Prospective Evaluation of Kidney Disease in Joubert Syndrome

Article

Leah R. Fleming, Daniel A. Doherty, Melissa A. Parisi, Ian A. Glass, Joy Bryant, Roxanne Fischer, Baris Turkbey, Peter Choyke, Kailash Daryanani, Meghana Vemulapalli, James C. Mullikin, May Christine Malicdan, Thierry Vilboux, John A. Sayer, William A. Gahl, and Meral Gunay-Aygun

Abstract Background and objectives Joubert syndrome is a genetically heterogeneous ciliopathy associated with .30 genes. Due to the number of contributing authors, The characteristics of kidney disease and genotype-phenotype correlations have not been evaluated in a large the affiliations are cohort at a single center. provided in the Supplemental Design, setting, participants, & measurements We evaluated 97 individuals with Joubert syndrome at the National Material. Institutes of Health Clinical Center using abdominal ultrasonography, blood and urine chemistries, and DNA Correspondence: sequencing. Dr. Meral Gunay- Aygun, National Results Patients were ages 0.6–36 years old (mean of 9.067.6 years old); 41 were female. Mutations were identified Human Genome TMEM67 C5orf42 CC2D2A CEP290 Research Institute, in 19 genes in 92 patients; two thirds of the mutations resided in six genes: , , , , National Institutes of AHI1,andKIAA0586. Kidney disease was detected in 30%, most commonly in association with the following Health, 10 Center genes: CEP290 (six of six), TMEM67 (11 of 22), and AHI1 (three of six). No kidney disease was identified in patients Drive, Building 10, with mutations in C5orf42 (zero of 15) or KIAA0586 (zero of six). Prenatal ultrasonography of kidneys was normal Room 10C10310C103, fi Bethesda, MD 20892- in 72% of patients with kidney disease. Speci c types of kidney disease included nephronophthisis (31%), an 1851. Email: overlap phenotype of autosomal recessive polycystic kidney disease/nephronophthisis (35%), unilateral [email protected] multicystic dysplastic kidney (10%), and indeterminate-type cystic kidney disease (24%). Early-onset hypertension occurred in 24% of patients with kidney disease. Age at ESRD (n=13) ranged from 6 to 24 years old (mean of 11.364.8 years old).

Conclusions Kidney disease occurs in up to one third of patients with Joubert syndrome, most commonly in those with mutations in CEP290, TMEM67,andAHI1. Patients with mutations in C5orf42 or KIAA0586 are less likely to develop kidney disease. Prenatal ultrasonography is a poor predictor of kidney involvement in Joubert syndrome. Unilateral multicystic dysplastic kidney and autosomal recessive polycystic kidney disease–like enlarged kidneys with early-onset hypertension can be part of the Joubert syndrome kidney phenotype. Clin J Am Soc Nephrol 12: 1962–1973, 2017. doi: https://doi.org/10.2215/CJN.05660517

Introduction use of the term Joubert syndrome and related disor- Fibrocystic kidney disease is a common feature in ders, which include Senior–Løken (nephronophthisis many ciliopathies, including Meckel, Bardet–Biedl, and retinal degeneration) syndrome and COACH and Joubert syndromes (1,2). First described in 1969 (colobomas, “oligophrenia” for cognitive impairment, (3), Joubert syndrome (OMIM 213300) is defined on ataxia, cerebellar vermis hypoplasia, and hepatic the basis of a characteristic set of cerebellum and fibrosis) syndrome (8). For simplicity, the term Joubert midbrain abnormalities that collectively result in the syndrome refers to all patients with the “molar diagnostic “molar tooth sign” on axial brain images tooth sign,” including patients with or without non- (4) (Figure 1). The incidence of Joubert syndrome is neurologic disease (6). In this article, we use Joubert estimated to range between 1:80,000 and 1:100,000 (5). syndrome to include Senior–Løken and COACH Core clinical features of Joubert syndrome observed syndromes. shortly after birth include hypotonia and episodic Like other syndromic ciliopathies, Joubert syn- tachypnea and/or apnea followed later by develop- drome is characterized by extreme genetic heteroge- mental delay, ocular motor apraxia, truncal ataxia, neity, with .30 causative genes (5). The frequency of and speech apraxia (5,6). Most patients have extra- kidney disease in individual gene–specificsubtypesof neurologic involvement that can include polydactyly, Joubert syndrome remains unknown; estimates range chorioretinal colobomas, retinal degeneration, con- from 25% to 27% (9,10). Most publications on kidney genital hepatic fibrosis, and/or fibrocystic kidney involvement in the disease have been limited to small disease (5,7). This clinical heterogeneity led to the case series or retrospective multicenter reports on the

1962 Copyright © 2017 by the American Society of Nephrology www.cjasn.org Vol 12 December, 2017 Clin J Am Soc Nephrol 12: 1962–1973, December, 2017 Kidney Disease in Joubert Syndrome, Fleming et al. 1963

basis of questionnaires and medical records review (7,9,11– correlations between genotype and kidney disease remain 13). In 1992, Saraiva and Baraitser (9) claimed that kidney largely unknown (5,15,16). disease and retinal disease in Joubert syndrome were Here, we describe the characteristics of kidney disease in closely linked and that only patients with retinal dystrophy 97 patients with Joubert syndrome prospectively evaluated were at risk for fibrocystic kidney disease. However, many at a single center. We detail kidney ultrasonography, mag- individuals with Joubert syndrome and kidney disease but netic resonance imaging (MRI) of the kidneys, and kidney no retinal degeneration have since been reported (5,7). function tests in the context of the molecular genetic diagnosis – Although the nephronophthisis in Senior Løken syndrome as well as other organ involvement, including liver disease (14) and cystic dysplastic kidneys in Arima syndrome (7) and ophthalmologic features. are relatively well described, the kidney disease in other patients with Joubert syndrome is often reported in non- specific terms, noting only absence or presence of “kidney Materials and Methods cysts” and neglecting data on features, such as BP, kidney Patients function, and delineation of the exact type of the fibrocytic All patients were prospectively evaluated at the National kidney disease. Although many of the over 30 causative Institutes of Health (NIH) Clinical Center between 2003 Joubert syndrome genes, including CEP290, NPHP1, AHI1, and 2014 under the intramural NIH research protocol OFD1, RPGRIP1L, CC2D2A, TMEM67, TMEM216, TMEM138, “Clinical and Molecular Investigations Into Ciliopathies” and TMEM237, have been associated with kidney disease, (www.clinicaltrials.gov; trial NCT00068224) approved by

Figure 1. | Clinical diagnosis of Joubert syndrome is made based on the characteristic “molar tooth sign” on brain imaging. Axial brain magnetic resonance imaging (MRI) obtained at the pontomesencephalic junction showing (A) the molar tooth sign (circle) and hypoplastic cerebellar vermis (arrows) compared with (C) normal. Midsagittal brain MRI showing (B) hypoplasia and dysplasia of the cerebellar vermis (circle) and enlarged fourth ventricle (asterisk) with rostral displacement of the fastigium compared with (D) normal. 1964 Clinical Journal of the American Society of Nephrology

