Ophthalmic Genetics

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Chorioretinal dystrophy, hypogonadotropic hypogonadism, and cerebellar ataxia: Boucher- Neuhauser syndrome due to a homozygous (c.3524C>G (p.Ser1175Cys)) variant in PNPLA6

Mustafa Doğan, Recep Eröz & Emrah Öztürk

To cite this article: Mustafa Doğan, Recep Eröz & Emrah Öztürk (2021) Chorioretinal dystrophy, hypogonadotropic hypogonadism, and cerebellar ataxia: Boucher-Neuhauser syndrome due to a homozygous (c.3524C>G (p.Ser1175Cys)) variant in PNPLA6 gene, Ophthalmic Genetics, 42:3, 276-282, DOI: 10.1080/13816810.2021.1894461 To link to this article: https://doi.org/10.1080/13816810.2021.1894461

Published online: 02 Mar 2021.

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Full Terms & Conditions of access and use can be found at https://www.tandfonline.com/action/journalInformation?journalCode=iopg20 OPHTHALMIC GENETICS 2021, VOL. 42, NO. 3, 276–282 https://doi.org/10.1080/13816810.2021.1894461

RESEARCH REPORT Chorioretinal dystrophy, hypogonadotropic hypogonadism, and cerebellar ataxia: Boucher-Neuhauser syndrome due to a homozygous (c.3524C>G (p.Ser1175Cys)) variant in PNPLA6 gene Mustafa Doğan a, Recep Eröz b, and Emrah Öztürk c aDepartment of Medical Genetics, Malatya Turgut Özal University Medical Faculty, Malatya, Turkey; bDepartment of Medical Genetics, Duzce University Medical Faculty, Duzce, Turkey; cDepartment of Ophthalmology, Malatya Training and Research Hospital, Malatya, Turkey

ABSTRACT ARTICLE HISTORY Purpose: The current study aims to raise awareness of Boucher – Neuhauser syndrome (BNHS) that occurs Received January 01, 2021 as a rare phenotype due to biallelic pathogenic variants in the PNPLA6 gene. Revised January 30, 2021 Methods: Detailed family histories and clinical data were recorded. Whole exome sequencing was Accepted February 15, 2021 performed and co-segregation analysis of the family was done by sanger sequencing. Also, review of 28 KEYWORDS molecularly confirmed patients with BNHS from the literature was evaluated. Boucher-Neuhauser Results: We identified a missense homozygous variant (c.3524 C > G (p.Ser1175Cys)) in the PNPLA6 gene, syndrome; PNPLA6 gene; which explains the phenotype of the patient and neurologic, ophthalmologic, endocrine, and genetic chorioretinal dystrophy evaluations established a diagnosis of BNHS. Symptoms, ethnicity, clinical and genetic findings of 28 molecularly confirmed patients with BNHS from the literature were also presented. Conclusion: We present the main findings of a Turkish family with BNHS together with detailed clinical and genetic profiles of patients diagnosed as BNHS that have been molecularly confirmed in the literature so far.

Background Gordon Holmes syndrome, spastic paraplegia, retinal degeneration, Oliver-McFarlane syndrome, and Laurence- PNPLA6 (Patatin-like domain containing 6) Moon syndrome. The presence of similar symptoms in consists of 34 exons on 19p13.2 and encodes associated phenotypes makes clinical diagnosis difficult, so a phospholipase called neuropathy target careful clinical evaluation is required to differentiate (NTE) (NM_001166111.2). It is a highly conserved phospholi­ PNPLA6 phenotypes (6). BNHS (MIM215470) is a rare pase that deacetylates intracellular phosphatidylcholine to pro­ syndrome characterized by the triad of early-onset autoso­ duce glycerophosphocholine and is involved in the mal recessive slowly progressive ataxia, hypogonadotropic development of axons in the brain, eye, and pituitary gland hypogonadism, and chorioretinal degeneration (7,8). Since (1). Hufnagel et al. examined PNPLA6 expression during both BNHS was first reported in the ophthalmic literature, to the human central nervous system and retinal development and best of our knowledge, a total of 27 patients with biallelic revealed PNPLA6 expression in embryonic tissues of the devel­ pathogenic variants in the PNPLA6 gene have been oping brain, neural retina, retinal pigment epithelium, choroid, reported worldwide (9). In the current study, we present anterior and posterior pituitary, cerebellum, and ventricular the clinical and genetic findings of a Turkish family with regions (2). NTE as an enzyme involved in lipid metabolism BNHS together with detailed clinical and genetic profiles of that is critical to the stability of cell membranes. Mutations in patients with BNHS that have been molecularly confirmed this region can disrupt the enzymatic activity of the PNPLA6 in the literature so far. protein, which can impair the formation of synaptic connec­ tions in neuronal networks and intercellular interactions with cellular organelles (3). When a mutation occurred, it can cause Material and methods a spectrum of disorders that range from intellectual disabilities to ataxia in humans (4). However, little has been revealed about Individual informed consents for medical examinations, geno­ the structure–function relationships between the domains of mic analyses, and case presentations were obtained from the the NTE protein, its enzymatic activity, and interactions with patient and his parents; the procedures adhered to the cellular organelles (5). Symptoms and clinical findings of the Declaration of Helsinki and the relevant studies approved by patients with Boucher-Neuhäuser syndrome(BNHS) are given the institutional review board. A comprehensive clinical history in Table 1. was taken, and neurological, ophthalmologic, as well as endocri­ Pathogenic variants detected in the PNPLA6 gene have nologic examinations were performed on all members of the been associated with very rarely phenotypes such as BNHS, family. The family pedigree has been shown in Figure 1.

