Cystine Crystals Research Articles

Examination of corneal deposits in nephropathic cystinosis using in vivo confocal microscopy and anterior segment optical coherence tomography: an age-dependent cross sectional study

BackgroundPresence of corneal cystine crystals is the main ocular manifestation of cystinosis, although controversial findings concerning the corneal layer with the highest density have been reported. The aim of this study was the analysis of the characteristics of crystal arrangement in different corneal layers and the assessment of corneal morphological changes with age.MethodsA cross sectional study was carried out in three children and three adults who had nephropathic cystinosis and corneal cystine depositions. All patients underwent a comprehensive ophthalmological examination including best corrected distance visual acuity, slit-lamp examination, in vivo confocal microscopy and anterior segment optical coherence tomography. An evaluation of the depth of crystal deposits and crystal density in different corneal layers was also performed. Due to the low number of subjects no statistical comparison was performed.ResultsAnterior segment optical coherence tomography images revealed deposition of hyperreflective crystals from limbus to limbus in each patient. Crystals appeared as randomly oriented hyperreflective, elongated structures on in vivo confocal microscopy images in all corneal layers except the endothelium. In children the deposits occurred predominantly in the anterior stroma, while in adults, the crystals were mostly localized in the posterior corneal stroma with the depth of crystal deposition showing an increasing tendency with age (mean depth of crystal density was 353.17 ± 49.23 μm in children and it was 555.75 ± 25.27 μm in adults). Mean crystal density of the epithelium was 1.47 ± 1.17 (median: 1.5; interquartile range: 0.3–2.4). Mean crystal density of the anterior and posterior stroma of children and adults was 3.37 ± 0.34 (median: 3.4; interquartile range: 3.25–3.55) vs. 1.23 ± 0.23 (median: 1.2; interquartile range: 1.05–1.35) and 0.76 ± 0.49 (median: 0.7; interquartile range: 0.4–1.15) vs. 3.63 ± 0.29 (median: 3.7; interquartile range: 3.45–3.8), respectively. Endothelium had intact structure in all cases. Some hexagonal crystals were observed in two subjects.ConclusionsIn vivo confocal microscopy and anterior segment optical coherence tomography confirmed an age-related pattern of crystal deposition. In children, crystals tend to locate anteriorly, while in adults, deposits are found posteriorly in corneal stroma.

CRISPR/Cas9 engineering of albino cystinuria Type A mice.

Cystinuria Type A is a relatively common genetic kidney disease occurring in 1 in 7,000 people worldwide that results from mutation of the cystine transporter rBAT encoded by Slc3a1. We used CRISPR/Cas9 technology to engineer cystinuria Type A mice via genome editing of the C57BL/6NHsd background. These mice are an improvement on currently available models as they are on a coisogenic genetic background and have a single defined mutation. In order to use albinism to track Cas9 activity, we co-injected gRNAs targeting Slc3a1 and tyrosinase (Tyr) with Cas9 expressing plasmid DNA into mouse embryos. Two different Slc3a1 mutational alleles were derived, with homozygous mice of both demonstrating elevated urinary cystine levels, cystine crystals, and bladder stones. We used whole genome sequencing to evaluate for potential off-target editing. No off-target indels were observed for the top 10 predicted off-targets for Slc3a1 or Tyr. Therefore, we used CRISPR/Cas9 to generate coisogenic albino cystinuria Type A mice that could be used for in vivo imaging, further study, or developing new treatments of cystinuria.

Infantile nephropathic cystinosis: Clinical features and outcome

Background: Nephropathic infantile cystinosis, the most common cause of renal Fanconi syndrome, presents in early infancy with impaired growth, polyuria and polydipsia, and progresses to end stage renal disease during the first decade. Diagnosis is based on corneal examination for cystine crystals, leukocyte cystine content and genetic testing of the CTNS gene. Information on clinical features and genotype of Indian children with cystinosis is limited. Methods: We describe clinical features, renal outcomes and genetic variants in the CTNS gene in Indian children with cystinosis. Results: We included 19 patients with cystinosis from 17 families predominantly presenting with poor growth (95%), polyuria (84%) and refractory rickets (74%). Cystine crystals were present in 84%. Fanconi syndrome was common; two had nephrocalcinosis and 9 presented with eGFR <60 ml/ min/ 1.73 m2. Genetic analysis, performed in 11 families, (12 patients) showed 8 variants. Five were reported, pathogenic variants, one was likely pathogenic and two were rare or novel variants of unknown significance. Conclusion: The p.Thr7PhefsTer7 variant, common to five unrelated patients, might be a population hotspot. Eight (42%) patients have been initiated on therapy with cysteamine. Studies with prolonged follow-up are necessary to define renal and extrarenal outcomes and the effect of cysteamine in Indian children.