the National Human Genome Research Institute Institu- pediatric clinical geneticist (M.G.-A.), as a group to tional Review Board. The study was advertised by the achieve consensus (17). Joubert Syndrome & Related Disorders Foundation. Pa- tients and/or their parents gave written, informed consent. Molecular Inversion Probes, Whole Exome, and Enrollment required a clinical diagnosis of Joubert Sanger Sequencing “ syndrome on the basis of the pathognomonic molar tooth The coding exons of 27 genes associated with Joubert sign” on brain MRI. For recruitment, the study was syndrome were sequenced by combining a molecular advertised to individuals with Joubert syndrome and their inversion probe capture method and next generation families by the Joubert Syndrome and Related Disorders sequencing as previously described (20). In addition, Foundation as a natural history study aiming to describe exome sequencing was performed using the HiSeq2000 the individual organ system involvement in Joubert syn- (Illumina, San Diego, CA) (21) and analyzed as previously drome, including kidney disease. To minimize ascertain- described (20). Additional details are in Supplemental ment bias, all travel, lodging, and other participation costs Material. as well as clinical and laboratory evaluations were spon- sored by the NIH. A total of 120 patients from 105 families Statistical Methods who reported to be diagnosed with Joubert syndrome on Ranges, averages, and SDs were calculated. Serum the basis of brain MRI applied. Brief phone interviews cystatin C was not available in four children (Supplemental ’ fi answering parents questions on risks and bene ts of the Table 1, patients 26, 31, 82, and 83) due to problems in study evaluations were performed by the senior investi- handling of blood specimens. For these patients, eGFR was gator (M.G.-A.) on all 120 individuals with Joubert syn- calculated using an alternative formula on the basis of drome from 105 families; no patients were excluded at this height and serum creatinine [eGFR =0.4133 (height in stage. Fifteen patients could not travel to the NIH Clinical centimeters/Scr)] (22). Center in Bethesda, Maryland, in some patients because of the severity of neurologic symptoms. The remaining 105 patients with Joubert syndrome from 90 families under- Results went week-long clinical evaluations at the NIH Clinical Patient Cohort Center. Eight patients from six families (including three Supplemental Table 1 shows the molecular genetic patients with the classic kidney and retina findings of findings in 97 patients as well as their major clinical Senior–Løken syndrome) were excluded, because they did features, including kidney disease, retinal degeneration, not have the “molar tooth sign” on our review of their brain MRI images (17). The remaining 97 patients were included in this study. All patients were evaluated by the senior investigator Table 1. Comparison of patients with Joubert syndrome with (M.G.-A.); evaluations included past medical history, and without kidney disease family history, physical examination, review of past medical records, review of brain magnetic resonance images, Kidney Disease No Kidney Disease high-resolution complete abdominal ultrasonography (all No. of patients 29/97 (30%) 68/97 (70%) patients), abdominal MRI (13 patients who were able Age, yr, mean6SD 9.666.5 8.868.0 to cooperate for the study without sedation), complete Females, males 11, 18 30, 38 a ophthalmologic examination, and comprehensive blood and Gene TMEM67 urine chemistries. Because abdominal MRI was performed in 11 11 CEP290 61 only 13 patients, ultrasonography-based kidney length, AHI1 33 which was available for all patients, was used to evaluate CC2D2A 28 kidney size. INPP5E 22 3 NPHP1 10 eGFR was calculated using pediatric [eGFR =39.8 RPGRIP1L 0.456 0.418 10 (height (meters)/Scr) (1.8/cystatin C) (30/ TMEM216 11 0.079 men 0.179 BUN) 1.076 (height (meters)/1.4) ] (18) and adult TMEM237 10 2 2 [eGFR =1753 (Scr) 1.1543 (age) 0.2033(0.742 if a woman) 3 C5orf42 015 (1.212 if black)] (19) formulas. Blood samples for DNA were KIAA0586 06 MKS1 05 collected from all patients and parents when available. TMEM231 fi 03 Kidney disease was de ned on the basis of eGFR and/or CSPP1 02 abnormal findings on kidney ultrasonography, including KIAA0753 02 increased echogenicity, increased or decreased kidney OFD1 02 CELSR2 size, and/or cystic changes. Liver disease was defined by 01 KIF7 01 elevated liver enzymes and/or increased echogenicity of B9D1 01 the liver and/or splenomegaly on abdominal ultrasonog- Unknown 1 4 raphy. Retinal degeneration was on the basis of typical Polydactyly 2/29 (7%) 10/68 (15%) findings on retinal examination after dilation of pupils. Coloboma 13/29 (45%) 17/68 (25%) All available brain magnetic resonance images were Retinal degeneration 11/29 (38%) 16/68 (24%) Liver involvement 18/29 (62%) 24/68 (35%) qualitatively evaluated by a team of five physicians, including two pediatric neuroradiologists, one adult aNumber of patients with and without kidney disease. neuroradiologist, one pediatric neurologist, and one lnJA o eho 2 9217,Dcme,2017 December, 1962–1973, 12: Nephrol Soc Am J Clin

Table 2. Characteristics of kidney disease in 29 patients with Joubert syndrome and kidney involvement

Patient Sex/ No./ Speciﬁc Age at Kidney Length GFR, Family Prenatal Laterality of Early-Onset Kidney Type of NIH Gene NIH Kidney US at NIH evaluationa ml/min No./ Ultrasonography Kidney Disease Hypertensionb Histopathology Kidney Evaluation, (Right, Left in SD) per 1.73 m2 NIH Ciliopathy Disease yr No.

3/3/97 m/3.9 TMEM67 Prenatal US normal. Diffuse moderate Bilateral 23.6, 24.3 28 (12)c + NP ARPKD-NPHP Oligohydramnios hyperechogenicity, at delivery, loss of CMD, enlarged kidneys palpable small cysts in at birth cortex and medulla 4/4/548 m/4.5 TMEM67 N Loss of CMD Bilateral 20.1, +1.5 146 2 NP NPHP 5/5/216 f/4.9 TMEM67 18 wk, Several cysts in Bilateral 20.2, at mean 69 (12)c + NP ARPKD-NPHP oligohydramnios, left kidney, enlarged normal diffusely hyperechoic echogenicity cystic kidneys (affected and CMD sibling’s prenatal US showed large cystic kidneys, oligohydramnios) 8/7/271 f/6.7 TMEM67 20 wk, enlarged Diffuse moderate Bilateral +0.6, 20.2 27 (8)c + Extracted right ARPKD-NPHP diffusely hyperechoic hyperechogenicity, kidney: kidneys with loss of CMD, histopathologic scattered cysts in ﬁndings cysts, no oligohydramnios cortex and consistent with medulla “autosomal recessive polycystic kidney disease,” interstitial nephritis, and glomerulosclerosis 9/7/272 m/9.3 TMEM67 N Tx Bilateral Tx Tx (7) + Extracted right ARPKD-NPHP kidney: moderate

chronic interstitial 1965 al. et Fleming Syndrome, Joubert in Disease Kidney nephritis, glomerulosclerosis, tubular atrophy, and nephrocalcinosis as well as cysts, mostly accumulated at the CM junction but also scattered throughout the cortex and medulla 10/8/557 m/6.8 TMEM67 N Diffuse mild Bilateral 22.5, 21.9 129 2 NP NPHP hyperechogenicity, loss of CMD

11/8/559 m/17 TMEM67 N Diffuse moderate Bilateral 20.5, 21.3 59 + NP ARPKD-NPHP hyperechogenicity, loss of CMD, medullary cysts 96Ciia ora fteAeia oit fNephrology of Society American the of Journal Clinical 1966 Table 2. (Continued)

13/10/238 m/8.2 TMEM67 19 wk, enlarged Diffuse moderate Bilateral +2.6, +2.3 38 (10)c + NA ARPKD-NPHP diffusely hyperechoic hyperechogenicity, kidneys with loss of CMD, loss of CMD, cysts in cortex progressive and medulla oligohydramnios 16/13/303 f/15 TMEM67 N Tx Bilateral Tx Tx (13) + Extracted right kidney ARPKD-NPHP (51 g, 8.534.43 2.8 cm); extensive glomerulosclerosis, tubular atrophy, and interstitial ﬁbrosis with severe chronic inﬂammation without any evidence of dysplasia. A few thin-walled tubular cysts at CM junction 17/14/252 f/15 TMEM67 N (affected Multiple cysts Bilateral 21.8, 20.9 83 2 NP Cystic, unable sibling’s in cortex to classify prenatal and medulla US showed of right polycystic kidney, normal kidneys) echogenicity and CMD 18/15/309 m/17 TMEM67 N Multiple cysts Bilateral 20.2, +1.3 106 2 NP Cystic, unable in cortex to classify and medulla, normal echogenicity and CMD 40/32/575 f/2.3 CC2D2A N Medullary Bilateral 21.8, 20.5 119 2 NP NPHP hyperecogenicity 43/35/185 m/3.6 CC2D2A N Loss of CMD Bilateral 23.4, 22.0 66 2 NP NPHP 48/39/480 m/0.9 CEP290 N Multiple cysts in Bilateral 21.7, 22.8 90 2 NP Cystic, unable cortex and to classify medulla of right kidney, normal echogenicity and CMD 49/40/552 m/4.3 CEP290 N Multiple cysts Bilateral 20.7, 21.4 115 2 NP Cystic, unable in cortex to classify and medulla of both kidneys, normal echogenicity and CMD 50/41/373 m/4.4 CEP290 20 wk, left Left multicystic Unilateral (left) +1.9, atrophic MCDK 55 (6)c 2 NP Unilateral multicystic dysplastic MCDK dysplastic kidney without kidney; right any normal kidney normal parenchyma, several cortical cysts in right kidney lnJA o eho 2 9217,Dcme,2017 December, 1962–1973, 12: Nephrol Soc Am J Clin Table 2. (Continued)