CONTACT Mustafa Doğan [email protected] Department of Medical Genetics, Malatya Malatya Turgut Özal University Training and Research Hospital, Malatya, Turkey. © 2021 Taylor & Francis Group, LLC OPHTHALMIC GENETICS 277

Table 1. Symptoms and clinical findings of the patients with Boucher-Neuhäuser performed with Cytoscan Optima Assay (Affymetrix). Due to syndrome. the absence of any genetic anomalies in these tests, whole exome N(%) sequencing was performed by capture of the coding regions and Night Blindness 7 (%25) splice sites of targeted using the Illumina SureSelect V6 Initial eyes complaints Strabismus and nystagmus 1(%3.6) Vision loss 14(50%) Exome kit. After library enrichment and quality control, the Not available 6 (%21.4) samples were sequenced using the Illumina HiSeq4000 instru­ First Symptoms Ataxia 8(%28.6) ment with 100bp paired-end reads at an average sequencing Delayed puberty 9(%32.1) Night Blindness 4(%14.3) depth of 100x. Raw reads were quality trimmed with Vision loss 7(%25) Trimmomatic. Surviving high quality reads were mapped to Chorioretinal dystrophy Yes 27(%96.4) reference (hg19) with using BWA (Burrows– No 1(%3.6) Night Blindness Yes 8(%28.6) Wheeler Alignment Tool). Picard, a command line java tool to No 4(%14.3) handle SAM and BAM files, was used to mark duplicate reads Not available 16(57.1%) with appropriate flags. GATK (Genome Analysis Toolkit), Cranial MRI Atrophy cerebellum 20 (%71.4) Normal 3 (%10.7) a software package to analyze high-throughput sequencing Not available 5 (17.9%) data, were used to call SNPs and indels. Following modules of Cerebellar ataxia Yes 24 (85.7%) GATK software package were used in respective order; Realigner No 4 (14.3%) Hypogonadotropic hypogonadism Yes 28 (100%) Target Creator, IndelRealigner, Base Recalibrator, Print Reads, No 0 (0%) Haplotypecaller, Select Variants, Variant Filtration, Combine Intellectual disability Yes 8 (28.6%) Variants to call and filter SNPs and indels. Annotation of No 20 (71.4%) Trichomegaly Yes 0 (0%) detected variants was performed using Illumina BaseSpace No 28 (100%) Variant Interpreter, InterVar, Franklin, VarSome, ClinVar, Short Stature Yes 8 (28.6%) OMIM, and Pubmed. Variants with a frequency higher than No 9(%32.1) Not available 11 (39.3%) 0.5% were filtered out. dbNSFP (contains SIFT, PolyPhen-2, Zygosity Homozygote 6 (21.4%) LRT, Mutation Taster) was used to predict the pathogenicity Compound heterozygote 22 (78.6%) about the deleteriousness of variants. Furthermore, genes associated with eye diseases (listed below) were reevaluated. RB1, PGAP1, CLPB, IFT172, RAX, Genomic DNA was extracted from peripheral leukocytes of fresh SLC4A4, GJA1, TBK1, GNAT2, GJA3, CSPP1, CYP1B1, blood samples taken from the index patient and karyotype HOXA1, LRAT, GJA8, GNAT1, GPR179, PRKCG, MECR, analysis was performed. Array comparative genomic hybridiza­ CACNA2D4, RBP4, RBP3, IFT81, TMEM126A, HOXB1, tion of the patient was done and microarray study was FREM1, ATF6, FREM2, OPTN, RD3, CFH, ABCB6, PLA2G5,

Figure 1. Pedigree of the Family with autosomal recessive Boucher – Neuhauser syndrome (BNHS). The pedigree comprises 18 members with three generations (8 females, 10 males). III-2 is proband and both II-4 and II-7 are the parents. III-1 and III-3 are siblings of the proband (a). The brain imaging of index case: The cerebellum’s severe atrophy can be seen on sagittal T2-weighted and axial T1-weighted MRI (b). Sanger sequencing traces of the Family showing the presence of the c.3524 C > G(p. Ser1175Cys) on the PNPLA6 gene in a homozygous state in the proband (c), heterozygous state in father (d), heterozygous state in mother (e), heterozygous state in sister (f) and normal in the brother (g). An illustration of PNPLA6 gene structure along with relative positions of all known pathogenic variants and occurring amino acid changes (including our c.3524 C > G(p.Ser1175Cys) missense variant in exon 30 of PNPLA6 gene) in their distribution across the protein (h). 278 M. DOĞAN ET AL.