Impaired auditory sensory memory in Cystinosis despite typical sensory processing: A high-density electrical mapping study of the mismatch negativity (MMN).

Cystinosis, a genetic rare disease characterized by cystine accumulation and crystallization, results in significant damage in a multitude of tissues and organs, such as the kidney, thyroid, eye, and brain. While Cystinosis' impact on brain function is relatively mild compared to its effects on other organs, the increased lifespan of this population and thus potential for productive societal contributions have led to increased interest on the effects on brain function. Nevertheless, and despite some evidence of structural brain differences, the neural impact of the mutation is still not well characterized. Here, using a passive duration oddball paradigm (with different stimulus onset asynchronies (SOAs), representing different levels of demand on memory) and high-density electrophysiology, we tested basic auditory processing in a group of 22 children and adolescents diagnosed with Cystinosis (age range: 6-17 years old) and in neurotypical age-matched controls (N=24). We examined whether the N1 and mismatch negativity (MMN) significantly differed between the groups and if those neural measures correlated with verbal and non-verbal IQ. Individuals diagnosed with Cystinosis presented similar N1 responses to their age-matched peers, indicating typical basic auditory processing in this population. However, whereas both groups showed similar MMN responses for the shortest (450ms) SOA, suggesting intact change detection and sensory memory, individuals diagnosed with Cystinosis presented clearly reduced responses for the longer (900ms and 1800ms) SOAs. This could indicate reduced duration auditory sensory memory traces, and thus sensory memory impairment, in children and adolescents diagnosed with Cystinosis. Future work addressing other aspects of sensory and working memory is needed to understand the underlying bases of the differences described here, and their implication for higher order processing.

CTNS\xa0mRNA molecular analysis revealed a novel mutation in a child with infantile nephropathic cystinosis: a case report

BackgroundCystinosis is an autosomal recessive lysosomal storage disorder characterized by accumulation of cystine in lysosomes throughout the body. Cystinosis is caused by mutations in the CTNS gene that encodes the lysosomal cystine carrier protein cystinosin. CTNS mutations result in either complete absence or reduced cystine transporting function of the protein. The diagnosis of nephropathic cystinosis is generally based on measuring leukocyte cystine level, demonstration of corneal cystine crystals by the slit lamp examination and confirmed by genetic analysis of the CTNS gene.Case presentationA boy born to consanguineous Caucasian parents had the characteristic clinical features of the infantile nephropathic cystinosis including renal Fanconi syndrome (polydipsia/polyuria, metabolic acidosis, hypokalemia, hypophosphatemia, low molecular weight proteinuria, glycosuria, cystine crystals in the cornea) and elevated WBC cystine levels. Initially we performed RFLP analysis of the common in the Northern European population 57-kb deletion of proband’s DNA, then a direct Sanger sequencing which revealed no mutations in the coding part of the CTNS gene. To confirm the diagnosis we performed RT-PCR analysis of total RNA obtained from patient-derived fibroblasts in combination with cDNA sequencing. This revealed the skipping of exon 4 and exon 5 in the CTNS in our patient. Therefore, we detected a novel 9-kb homozygous deletion in the CTNS gene at genomic DNA level, spanning region from intron 3 to intron 5. In order to identify the inheritance pattern of the deletion we analyzed DNA of proband’s mother and father. Both parents were found to be heterozygous carriers of the CTNS mutation.ConclusionsAnalysis of CTNS gene transcript allowed to identify a large homozygous deletion in the patient with infantile nephropathic cystinosis. Mutational detection at RNA level may be an efficient tool to establish the genetic defect in some cystinosis patients.

Ophthalmic Outcome in a Belgian Cohort of Cystinosis Patients Treated with a Compounded Preparation of Cysteamine Eye Drops: Retrospective Analysis.