52/43/412 m/10 CEP290 N Multiple medullary Bilateral 20.9, 20.9 35 2 NP NPHP cysts, normal echogen 53/44/213 f/13 CEP290 19 wk, Tx Bilateral Tx Tx (9) 2 NA NPHP hyperechogenic kidneys (resolved in third trimester) 54/45/441 f/24 CEP290 N Multiple medullary Bilateral +0.3, 20.9 21 (24)c 2 NP NPHP cysts, moderate hyperechogenicity 56/47/517 m/3.3 AHI1 19 wk, left Left kidney Unilateral (left) +1.7, atrophic MCDK 108 2 NP Unilateral multicystic could not be MCDK dysplastic visualized, kidney; right kidney right kidney normal normal 59/50/540 f/18 AHI1 N Left kidney with Asymmetrical +1.9, 20.7 92 2 NP Cystic, unable several large (left earlier to classify, cysts, right onset than right) (asymmetric) kidney normal size with multiple cysts and moderate hyperechogenicity 60/51/228 f/21 AHI1 N Left multicystic Unilateral (left) +4.3, atrophic MCDK 219 2 NP Unilateral dysplastic MCDK kidney, right kidney normal (14.9 cm) 72/63/372 m/3.2 INPP5E 8 mo, Multiple medullary Bilateral +3.9, +3.7 137 2 NP ARPKD-NPHP inyDsaei obr ydoe lmn ta.1967 al. et Fleming Syndrome, Joubert in Disease Kidney oligohydramnios cysts, loss of CMD, moderate hyperechogenicity 73/64/352 m/15 INPP5E N Tx Bilateral Tx Tx (6) 2 NP NPHP 79/67/438 m/3.4 NPHP1 N Medullary cysts Bilateral 21.0, +1.3 117 2 NP NPHP and echogenic foci in medullary pyramids 80/68/396 f/9 TMEM216 N N Bilateral +0.5, 21.64 52 2 NA Unable to classify (associated with bladder dysfunction) 88/76/360 m/21 RPGRIP1L N Diffuse moderate Bilateral +2.0, +1.3 33 2 NP ARPKD-NPHP hyperechogenicity, loss of CMD, cysts in medulla and cortex 89/77/474 f/4.5 TMEM237 20 wk, enlarged Diffuse moderate Bilateral +1.3, 21.8 52 2 NP ARPKD-NPHP (three times hyperechogenicity, normal size) loss of CMD, kidneys with markedly echogenic foci increased echogenicity and cysts in medulla and cortex 1968 Clinical Journal of the American Society of Nephrology

coloboma, liver disease, and polydactyly. There were 41 females and 56 males. Ages ranged from 0.6 to 36 years c ﬁ old (mean of 9.067.6 years old) (Supplemental Table 1). Kidney Disease Speci Type of

to classify Nine families had two siblings (families 6, 7, 8, 18, 24, 29, 36, 65, and 78) and two families had three siblings with Joubert syndrome (families 16 and 66) evaluated at the NIH. In addition, four other families were represented in this study by a single patient (Table 1, families 5, 14, 34, and 44), and family 66 had another affected deceased Kidney child. Histopathology The molecular genetic cause was identiﬁed in 19 different NP Cystic, unable Joubert syndrome genes in all but ﬁve families; detailed

b molecular genetic ﬁndings of this cohort were reported separately (20) (Supplemental Table 1). Mutations for two 2 thirds of the families resided in six genes: TMEM67 (20%),

Early-Onset C5orf42 CC2D2A CEP290 AHI1

Hypertension (14%), (11%), (8%), (7%), and KIAA0586 (7%). Each of the remaining 13 genes accounted 2

c for a small proportion of the families ranging from 1% to 6%: MKS1 (6%), INPP5E (4%), NPHP1 (1%), TMEM216 (2%), GFR, ml/min OFD1 CSPP1 TMEM231 KIF7 B9D1 per 1.73 m (2%), (2%), (1%), (1%), (1%), RPGRIP1L (1%), TMEM237 (1%), KIAA0753 (1%), and a CELSR2 (1%).

0.5 34 (15) Retinal degeneration was detected in 20%, coloboma was 2 detected in 30%, polydactyly was detected in 13%, and liver 0.5.

2 involvement was detected in 43% (Supplemental Table 1). Kidney Length (Right, Left in SD) at NIH evaluation Kidney Disease Kidney involvement was identiﬁed in 29 patients; in the remaining 68 patients, kidney ultrasonography and eGFR were normal, and there was no hypertension or chronic

Laterality of anemia. (Tables 1 and 2). Four patients with kidney disease Kidney Disease

Bilateral (Table 2, patients 9, 16, 53, and 73) underwent kidney transplantation before their NIH evaluation. Of the remaining 25 patients with kidney involvement, 24 had abnormalities on kidney ultrasonography; 13 of these 24 had decreased eGFR (Table 2). One patient had decreased eGFR with normal kidney ultrasonography. NIH Kidney US

especially in the cortex, single cortical cyst in right kidney Kidney Transplantation At the time that this paper was written, 13 of the 29 patients with Joubert syndrome and kidney involvement had reached ESRD at ages 6–24 years old (mean of 11.364.8 years old); seven patients have undergone transplantation after their NIH evaluation (Table 2, patients 3, 5, 8, 13, 50, 54, and 97), and two other patients (Table 2, patients 52 and Prenatal 80) reached ESRD at age 12 years old but have not yet been Ultrasonography transplanted.

Kidney-Related Findings on Prenatal Ultrasonography Prenatal kidney ultrasonography was abnormal in only

Gene eight (28%) of the 29 patients with kidney disease (Table 2, patients 5, 8, 13, 50, 53, 56, 72, and 89). In two other affected individuals (Supplemental Table 1, patients 17 and 42), prenatal ultrasonography was normal, but their mothers yr NIH Sex/ Age at had other affected pregnancies with enlarged diffusely Evaluation, cystic kidneys, resulting in fetal demise. In four patients (Table 2, patients 5, 8, 13, and 89), enlarged hyperechogenic kidneys with or without discrete cysts and/or oligohy-

No. – No./ No./ dramnios were seen at gestational age 19 20 weeks. The Family Patient Hypertension diagnosed before any decrease in GFR. Kidney length on the basis of US measurement. These patients received renal transplantation after the NIH visit; ages at transplantation in years are indicated in parentheses. mother of patient 5 had another pregnancy with similar Table 2. (Continued) NIH, National Institutes ofdisease-nephronophthisis; Health; US, N, renal normal; ultrasonography; NPHP,dysplastic nephronophthisis; m, kidney. male; f, CMD,a female; corticomedullary Tx, differentiation; transplanted (age NP,b at not transplantation performed; ARPKD-NPHP, in years); autosomalc CM, recessive corticomedullary; polycystic NA, kidney not available; MCDK, multicyctic NIH Ciliopathy 97/84/358 m/9.1 Unknown N Moderate hyperechogenicity ultrasonography ﬁndings at 19 weeks. In patient 53, who Clin J Am Soc Nephrol 12: 1962–1973, December, 2017 Kidney Disease in Joubert Syndrome, Fleming et al. 1969