RDH12, RDH11, TTPA, ADAMTS18, HESX1, PROM1, PLK4, PDE6C, PDE6B, PDE6A, IGBP1, CRYGS, TIMM8A, LIM2, JAG1, FZD4, ABCA4, SLC16A12, NR2F1, BBIP1, GNPTG, NAA10, EYA1, PDE6H, PDE6G, PCARE, LCA5, NYX, GJB2, ARL2BP, GJB6, MKS1, PPT1, UNC119, CHM, GLI2, CRYGC, SLC4A7, CRYGD, PIKFYVE, CRYGB, ERCC1, PQBP1, SHH, TCTN3, TCTN2, C1QTNF5, TCTN1, FLVCR1, ERCC2, ERCC5, ERCC6, VIM, HCN1. TIMP3, TEAD1, OFD1, CHST6, WDR36, ELOVL4, MYOC, Rare variants were classified according to the ACMG/AMP CRPPA, IFT140, ROM1, PAX6, FAM126A, PAX2, COL2A1, variant interpretation framework (10). Sanger confirmation of ZEB1, PXDN, RARB, PIGL, CRYAA, RGR, CRYAB, CRB1, the detected variant with the family members was done by GRN, OAT, CTDP1, COL11A1, NTF4, TMEM67, CHN1, using an ABI PRISM 3130 autosequencer (Applied RHO, GCNT2, SPP2, MFN2, ZNF469, HARS, JAM3, GSN, Biosystems). For this study; 17 publications reporting 27 WHRN, AGK, WDR19, AGBL5, TTLL5, AGBL1, NMNAT1, patients with a molecularly confirmed diagnosis of BNHS SMOC1, ALMS1, CEP41, GUCA1B, GUCA1A, BEST1, were evaluated. The main findings of the cases described in ABHD12, RIMS1, CDH3, EFEMP1, SALL2, TUBB3, SALL4, the literature and our own index case (i.e. a total of 28 cases) CEP250, NEK2, MTPAP,PITX2, PITX3, SLC25A46, MITF, are summarized in Table 2. KERA, LEMD2, MAF, DGKQ, MAK, ADAM9, FSCN2, SNRNP200, FTL, COL17A1, ARL6, ARL3, TACSTD2, Statistical analysis SPATA7, NEUROD1, PORCN, LOXHD1, ATXN7, TTC21B, PGK1, NDP, SLC38A8, ZNF423, IMPG1, IMPG2, STRA6, All data were evaluated using SPSS 22.0 (SPSS, Inc., Chicago, WFS1, SRD5A3, SLC4A11, RAX2, BFSP2, BFSP1, LRIT3, Illinois, USA) statistical software. We used descriptive statistics TUBGCP6, TPP1, REEP6, TUBGCP4, ACO2, CEP164, for all data of the study. Descriptive statistics for variables were POMT1, TSPAN12, RLBP1, KIAA1549, OPA1, ZNF408, given as mean, standard deviation, n, and percentage. OPA3, IDH3B, CNGA1, CNGA3, CEP290, IDH3A, IQCB1, UNC45B, ADGRV1, VPS13B, FOXL2, RP1, MFRP, RRM2B, Results RP2, DNAJC5, CNGB3, ZNF513, RP9, CNGB1, SLC24A1, PRPS1, CANT1, PRCD, USH1C, CRX, LRP5, SEMA3E, The proband (Figure 1a, III: 2) is a 30-year-old male with TULP1, TTR, ARL13B, OR2W3, NHS, IARS2, RAB28, hypogonadotropic hypogonadism was referred to our medical DRAM2, CNNM4, TUB, SEMA4A, KCNJ13, MERTK, genetics clinic for genetic diagnosis. In general examination, USH2A, GFER, RPGRIP1, FRAS1, USH1G, IMPDH1, the patient height was 163 cm and weighted 71 kg. According MAB21L2, RAB18, BCOR, TRIM32, MMACHC, C12orf57, to the story taken from the parents, his prenatal history was PANK2, CISD2, ACTB, PEX26, SOX2, RS1, KIF21A, HMCN1, unremarkable and no pregnancy complications were reported. SIL1, C12orf65, EYS, SOX5, ACVR1, KRT3, P3H2, ADGRA3, He was the second child of consanguineous parents of Turkish MYO7A, OVOL2, MIP, COL8A2, WDPCP, EPHA2, CERKL, descent and he has a healthy sister and a brother. Neuromotor MTTP, PEX11B, PCDH15, TRPM1, INPP5E, DHX38, BBS2, retardation was identified and there were impaired cognitive BBS1, TREX1, SLC33A1, CPLANE1, GDF3, GDF6, RPE65, functions. The patient had complaints of balance disorder and COL9A1, ITM2B, VSX1, VSX2, INVS, COL18A1, ROBO3, difficulty in walking that started around the age of 5 years old. FOXE3, TENM3, SLC7A14, HCCS, KIF11, SIPA1L3, AUH, The tandem gait was incompetent, ataxic and it was noted that SH3PXD2B, BBS9, CTSF, BBS7, CC2D2A, CTSD, BBS5, BBS4 these complaints gradually increased. On neurological exam­ PCYT1A, KIF7, TOPORS, BBS10, BBS12, PITPNM3, CLDN19, ination, there were jerk nystagmus and cerebellar ataxia but RDH5, KIZ, HMX1, SDCCAG8, YAP1, FOXC1, GUCY2D, there were no neurological defects as peripheral neuropathy or CRYBA2, CRYBA1, CRYBA4, EMC1, CTNNA1, HSF4, spasticity. It was observed that T2 flair hyperintense signal PEX16, PEX19, CRYBB1, CRYBB3, CRYBB2, KRT12, PEX10, increases in the posterior leg of the internal capsule and there PEX12, VAX1, PEX13, PEX14, RP1L1, TTC8, KLHL7, was atrophic dilatation in the cerebellar foils in cranial MR. RAB3GAP2, CTNNB1, PRPF31, B9D1, B9D2, RTN4IP1, He was diagnosed with hypogonadotropic hypogonadism in PNPLA6, RAB3GAP1, PRDM5, CIB2, NRL, MSMO1, LOXL1, 15 years ago at the age of 15 years. He was treated intermit­ GRIP1, CAPN5, RGS9, SBF2, PHYH, OCRL, RPGRIP1L, tently with testosterone, human chorionic gonadotropin but TRNT1, CFAP410, PRSS56, LZTFL1, KCTD7, DHDDS, no significant growth was achieved in the testes and penis. PRPF4, PRPF6, FAM161A, TBC1D20, RGS9BP, COL4A1, Gonadotropin-releasing hormone (GnRH) stimulation test TMEM138, PRPF3, CHMP4B, LMX1B, AIPL1, UBIAD1, was performed in the patient whose luteinizing hormone CABP4, LTBP2, ADIPOR1, CACNA1F, PRPF8, GRK1, (LH), follicle-stimulating hormone (FSH), and testosterone KCNV2, NPHP1, CDH23, NPHP3, NPHP4, OTX2, PDZD7, levels were below normal, but there was no obvious increase TFAP2A, ACBD5, CAV1, TDRD7, BMP7, TMEM231, in FSH and LH. In scrotal USG, both testicles were atrophic PHOX2A, C8orf37, BMP4, AHI1, AMACR, COL5A1, PRPH2, (right testis 14.5x11x22, left testis 14x9x20 mm) and spermio­ TMEM237, MIR184, IFT27, GNB3, CRYL1, TCF4, TGFBI, gram result was azospermic. GALK1, RERE, NGLY1, CHD7, NR2E3, HK1, RPGR, MFSD8, FYCO1, SIX6, ZIC2, GRM6, CA2, CYP4V2, SIX3, CA4, Ophthalmologic assessments ABCD1, POLG, TGIF1, HGSNAT, MKKS, ABCC6, ARHGEF18, PEX2, PEX1, POMGNT1, LSS, DCN, ALDH1A3, A 30-year-old Turkish male patient, who defined his first eye PEX7, VCAN, PEX3, TMEM216, PEX6, CDHR1, PEX5, SAG, complaints as strabismus and nystagmus beginning around the NDUFS1, CLRN1, DTHD1, MVK, LAMA1, LCAT, SPG7, age of 6, was evaluated for impaired vision. Blurred vision and CLN8, CLN6, CLN5, ATOH7, CLN3, PDE6D, POC1B, night blindness were first noted around the age of 18. The OPHTHALMIC GENETICS 279

