Iatrogenic Cushing’s Syndrome Due to Topical Ocular Treatment Daisuke Fukuhara, MD, PhD,a Toshihiko Takiura, MD, a Hiroshi Keino, MD, PhD, b Annabelle A. Okada, MD, PhD,b Kunimasa Yan, MD, PhDa

Iatrogenic Cushing’s syndrome (CS) is a severe adverse effect of systemic abstract glucocorticoid (GC) therapy in children, but is extremely rare in the setting of topical ocular GC therapy. In this article, we report the case of a 9-year- old girl suffering from idiopathic uveitis who developed CS due to topical ocular GC treatment. She was referred to the ophthalmology department with a complaint of painful eyes, at which time she was diagnosed with bilateral iridocyclitis and started on a treatment of betamethasone sodium phosphate eye drops. Six months after the initiation of topical ocular GC treatment, she was referred to our pediatric department with stunted growth, truncal obesity, purple skin striate, buffalo hump, and moon face. Because her serum cortisol and plasma adrenocorticotropic hormone levels were undetectable, she was diagnosed with iatrogenic CS. After the Departments of aPediatrics, and bOphthalmology, Kyorin doses of topical ocular GC were reduced, the clinical symptoms of CS were University School of Medicine, Mitaka, Tokyo, Japan improved. The fact that the amount of topical ocular GC with our patient was Dr Fukuhara conceptualized and designed the apparently less than that of similar previous cases tempted us to perform study and drafted the initial manuscript; genetic analysis of her NR3C1 gene. We found that our patient had a single Dr Takiura carried out the initial analyses and heterozygous nucleotide substitution in the 3′ untranslated region of the reviewed and revised the manuscript; Drs Keino and Okada reviewed and revised the manuscript; NR3C1 gene, which may explain why she developed CS. However, additional Dr Yan oversaw the data and critically reviewed investigations are required to determine if our findings can be extrapolated the manuscript; and all authors approved the to other patients. In conclusion, clinicians should be aware that even fi nal manuscript as submitted and agree to be accountable for all aspects of the work. extremely low doses of topical ocular therapy can cause DOI: 10.1542/peds.2016-1233 iatrogenic CS. Accepted for publication Oct 10, 2016 Address correspondence to Kunimasa Yan, MD, PhD, Department of Pediatrics, Kyorin University School Iatrogenic Cushing’s syndrome different and depends on genetic of Medicine, 6-20-2 Shinkawa, Mitaka, Tokyo 181- (CS) is a severe adverse effect of and acquired factors. 3 The actions 8611, Japan. E-mail: [email protected] glucocorticoid (GC) therapy in both of GCs are mediated by the GC PEDIATRICS (ISSN Numbers: Print, 0031-4005; Online, children and adults, and is mostly receptor (GR), which acts as a 1098-4275). caused by long-term and abundant ligand-activated transcription Copyright © 2017 by the American Academy of systemic GC administration. Topical factor regulating the transcription Pediatrics GC treatment with nasal drops or of thousands of GC-responsive FINANCIAL DISCLOSURE: The authors have ointment rarely induces iatrogenic genes in a positive or negative indicated they have no fi nancial relationships CS. 1, 2 GC treatment via eye drops fashion. 4 It is currently known that relevant to this article to disclose. is commonly used to treat ocular 4 functionally characterized single- FUNDING: No external funding. inflammatory diseases, including nucleotide polymorphisms (SNPs) POTENTIAL CONFLICT OF INTEREST: The authors juvenile idiopathic uveitis. This of GR (9β, ER22/23EK, BclI and have indicated they have no potential confl icts of interest to disclose. treatment causes local adverse N363S) modulate GC sensitivity. 3, 5 effects, such as glaucoma or cataracts. On the other hand, 1 patient However, iatrogenic CS is extremely exhibiting manifestations of GC To cite: Fukuhara D, Takiura T, Keino H, et al. rare. hypersensitivity caused by a novel Iatrogenic Cushing’s Syndrome Due to Topical The sensitivity to endogenous NR3C1 gene mutation has been Ocular Glucocorticoid Treatment. Pediatrics. 2017; 139(2):e20161233 and synthetic GCs is individually described previously.6 In this article,