required kidney transplantation at age 9 years old, prenatal eGFR (Table 2). Patient 9 was screened at age 3 years old ultrasonography at 19 weeks showed hyperechogenic after his sister’s diagnosis of “polycystic kidneys”;hehad kidneys that returned to normal on repeat ultrasonography cardiomyopathy secondary to undiagnosed early-onset in the third trimester. Patient 72 had polyhydramnios severe hypertension, which resolved after treatment. during the eighth month of pregnancy. In patient 3, prenatal ultrasonography was normal, but oligohydramnios was diagnosed at delivery; the infant’s kidneys were palpable Discussion at birth. In two patients (50 and 56), the kidney abnormality The frequency of kidney disease in our Joubert syndrome was unilateral; ultrasonography at the 20th week of gestation cohort was 30%, slightly higher than published estimates of – fl in each infant showed left multicystic dysplastic kidney, and 25% 27% (9,10), perhaps a re ection of our comprehensive fi the right kidney was normal. evaluations performed speci cally to detect kidney disease. The percentage of individuals with kidney disease may also increase with age; our cohort’s mean age was 9 years Kidney Imaging Findings at the NIH Clinical Center old. The most common abnormality on kidney ultrasonog- Nephronophthisis was present in 31% of our patients raphy was increased echogenicity involving both cortex with Joubert syndrome and kidney disease, and it re- and medulla, resulting in loss of corticomedullary differ- sembled the kidney disease of nonsyndromic nephronophthisis entiation with or without discrete cysts (11 of 24 patients (23,24). Clinically, kidney size was normal or small, with with abnormal ultrasonography). In five of these 11 increased echogenicity and a few cysts at the cortico- patients (Table 2, patients 3, 5, 8, 13, and 89), the kidneys medullary junction. Of the nine patients with Joubert were perinatally enlarged; one of these patients still had syndrome and nephronophthisis, two had Senior–Løken enlarged kidneys at the time of NIH evaluation at age 8 syndrome with retinal degeneration due to mutations in years old (Table 2, patient 13). In the remaining four, CEP290 andINPP5E; the seven patients with nephronoph- kidney size normalized or became smaller over time thisis and no retinal disease had mutations in CC2D2A, (Figure 2, A and B). Patient 60 with left multicystic TMEM67, and NPHP1. dysplastic kidney had an enlarged right kidney likely A hybrid autosomal recessive polycystic kidney disease due to compensatory hypertrophy; in the remaining 24 (PKD)-nephronophthisis pattern of kidney disease was patients with kidney disease, kidney size was either normal present in 35% of patients with Joubert syndrome and or small (Table 2). In four patients (Table 2, patients 40, 52, kidney disease. One characteristic feature of autosomal 54, and 79), cysts and/or hyperechogenicity were confined recessive PKD (25) but not nephronophthisis (24), present to medulla (Figure 2D). Asymmetric kidney disease was in seven patients, was early-onset severe hypertension (before identified in four patients (Table 2, patients 50, 56, 59, and any decrease in eGFR). Another feature of autosomal recessive 60), including three with left multicystic dysplastic kidney PKD, enlarged kidneys, was seen in five patients, making the (Figure 2, E and F). Patient 56’s nuclear scan at age 12 early differentiation of Joubert syndrome and autosomal months old failed to show any kidney function of the left recessive PKD challenging. In fact, three patients (8,9,and kidney (Figure 2F), and ultrasonography at 19 months 13) were originally diagnosed as having autosomal recessive showed a normal right kidney and a small left-sided PKD and carried this diagnosis until their NIH evaluation hypoechoic mass that may have represented an atrophic under our autosomal recessive PKD study at ages 6, 9, and 8 kidney; his kidney function and voiding cystourethrogram ’ years old; at that time, their mild oculomotor apraxia and were normal. Patient 59 s yearly ultrasonography exami- developmental delays made us suspect Joubert syndrome. An nations between ages 1 and 7 years old were completely fi additional patient (patient 3) was misdiagnosed as having normal. At age 7 years old, the only abnormal nding was a autosomal recessive PKD in addition to his known diagnosis small cortical cyst at the upper pole of the left kidney; of Joubert syndrome, which was the cause of his polycystic annual ultrasonography imaging between ages 7 and 13 kidneys. Before making a prenatal diagnosis of autosomal years old remained otherwise normal. At age 17 years old, recessive PKD, cerebellar abnormalities and polydactyly both kidneys were hyperechogenic; the left kidney (length should be excluded, because renal disease in Joubert syn- 2 at 0.7 SD) had several large cysts in the upper pole, drome can be indistinguishable from autosomal recessive whereas the right kidney was relatively larger (length at PKD. Only 13% of patients with Joubert syndrome have +1.9 SD), with a few small cortical cysts. polydactyly (Table 1), and therefore, a careful evaluation for brain anomalies is essential for prenatal differentiation of Kidney Histopathology Findings Joubert syndrome from autosomal recessive PKD. In addi- Histopathology of three kidney explants (Table 2, pa- tion, if a child diagnosed as having autosomal recessive tients 8, 9, and 16) showed severe tubular atrophy, in- PKD manifests developmental delay and/or ocular motor fi fl terstitial brosis, chronic in ammation, variable degrees of apraxia, a brain MRI should be performed, because patients glomerulosclerosis, and nephrocalcinosis as well as cysts, with Joubert syndrome with mild cerebellar involvement mostly accumulated at the corticomedullary junction but may be misdiagnosed as having autosomal recessive PKD. also scattered throughout the cortex and medulla. The combination of coloboma, kidney disease, and polydactyly strongly suggests a ciliopathy, specifically Joubert Early-Onset Hypertension syndrome. Molecular studies can also be helpful, because Of the 29 patients with kidney disease, seven had early- seven of the ten patients in the autosomal recessive PKD- onset hypertension diagnosed shortly after birth or within nephronophthisis group had mutations in TMEM67 and the first years of life before any measurable decrease in enlarged, autosomal recessive PKD–like kidneys; indeed, 1970 Clinical Journal of the American Society of Nephrology

Figure 2. | Kidney disease in patients with Joubert syndrome include autosomal recessive PKD-like disease and unilateral multicytic dysplastic kidneys. (A) Ultrasonographic image of patient 9 at age 4 years old showing enlarged and hyperechogenic left kidney (outlined by the dots) with loss of corticomedullary differentiation. (B) Patient 9’s kidney length (bold red line; average of right and left) plotted against age in comparison with normal mean (bold gray line) and +2 SD (gray line). (C) High-resolution ultrasonographic image of patient 88’s left kidney displaying cysts in cortex. (D) Ultrasonography image of patient 40’s left kidney showing medullary hyperechogenicity. (E) T2-weighted coronal magnetic resonance image of patient 60 showing a left multicystic dysplastic kidneyand normal right kidney.(F) 99mTc nuclear scan (coronal image) of patient 56 at age 12 months old showing no function of his left multicystic dysplastic kidney. This patient’s prenatal ultrasonography at 19 weeks of gestation showed a left multicystic dysplastickidneyand a normal right kidney.On ultrasonographyat age 3 years old, his left kidney could not be visualized, but his right kidney was normal. mutations in TMEM67 also cause Meckel syndrome, an- part of many syndromes, asymmetric kidney disease is other ciliopathy with cystic kidney disease (1). unexpected in ciliopathies (1,23). We are aware of multi- Our most surprising ﬁnding was unilateral multicystic cystic dysplastic kidney reported in Joubert syndrome in dysplastic kidney in three patients with Joubert syndrome the medical literature: one Iranian patient with Joubert (patients 50, 56, and 60), two of whom had completely syndrome and right-sided multicystic dysplastic kidney normal contralateral kidney and normal eGFR at ages 3 (26) and one with either unilateral kidney agenesis (7) or (patient 56) and 20 years old (patient 60) (Figure 2, E and F, unilateral multicystic dysplastic kidney misdiagnosed as Table 2). Both of these patients had mutations in AHI1. kidney agenesis due to resolution of the atrophic kidney Although unilateral multicystic dysplastic kidney occurs as (27). Of note, patient 60 in our cohort was part of a Clin J Am Soc Nephrol 12: 1962–1973, December, 2017 Kidney Disease in Joubert Syndrome, Fleming et al. 1971