Table 2. Ethnicity, clinical and genetic findings of patients with Boucher-Neuhäuser syndrome. PNPLA6 gene Variant One Variant Two Age/ S Eth Clin Fin cDNA (AA) Dmn VT cDNA (AA) Dmn VT RN 30/M Tur HH, CD, At, SS, c.3524 C > G(p.Ser1175Cys) EST Mis c.3524 C > G(p.Ser1175Cys) EST Mis PST mID 28/M Chn HH, CD, At c.3547 C > T(p.Arg1183Trp) EST Mis c.1841 + 3A>G cNMP Splc (9) (r.1841_1842ins1841 + 1_1841 + 185) 28/F Chn HH, CD c.3436 G > A(p.Ala1146Thr) EST Mis c.2212–10A>G cNMP Splc (9) 11/M Chn HH,CD c.2266 C > T(p.Gln756*) - Non c.3436 G > A(p.Ala1146Thr) EST Mis (9) 25/F Chn/ HH, CD, SS c.1238_1239insC(p.Leu414Serfs*28) - Ins c.3130A>G(p.Thr1044Ala) EST Mis (9) Cau 39/M Chn HH, CD c.3386 G > T(p.G1129V) EST Mis c.3534 G > C(p.W1178C) EST Mis (11) 32/M Ger HH, CD, At c.288 T > G(p.Y96*) - Non c.865 C > G(p.R289G) cNMP Mis (12) 42/M Ger HH, CD, ataxia, SS c.343–2A > T - Splc c.4075 C > T(p.R1359W) - Mis (12) 44/F Eur HH, CD, At, SS, c.2212–1 G > C(p.V738Qfs*98) cNMP Splc c.3328 G > A(p.V1110M) EST Mis (6) mID 42/F Eur HH, At, mID c.2212–1 G > C(p.V738Qfs*98) cNMP Splc c.3328 G > A(p.V1110M) EST Mis (6) 56/F Eur HH, CD, At, mID c.3173 C > T(p.T1058I) EST Mis c.3173 C > T(p.T1058I) EST Mis (6) 55/M Eur HH, CD, At, mID c.3173 C > T(p.T1058I) EST Mis c.3173 C > T(p.T1058I) EST Mis (6) 53/F Eur HH, CD, At, mID c.3173 C > T(p.T1058I) EST Mis c.3173 C > T(p.T1058I) EST Mis (6) 48/F Eur HH, CD, At, mID c.3173 C > T(p.T1058I) EST Mis c.3173 C > T(p.T1058I) EST Mis (6) 61/M Eur HH, CD, At c.1732 G > T(p.G578W) cNMP Mis c.3197 T > C(p.F1066S) EST Mis (6) 57/M Eur HH, CD, At c.1732 G > T(p.G578W) cNMP Mis c.3197 T > C(p.F1066S) EST Mis (6) 26/M Eur HH, CD, At c.3134 C > T(p.S1045L) EST Mis c.3365 C > T(p.P1122L) EST Mis (6) 28/M Jap HH, CD, At c.3523_3524insTGTCCG EST Mis c.2923A > G p.T975A EST Mis (13) (p.1175_1176insVS) 48/F Jap HH, CD, At c.3534 G > C(p.W1178C) EST Mis c.3534 G> Cp.W1178C EST Mis (14) 59/F Amr HH, CD, At c.3519 C > G(p.S1173R) EST Mis c.3134 C > T(p.S1045L) EST Mis (15) 25/M Dut HH, CD, At, SS c.3439 C > T(p.R1147C) EST Mis c.1270_1271insG(p.D424Gfs*18) - Ins (16) 23/F Dut HH, CD, At, SS c.3439 C > T(p.R1147C) EST Mis c.1270_1271insG(p.D424Gfs*18) - Ins (16) 40/M Amr HH, CD, At c.3084_3085insGCCA(p.R1031Qfs*38) EST Ins c.3548 G > A(p.R1183Q) EST Mis (17) 25/F Amr HH, CD, At, SS, c.1431 T > A(p.C477*) - Non c.3034 G > A(p.G1012S) EST Mis (18) mID 55/M Cau HH, CD, ataxia c.3084_3085insGCCA(p.R1031Qfs*38) EST Ins c.4076 G > A(p.R1359Q) - Mis (19) 70/F Cau HH, CD, At c.3084_3085insGCCA(p.R1031Qfs*38) EST Ins c.4076 G > A(p.R1359Q) - Mis (19) 19/M Brz HH, CD, At, SS c.3531 G > A(p.W1177*) EST Non c.2441 T > C(p.L814P) - Mis (20) 51/F Can HH, CD, At c.644 T > A(p.V215D) cNMP Mis c.3404 G > A(p.R1135Q) EST Mis (21) S: Sex, M: MaleEth: ethnicity, F: Female, VT: Variant Type, Cau: Caucasian, Amr: American, Jap: Japan, Chn: Chinese, Eur: European, Tur: Turkish, Brz: Brazilian, Ger: German, Dut: Dutch Can: Canadian, Mis:Missense, Splc: Splice site, Ins: Insertion, Non: Nonsense, EST: Phospholipide esterase domain, Dmn: Domain, cDNA complementary DNA, AA: Amino acid, Tur: Turkish, Ex: Exon, Ref: Reference HH: Hypogonadotropic hypogonadism CD: Chorioretinal dystophy, At: Ataxia SS: Short stature MID: Mild intellectual disability, PST: Present Study, Clin Fin: Clinical Findings, RN:Reference Number