Downloaded from www.aappublications.org/news by guest on September 30, 2021 PEDIATRICS Volume 139 , number 2 , February 2017 :e 20161233 CASE REPORT we report the case of a 9-year-old girl on her physical findings and low NR3C1 gene, rs13306585, which is suffering from idiopathic uveitis who endogenous cortisol and ACTH levels, located in the 3′ untranslated region developed CS due to topical ocular we diagnosed her with CS, which was (3′-UTR). No other mutations or GC treatment but had only a single most likely due to excess absorption polymorphisms were identified. nucleotide variation rather than such of ocular GC. Therefore, we started SNPs or mutation. oral methotrexate treatment to reduce the amount of eye drops of DISCUSSION betamethasone sodium phosphate. In this article, we report an extremely CASE REPORT Consequently, her weight gain rare case of iatrogenic CS due to A 9-year-old girl was referred to the decreased while her height increased topical ocular GC treatment. Synthetic ophthalmology department with to a typical SD ( Fig 1D). Six months GCs are one of the most important a complaint of painful eyes. Her after the initiation of methotrexate and widely used drugs in both adults medical history was unremarkable, treatment, she exhibited morning and children. They are used to treat a without any ocular injury or serum cortisol and plasma ACTH variety of disorders, such as allergies surgery. In addition, she did not levels of 10.1 μg/dL and 42.6 pg/mL, and dermatological, inflammatory, complain of any joint pain or bowel respectively, as well as no clinical and autoimmune diseases, because symptoms. She was diagnosed with symptoms of CS ( Fig 1C). they have a striking antiinflammatory bilateral iridocyclitis and started and immunosuppressive effect. All Amplifi cation and Sequencing of the on a treatment of betamethasone clinicians are aware that prolonged NR3C1 Gene sodium phosphate eye drops (0.1% use of systemic GCs at high doses solution). The frequency of eye drops Written informed consent was results in severe adverse effects. was increased to up to 6 times per obtained from the parents of the However, topical GC therapy day due to uncontrollable ocular patient. We performed genotyping uncommonly results in those adverse inflammation. She was referred to of the coding sequence, which effects, and ocular GC therapy is an our pediatric department after 6 comprised exons 2 through 9, extremely rare cause compared with months of treatment, at which time and the intron–exon junctions of nasal and dermatological therapies. 1, 2 she presented with purple skin the NR3C1 gene, including the 4 β There are 2 major potential routes striae. She was not taking any oral or functional GR SNPs (9 , rs6198; to absorb topical ocular GCs into parenteral GCs, nor was she taking ER22/23EK, rs6189, and rs6190; circulation. One is the conjunctiva, any other medication. BclI, rs41423247; and N363S, rs6195) using Sanger sequencing as which is thin and vascular, and Physical examination at our described below. Genomic DNA was facilitates rapid diffusion, and outpatient clinic revealed that she isolated from peripheral blood by the other is the nasal mucosa via was afebrile with a heart rate of 80 using the DNeasy Blood & Tissue Kit the lacrimal drainage system. In beats per minute and (Qiagen, Hilden, Germany, catalog addition, the topically applied GC, of 104/60 mm Hg. Her body weight number 69504). These sequences which penetrates the eye via the and height were 49.6 kg (+1.7 SD) were validated via polymerase cornea, can also be absorbed into and 140.8 cm (–0.3 SD), respectively. chain reaction amplification by circulation. In this process, the lipid- Her physical signs included truncal using TaKaRa Ex Taq (TaKaRa rich corneal epithelium works as a obesity, buffalo hump, moon face, Bio, Inc, Otsu, Shiga, Japan, catalog barrier to intraocular penetration and femoral skin striae, which are number RR001A) according to the for hydrophilic derivatives, such as representative clinical findings of manufacturer’s protocol. Primers betamethasone sodium phosphate. CS ( Fig 1B). These findings were were designed to amplify each locus However, the presence of intraocular not observed the previous year ( Fig (Table 1). The PCR products were inflammation is suggested to 7 1A). She was at a prepubertal stage, analyzed via Sanger sequencing by accelerate the penetration of GCs. defined to be grade 1 on the Tanner using the 3500 Dx Genetic Analyzer Therefore, there is a possibility scale. At this time, she was diagnosed (Applied Biosystems, Foster City, that the concentrations of GCs were with stunted growth ( Fig 1D). CA) according to the manufacturer’s relatively high in the blood in our patient. Laboratory findings revealed no protocol. abnormal data with the peripheral Although the potential systemic blood, biochemistry, or urinalysis. effects of topical GCs are less Her serum cortisol and plasma RESULTS clearly defined, a 50% decrease in adrenocorticotropic hormone (ACTH) We identified a single heterozygous endogenous GC production was levels at 8.00 AM were <1.0 μg/dL A to G nucleotide substitution at observed in male volunteers given and <2 pg/mL, respectively. Based position 3584 in exon 9 of the 0.1% dexamethasone drops