previously reported series (family K8062) (28). Recently, Bardet–Biedl syndromes may be due to the variation in the mutations in HNF1b and TBX18 have been identified in expertise of the prenatal ultrasonography specialists as well rare families with multicystic dysplastic kidney (29–31). as differences in the nature of kidney disease among Our patients, however, had mutations in CEP290 and AHI1 different ciliopathies. as single-gene causes of syndromic multicystic dysplastic We can derive several important conclusions from this kidney associated with Joubert syndrome. study. First, kidney disease affects up to one third of Modifier genes may influence the occurrence of kidney patients with Joubert syndrome and is more common in disease in Joubert syndrome (32). All 11 sibling sets those with pathogenic variants in CEP290, TMEM67,and evaluated at the NIH were concordant for kidney disease. AHI1 genes; it was not observed in patients with mutations However, in two families, there was history of a deceased in C5orf42 or KIAA0586. Second, the severity and progres- sibling who was discordant for presence of kidney disease sion of kidney disease seemed comparable across Joubert (Supplemental Table 1, Table 3, families 34 and 66). In syndrome genotypes, with most patients requiring kidney family 34 (Supplemental Table 1), patient 42 had no transplantation in childhood or young adulthood. Third, significant kidney disease at age 3.2 years old, despite prenatal ultrasonography does not predict kidney disease having an affected brother who passed away perinatally well in Joubert syndrome; it was abnormal in only 28% of due to cystic kidneys. Similarly, in family 66, a fourth child patients with Joubert syndrome and kidney disease. More- passed away at birth with polycystic kidneys (33), whereas over, the prenatal kidney ultrasonography findings in three other affected siblings (Supplemental Table 1, patients Joubert syndrome are indistinguishable from autosomal 76–78) had normal kidney imaging and function at ages recessive PKD, and therefore, fetuses with hyperechogenic 15 months old and 4 years and 9 months old (twins). It is kidneys should have careful imaging for other abnormal- possible that these patients may develop kidney disease as ities, especially brain anomalies; in addition, polydactyly, if they get older. present, can be a helpful sign of Joubert syndrome, Prenatal ultrasonography was abnormal in 28% of although its absence does not exclude the condition. patients with Joubert syndrome and kidney disease, Fourth, early-onset hypertension, before kidney function representing a surprisingly low prenatal detection rate of is decreased, occurred in 24% of patients with Joubert kidney disease in Joubert syndrome. In another multisys- syndrome and kidney disease, particularly those with tem ciliopathy, Bardet–Biedl syndrome (34), Forsythe et al. enlarged, autosomal recessive PKD–like, diffusely hyper- (34) reported a prenatal detection rate of 64% for kidney echogenic kidneys. Fifth, in families with multiple children disease. This discrepancy in prenatal detection rates of with Joubert syndrome, siblings are largely concordant for kidney disease of patients with Joubert syndrome and kidney disease, although exceptions exist. On the basis of

Table 3. Molecular genetic and clinical ﬁndings of the 11 sibling sets with Joubert syndrome

NIH Patient Family Age, Retinal Liver Kidney Ciliopathy Sex Gene Polydactyly Coloboma No. No. yr Dystrophy Disease Disease No.

6 6 302 m 5.1 TMEM67 2 + 2 + 2 7301m9.52 + 2 + 2 8 7 271 f 6.7 TMEM67 222++ 9272m9.3222++ 10 8 557 m 6.8 TMEM67 2 + 2 ++ 11 559 m 16.7 2 + 2 ++ 19 16 562 m 22.9 TMEM67 2 + 2 + 2 20 561 m 24.9 2 + 2 + 2 21 560 m 29.6 2 + 2 + 2 23 18 488 m 0.6 C5orf42 22222 24 487 f 5.3 22222 30 24 501 m 10 C5orf42 + 22+ 2 31 500 f 11.8 + 22+ 2 36 29 482 f 24.5 C5orf42 + 22+ 2 37 481 m 27.6 + 22+ 2 44 36 577 f 13.1 CC2D2A 22222 45 576 f 15.7 22222 74 65 7504 f 19 INPP5E 22+ 22 75 7503 f 21 22+ 22 76a 66 520 f 1.2 TMEM231 22+/2 +/22 77 518 f 4.8 22+ 22 78 519 f 4.8 22+ 22 90 78 390 f 1.9 KIAA0753 22222 91 389 m 7 22222

NIH, National Institutes of Health; m, male; f, female. aBecause of the young age of this patient, her retinal examination was not conclusive, and liver involvement classification, which was made on the basis of liver hyperechogenicity, was not definitive, because liver can be physiologically hyperechogenic during the first years of life. 1972 Clinical Journal of the American Society of Nephrology

these conclusions, we make several recommendations for Bird TD, Dolan CR, Fong CT, Smith RJH, Stephens K, Seattle, WA, surveillance. All patients with Joubert syndrome should University of Washington, 1993 6. Romani M, Micalizzi A, Valente EM: Joubert syndrome: Con- have BP measurement, complete abdominal ultrasonogra- genital cerebellar ataxia with the molar tooth. Lancet Neurol 12: phy, liver and kidney function tests, and complete blood 894–905, 2013 counts at the time of diagnosis and annually thereafter. 7. Satran D, Pierpont ME, Dobyns WB: Cerebello-oculo-renal Patients with Joubert syndrome and unilateral multicystic syndromes including Arima, Senior-Lo¨ken and COACH syn- dysplastic kidney should be radiologically assessed for dromes: More than just variants of Joubert syndrome. Am J Med Genet 86: 459–469, 1999 other congenital anomalies of the kidney and urinary tract, 8. Gleeson JG, Keeler LC, Parisi MA, Marsh SE, Chance PF, Glass IA, such as vesicoureteral reflux or contralateral ureteropelvic Graham Jr. JM, Maria BL, Barkovich AJ, Dobyns WB: Molar junction obstruction (27,35,36). tooth sign of the midbrain-hindbrain junction: Occurrence in Although this study represents the largest number of multiple distinct syndromes. Am J Med Genet A 125A: 125–134, subjects with Joubert syndrome evaluated at a single 2004 9. Saraiva JM, Baraitser M: Joubert syndrome: A review. Am J Med center, the size of the cohort is still relatively small. Genet 43: 726–731, 1992 Gene-phenotype correlations were not possible, except 10. Bachmann-Gagescu R, Dempsey JC, Phelps IG, O’Roak BJ, for the most common genetic causes. Our cohort is likely Knutzen DM, Rue TC, Ishak GE, Isabella CR, Gorden N, Adkins J, enriched for patients with Joubert syndrome and kidney Boyle EA, de Lacy N, O’Day D, Alswaid A, Ramadevi A R, ` and liver diseases, because the comprehensive evaluations Lingappa L, Lourenço C, Martorell L, Garcia-Cazorla A, Ozyu¨rek H, Haliloglu G, Tuysuz B, Topçu M, Chance P,Parisi MA, Glass IA, performed at the NIH Clinical Center may have attracted Shendure J, Doherty D; University of Washington Center for patients with these manifestations. However, our study Mendelian Genomics: Joubert syndrome: A model for untangling also offered other investigations, including neurocognitive recessive disorders with extreme genetic heterogeneity. JMed testing, DNA sequencing, sleep studies, and echocardio- Genet 52: 514–522, 2015 11. Hildebrandt F, Nothwang HG, Vossmerbaümer U, Springer C, grams, which may have minimized this bias. Patients with Strahm B, Hoppe B, Keuth B, Fuchshuber A, Querfeld U, Neuhaus Joubert syndrome and severe disease, such as those who TJ, Brandis M: Lack of large, homozygous deletions of the required mechanical ventilation, may be under-represented, nephronophthisis 1 region in Joubert syndrome type B. APN Study because they could not travel to Bethesda, Maryland. In Group. Arbeitsgemeinschaft fu¨rPa¨diatrische Nephrologie. Pe- addition, the overall frequency of kidney disease as well as other diatr Nephrol 12: 16–19, 1998 12. Steinlin M, Schmid M, Landau K, Boltshauser E: Follow-up in extraneurologic manifestations may have been underestimated children with Joubert syndrome. Neuropediatrics 28: 204–211, by this cross-sectional study, because these manifestations may 1997 developasthepatientgetsolder. 13. Boycott KM, Parboosingh JS, Scott JN, McLeod DR, Greenberg CR, Fujiwara TM, Mah JK, Midgley J, Wade A, Bernier FP, Acknowledgments Chodirker BN, Bunge M, Innes AM: Meckel syndrome in the Hutterite The authors thank the Joubert Syndrome and Related Disorders population is actually a Joubert-related cerebello-oculo-renal syndrome. Am J Med Genet A 143A: 1715–1725, 2007 Foundation for their extensive support and the individuals with 14. Warady BA, Cibis G, Alon U, Blowey D, Hellerstein S: Senior- Joubert syndrome and their families who generously participated in Loken syndrome: Revisited. Pediatrics 94: 111–112, 1994 this investigation. 15. Sayer JA, Otto EA, O’Toole JF, Nurnberg G, Kennedy MA, Becker D.A.D. was supported by grant R01NS064077, the University of C, Hennies HC, Helou J, Attanasio M, Fausett BV,Utsch B, Khanna H, Liu Y, Drummond I, Kawakami I, Kusakabe T, Tsuda M, Ma L, Washington Intellectual and Developmental Disabilities Research Lee H, Larson RG, Allen SJ, Wilkinson CJ, Nigg EA, Shou C, Lillo C, Center, grant U54HD083091 Genetics Core 6845 and subproject Williams DS, Hoppe B, Kemper MJ, Neuhaus T, Parisi MA, Glass 6849, and private donations from families of children with Joubert IA, Petry M, Kispert A, Gloy J, Ganner A, Walz G, Zhu X, Goldman syndrome. This research was supported by the Intramural Research D, Nurnberg P, Swaroop A, Leroux MR, Hildebrandt F: The Program of the National Human Genome Research Institute and the centrosomal protein nephrocystin-6 is mutated in Joubert syndrome and activates transcription factor ATF4. 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Gunay-Aygun M, Font-Montgomery E, Lukose L, Tuchman M, Graf J, Bryant JC, Kleta R, Garcia A, EdwardsH, Piwnica-Worms K, See related editorial, “A Perspective on Inherited Kidney Disease: Adams D, Bernardini I, Fischer RE, KrasnewichD, Oden N, Ling A, ” – Quezado Z, Zak C, Daryanani KT, Turkbey B, Choyke P, Guay- Lessons for Practicing Nephrologists, on pages 1914 1916. Woodford LM, Gahl WA: Correlation of kidney function, volume and imaging findings, and PKHD1 mutations in 73 patients with This article contains supplemental material online at http://cjasn. autosomal recessive polycystic kidney disease. Clin J Am Soc asnjournals.org/lookup/suppl/doi:10.2215/CJN.05660517/-/ Nephrol 5: 972–984, 2010 DCSupplemental. Prospective Evaluation of Kidney Disease in Joubert Syndrome