visual acuity of the patient progressively decreased by age of 30. found that could explain the patient’s retinal findings. It was At the initial exam, the best-corrected visual acuity was hand confirmedby Sanger sequencing and parents were shown to be movements in both eyes, with a − 7.00 spherical equivalent carriers for this change. Family pedigree, sequencing view of refractive correction. Intraocular pressure and anterior seg­ the variant in PNPLA6 gene, the brain imaging of index case ment examination were normal in both eyes. Fundus examina­ and an illustration of PNPLA6 gene structure along with rela­ tion revealed diffuse chorioretinal atrophy with central retinal tive positions of all known pathogenic variants and occurring atrophy and pigmentary clusters (Figure 2a-b). Optical coher­ amino acid changes (including our c.3524 C > G(p. ence tomography had shown retinal thinning, loss of the Ser1175Cys) missense variant) are given in Figure 1. This layered retinal architecture, marked loss of retinal pigment variant does not find in gnomAD exomes and genomes and epithelium, and choriocapillaris loss (Figure 2c-d). The patient has not been reported before in ClinVar. Sequence- and struc­ had full extraocular movements bilaterally, with jerk nystag­ ture-based prediction tools further support the pathogenicity mus. The previous examination findings of our patient were of this variant and considering family segregation analysis searched from the electronic database. It was observed that the results, we evaluated this variant as pathogenic according to spherical equivalent increased progressively and visual acuity ACMG guidelines (10). The entire exome dataset including progressively decreased. Interestingly, the patient’s fundus Genome Aggregation Database (gnomAD), conservation findings were evaluated in favor of retinitis pigmentosa in the score (GERP), predictions of pathogenicity based on ACMG examinations performed by five different physicians at differ­ recommendations is shown in Table 4. ent years (Table 3). Our index patient was studied by whole exome sequencing Discussion and a missense, homozygous variant (c.3524 C > G (p. Ser1175Cys)) in PNPLA6 gene, which could explain the PNPLA6 related disorders have varying clinical phenotypes patients’ clinical status was detected. No other variant was and characterized by a variable spectrum such as cerebellar 280 M. DOĞAN ET AL.

Figure 2. Color fundus photographs of right and left eye (a-b); Both eyes illustrate diffuse chorioretinal atrophy and pigmentary clusters (black arrows). Optical coherence tomography (OCT) images of right and left eye (c-d); Both eyes show retinal thinning, loss of the layered retinal architecture, marked loss of retinal pigment epithelium, and choriocapillaris loss. Because of severe nystagmus and vision loss, higher quality fundus photographs and OCT images could not be taken.

