Use of Whole-Exome Sequencing for Diagnosis of Limb-Girdle Muscular Dystrophy: Outcomes and Lessons Learned

Supplementary Online Content

Ghaoui R, Cooper ST, Lek M, et al. Use of whole-exome sequencing for diagnosis of limb-girdle muscular dystrophy: outcomes and lessons learned. JAMA Neurol. Published online October 5, 2015. doi:10.1001/jamaneurol.2015.2274.

eTable 1. List of known myopathy genes eTable 2. Results of 27 families with likely pathogenic changes in candidate genes eTable 3. Variants of uncertain significance in two families

This supplementary material has been provided by the authors to give readers additional information about their work.

Downloaded From: https://jamanetwork.com/ on 09/29/2021 eTable 1. List of known myopathy genes ABHD5 ETFB POMGNT1

ACADVL ETFDH POMT1

ACTA1 FHL1 POMT2 ACVR1 FKRP PRKAG2

AGL FKTN PTPLA AGRN FLNC PTRF ALG2 GAA PYGM ALG13 GBE1 RAPSN ALG14 GFPT1 RBCK1 ANO5 GMPPB RYR1 B3GALNT2 GNE SCN4A

BAG3 GTDC2(POMGNT2) SERCA1

BIN1 GYG1 SEPN1 CAPN3 GYS1 SGCA CAV3 HNRPDL SGCB

CFL2 HSPG2 SGCD CHAT ISCU SGCG CHKB ISPD SLC22A5

CHRNA1 ITGA7 SLC25A20

CHRNB1 KLHL9 SMCHD1

CHRND KLHL40 SPEG CHRNE KLHL41 STAC3 CHRNG LAMA2 STIM1 CLCN1 LAMB2 SYNE1 CNTN1 LAMP2 SYNE2 COL6A1 LARGE TCAP

COL6A2 LDB3 TIA1

Downloaded From: https://jamanetwork.com/ on 09/29/2021 COL6A3 LDHA TMEM43

COLQ LMNA (Lamin A/C) TNNT1

CPT2 LMOD3 TNPO3 CRYAB LPIN1 TPM2 DAG1 MATR3 TOR1AIP1

DES MEGF10 TPM3

DMD MSTN TK2 DMPK MTM1 TRAPPC11

DNAJB6 MUSK TRIM32

DNM2 MYBPC3 TRIM63

DOK7 MYH7 TTN DOLK MYOT VAPB DPAGT1 NEB VCP

DPM1 OBSCN DPM2 PABPN1

DPM3 PDE9A DYSF PFKM ECEL1 PGM1 EMD PHKA1 ENO3 PLEC ETFA PNPLA2

List of known myopathy genes that were used to screen LGMD families for sequence variants using the xBrowse web server.

Downloaded From: https://jamanetwork.com/ on 09/29/2021 eTable 2. Results of 27 families with likely pathogenic changes in candidate genes Fa Age at Distrib Additiona Organ Maxi Biopsy Causa mily Present ution of l clinical involveme mum Findings tive ation weakne character nt CK gene (yrs) ss istics (U/L) 1 2 Proxim Lumbar Nil 403 Dystrophic Lamin al UL lordosis features, A/C and LL and tight rimmed (LMN tendo- vacuoles A) Achilles 2 9 Proxim Elbow and Nil 1200 Dystrophic Lamin al UL tendo- features with A/C and LL Achilles increased (LMN contractur internal A) es nuclei and moth eaten fibres 3 9 Proxim Toe Nil 7000 Dystrophic, FKRP al LL walking reduced and tight alpha- tendo- dystroglycan Achilles staining 4 3 Proxim Contractur Nil 5000 Dystrophic, FKRP al UL es at the reduced and LL elbows alpha- and knees dystroglycan and staining scoliosis. 5 47 Proxim Posterior Nil 9000 Myopathic ANO5 al UL lower features and LL limb muscle weakness and atrophy 6 21 Proxim Myalgias Nil 23000 Basophillic ANO5 al LL with regenerating exercise fibres and mildly increased central nuclei 7 21 Proxim Distal Nil 7402 Necrotising DYSF al LL weakness myopathy and dysferlin deficiency on IHC

Downloaded From: https://jamanetwork.com/ on 09/29/2021 8 16 Proxim Calf Nil 4000 Dystrophic DYSF al LL atrophy feature and almost complete absence of dysferlin on IHC/WB 9 3 Proxim Calf Nil 11,000 Dystrophic CAPN al UL hypertrop with some 3 and LL hy and patchy tight inflammatory Tendo- interstitial achilles infiltrate 10 10 Proxim Scapular Nil 274 End stage CAPN al UL winging muscle 3 and LL 11 2 Proxim Myalgias Nil 8000 Dystrophic, FKTN al LL with severely exercise reduced alpha-DG on IHC and absent on WB 12 10 Proxim Developm Cardiomyo 104 Dystrophic TOR1 al UL ental pathy with presence AIP1 and LL delay in of both type 1 motor and 2 milestones angulated facial atrophic weakness fibres 13 37 Proxim Abnormal Nil N/A Dystrophic, GTDC al UL eye many internal 2/ and LL movement nuclei. IHC- POMG s, reduced NT2 cognitive antibodies deficits againts alpha- DG. 14 20 Proxim Developm Nil 1400 Dystrophic, GMPP al UL ental reduced B and LL delay and patchy cognitive staining for deficits alphaDG 15 10 Proxim Mild Nil 413 Dystrophic LAMA al UL facial features, 2 and LL weakness, increase in ptosis, central nuclei mild and marked