Leah R. Fleming MD1,2, Daniel A. Doherty MD, PhD3,4, Melissa A. Parisi MD, PhD5, Ian A.

Glass MD3,4, Joy Bryant RN1, Roxanne Fischer MS1, Baris Turkbey MD6, Peter Choyke MD6,

Kailash Daryanani,7 Meghana Vemulapalli MsC8, James C. Mullikin PhD,8, NISC Comparative

Sequencing Program8, May Christine Malicdan MD, PhD1,9, Thierry Vilboux PhD,1,10, John A.

Sayer MD, PhD11, William A Gahl MD, PhD,1,8,12, Meral Gunay-Aygun MD.1,12,13

Affiliations

1. Section on Human Biochemical Genetics, Medical Genetics Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20892, USA 2. Department of Genetics and Metabolism, St. Luke’s Children’s Specialty Center, Boise, ID, 83712, USA 3. Department of Pediatrics, University of Washington, Seattle, WA 98195, USA 4. Center for Integrative Brain Research, Seattle Children's Research Institute, Seattle, WA 98195, USA 5. Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA 6. Molecular Imaging Program, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA 7. Radiology and Imaging Sciences, Clinical Center, National Institutes of Health, Bethesda, MD 8. NIH Intramural Sequencing Center (NISC), National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20892, USA 9. NIH Undiagnosed Diseases Program, Common Fund, Office of the Director, National Institutes of Health, Bethesda, MD 20892, USA 10. Inova Translational Medicine Institute, Falls Church, VA 22042, USA 11. Institute of Genetic Medicine, Newcastle University, International Centre for Life, Newcastle, UK 12. Office of the Clinical Director, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD 20892, USA 13. Johns Hopkins University School of Medicine, Department of Pediatrics and McKusick- Nathans Institute of Genetic Medicine, Baltimore, MD 21287, USA

Supplemental material is neither peer-reviewed nor thoroughly edited by CJASN. The authors alone are responsible for the accuracy and presentation of the material.

Supplementary Files

Prospective Evaluation of Kidney Disease in Joubert Syndrome.

Leah R. Fleming MD1,2, Daniel A. Doherty MD, PhD3,4, Melissa A. Parisi MD, PhD5, Ian A.

Glass3,4, Joy Bryant RN1, Roxanne Fischer MS1, Baris Turkbey MD6, Peter Choyke MD6,

Kailash Daryanani,7 Meghana Vemulapalli MsC8, James C. Mullikin PhD,8, NISC Comparative

Sequencing Program8, May Christine Malicdan MD, PhD1,9, Thierry Vilboux PhD,1,10, John A.

Sayer MD, PhD11, William A Gahl MD, PhD,1,8,12, Meral Gunay-Aygun MD1,12,13

Supplementary Methods

Molecular Genetics Methods

The 27 JS genes sequenced by combining a MIP capture method and next-generation sequencing were AHI1, ARL13B, B9D1, B9D2, C2CD3, C5orf42, CC2D2A, CEP290, CEP41, CSPP1,

IFT172, INPP5E, KIF7, MKS1, NPHP1, OFD1, RPGRIP1L, TCTN1, TCTN2, TCTN3, TMEM138,

TMEM216, TMEM231, TMEM237, TMEM67, TTC12B and ZNF423.(1) Whole exome sequencing performed using the HiSeq2000 (Illumina, San Diego, CA) employed 101-bp paired-end read sequencing. Confirmatory Sanger sequencing of gDNA and cDNA was performed as described.

The molecular genetic diagnosis was made based on the identification of variants in the known genes if one or more of the following criteria were met: 1. Two null alleles in a gene associated with JS, 2. Both alleles with variants previously reported to cause JS, 3. One allele with a variant previously reported to cause JS and another null allele, 4. One null allele and a variant predicted to be deleterious (PolyPhen2, CADD and Sift), 4. One allele with a variant previously reported to cause JS and a variant predicted to be deleterious (PolyPhen2, CADD and Sift) 5. Both alleles with variants predicted to be deleterious (PolyPhen2, CADD and Sift) and MAF <0.0001 (ExAC). Supplemental material is neither peer-reviewed nor thoroughly edited by CJASN. The authors alone are responsible for the accuracy and presentation of the material.