Table 3. The previous ocular examination findings of our patient obtained from Table 4. The entire exome dataset including Genome Aggregation Database the electronic database. (gnomAD), conservation score (GERP), predictions of pathogenicity based on Age (year) SE (R/L) BCVA (R/L), (Decimal) Fundus ACMG recommendations. 21 −2/-2 0.3/0.2 Bone-specile like lesions GENE PNPLA6 23 −3/-4 0.1/0.1 RP Transcript ID NM_001166111.2 24 −4/-5 0.1/0.1 RP dbSNP rs1555751592 27 −6/-6 2 m CF/0.05 RP Variant c.3524 C > G (p.Ser1175Cys) 29 −6.5/-6.5 2 m CF/1 m CF RP Variant location Exon 30 30* −7/-7 HM/HM Diffuse chorioretinal atrophy Variant type Missense *The last examination of the patient that performed by us. SE: spherical equiva­ MutationTaster Disease-causing lent, R: right, L: left, BCVA: best-corrected visual acuity, CF: counting finger, HM: PROVEAN Damaging hand movements, RP: retinitis pigmentosa SIFT Damaging gnomAD (exomes) Not found ClinVar Not found Conservation Conserved DANN score 0.9925 ataxia, chorioretinal dystrophy, hypogonadotropic hypogo­ GERP score NR: 4.9699; RS: 4.9699 ACMG classification Likely pathogenic nadism, peripheral neuropathy, hair anomalies, short stature ACMG pathogenicity criteria PM1, PM2, PP2, PP3, PP5 and intellectual disability. In 2014, Synofizik et al. found that gnomAD: Genome Aggregation Database, ACMG: The American College of pathogenic variants in the PNPLA6 gene were the primary Medical Genetics and Genomics cause of the BNHS (6). In the present study, we described a 30- year-old Turkish patient with clinical features of BNHS and identified a missense homozygous variant of conserved amino acid in the PNPLA6 gene which was absent in unaffectedfamily (S1175C) has different clinical manifestations, namely lack of members. He has been examined in various medical depart­ retinal findings in contrast to our patient. Considering that the ments, but definitediagnosis has not been established. No cases cause of diffuse chorioretinal degeneration in our patient may with similar features were described in his family history. To be another genetic reason, the exome analysis of the patient our knowledge, this variant has been previously described in was re-evaluated and no different variant was found to explain only one publication so far. Topaloglu et al. reported the same the patient’s retinal findings. In the literature, we noticed that variant in two sibling patients, 51 years old male and 46 years at least three patients were diagnosed with BNHS after the age old female, in 2014 (16). These patients had ataxia, hypogona­ of 50 with chorioretinal dystrophy (15,19). Since chorioretinal dotropic hypogonadism, but no ophthalmologic findings and findings may develop after midlife in patients with bialelic their clinics were compatible with Gordon-Holmes syndrome. pathogenic variants in PNPLA6 gene; we think it will be It is very interesting that another case with a similar genotype important to re-evaluate the two siblings identified by OPHTHALMIC GENETICS 281

Topaloglu et al. by performing detailed eye examinations. development and synaptic connections and in brain lipid meta­ Hufnagel et al. (2) predicted that various variants in the bolism (5,15). Particularly, choroideremia-like and pigmentary PNPLA6 gene may lead to activity changes in the neuropathy retinal changes should be kept in mind as BNHS’s character­ target esterase, causing more severe and early onset presenta­ istic ocular phenotype, which can be seen at an early age. Along tions, but we see that even the same variant may produce with these findings, ophthalmological evaluation is important different clinical findings. However, more research is needed in understanding BNHS and in distinguishing it from other to reveal why variants in the PNPLA6 gene lead to various hereditary visual disorders and ataxias. clinical presentations in different cell types of the animal According to the clinical data of patients with defined models. BNHS; the first clinical symptoms are visual loss and night Genotype-phenotype correlation with the variants detected blindness at a rate of 39.3%, ataxia at 28.6%, and delay in in the gene has not yet been demonstrated. However, Wu et al. puberty 32.1%. The rarity of the PNPLA6 gene-related disor­ reported that among patients with a variant in the PNPLA6 ders and the complex phenotype make the accurate diagnosis gene, retinal involvement was higher in patients with deleter­ of patients difficult. It is important to raise awareness among ious variants. They classified splicing, nonsense, insertion, and ophthalmologists since the presentation of the disease with deletion variants as “severe allele” and missense variant as vision problems in the early period is around 39.3%. It is “mild allele”. They also found that patients with chorioretinal observed that the development of ataxia in patients with dystrophy carried notably more variants in the patatin-like BNHS is mostly between the first and third decades of life phospholipase (Pat) domain (≥1 variant in Pat domain) than (24). However, a case with later onset ataxia (50 years) has those without the retinal phenotypes (9). Our patient had been described in the literature (15). It was seen that 8 out of 28 a biallelic missense variant with retinal findings in Pat domain. patients (28.6%) with molecularly confirmed BNHS presented Synofizicet al. reported that one of the two siblings in their 40s with gait ataxia in early childhood. The rate of presentation with the same biallelic variants (one deleterious and one mis­ with delayed puberty in our study appears to be 9/28 (32.1%). sense in Pat domain) diagnosed with BNHS had chorioretinal Hypogonadotropic hypogonadism is generally described in the dystrophy while the other did not (6). Yoon et al. detected two onset of puberty (25). When the clinical findings of BNHS deleterious variants in cNMP domain of the PNPLA6 gene in patients molecularly confirmed in the literature to be consid­ a patient with spastic paraplegia and without retinal findings ered, it was seen that 100% of the patients had hypogonado­ (22). As the number of cases increases, we will be able to reach tropic hypogonadism. Ocular findings and/or onset of ataxia more accurate results regarding the genotype–phenotype rela­ usually occur at an early age with BNHS and presentation at tionships of the PNPLA6 gene, which shows clinical hetero­ late ages is rare (19). From this point of view, it is important geneity. Ocular signs and symptoms may be the first and only that endocrinology specialists consider BNHS syndrome, finding for years (9), and these patients are usually diagnosed although it is one of the rare causes in the hypogonadotropic with non-syndromic retinal degeneration or retinitis pigmen­ hypogonadism etiology. It is stated that some patients with tosa. It was reported that most BNHS patients had chorioret­ BNHS may have cognitive impairment. When 28 BNHS inal atrophy and approximately half had pigment changes on patients were evaluated to date, cognitive impairment (as in fundoscopy (15). Our patient was being followed up with our patient) was present in 28.6% of the patients and progres­ a diagnosis of retinitis pigmentosa due to pigment changes in sive deterioration was not revealed (6,26). his fundoscopic examination. Although nyctalopia and pig­ To the best of the authors’ knowledge, this is the first family ment clusters suggest retinitis pigmentosa in our proband, with BNHS was identified in Turkey. The patients who with there was no waxy disc pallor in fundus or significant retinal BNHS were mostly reported as sporadic cases in families with no vessel weakening. In addition, the patient did not have poster­ consanguinity (22/28 (%78,5)). For this syndrome, which is ior subcapsular cataract, which is frequently seen in retinitis inherited as autosomal recessive, we think that as the number pigmentosa. There are cases in the literature who were followed of patients defined increases, it may be detected more in families up with the diagnosis of pigmentary retinopathy and/or chor­ with consanguinity. Today, with molecular genetic diagnosis, oideremia and were diagnosed with BNHS after molecular early prenatal diagnosis, and PGD (preimplantation genetic genetic analysis (17). Our patient had main ocular findings diagnosis) opportunities are offered to families. It also gives such as chorioretinal atrophy, pigmentary clusters, nystagmus, the patient and family the opportunity to provide genetic coun­ central vision loss, and the absence of optic atrophy which were seling about the risk of passing the disease to next generations. previously reported in the cases with BNHS and fundus find­ We think that this report will contribute to a better understand­ ings of the patient such as retinal thinning, loss of layered ing of the genetic background of BNHS patients. Further studies retinal structure, significant retinal pigment epithelium loss are needed on how the variants affect gene functions and how and choriocapillaris loss were also compatible with BNHS. the findings develop in the retina. As the number of patients Some of the patients with BNHS have also reported findings identified with BNHS increases, we will be able to obtain more such as retinal tubulation, intraretinal cysts, and isolated macu­ information about genotype-phenotype correlations. lar changes that were not present in our patient (12,17,21). However, it appears that regardless of the severity of the chor­ ioretinal degeneration observed in the reported cases, the Acknowledgments visual result can range from almost intact vision to blindness We are very grateful to all the individuals who participated in our study. (15,23). This may be attributed to the PNPLA6 domains in the This research did not receive any specific grant from funding agencies in studies and their complex role in maintaining neuronal the public, commercial, or not-for-profit sectors. 282 M. DOĞAN ET AL.