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Downloaded from www.aappublications.org/news by guest on September 30, 2021 PEDIATRICS Volume 139 , number 2 , February 2017 e3 (4.5 mg/day: estimated as TABLE 1 Primer Pairs for Genetic Analysis of the NR3C1 Gene 8 7 prednisolone) for 6 days. McGhee et al Locusa Forward Reverse suggest that intensive topical ocular Exon 2 GTTCGATACACTTTTGCCCT ACAAATAAGGCAGCACAACT GCs, such as 1.0% prednisolone AGAGAACCCCAAGAGTTCAGCA GTTCGACCAGGGAAGTTCAGAGT acetate and 0.1% dexamethasone, Exon 3 TAGGAGGCATCTTTTTGGGA TGCCCAAACTTAACACCATC seem to be absorbed into circulation Exon 4 TTTTGACCTCTCGACCTCAT AACCCATCCAGTTCTTCCTT at a level that can produce adrenal Exon 5 AATGTTTTTAGTCCCACGCA AACATTGACCTCTGCCATTC Exon 6 CATTTTCTGTTAGGGGTGCC GAGTGAGGAAACAACTGCAA suppression. For example, intensive Exon 7 TAGGATGGACTGCTGTGAAA TCTCTGTTTCTGCCATACCT dosing, such as hourly application per Exon 8 ATCCTGCATTTTCTCTTGGC AAGCACTTCCGTACAAAACA day, although totaling an empirical dose Exon 9 AGTTTGTGGATGTTGGTGATAG TGCAAAAATAGGGCGTTAGG of only 6 to 8 mg of prednisolone, AATTCCCCGAGATGTTAGCTGA GACAGATGGGAATGTGAAAATGGG may produce an inherently greater GTGGATGATGGTTGCAAAAGAC GGCAACCTATGAGATTCTGCAC GCATTCATACAGGCAGCGATG TGAATTCTGAAGGGAGCGTGG GC effect than the equivalent orally GGTGCCTAAGAAAACTGCTT TACCCTTTGCAGTAATTGGC administered dose because ocular Bcll SNP AGCAGAAGTACTAACAAAGAGC ATCTGAATTGGGGATGAGGT GC does not undergo first-pass a All exons include their intron–exon junctions. metabolism by the portal system. 7

There have only been 4 pediatric TABLE 2 Reports in the Literature of Iatrogenic Cushing’s Syndrome Caused by Topical Ocular cases previously reported in the Glucocorticoid literature with iatrogenic CS caused Report Sex Age, y Disease Ocular Steroid Dose,a Duration, by topical ocular GC therapy mg/d min 9–12 (Table 2). Case 1 9 Boy 11 Idiopathic 1% predonisolone 12 6 Case 1 was an 11-year-old boy uveitis acetate 10 who developed CS after 6 months Case 2 Boy 2 Congenital 1% predonisolone 6–24 24 corneal acetate of intensive topical ocular GC clouding therapy. Although this patient was Case 31 Boy 7.6 Orbital injury 2% dexamethasone 75 6 similar in age to our patient, the Case 412 Girl 0.5 Congenital 0.1% 7.5 2 dose of GC was slightly higher than cataract dexamethasone ours. Furthermore, this patient Our case Girl 9 Idiopathic 0.1% 3.75 6 uveitis betamethasone was also treated with posterior a sub-Tenon’s injections of 80-mg The dose is estimated to prednisolone. methylprednisolone acetate suspension every 6 weeks for the dose of GC was relatively high. performed genetic analysis of all the 6 months. After 6 months of these The patients in these cases received coding sequences of the NR3C1 gene injections, CS in this patient was topical GCs every 1 or 2 hours, to determine whether our patient aggravated. Case 2 was a 6-year-old whereas our patient received doses had any defects. We found that our girl with diminished growth from every 4 hours. Moreover, our patient patient had a single heterozygous the age of 2 years. This patient had was treated with the lowest dose of A to G nucleotide substitution ′ been taking higher dose of topical topical ocular GC compared with the in the 3 -UTR, rs13306585. This ocular GC compared to our patient. previous cases. variation has been reported in 5 of Case 3 was a 7.5-year-old boy who Not all individuals equally respond 5008 alleles, 4 of which are from had an accidental penetrating orbital to GC therapy, depending on both Japanese populations (http:// www. injury. He had been treated with an functionality and expression of the ncbi. nlm. nih. gov/ variation/ tools/ extremely high dose of topical ocular intracellular GR. Four functional 1000genomes/ ). It is well known GC, estimated as a dose of 75 mg/day GR SNPs and some mutations have that the 3′-UTR plays a critical role in of prednisolone. Finally, case 4 was been identified, and the minor translation and posttranscriptional a 5-month-old boy who developed alleles of the BclI and N363S SNPs gene modification. 15, 16 Indeed, one CS after 3 months of topical ocular and the D401H mutation are of the SNPs in the 3′-UTR of the GC therapy. Because this patient closely associated with increased NR3C1 gene, 9β, is known to cause was much younger than the others, GC sensitivity. 6, 13, 14 Therefore, we relative GC resistance. 4, 14 Based on

FIGURE 1 Continued Appearance of the patient before (A), at the onset of (B), and after (C) revealing clinical manifestations of Cushing’s syndrome. D, Longitudinal growth and weight curves of the patient. The arrows indicate the time points for her photographs in A–C. The dashed arrows indicate time points for the laboratory fi ndings of ACTH and cortisol.