Downloaded From: https://jamanetwork.com/ on 09/29/2021 elbow fibre splitting. contractur es; tight tendo- Achilles 16 40 Proxim Mild Cardiac 369 Dystrophic, LAMA al UL distal LL arrhythmia necrotic 2 and LL weakness fibres inclusions and sparse myofibrillar changes including an occasional core 17 2 Proxim Ankle Cardiac- 800 Not COL6 al UL contractur arrhythmia performed A1 and LL es s 18 5 Proxim Ankle Nil 361 Myopathic/dy COL6 al UL contractur strophic A3 and LL es process. Focal moderate fibrosis 19 2 Proxim Contractur Restrictive 123 Muscular COL6 al UL es-ankles lung dystrophy A3 and LL and long disease with finger prominence flexors of fibre splitting 20 10 Proxim Scoliosis, Restrictive 211 Dystrophic COL6 al UL orbicularis lung features. A3 and LL oris disease Focal weakness moderate and fibrosis. scapular winging 21 13 Proxim Reduced Nil 5416 Fibre type PYGM al UL exercise disproportion and LL intoleranc with type 1 e fibre hypotrophy 22 25 Proxim Myalgias Nil 2000 Dystrophic, CPT2 al LL with cold variation in temperatu fibre size. res One

Downloaded From: https://jamanetwork.com/ on 09/29/2021 regenerating fibre, presence of rimmed vacuoles 23 5 Proxim Fatiguabil Nil 1410 Dystrophic. DOK7 al UL ity and Some and LL facial internalised weakness nuclei, and scattered hyalinised fibres. Increased interstitial fibrous tissue and occasional foci of chronic inflammatory cell infiltrate 24 54 Proxim Head drop Cardiomyo 106 Dystrophic, ACTA1 al Ul and axial pathy occasional and LL weakness nemaline rods 25 30 Proxim Opthalmo Macro- 3000 Dystrophic STIM1 al Ul plegia, thrombocyt features with and LL facial and openia presence of neck tubular flexor aggregates weakness 26 38 Proxim Mild Nil 570 Dystrophic FLNC al UL distal features, with and LL weakness some rimmed in UL and vacuoles and LL unusual cytoplasmic inclusions. 27 7 Proxim Scapular Nil 126 Mild CHD7 al UL winging myopathic features, suggestive of a dystrophic process Results of 27 families with likely pathogenic changes in candidate genes. All families had proximal weakness. Clinical information was collected on age of onset, degree and pattern of muscle involvement, presence of facial weakness, family history, evidence of

Downloaded From: https://jamanetwork.com/ on 09/29/2021 muscle hypertrophy, cardiac and respiratory muscle involvement, intellectual impairment, joint contractures. Additional clinical characteristics for each family, maximum CK result and biopsy findings are noted. No biopsy was performed on proband of Family 17. CK=creatine kinase, DG=dystroglycan. RefSeq: Lamin A/C (LMNA) (NM_170707), FKRP (NM_024301.4), ANO5 (NM_213599), DYSF (NG_008694.1, NM_003494.3), CAPN3 (NM_000070.2), FKTN (NM_001079802.1), TOR1AIP1 (NM_001267578.1), GTDC2 (NM_032806.5), GMPPB (NM_021971.1), LAMA2 (NM_000426.3), COL6A1 (NM_001848.2), COL6A3 (NG_008676.1,NM_004369.3), PYGM (NM_005609.2), CPT2 (NM_000098.2), DOK7 (NM_173660.4), ACTA1 (NM_001100.3), STIM1 (NM_003156.3), FLNC (NM_001458.4), CHD7 (NM_017780.3).

Downloaded From: https://jamanetwork.com/ on 09/29/2021 eTable 3. Variants of uncertain significance in two families Fami Age at Distribut Additional Organ Maxim Biopsy Causat ly Presentat ion of clinical involvem um CK Findings ive ion (yrs) weaknes characteri ent (U/L) gene s stics 28 11 Proximal Lumbar Restrictiv 7000 Dystrophi ?MSTN UL and lordosis e lung c features, LL and disease type 1 bilateral hypotroph scapular y swinging, increased calf muscle internal hypertroph nuclei of y 40%, focal endomysi al fibrosis and regenerati ng fibres 29 40 Proximal Mild finger Nil 2492 Dystrophi ?ANO5 LL extension c features, weakness predomin ance of type 1 atrophic fibres and ring fibres. Variants of uncertain significance in two families. The proband of Family 28 was adopted and DNA was unavailable from the parents to confirm segregation. In Family 29, muscle biopsy was unavailable for further RNA sequence analysis and familial segregation could not be confirmed. CK=creatine kinase, DG=dystroglycan. RefSeq: MSTN (NG_009800.1, NM_005259.2) & ANO5 (NM_213599).

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