Funding hypogonadotropic hypogonadism: two novel cases and a review of 40 cases from the literature. J Neurol. 2015;262(1):194–202. There is no funding doi:10.1007/s00415-014-7555-9. 13. Kobayashi F, Kurihara Y, Nagasaka K, Iida H, Shindo K, Takiyama Y. A patient with cerebellar ataxia, hypogonadotropic ORCID hypogonadism and vitelliform macular dystrophy: Boucher- Neuhäuser syndrome. Cinical Neurol. 2010;50:98–102. Mustafa Doğan http://orcid.org/0000-0003-0464-6565 doi:10.5692/clinicalneurol.50.98. Recep Eröz http://orcid.org/0000-0003-0840-2613 14. Umehara T, Yaguchi H, Suzuki M, Isozaki E, Mochio S. Are Emrah Öztürk http://orcid.org/0000-0002-3590-3213 hypersegmented neutrophils a characteristic of Boucher- Neuhäuser syndrome? J Neurol Sci. 2010;295(1–2):128–30. doi:10.1016/j.jns.2010.05.005. Declaration of interest 15. Deik A, Johannes B, Rucker JC, Sanchez E, Brodie SE, Deegan E, Landy K, Kajiwara Y, Scelsa S. 2014. Compound heterozygous We have no conflicts of interest. We alone are responsible for the content PNPLA6 mutations cause Boucher– neuhäuser syndrome with and writing of this article. late-onset ataxia. J Neurol. 261(12):2411–23. doi:10.1007/s00415- 014-7516-3. References 16. Topaloglu AK, Lomniczi A, Kretzscmar D, Dissen GA, Kotan LD, Mcardle CA, Koç AF, Hamel BC, Guclu M, Papatya ED, et al. Loss- 1. Sogorb MA, Pamies D, Estevan C, Estévez J, Vilanova E. Roles of of-function mutations in PNPLA6 encoding neuropathy target NTE protein and encoding gene in development and neurodeve­ esterase underlie pubertal failure and neurological deficits in gor­ lopmental toxicity. Chem Biol Interact. 2016;259:352–57. don holmes syndrome. J Clin Endocrinol Metab. 2014;99 doi:10.1016/j.cbi.2016.07.030. (10):2067–75. doi:10.1210/jc.2014-1836. 2. Hufnagel RB, Arno G, Hein ND, Hersheson J, Prasad M, 17. O’Neil E, Serrano L, Scoles D, Cunningham KE, Han G, Chiang J, Anderson Y, Krueger LA, Gregory LC, Stoetzel C, Jaworek TJ, Bennett J, Aleman TS. 2019. Detailed retinal phenotype of et al. Neuropathy target esterase impairments cause Boucher-Neuhäuser syndrome associated with mutations in Oliver-McFarlane and Laurence-Moon syndromes. J Med Genet. PNPLA6 mimicking choroideremia. Ophthalmic Genet. 40 2015;52(2):85–94. doi:10.1136/jmedgenet-2014-102856. (3):267–75. doi:10.1080/13816810.2019.1605392. 3. Chang P, He L, Wang Y, Heier C, Wu Y, Huang F. 2019. 18. DeNaro BB, Gavazi ED, Rubaltelli DM, Freund KB, Lee W, Characterization of the interaction of neuropathy target esterase Yannuzzi LA, Tsang SH, Kang JJ. Chorioretinal changes in with the endoplasmic reticulum and lipid droplets. Biomolecules. 9 a genetically confirmed case of Boucher-Neuhäuser syndrome. (12):848. doi:10.3390/biom9120848. Physiol Behav. 2018;176:139–48. doi:10.1097/ICB.0000000000000769. 4. Sunderhaus ER, Law AD, Kretzschmar D. ER responses play a key 19. Langdahl JH, Frederiksen AL, Nguyen N, Brusgaard K, Juhl CB. role in Swiss-Cheese/Neuropathy Target Esterase-associated 2017. Boucher Neuhäuser Syndrome – a rare cause of inherited neurodegeneration. Neurobiol Dis. 2019;130:1–28. doi:10.1016/j. hypogonadotropic hypogonadism. A case of two adult siblings with nbd.2019.104520. two novel mutations in PNPLA6. Eur J Med Genet. 60(2):105–09. 5. Richardson RJ, Hein ND, Wijeyesakere SJ, Fink JK, Makhaeva GF. doi:10.1016/j.ejmg.2016.11.003. 2013. Neuropathy target esterase (NTE): overview and future. 20. Teive HA, Camargo CH, Sato MT, Shiokawa N, Boguszewski CL, Chem Biol Interact. 