IntroductionTreatment of the anterior segment problems in cystinosis is challenging as oral cysteamine is ineffective in the treatment of corneal problems because of its avascular structure. Although cysteamine eye drops have been formulated to counter this issue, the stability of cysteamine in these off-licensed formulations and treatment compliance are major problems. The aim of this retrospective study was to determine the efficacy of a compounded preparation of aqueous 0.5% cysteamine eye drops in the management of corneal complications of cystinosis.MethodsData of patients attending the multidisciplinary cystinosis clinic at the University Hospitals Leuven, Belgium between January 2015 and December 2018 were analyzed. All cystinosis patients were treated with the compounded preparation of aqueous 0.5% cysteamine eye drops and oral cysteamine.ResultsA total of 12 patients were treated with the compounded preparation of aqueous 0.5% cysteamine eye drops, of whom 75% were aged > 18 years (n = 9). The mean instillation frequency of the cysteamine eye drops was 3.3 drops/eye per day, and the mean number of hospital visits was two per year. All patients showed photophobia, > 30% corneal infiltration, blepharospasm, eye pain and conjunctival hyperemia during the study period. None of these symptoms improved with treatment with aqueous compounded 0.5% cysteamine eye drops. The corneal cystine crystal score was ≥ 2 in all patients at the last visit.ConclusionTreatment with the compounded preparation of aqueous 0.5% cysteamine eye drops, combined with oral cysteamine, was not effective in reducing corneal cystine crystal deposition and other ocular symptoms in these patients with cystinosis.FundingRecordati Rare Diseases.

Protection of Cystinotic Mice by Kidney-Specific Megalin Ablation Supports an Endocytosis-Based Mechanism for Nephropathic Cystinosis Progression.

Deletions or inactivating mutations of the cystinosin gene CTNS lead to cystine accumulation and crystals at acidic pH in patients with nephropathic cystinosis, a rare lysosomal storage disease and the main cause of hereditary renal Fanconi syndrome. Early use of oral cysteamine to prevent cystine accumulation slows progression of nephropathic cystinosis but it is a demanding treatment and not a cure. The source of cystine accumulating in kidney proximal tubular cells and cystine's role in disease progression are unknown. To investigate whether receptor-mediated endocytosis by the megalin/LRP2 pathway of ultrafiltrated, disulfide-rich plasma proteins could be a source of cystine in proximal tubular cells, we used a mouse model of cystinosis in which conditional excision of floxed megalin/LRP2 alleles in proximal tubular cells of cystinotic mice was achieved by a Cre-LoxP strategy using Wnt4-CRE. We evaluated mice aged 6-9 months for kidney cystine levels and crystals; histopathology, with emphasis on swan-neck lesions and proximal-tubular-cell apoptosis and proliferation (turnover); and proximal-tubular-cell expression of the major apical transporters sodium-phosphate cotransporter 2A (NaPi-IIa) and sodium-glucose cotransporter-2 (SGLT-2). Wnt4-CRE-driven megalin/LRP2 ablation in cystinotic mice efficiently blocked kidney cystine accumulation, thereby preventing lysosomal deformations and crystal deposition in proximal tubular cells. Swan-neck lesions were largely prevented and proximal-tubular-cell turnover was normalized. Apical expression of the two cotransporters was also preserved. These observations support a key role of the megalin/LRP2 pathway in the progression of nephropathic cystinosis and provide a proof of concept for the pathway as a therapeutic target.

Oh cystinosin: let me count the ways!

Nephropathic cystinosis is the most common genetic cause of a renal Fanconi syndrome and results from dysfunction of the lysosomal cystine-transporter protein cystinosin. The multiple organ dysfunctions of affected patients were thought to be related to the defective protein, with cystine crystal formation. However, such crystals were not always present when looked for. More recently, study of the biology of cystinosis has expanded to include many other cellular processes that may be pathogenic in the disease, and now galectin-3 can be added to those identified.

Investigation of Optimum pH and Temperature for &lt;em&gt;In-Vitro&lt;/em&gt; Crystallization of Urinary Cystine