1. Vilboux T, Doherty DA, Glass IA, Parisi MA, Phelps IG, Cullinane AR, Zein W, Brooks BP, Heller T, Soldatos A, Oden NL, Yildirimli D, Vemulapalli M, Mullikin JC, Nisc Comparative Sequencing P, Malicdan MC, Gahl WA, Gunay-Aygun M: Molecular genetic findings and clinical correlations in 100 patients with Joubert syndrome and related disorders prospectively evaluated at a single center. Genetics in medicine : official journal of the American College of Medical Genetics, 2017 of thematerial. Supplemental materialisneitherpeer-reviewednorthoroughlyeditedbyCJASN.Theauthorsalone areresponsiblefortheaccuracyandpresentation Table 39 38 741m2. C5orf42 27.6 m 481 37 34 33 32 642f2. C5orf42 24.5 f 482 36 35 150f1. C5orf42 11.8 f 500 31 159m1. TMEM67 16.7 TMEM67 m 6.8 m 559 14 557 13 12 11 10 051m1 C5orf42 10 m 501 30 26 25 18 348m06C5orf42 0.6 m 488 23 4487 24 41 40 15 29 28 TMEM67 29.6 TMEM67 m 24.9 TMEM67 m 560 22.9 m 561 562 22 21 20 19 27 16 17 7 . TMEM67 9.3 m TMEM67 9.5 272 TMEM67 m 5.1 271 m 301 302 9 8 7 6 5 4 3 1 2 Patient No 1S.Molecular 14** 5** 31 30 27 26 25 28 11 10 20 19 15 29 24 18 16 33 32 12 23 22 17 21 13 9 8 7 6 4 3 1 2 Family No 483 7 . CC2D2A 1.8 f 378 2 88C5orf42 18.8 C5orf42 14.1 m C5orf42 f 528 10.3 m 568 404 9 9C5orf42 19 m 491 5 TMEM67 9 TMEM67 m 8.2 m 255 238 459 458 355 TMEM67 17.4 m 309 4 . TMEM67 4.5 m 216 548 6 . TMEM67 0.9 m 361 9 . CC2D2A CC2D2A 3.5 2.3 m f 495 575 4 19TMEM67 11.9 f 542 7 C5orf42 8 m 534 471 432 523 4 . TMEM67 2.2 f 545 0 47TMEM67 14.7 f 303 252 7m39TMEM67 3.9 m 97 NIH Ciliopathyt No f f f f f f f f f f f Gender Geneticand 48TMEM67 14.8 62TMEM67 36.2 . TMEM67 7.8 TMEM67 6.7 TMEM67 4.9 . CC2D2 2.3 C5orf42 8.4 C5orf42 2.3 C5orf42 2.2 C5orf42 2.1 C5orf42 5.3 Age Gene Main A Clinical c.3975+4_3975+7del c.1784T>G; .772> c.2624C>T; p.(Ser875Phe) c.455C>T; p.(Ser152Phe) c.5737+2T>C c.2923C>T; p.(Gln975*) c.1784T>G; c.7477C>T; p.(Arg2493*) .Po37lf*7 c.8710C>T; c.7190del; p.(Pro2397Glnfs*37) c.297G>T; c.224‐2 A>T c.579_580del; c.579_580del; c.1081G>T; c.1843 T>C;p.(Cys615Arg) c.1843 T>C;p.(Cys615Arg) c.2802delA; c.2802delA; c.2522A>C; .Po37lf*7 c.8710C>T; c.7190del; p.(Pro2397Glnfs*37) c.2999G>T; c.510dup; p.(Leu171Serfs*8) c.1819del; p.(Tyr607Thrfs*6) c.1126C>G; c.1784T>G; c.7988_7989del; c.769A>G; c.1674+3A>G .322u c.2661+5G>A c.7988_7989del; c.2322+2dup c.748G>A, c.755T>C; p.(Met252Thr) c.248‐4_248 c.1538A>G; .Vl07hf*)c.4667A>T; p.(Asp1556Val) c.4741A>G; c.3289del; p.(Val1097Phefs*2) c.1538 A>G; c.1819del; p.(Tyr607Thrfs*6) c.755T>C; p.(Met252Thr) c.755T>C; p.(Met252Thr) c.755T>C; p.(Met252Thr) c.622A>T; c. 2498 Findings T>C;p.(Ile833Thr) p.(Arg208*)

p.(Gly250Arg) p.(Met257Val) p.(Lys99Asn) p.(Gln841Pro) p.(Leu595*) p.(Leu595*) p.(Glu361*) p.(Trp1000Leu) p.(Leu595*) p.(Tyr513Cys) p.(Thr1581Ala) .An3Mts2)Not identified Not identified p.(Asn935Metfs*25) p.(Asn935Metfs*25) p.(Thr513Cys) ‐3insAAGTTTT .Gy9Ies1)c.950C>G; p.(Thr317Arg) c.950C>G; p.(Thr317Arg) p.(Gly195Ilefs*13) p.(Gly195Ilefs*13) of97Joubert Mutation 1 .Gy63lf*0 c.7988_7989del; p.(Gly2663Alafs*40) p.(Gly2663Alafs*40) .Gy63lf*0 c.7988_7989del; p.(Gly2663Alafs*40) p.(Gly2663Alafs*40)

SydromePatients c.8263_8264insG; c.8710C>T; c.4667A>T; c.8263_8264insG; c.2661+5G>A c.2334G>A; c.622A>T; p.(Arg208*) c.3001A>G; p.(Thr1001Ala) c.5348C>A; p.(Ala1783Asp) c.3452T>C; c.2322+2dup c.8263dup; p.(Thr2755Asnfs*8) c.8710C>T; Not Not c.1843T>C; p.(Cys615Arg) c.3347C>T; c.1843 T>C;p.(Cys615Arg) c.1843 c.1843 c.7817T>A; p.(Leu2606*) c.245C>G; p.(Pro82Arg) c.245C>G; p.(Pro82Arg) c.245C>G; p.(Pro82Arg) c.2879C>T; c.2140_2142del; c.2122G>A; c.1351C>T; 2498 c. identified identified T>C;p.(Cys615Arg) T>C;p.(Cys615Arg) T>C;p.(Ile833Thr) Evaluated p.(Arg2904*) p.(Arg2904*) p.(Arg2904*) p.(Ala960Val) p.(Val1151Ala) p.(Arg2904*) p.(Thr1116Met) p.(Arg451*) p.(Asp1556Val) p.(Gln376Glu) p.(Trp778*) p.(Ala708Thr) p.(Ser715del) p.(Thr2755Serfs*8) p.(Thr2755Serfs*8) at theNIH and Mutation 2 c.8855+1G>A Clinical Center. + + + + + ‐‐ ‐‐ ‐‐ ‐‐ ‐‐ ‐ ‐ ‐‐ ‐ ‐‐ ‐‐ ‐ ‐ ‐ ‐‐ ‐ ‐ ‐‐ ‐‐ ‐ ‐ ‐ ‐‐ ‐‐ ‐‐ ‐ ‐ ‐ ‐ ‐‐ ‐ ‐‐ ‐ ‐‐ ‐ ‐ Polydactyly + + + + + + + + ++ + + + + + + + + + + ‐ ‐ ‐ ‐ ‐ Coloboma ‐ ‐ ‐ ‐‐‐ ‐‐‐ ‐ ‐‐‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐‐‐ ‐‐‐ ‐‐‐ ‐ ‐‐‐ ‐‐ ‐ ‐ ‐‐‐ ‐‐ ‐ ‐‐‐ ‐ ‐ ‐ ‐‐‐ ‐ ‐ ‐ ‐‐‐ ‐ ‐ Retinal Dystrophy + + + + + + ++ ++ ++ + ++ ++ + + ++ ++ + ++ + + + + + + ++ + + ++ Liver Disease + + ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ ‐ Kidney Disease Supplemental material is neither peer-reviewed nor thoroughly edited by CJASN. The authors alone are responsible for the accuracy and presentation of the material.