203(1):238–44. doi:10.1016/j.cbi.2012.10.024. Raskin S, Buck C, Seminara SB, Munhoz RP. 2018. Different 6. Synofzik M, Gonzalez MA, Lourenco CM, Coutelier M, Haack TB, cerebellar ataxia phenotypes associated with mutations of the Rebelo A, Hannequin D, Strom TM, Prokisch H, Kernstock C, PNPLA6 gene in Brazilian patients with recessive ataxias. et al. PNPLA6 mutations cause Boucher-Neuhäuser and Gordon Cerebellum. 17(3):380–85. doi:10.1007/s12311-017-0909-y. Holmes syndromes as part of a broad neurodegenerative spectrum. 21. Donaldson L, Tarnopolsky MA, Martin JA, Rodriguez AR. 2020. Brain. 2014;137(1):69–77. doi:10.1093/brain/awt326. Severe chorioretinal atrophy in Boucher-Neuhauser syndrome. 7. Boucher BJ, Gibberd FB. Familial ataxia, hypogonadism, and ret­ Can J Ophthalmol. 55(1):26–28. doi:10.1016/j.jcjo.2019.07.001. inal degeneration. Acta Neurol Scand. 1969;45(4):507–10. 22. Yoon G, Baskin B, Tarnopolsky M, Boycott KM, Geraghty MT, doi:10.1111/j.1600-0404.1969.tb01261.x. PMID: 5806782. Sell E, Goobie S, Meschino W, Banwell B, Ray PN. 2013. 8. Neuhauser G, Opitz JM. Autosomal recessive syndrome of cere­ Autosomal recessive hereditary spastic paraplegia - Clinical and bellar ataxia and hypogonadotropic hypogonadism. Clin Genet. genetic characteristics of a well-defined cohort. Neurogenetics. 14 1975;7(5):426–34. doi:10.1111/j.1399-0004.1975.tb00353.x. (3–4):181–88. doi:10.1007/s10048-013-0366-9. 9. Wu S, Sun Z, Zhu T, Weleber RG, Yang P, Wei X, Pennesi ME, 23. Rump P, Hamel BC, Pinckers AJ, Van Dop PA. 1997. Two sibs with Sui S. Novel variants in PNPLA6 causing syndromic retinal chorioretinal dystrophy, hypogonadotrophic hypogonadism, and dystrophy. Exp Eye Res. 2020 July:108327. doi:10.1016/j. cerebellar ataxia: Boucher-Neuhauser syndrome. J Med Genet. 34 exer.2020.108327. (9):767–71. doi:10.1136/jmg.34.9.767. 10. Richards S, Aziz N, Bale S, Bick D, Das S, Gastier FJ, Grody WW, 24. Ling H, Unnwongse K, Bhidayasiri R. 2009. Complex movement Hedge M, Lyon E, Spector E, et al. Standards and guidelines for the disorders in a sporadic Boucher–Neuhauser syndrome: phenotypic interpretation of sequence variants: a joint consensus recommen­ manifestations beyond the triad. Mov Disord. 24(15):2304–06. dation of the American college of medical genetics and genomics doi:10.1002/mds.22831. and the association for molecular pathology. Genet Med. 2015;17 25. Braslavsky D, Grinspon RP, Ballerini MG, Bedecarrás P, Loreti N, (5):405–23. doi:10.1038/gim.2015.30. Bastida G, Ropelato MG, Keselman A, Campo S, Alberto RR, et al. 11. Zheng R, Zhao Y, Wu J, Wang Y, Liu JL, Zhou ZL, Zhou XT, Hypogonadotropic hypogonadism in infants with congenital hypo­ Chen DN, Liao WH, Li JD. A novel PNPLA6 compound hetero­ pituitarism: a challenge to diagnose at an early stage. Horm Res zygous mutation identified in a Chinese patient with Boucher- Paediatr. 2015;84(5):289–97. doi:10.1159/000439051. Neuhäuser syndrome. Mol Med Rep. 2018;18(1):261–67. 26. Kate MP, Kesavadas C, Nair M, Krishnan S, Soman M, Singh A. doi:10.3892/mmr.2018.8955. Late-onset Boucher–Neuhauser syndrome (late BNHS) associated 12. Tarnutzer AA, Kahlert CG, Timmann D, Chang DI, Harmuth F, with white-matter changes: a report of two cases and review of Bauer P, Strauman D, Synofizik M. Boucher–Neuhäuser syn­ literature. J Neurol Neurosurg Psychiatry. 2011;82(8):888–91. drome: cerebellar degeneration, chorioretinal dystrophy and doi:10.1136/jnnp.2009.196790.