Cystinuria contributes in formation of urinary stones. But, it has been reported that cystinuria is diagnosed when someone experiences with cystine stones. Therefore, early diagnosis of this condition is important. Thus, the objective of the study was to determine the optimum pH and temperature for crystallization of urine cystine in-vitro . Cystinuria solutions were prepared with the concentrations of 40, 60, 70, 75, 80, 90, 100 and 120 mg/dL. The pH of each solution was changed with the addition of acetic acid. Then solutions were exposed to temperature +4°C and 37°C, for 15, 30 and 45min. The sediments were observed microscopically for cystine crystals formation. Then acetone was added to cystinuria with the ratio of cystinuria:acetone, 8:1, 4:1, 2:1 and 1.1 and pH was altered with acetic acid and were subjected to +4 °C and 37 °C, for 15, 30 and 45 minutes and sediment was observed for cystine crystals under the microscope. Cystine crystallization had been occurred in the cystinuria of ≥100 mg/dL at pH 5 at 37 ° C and +4 °C, 30min after the addition of acetic acid whereas with the addition of acetone at cystinuria of ≥75mg/dL at pH 5 in both 37°C and at +4°C, 30min after the addition of acetic acid. The number of cystine crystals per High Power Field (HPF) was highest where cystinuria:acetone was 8:1. The optimum conditions for cystine crystallization is at pH 5, 37 °C and +4 °C, 30min after acidifying with acetic acid at the minimum concentration of 100 mg/dL of cystinuria. With the addition of acetone, at the ratio of cystinuria:acetone 8:1 with minimum concentration of cystinuria of 75 mg/dL. KEYWORDS: Cystine, Crystallization, Acetic acid, Acetone, Temperature, pH

Protective effect of salvianolic acid B against oxidative injury associated with cystine stone formation

The aim of this study was to investigate the role of oxidative stress in cystine crystal formation and whether salvianolic acid B, a natural antioxidant, could prevent cystine-mediated oxidative injury in vivo and in vitro. The levels of oxidative stress and antioxidase activity in cystine stone patients were assessed. Then, the oxidative stress exerted by cystine on human kidney-2 (HK-2) cell viability and biochemical parameters including antioxidase activity and antioxidant protein expression were evaluated, and the protective action of salvianolic acid B was also examined. Finally, salvianolic acid B was tested to determine whether it could prevent or reduce renal crystal formation in Slc7a9 knockout mice. The activity levels of superoxide dismutase (SOD) and glutathione peroxidase (GPx) were decreased, and the amount of malondialdehyde (MDA) was increased in patients with cystine stones compared with people without cystine stones (p < 0.05). Significant reductions in cell viability, antioxidase activity and antioxidant protein expression levels were found in the cystine group compared with controls. However, such oxidative injuries were prevented by salvianolic acid B. In the animal study, loose crystals with white spots were seen in the renal parenchyma, bilateral renal pelvis and bladders in the Slc7a9 knockout group. In contrast, no renal crystals were seen in the control group, and markedly fewer crystals with significantly higher antioxidase activity and diminished oxidative stress were detected in the salvianolic acid B group. Cystine cytotoxicity in vitro and cystine stone formation in vivo were associated with oxidative stress, and salvianolic acid B could protect against cystine stone-induced injury.

A deletion in the Ctns gene causes renal tubular dysfunction and cystine accumulation in LEA/Tohm rats

The Long-Evans Agouti (LEA/Tohm) rat has recently been established as a new rat model of type 2 diabetes. The onset of diabetes mellitus was observed only in male LEA/Tohm rats; however, urinary glucose appeared before the onset of diabetes. To clarify the genetic basis of urinary glucose, we performed genetic linkage analysis using (BN × LEA) F2 intercross progeny. A recessively acting locus responsible for urinary glucose excretion (ugl) was mapped to a 7.9 Mb region of chromosome 10, which contains the cystinosin (Ctns) gene. The Ctns gene encodes the cystine transporter, which transports cystine out of the lysosome and is responsible for nephropathic cystinosis in humans. Sequence analysis identified a 13-bp deletion in the Ctns gene, leading to a truncated and loss-of-function protein, which cause cystine accumulation in various tissues. We also developed a novel congenic rat strain harboring the Ctnsugl mutation on the F344 genetic background. Phenotypic analysis of F344-Ctnsugl rats indicated that the incidence of urinary glucose was 100% in both males and females at around 40 weeks of age, and marked cystine accumulation was observed in the tissues, as well as remarkable renal lesions and cystine crystals in the lysosomes of the renal cortex. Furthermore, treatment with cysteamine depleted the cystine contents in F344-Ctnsugl rat embryonic fibroblasts. These results indicated that the F344-Ctnsugl rat provides a novel rat model of cystinosis, which allows not only a better understanding of the pathogenesis and pathophysiology of cystinosis but will also contribute to the development of new therapies.

The Ocular Status of Cystinosis Patients Receiving a Hospital Pharmacy-Made Preparation of Cysteamine Eye Drops: A Case Series.