Downloaded from www.aappublications.org/news by guest on September 30, 2021 e4 FUKUHARA et al the results of the gene analysis, we to topical application: toxicity of topical and periocular speculated that rs13306585 may case report and review literature. corticosteroid therapy in an 11-year- influence NR3C1 posttranscriptional Endocrine. 2010;38(3):328–334 old male with posterior uveitis. modifications, leading to increased 3. Quax RA, Manenschijn L, Koper JW, Am J Ophthalmol. 2000;130(2):240–241 GC sensitivity, as was noted in the et al. Glucocorticoid sensitivity in health 10. Steelman J, Kappy M. Adrenal present case. Additional experiments, and disease. Nat Rev Endocrinol. suppression and growth retardation including studies regarding 2013;9(11):670–686 from ocular . epigenetic NR3C1 gene regulation, 4. Nicolaides NC, Galata Z, Kino T, J Pediatr Ophthalmol Strabismus. may enable us to understand why Chrousos GP, Charmandari E. The 2001;38(3):177–178 this patient developed CS. human glucocorticoid receptor: 11. Messina MF, Valenzise M, Aversa molecular basis of biologic function. S, Arrigo T, Luca F. Iatrogenic . 2010;75(1):1–12 Cushing syndrome caused by ocular 5. W üst S, Van Rossum EF, Federenko in a child. BMJ Case ABBREVIATIONS IS, Koper JW, Kumsta R, Hellhammer Rep. 2009;2009 ACTH: adrenocorticotropic DH. Common polymorphisms in 12. Scherrer KS, Weitz M, Eisenack J, hormone the glucocorticoid receptor gene Truffer B, Konrad D. Cushing syndrome CS: Cushing’s syndrome are associated with adrenocortical after bilateral lensectomy. Eur J GC: glucocorticoid responses to psychosocial Pediatr. 2015;174(3):399–401 GR: glucocorticoid receptor stress. J Clin Endocrinol Metab. 13. Oakley RH, Cidlowski JA. The biology 2004;89(2):565–573 SNP: single-nucleotide of the glucocorticoid receptor: new polymorphism 6. Charmandari E, Ichijo T, Jubiz W, signaling mechanisms in health 3′ UTR: 3 ′ untranslated region et al. A novel point mutation in the and disease. J Allergy Clin Immunol. amino terminal domain of the human 2013;132(5):1033–1044 glucocorticoid receptor (hGR) gene 14. Manenschijn L, van den Akker enhancing hGR-mediated gene EL, Lamberts SW, van Rossum REFERENCES expression. J Clin Endocrinol Metab. EF. Clinical features associated 2008;93(12):4963–4968 1. Perry RJ, Findlay CA, Donaldson with glucocorticoid receptor MD. Cushing’s syndrome, growth 7. McGhee CN, Dean S, Danesh-Meyer polymorphisms. An overview. Ann N Y impairment, and occult adrenal H. Locally administered ocular Acad Sci. 2009;1179:179–198 suppression associated with corticosteroids: benefi ts and risks. 15. Jia J, Yao P, Arif A, Fox PL. Regulation intranasal steroids. Arch Dis Child. Drug Saf. 2002;25(1):33–55 and dysregulation of 3'UTR-mediated 2002;87(1):45–48 8. Burch PG, Migeon CJ. Systemic translational control. Curr Opin Genet 2. Tempark T, Phatarakijnirund V, absorption of topical steroids. Arch Dev. 2013;23(1):29–34 Chatproedprai S, Watcharasindhu Ophthalmol. 1968;79(2):174–176 16. Schwerk J, Savan R. Translating the S, Supornsilchai V, Wananukul S. 9. Ozerdem U, Levi L, Cheng L, Song untranslated region. J Immunol. Exogenous Cushing’s syndrome due MK, Scher C, Freeman WR. Systemic 2015;195(7):2963–2971

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Downloaded from www.aappublications.org/news by guest on September 30, 2021 Iatrogenic Cushing's Syndrome Due to Topical Ocular Glucocorticoid Treatment Daisuke Fukuhara, Toshihiko Takiura, Hiroshi Keino, Annabelle A. Okada and Kunimasa Yan Pediatrics originally published online January 19, 2017;

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