases without having data from their patients to support such a claim. Although the current surgical approaches for congenital cataracts have adverse events and several limitations that have been identified in controlled long-term studies, Lin et al.5 do not provide sufficient shortterm or long-term supportive evidence to support their statement that

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BRIEF COMMUNICATIONS ARISING their approach may afford an infant eye a good chance of an improved visual outcome. The study is limited by varied follow-up, poor visual acuities, insufficient details regarding adverse events, and a lack of information about the need for additional surgeries. These limitations should alert us to the need to proceed cautiously. Using such an unproven experimental therapy in both eyes of a patient is not appropriate. We all support innovation and disruptive technology. Yet, at the same time, it is necessary to pay careful attention to details including thorough documentation and achievement of long-term milestones to support the conclusions. One should be even more vigilant when proposing a new therapy for our youngest patients, who represent the most vulnerable population. It is important to report both study results and limitations in a clear and balanced fashion, particularly to parents who desperately want the best possible outcome for their infants. Demetrios G. Vavvas1,2, Thaddeus P. Dryja3, M. Edward Wilson4, Timothy W. Olsen5, Ankoor Shah1,6, Ula Jurkunas1,2, Roberto Pineda1,2, Vasiliki Poulaki1,7, Sotiria Palioura8, Peter Veldman9, Javier Moreno-Montañés10, Maria D. Pinazo-Duran11, José Carlos Pastor12, Miltiadis Tsilimbaris13, Douglas Rhee14, Kathryn Colby9, David G. Hunter1,6, Solon Thanos15, Taiji Sakamoto16, Louis R. Pasquale1,2, Joan W. Miller1,2, Deborah VanderVeen1,6 & Scott R. Lambert17 1 Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, USA. 2 Department of Ophthalmology, Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, USA. email: [email protected]; [email protected]. edu; [email protected]

Molecular Ophthalmobiology Group at the Department of Ophthalmology, University of Valencia, Valencia, Spain. 12 Department of Ophthalmology, Hospital Clinico Universitario and IOBA (Eye Institute) University of Valladolid, Valladolid, Spain. email: [email protected] 13 Department of Ophthalmology, University of Crete, Crete, Greece. 14 Department of Ophthalmology, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA. 15 Institute of Experimental Ophthalmology, Westfalian Wilhelms-University of Münster, Albert-Schweitzer Campus 1, D15, 48149 Münster, Germany. 16 Department of Ophthalmology, Kagoshima University, Kagoshima, Japan. 17 Department of Ophthalmology, Stanford University School of Medicine, Stanford, California, USA. email: [email protected] Received 13 April 2016; accepted 23 January 2018. 1. 2. 3. 4. 5. 6. 7. 8.

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Ocular Pathology Service, Massachusetts Eye and Ear Infirmary, Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts, USA. 4 Albert Florens Storm Eye Institute, Medical University of South Carolina, Charleston, South Carolina, USA. email: [email protected] 5 Department of Ophthalmology, Mayo Clinic, Rochester, Minnesota, USA. email: [email protected] 6 Department of Ophthalmology, Boston Children’s Hospital, Boston, Massachusetts, USA. email: [email protected] 7 Department of Ophthalmology, Veterans Affairs Hospital, Boston University, Boston, Massachusetts, USA. 8 Department of Ophthalmology, Bascom Palmer Eye Institute, University of Miami, Miami, Florida, USA. 9 Department of Ophthalmology, University of Chicago, Chicago, Illinois, USA. 10 Department of Ophthalmology, University of Navarra, Pamplona, Spain. 11 Ophthalmic Research Unit “Santiago Grisolía” and Cellular and

9. 10. 11. 12. 13. 14.

Wu, X., Long, E., Lin, H. & Liu, Y. Prevalence and epidemiological characteristics of congenital cataract: a systematic review and meta-analysis. Sci. Rep. 6, 28564 (2016). Lambert, S. R. et al. Is there a latent period for the surgical treatment of children with dense bilateral congenital cataracts? J. AAPOS 10, 30–36 (2006). Young, M. P., Heidary, G. & VanderVeen, D. K. Relationship between the timing of cataract surgery and development of nystagmus in patients with bilateral infantile cataracts. J. AAPOS 16, 554–557 (2012). Sukhija, J., Ram, J., Gupta, N., Sawhney, A. & Kaur, S. Long-term results after primary intraocular lens implantation in children operated less than 2 years of age for congenital cataract. Indian J. Ophthalmol. 62, 1132–1135 (2014). Lin, H. et al. Lens regeneration using endogenous stem cells with gain of visual function. Nature 531, 323–328 (2016); corrigendum 541, 558 (2017). Cvekl, A. & Ashery-Padan, R. The cellular and molecular mechanisms of vertebrate lens development. Development 141, 4432–4447 (2014). Piatigorsky, J. Lens differentiation in vertebrates. A review of cellular and molecular features. Differentiation 19, 134–153 (1981). Augusteyn, R. C. On the growth and internal structure of the human lens. Exp. Eye Res. 90, 643–654 (2010). Hanna, C. & O’Brien, J. E. Cell production and migration in the epithelial layer of the lens. Arch. Ophthalmol. 66, 103–107 (1961). Harding, C. V., Hughes, W. L., Bond, V. P. & Schork, P. Autoradiographic localization of tritiated thymidine in wholemount preparations of lens epithelium. Arch. Ophthalmol. 63, 58–65 (1960). Mikulicich, A. G. & Young, R. W. Cell proliferation and displacement in the lens epithelium of young rats injected with tritiated thymidine. Invest. Ophthalmol. 2, 344–354 (1963). Šikic´, H., Shi, Y., Lubura, S. & Bassnett, S. A stochastic model of eye lens growth. J. Theor. Biol. 376, 15–31 (2015). Smith, P. Diseases of crystalline lens and capsule. 1. On the growth of the crystalline lens. Trans. Ophthalmol. Soc. UK 3, 79–99 (1883). Miyake, K., Ota, I., Miyake, S. & Horiguchi, M. Liquefied aftercataract: a complication of continuous curvilinear capsulorhexis and intraocular lens implantation in the lens capsule. Am. J. Ophthalmol. 125, 429–435 (1998).

Author Contributions D.G.V., T.P.D., M.E.W., T.W.O., L.R.P., J.W.M., D.V. and S.R.L. conceived, wrote and edited the manuscript. A.S., U.J., R.P., V.P., S.P., P.V., J.M.-M., M.D.P.-D., J.C.P., M.T., D.R., K.C., D.G.H., S.T. and T.S. reviewed and edited the manuscript. Competing Financial Interests Declared none. doi:10.1038/nature26149

Liu et al. reply Replying to: D. G. Vavvas et al. Nature 556, http://dx.doi.org/10.1038/nature26149 (2018); A. L. Solebo, C. J. Hammond & J. S. Rahi Nature 556, http://dx.doi.org/10.1038/nature26148 (2018)

In the accompanying Comments1,2, Vavvas et al. describe issues regarding the normality of lens regeneration and Solebo et al. describe improving outcomes of congenital cataract. We welcome these Comments on our paper.

In our Article3, we did not claim that a completely normal lens was regenerated. Instead, our hypothesis was that current surgical methods for paediatric cataract, namely anterior continuous curvilinear capsulorhexis, may impair the integrity of lens epithelial stem cells (LECs). 5 A P R I L 2 0 1 8 | VO L 5 5 6 | NAT U R E | E 3