IntroductionInfantile nephropathic cystinosis (INC) is an autosomal recessive lysosomal disorder in which patients develop deposits of cystine crystals in their kidneys and corneas from a young age.MethodsWe conducted a retrospective analysis of children with INC seen by ophthalmologists at the Manchester Royal Eye Hospital between 2002 and 2018, to evaluate clinical findings, symptoms and treatment.ResultsTwenty-two children diagnosed with INC from age 0 (prenatally) to 11 years were assessed. All evaluable patients had corneal cystine crystal deposits, and 15 had mild to moderate photophobia. Ten patients had other ocular conditions including blepharitis/chalzion (n = 6), swollen optic nerve (n = 3), punctate epitheliopathy (n = 3), corneal scarring (n = 1),and elevated intraocular pressure (n = 2). Confocal imaging identified nerve abnormalities in two patients (enlarged corneal nerve + abnormal-looking tortuous nerves in one patient and beaded nerves in the sub-basal plexus in the other), both of whom had significant crystal deposition in the anterior stroma. Visual acuity was relatively unaffected. All 22 patients were receiving oral cysteamine, and 21 were applying cysteamine eye drops (galenic preparation of 0.55% concentration, compounded by a hospital pharmacy). Recommended application frequency was at least eight times per day in all patients with dosing information available.ConclusionsThis case series of patients with INC highlights the consistent pattern of corneal cystine crystal deposition, which is universally present from a young age in this condition, and the high incidence of photophobia even in young children. Corneal manifestations of INC persisted despite frequent administration of the hospital pharmacy-made eye drop preparation. Reasons for this lack of efficacy may include the lag period between diagnosis and first prescription of cysteamine eye drops and the difficulty in maintaining rigorous compliance with this treatment. In addition, the challenge for patients of maintaining optimal storage conditions may adversely affect the stability and efficacy of cysteamine within this preparation.FundingEditorial assistance was funded by Orphan Europe Ltd.

Cystinuria: genetic aspects, mouse models, and a new approach to therapy.

Cystinuria, a genetic disorder of cystine transport, is characterized by excessive excretion of cystine in the urine and recurrent cystine stones in the kidneys and, to a lesser extent, in the bladder. Males generally are more severely affected than females. The disorder may lead to chronic kidney disease in many patients. The cystine transporter (b0,+) is a heterodimer consisting of the rBAT (encoded by SLC3A1) and b0,+AT (encoded by SLC7A9) subunits joined by a disulfide bridge. The molecular basis of cystinuria is known in great detail, and this information is now being used to define genotype-phenotype correlations. Current treatments for cystinuria include increased fluid intake to increase cystine solubility and the administration of thiol drugs for more severe cases. These drugs, however, have poor patient compliance due to adverse effects. Thus, there is a need to reduce or eliminate the risks associated with therapy for cystinuria. Four mouse models for cystinuria have been described and these models provide a resource for evaluating the safety and efficacy of new therapies for cystinuria. We are evaluating a new approach for the treatment of cystine stones based on the inhibition of cystine crystal growth by cystine analogs. Our ongoing studies indicate that cystine diamides are effective in preventing cystine stone formation in the Slc3a1 knockout mouse model for cystinuria. In addition to crystal growth, crystal aggregation is required for stone formation. Male and female mice with cystinuria have comparable levels of crystalluria, but very few female mice form stones. The identification of factors that inhibit cystine crystal aggregation in female mice may provide insight into the gender difference in disease severity in patients with cystinuria.

The effect of selenium supplementation on cystine crystal volume in patients with cystinuria.

Background: Cystinuria as an autosomal recessive sickness is a relatively rare disease. Formation of cystine stones indicates cystinuria. Few studies are considered the cysteine crystal volume in management of cystinuria. Selenium may inhibit organization of crystal stone, growth and stone aggregation. Since the role of selenium on inhibition of cystine crystal formation is not known, the aim of this study was to evaluate the effect of selenium supplementation on cystine crystals volume in patients with cystinuria.Materials and methods: This double-blind clinical trial study was conducted on 48 patients in Al-Zahra hospital, Isfahan, Iran from 2015 to 2017. These patients received selenium (200 mg/ daily) for 6 weeks. The urine crystal volume was evaluated before and after treatment. Data were entered SPSS and analyzed by Paired sample T test, Spearman and Pearson coefficient correlation. P- value < 0.05 was considered significant.Results: In current study, mean cystine crystal volume before and after treatment was 6787.4 ± 11902.6 and 3110.9 ± 7225.4, respectively. Significant difference was observed before and after treatment in terms of cystine crystal volume (p < 0.001). No relation was observed between the mean cystine crystal volume with sex, age and type of medical procedures (p > 0.05).Conclusion: In this study, selenium treatment affected cystine crystal volume. It seems that selenium had the potential value to alleviate the volume of cystine crystal. Therefore, since reducing of cystine crystal volume decreases crystal formation, selenium may be effective to cure patients with cystinuria. However, age, sex and type of medical procedures did not affect cysteine crystal volume.

