Clinical improvements after treatment with a low‑valine and low‑fat diet in a pediatric patient with enoyl‑CoA hydratase, short chain 1 (ECHS1) deficiency

Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 h ps://doi.o g/10.1186/s13023-022-02468-6 RESEARCH Clinical imp o emen s a e  ea men wi halow- aline andlow- a die inapedia ic pa ien wi henoyl-CoA hyd a ase, sho chain 1 (ECHS1) de iciency Sil ia Pa a1, Ka he ine Flo es‑Rojas1,2, Angel Gil3,4,5* , Edua do López‑Laso2,6,7, Lau a Ma i‑Sánchez8,9, Heydi Baide‑Mai ena10,11, Belén Pé ez‑Dueñas10,11 and Me cedes Gil‑Campos1,2,5 Abs ac Backg ound: Enoyl‑CoA hyd a ase sho ‑chain 1 (ECHS1) is a key mi ochond ial enzyme ha is in ol ed in aline ca abolism and a y acid be a‑oxida ion. Mu a ions in he ECHS1 gene lead o enzyma ic de iciency, esul ing in he accumula ion o ce ain in e media es om he aline ca abolism pa hway. This dis up s he py u a e dehyd ogenase complex and he mi ochond ial espi a o y chain, wi h consequen cellula damage. Pa ien s p esen wi h a a iable age o onse and a wide spec um o clinical ea u es. The Leigh synd ome pheno ype is he mos equen ly epo ed o m o he disease. He ein, we epo a case o a male wi h ECHS1 de iciency who was diagnosed a 8 yea s o age. He p esen ed se e e dys onia, hype lo dosis, mode a e o se e e kyphoscoliosis, g ea di icul y in walking, and se e e dysa h ia. A aline‑ es ic ed and o al a ‑ es ic ed die was conside ed as a he apeu ic op ion a e he gene ic diagnosis. An a ailable o mula ha es ic ed b anched‑chain amino acids and especially es ic ed aline was used. We also es ic ed animal p o ein in ake and p o ided a low‑ a die ha was pa icula ly low in dai y a . Resul s: This p o ein‑ and a ‑ es ic ed die was ini ia ed wi h adequa e ole ance and adhe ence. A e h ee yea s, he pa ien no iced an imp o emen in dys onia, especially in walking. He cu en ly equi es minimal suppo o walk o s and. The e o e, he has enhanced his au onomy o go o school o es ablish a ca ee o himsel . His quali y o li e and mo i a ion o ea men ha e g ea ly inc eased. Conclusions: The e is s ill a subs an ial lack o knowledge abou his a e diso de , especially knowledge abou u u e e ec i e ea men s. Howe e , ea ly diagnosis and ea men wi h a aline‑ and a ‑ es ic ed die , pa icula ly dai y a ‑ es ic ed die , appea ed o limi disease p og ession in his pa ien wi h ECHS1 de iciency. Keywo ds: Child en, Enoyl‑CoA hyd a ase, Leigh synd ome, Die , Valine © The Au ho (s) 2022. Open Access This a icle is licensed unde a C ea i e Commons A ibu ion 4.0 In e na ional License, which pe mi s use, sha ing, adap a ion, dis ibu ion and ep oduc ion in any medium o o ma , as long as you gi e app op ia e c edi o he o iginal au ho (s) and he sou ce, p o ide a link o he C ea i e Commons licence, and indica e i changes we e made. The images o o he hi d pa y ma e ial in his a icle a e included in he a icle’s C ea i e Commons licence, unless indica ed o he wise in a c edi line o he ma e ial. I ma e ial is no included in he a icle’s C ea i e Commons licence and you in ended use is no pe mi ed by s a u o y egula ion o exceeds he pe mi ed use, you will need o ob ain pe mission di ec ly om he copy igh holde . To iew a copy o his licence, isi h p:// c ea i eco mmons. o g/ licen ses/ by/4. 0/. The C ea i e Commons Public Domain Dedica ion wai e (h p:// c ea i eco mmons. o g/ publi cdoma in/ ze o/1. 