42 34** 446 m 3.2 CC2D2A c.3289del; p.(Val1097Phefs*2) c.3851G>A; p.(Arg1284His) ++‐‐‐ 43 35 185 m 3.6 CC2D2A c.3145C>T; p.(Arg1049*) c.3347C>T; p.(Thr1116Met) ‐‐‐ ++ 44 577 f 13.1 CC2D2A c.3743_3746dup; p.(Pro1250Glyfs*11) c.3989G>A; p.(Arg1330Gln) ‐‐‐‐‐ 36 45 576 f 15.7 CC2D2A c.3743_3746dup; p.(Pro1250Glyfs*11) c.3989G>A; p.(Arg1330Gln) ‐‐‐‐‐ 46 37 565 f 15.4 CC2D2A c.1017+1G>A c.4600T>G; p.(Leu1534Val) ‐‐+ + ‐ 47 38 364 f 16.3 CC2D2A c.3055C>T; p.(Arg1019*) c.3288G>C; p.(Gln1096His) ‐‐‐‐‐ 48 39 480 m 0.9 CEP290 c.5182G>T; p.(Glu1728*) c.5668G>T; p.(Gly1890*) ‐‐‐‐+ 49 40 552 m 4.3 CEP290 c.4723A>T; p.(Lys1575*) c.4966G>T; p.(Glu1656*) ‐‐+ ‐ + 50 41 373 m 4.4 CEP290 c.4882C>T; p.(Gln1628*) c.6072C>A; p.(Tyr2024*) ‐‐+ ++ 51 42 455 m 6.9 CEP290 c.1623+1G>A c.5668G>T; p.(Gly1890*) ‐‐+ ‐‐ 52 43 412 m 10 CEP290 c.1666dup; p.(Ile556Asnfs*20) c.5344C>T; p.(Arg1782*) ‐‐+ ‐ + 53 44** 213 f 13.2 CEP290 c.5611_5614del; p.(Gln1871Valfs*2) c.4882C>T; p.(Gln1628*) ‐ ++‐ + 54 45 441 f 23.5 CEP290 c.5668G>T; p.(Gly1890*) c.5356_5571del; p.(Glu1786_Leu1857del) ‐ ++‐ + 55 46 513 m 1.9 AHI1 c.1997A>T; p.(Asp666Val) c.2297G>A; p.(Gly766Glu) ‐‐‐‐‐ 56 47 517 m 3.3 AHI1 c.662C>A; p.(Ser221*) c.1583C>G; p.(Ser528*) ‐‐+ ++ 57 48 472 m 5 AHI1 c.1976A>T; p.(Asp659Val) c.2212C>T; p.(Arg738*) ‐‐+ ‐‐ 58 49 574 m 5.1 AHI1 c.910dup; p.(Thr304Asnfs*6) c.2105C>T; p.(Thr702Met) ‐‐+ ‐‐ 59 50 540 f 17.9 AHI1 c.1115A>G; p.(Asp372Gly) c.2173T>C; p.(Trp725Arg) ‐‐+ ++ 60 51 228 f 20.9 AHI1 c.736A>T; p.(Lys246*) c.2495T>G; p.(Leu832*) ‐‐+ ++ 61 52 443 f 2.2 KIAA0586 c.392del; p.(Arg131Lysfs*4) c.1254‐1G>C ‐‐‐‐‐ 62 53 368 m 4 KIAA0586 c.392del; p.(Arg131Lysfs*4) c.1000C>T; p.(Gln334*) ‐ + ‐‐‐ 63 54 494 m 4.4 KIAA0586 c.94dup; p.(His32Profs*8) c.586‐350_1129+1117del ‐ + ‐‐‐ 64 55 579 m 4.4 KIAA0586 c.392del; p.(Arg131Lysfs*4) c.586‐350_1129+1117del ‐‐‐‐‐ 65 56 507 m 4.7 KIAA0586 c.831C>T; p.(Leu277Leu) c.586‐350_1129+1117del ‐‐‐‐‐ 66 57 531 m 13.6 KIAA0586 c.392del; p.(Arg131Lysfs*4) c.961+1G>A ‐‐‐‐‐ 67 58 502 m 1.5 MKS1 c.1389G>T; p.(Arg463Arg) c.493C>T; p.(Arg165Cys) ‐‐‐‐‐ 68 59 397 m 1.7 MKS1 c.950G>A; p.(Gly317Glu) c.1115_1117del; p.(Ser372del) + ‐ + ‐‐ 69 60 537 m 2 MKS1 c.417G>A; p.(Glu139Glu) c.1476T>G; p.(Cys492Trp) + ‐ + + ‐ 70 61 510 m 8 MKS1 c.1408‐36_1408‐6del c.1387C>G; p.(Arg463Gly) ‐‐+ ‐‐ 71 62 573 m 14 MKS1 c.417G>A; p.(Glu139Glu) c.1208C>T; p.(Ser403Leu) ‐‐+ ‐‐ 72 63 372 m 3.2 INPP5E c.473delG; p.(Gly158Valfs*40) c.1304G>A; p.(Arg435Gln) ‐ ++‐ + 73 64 352 m 14.5 INPP5E c.1784_1787del; p.(Val595Glyfs*21) c.1862G>A; p.(Arg621Gln) ‐‐+ ++ 74 7504 f 19 INPP5E ‐‐+ 65 c.1565G>C; p.(Gly522Ala) c.1565G>C; p.(Gly522Ala) ‐‐ 75 7503 f 21 INPP5E c.1565G>C; p.(Gly522Ala) c.1565G>C; p.(Gly522Ala) ‐‐+ ‐‐ 76 520 f 1.2 TMEM231 c.712G>A; p.(Asp238Asn) g.18555_19148conNG_026383.1:g.2718_3301; p.(Ile224Leufs*5) ‐‐‐ + ‐ 7766¥ 518 f 4.8 TMEM231 c.712G>A; p.(Asp238Asn) g.18555_19148conNG_026383.1:g.2718_3301; p.(Ile224Leufs*5) ‐‐+ ‐‐ 78 519 f 4.8 TMEM231 c.712G>A; p.(Asp238Asn) g.18555_19148conNG_026383.1:g.2718_3301; p.(Ile224Leufs*5) ‐‐+ ‐‐ 79 67 438 m 3.4 NPHP1 290 kb common deletion 290 kb common deletion ‐‐‐‐+ 80 68 396 f 9 TMEM216 c.218G>T; p.(Arg73Leu) c.253C>T; p.(Arg85*) + ‐ ‐ ++ 81 69 408 m 12 TMEM216 c.218G>T; p.(Arg73Leu) c.218G>T; p.(Arg73Leu) ++‐‐‐ 82 70 3331 m 4.3 OFD1 c.2656del; p.(Gln886Lysfs*2) Not applicable as X‐linked ‐‐+ ‐‐ 83 71 452 m 12.3 OFD1 c.149A>G; p.(His50Arg) Not applicable as X linked ‐‐‐ + ‐ 84 72 466 f 2.3 CSPP1 c.2244_2245del; p.(Glu750Glyfs*30) c.2280del; p.(Glu761Lysfs*35) ‐‐+ ‐‐ 85 73 387 f 3.5 CSPP1 c.1835_1845del; p.Arg612Thrfs*2 c.2814‐1G>C ‐‐‐‐‐ 86 74 409 m 2.1 KIF7 c.3505C>T; p.(Gln1169*) c.3505C>T; p.(Gln1169*) ‐‐‐‐‐ 87 75 400 m 4.8 B9D1 c.95A>G; p.(Tyr32Cys) c.466C>T; p.(Arg156Trp) ‐‐‐‐‐ 88 76 360 m 21.4 RPGRIP1L c.671_677del; p.(Glu224Glyfs*7) c.1340T>C; p.(Leu447Ser) and c.3790G>T; p.(Asp1264Tyr) + + ‐‐+ 89 77 474 f 4.5 TMEM237 c.76C>T; p.(Gln26*) c.943+1G>T ‐ ++‐ + 90 390 f 1.9 KIAA0753 c.769A>G; p.(Arg257Gly) c.2359‐1G>C ‐‐‐‐‐ 78 Supplemental material is neither peer-reviewed nor thoroughly edited by CJASN. The authors alone are responsible for the accuracy and presentation of the material. 78 91 389 m 7 KIAA0753 c.769A>G; p.(Arg257Gly) c.2359‐1G>C ‐‐‐‐‐ 92 79 449 f 7.9 CELSR2 c.1150G>A; p.(Ala384Thr) c.6908C>T; p.(Thr2303Met) ‐‐‐ + ‐ 93 80 393 m 1.7 Unknown Not identified Not identified ++‐ + ‐ 94 81 578 f 2.3 Unknown Not identified Not identified ‐ + ‐‐‐ 95 82 570 m 4.7 Unknown Not identified Not identified ‐‐‐‐‐ 96 83 419 f 2 Unknown Not identified Not identified ‐‐+ ‐‐ 97 84 358 m 9.1 Unknown Not identified Not identified ‐‐‐ ++ Patients are grouped by gene and within each gene patients are sorted from the youngest to oldest. **These 4 families had history of an affected deceased sibling. ¥This family had a stillborn son with polycystic kidneys after their evaluation at NIH.