In vitro and ex vivo implantation of cystine crystals and treatment by contact lens

Cystinosis is a rare genetic disorder order affecting the transport of the amino acid cystine across the lysosomal membrane, leading to crystal formation throughout the body. The formation of these crystals in the cornea leads to photophobia and other ocular complications. The current treatment is to use eye drops of a thiol called cysteamine, delivered at a rate about 8–12 drops per day due to very low bioavailability intrinsic to eye drops. This paper examines an alternative and possibly more effective method of using contact lenses, which are suggested to have a bioavailability of around 50%, to deliver cysteamine to the cornea. This paper focuses on first designing in vitro, ex vivo and in vivo rabbit models of cornea containing cystine crystals and testing for feasibility of crystal dissolution by using contact lenses. In each model, cystine crystals were formed by loading cysteine into the cornea followed by oxidation to form cystine crystals. In some cases, cystine was directly transported into the cornea to form the crystals. The in vitro model—using a cystine-laden hydroxyl ethyl methacrylate hydrogel in the shape of a cornea—demonstrated an effectiveness of roughly 52% of loaded cysteamine reaching the “hydrogel cornea.” The ex vivo model—using rabbit cadaver eyes impregnated with cystine crystals—again showed a near 50% bioavailability—using 27 ± 2 lenses to lead to crystal dissolution when 13 lenses were needed if 100% bioavailability could be reached. The in vivo studies showed that exposing the cornea of a normal animal can lead to crystal formation but the crystals dissolve rapidly likely due to clearance through the aqueous humor. Thus, efficacy of eluting contacts at dissolving crystals could not be tested in vivo. These results suggest that cysteamine loaded contact lens could transport cysteamine into the cornea at higher bioavailability compared to the eye drops and thus be potentially effective at treating cystinosis.

Latest Clinical Approaches in the Ocular Management of Cystinosis: A Review of Current Practice and Opinion from the Ophthalmology Cystinosis Forum.

Cystinosis, a rare autosomal recessive disease caused by intracellular cystine accumulation, occurs in an estimated 1/100,000–200,000 live births. Ocular non-nephropathic cystinosis is typically diagnosed during adulthood, when patients present with corneal crystal deposition and no systemic involvement. Due to the rarity of the condition, diagnosis is often delayed and can have a significant impact on the overall prognosis of the disease. Early diagnosis is therefore imperative to ensure successful treatment and improve quality of life, as most of its clinical manifestations can be prevented or delayed. Early detection strategies and practical approaches for the ocular management of cystinosis were discussed during the Ophthalmology Cystinosis Forum, a 1-day meeting held in Berlin, Germany during June 2017. Recommendations for early detection comprise ophthalmic assessment, including self- and clinician-assessed recording of photophobia, and visual acuity, slit-lamp examination and tonometry ophthalmic examinations. In vivo confocal microscopy and anterior segment optical coherence tomography were highlighted as valuable techniques in evaluating cystine crystals in the cornea, in vivo and non-invasively. The mainstay of ocular cystinosis treatment is the cystine-depleting aminothiol cysteamine. Indeed, early treatment with and strict adherence to cysteamine therapy has a considerable impact on the long-term prognosis of ocular cystinosis. In rare diseases such as ocular cystinosis, standardised guidelines and recommendations for detection, patient care and follow-up assessments are essential. Such guidelines provide a support tool for healthcare professionals caring for ocular cystinosis patients. Multidisciplinary teams (MDTs) are essential for delivering gold standard care and improving quality of life for patients and their families. This review paper highlights current early detection policies, clinical treatment strategies and practical approaches for the ocular management of cystinosis, including implementing a cystinosis MDT. Additionally, discussions of the Ophthalmology Cystinosis Forum held in 2017 are summarised.FundingOrphan Europe.Plain Language SummaryPlain language summary available for this article.