0/) applies o he da a made a ailable in his a icle, unless o he wise s a ed in a c edi line o he da a. Backg ound Enoyl-CoA hyd a ase, sho -chain 1 (ECHS1), EC 4.2.1.17, is also known as SCEH. I is a p o ein encoded by gene ID 1892, which is loca ed on ch omosome 10. The gene p oduc is a membe o he hyd a ase/isome ase supe amily ha localizes o he mi ochond ial ma ix. I is b oadly exp essed in he li e , kidney, and adipose is- sue, and i s exp ession in 22 o he issues, including he Open Access *Co espondence: agil@ug .es 3 Depa men o Biochemis y and Molecula Biology II, Ins i u e o Nu i ion and Food Technology “José Ma aix,” Biomedical Resea ch Cen e , Pa que Tecnológico de la Salud, Uni e si y o G anada, A enida del Conocimien o s/n, A milla, 18100 G anada, Spain Full lis o au ho in o ma ion is a ailable a he end o he a icle Page 2 o 10 Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 whole gas oin es inal ac , b ain, and nume ous sec e- o y glands, has been epo ed. T ansc ip a ian s u i- lizing al e na i e ansc ip ion ini ia ion si es ha e been desc ibed [1]. ECHS1 ca alyzes he hyd a ion o he double bond be ween he second and hi d ca bons o 2- ans-enoyl- coenzyme A (CoA) in e media es o L-3-hyd oxy-acyl- CoAs [2, 3]. I ca alyzes se e al me abolic pa hways. In pa icula , his c o onase ca alyzes he second s ep o be a-oxida ion o a y acids [4] and plays a key ole in aline ca abolism. Fu he mo e, i akes pa in he ou h s ep o his me abolic pa hway by ca alyzing he hyd a- ion o me hac ylyl-CoA o 3-hyd oxyisobu y yl-CoA and he con e sion o ac yloyl-CoA o 3-hyd oxyp o- pyl-CoA [5, 6]. ECHS1 is also in ol ed in he me abo- lism o isoleucine in he ans o ma ion o iglyl-CoA o 2-me hyl-3-hyd oxybu y yl-CoA (Fig.1). Mu a ions in he ECHS1 gene (OMIM* 602292) esul in ECHS1 de iciency, which is a a e au osomal ecessi e diso de ha was i s epo ed by Pe e s e al. [7] as a cause o Leigh synd ome (OMIM* 256000). Bo h he age a onse and clinical mani es a ions a y g ea ly om pa ien o pa ien . Masnada e al. [8] dis inguished ou pheno ypes o ECHS1 de iciency based on clinical and neu ological ea u es. The i s is o se e e neona al onse wi h a apidly a al cou se and whi e ma e changes. The second is a se e e in an ile p esen a ion wi h slowe neu ological de e io a ion and deep g ay nuclei degene - a ion, de elopmen al delay, py amidal and ex apy ami- dal signs, eeding di icul ies, and op ic a ophy. The hi d pheno ype is ano he slowly p og essi e ju enile o m ha is simila o he p e ious one bu wi h less se e i y and mainly basal ganglia in ol emen . The ou h pheno- ype is a less common o m o p esen a ion ha is based on pa oxysmal exe cise-induced dys onia wi h a no mal neu ological examina ion be ween hese a acks and isola ed degene a ion o he pallidum on magne ic eso- nance imaging [8]. A cha ac e is ic magne ic esonance imaging (MRI) pa e n o basal ganglia lesions has been epo ed, wi h small cys s in he pu amen and pallidum [9]. Ca diac in ol emen [4] and cu is laxa a e also clini- cal ea u es o his disease [10]. De iciency o ECHS1 usually leads o he accumula- ion o me hac ylyl-CoA and ac yloyl-CoA. These ha m- ul subs a es a e highly eac i e wi h sul hyd yl g oups and end o o m adduc s wi h cys eine o cys eamine [6, 7, 11]. High concen a ions o hese in e media e me aboli es seem o cause dys unc ion in he py u a e dehyd ogenase complex (PDC) and he mi ochond ial espi a o y chain, esul ing in cellula damage [12]. Addi ionally, inc eased exc e ion o 2-me hyl-2,3-dihy- d oxybu y ic acid in u ine is equen ly obse ed [13]. S-(2-ca boxyp opyl)-cys eine, S-(2-ca boxyp opyl)- cys eamine and N-ace yl-S-(2-ca boxyp opyl)-cys eine Fig. 1 Illus a ion o he sho ‑chain a y acid, odd‑chain a y acid and aline ca abolic pa hways, showing he o ma ion o ce ain me aboli es as a esul o sho ‑chain enoyl‑CoA hyd a