Potential role of stromal collagen in cystine crystallization in cystinosis patients

Cystinosis is a genetic disease that leads to the accumulation of intracellular cystine crystals in all organs including cornea due to the loss of cystine efflux transporters in the lysosome of the cells. While the mechanism for formation of intracellular cystine is well understood for most organs, it does not explain many observations for crystal accumulation in the cornea of cystinosis patients. First, the crystals in cornea are extracellular and needle-like with several hundred microns length which is in sharp contrast with the rectangular or hexagonal crystals found throughout other organs. Second, these crystals are arranged parallel to the stromal collagen, which is a unique to the cornea. Third, crystal growth in the cornea reaches a saturation point after where no further crystallization occurs. We propose a hypothesis supported by in vitro and ex vivo data to explain these observations. We hypothesize that the stroma crystals form extracellularly due to the ionic interactions between the cystine diffusing into the eye and collagen fibrils present in the stroma. We examine cystine crystal growth both with in vitro polymer solutions and ex vivo in rabbit cadaver eyes to show that negatively charged polymers lead to the formation of more cystine precipitation in aqueous solution and that needle-like cystine crystal formation can be observed only in presence of certain polyelectrolytes including collagen. This proposed mechanism explains many of the yet unanswered questions but it needs further support from in vivo studies. The improved understanding could lead to improved treatment of corneal cystinosis.

Novel approaches for improving stability of cysteamine formulations

Cystinosis is a genetic disorder that leads to the formation of cystine crystals in many organs in the body including cornea. Ocular manifestation of this disease is treated by eye drops of cysteamine which can easily oxidize into its disulfide cystamine. The rapid oxidation limits the shelf life as well the duration during which the drug can be used after opening the eye drop bottle. We evaluate two approaches of preventing the oxidation of cysteamine with the goal of increasing the time of use after opening the bottle to one month. The first approach integrates antioxidants such as catalase enzyme and vitamins C and E into the aqueous solution. Results show that catalase is the most effective additive as it decreases the oxidation rate by 58%, which on its own is not sufficient to reach targeted one month stability. The second approach focuses on incorporating diffusion barriers to prevent oxygen from reaching the cysteamine solution. This was accomplished by two methods: formulation of a hydrophobic layer which floats on the surface of the aqueous solution and integration of OMAC® oxygen-resistant material into the eye drop bottle. Both methods delay the onset of cysteamine degradation and decrease the rate of degradation. In particular, an eye drop bottle with three layers of OMAC® has less than 10% degradation after one month of opening the bottle and withdrawing a drop each day. By integrating all three methods, we designed a system where >90% of cysteamine remains in the active form for 70 days after opening the bottle. In addition, we examine the use of OMAC® as heat-sealed pouches for storage of cysteamine eye drop bottles during packaging to eliminate the need for the current approach of freezing the formulation during shipping. The results show that such heat-sealed pouches would keep cysteamine stable for over one year at ambient conditions.

Nephropathic Cystinosis: Symptoms, Treatment, and Perspectives of a Systemic Disease.

Cystinosis is a rare autosomal recessive lysosomal storage disorder caused by mutations in the CTNS gene. Main dysfunction is a defective clearance of cystine from lysosomes that leads to accumulation of cystine crystals in every tissue of the body. There are three different forms: infantile nephropathic cystinosis, which is the most common form, juvenile nephropatic, and non-nephropathic cystinosis. Mostly, first symptom in infantile nephropathic cystinosis is renal Fanconi syndrome that occurs within the first year of life. Another prominent symptom is photophobia due to corneal crystal deposition. Cystine depletion therapy with cysteamine delays end-stage renal failure but does not stop progression of the disease. A new cysteamine formulation with delayed-release simplifies the administration schedule but still does not cure cystinosis. Even long-term depletion treatment resulting in bypassing the defective lysosomal transporter cannot reverse Fanconi syndrome. A future perspective offering a curative therapy may be transplantation of CTNS-carrying stem cells that has successfully been performed in mice.

Cysteamine hydrochloride eye drop solution for the treatment of corneal cystine crystal deposits in patients with cystinosis: an evidence-based review.

Cystinosis is a rare, autosomal recessive disorder leading to defective transport of cystine out of lysosomes. Subsequent cystine crystal accumulation can occur in various tissues, including the ocular surface. This review explores the efficacy of cysteamine hydrochloride eye drops in the treatment of corneal cystine crystal accumulation and its safety profile.