ase 1 (ECHS1) de iciency. The concep ha 2,3‑dihyd oxy‑2‑me hylbu y ic acid o igina es om ac yloyl‑CoA was indi ec ly in e ed based on Pe e s e al. [6] Page 3 o 10 Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 ha e also been p oposed as use ul diagnos ic ma ke s [2, 14]. O he pa ien s p esen wi h inc eased plasma le els o lac a e and py u a e in a conse ed a io [15] (Fig.1). O e 50 pa ien s om mo e han 35 amilies ha e been epo ed o da e [9, 16]. Whole-exome sequencing (WES) has inc eased he ecogni ion o pa hogenic a ian s in ECHS1. Mos o hese a ian s a e missense a ian s [3], sugges ing ha en i e loss-o - unc ion mu a ions may be incompa ible wi h li e [17]. The majo i y o pa ien s wi h ECHS1 de iciency a e ini ially diagnosed by hei clinical ea u es and do no unde go molecula diagnosis un il la e in li e. They usu- ally ecei e s anda d ea men s ha a e used o o he mi ochond ial diseases. Anecdo al epo s o ea men wi h N-ace ylcys eine o de oxi y he eac i e me aboli es o me hac ylyl-CoA and ac yloyl-CoA ha e been pub- lished [18]. A aline- es ic ed die is a a ional biochem- ical app oach. Recen ly, a aline- es ic ed die appea ed o be bene icial o i e pa ien s wi h ECHS1 de ec s, imp o ing he biochemical and clinical mani es a ions o hese diso de s [19]. The adminis a ion o co ac o s, i.e., coenzyme Q10, i amins C and E, ibo la in, c ea ine monohyd a e and o he an ioxidan s, ha a e used in mi ochond ial diso de s wi h limi ed e idence o ben- e i has also been conside ed [18, 20, 21]. Fu he mo e, es ic ing he consump ion o speci ic a y acids has been es ed in se e al FAO diso de s [22]; his app oach aims o educe he syn hesis o oxic in e media es [23]. In addi ion, al hough high- a , educed-ca bohyd a e ke ogenic die s ha e been used in some cases o mod- e a e ECHS1 de iciency, hey ha e been ine ec i e in imp o ing he symp oms in se e e cases o he disease [24, 25]. Based on he p e ious backg ound, he e, we epo he case o an ECHS1-de icien child and how he use o a aline- and a - es ic ed die pa ially imp o ed his condi ion. Ma e ials andme hods Pa icipan The pa ien was a boy wi h ECHS1 de iciency who was diagnosed in 2017 in he Pedia ic Neu ology Uni o a hi d-le el hospi al in Spain. He was he i s bo n om a se o wins and was deli e ed by scheduled cesa ean sec- ion in 2006. He was concei ed ia he in i o e iliza- ion o wo o he mo he ’s eggs and he p egnancy was une en ul. His win sis e is heal hy. His pa en s a e also heal hy, and hei ma iage is no consanguineous. He had no mal neu ological de elopmen up o 15 mon hs o age. Ten days a e ecei ing a accina- ion, he su e ed om neu ological eg ession, wi h acu e a axia onse , gai loss, and py amidal signs. In addi ion, se e e dys onia a ec ed his daily ac i i ies, including walking, handling, and speech. Gene ally, he was smiling, happy, and engaged in good social in e ac ions. Feeding and sleep we e bo h ade- qua e, and he heigh - o-weigh a io was no mal. He ecei ed speech he apy, physical he apy, and psycho- pedagogical suppo . Molecula gene ic analysis This pa ien was diagnosed om a mul icen e p o- jec o pa ien s wi h basal ganglia diseases o unknown cause [9]. The ECHS1 a ian s we e alida ed and seg- ega ed using Sange sequencing. The gene ic analysis o a whole-blood sample iden i ied wo he e ozygous a ian s, c.123_124del p.(Gly42Glu s*3) and c.830C > T p.(Th 277Ile), in he ECHS1 gene (NM_004092.3, sequencing). The e o e, he esul was epo ed o be compa ible wi h he diagnosis o Leigh synd ome. Genomic DNA was ex ac ed om pe iphe al blood samples using s anda d me hods. The sequencing o all exons and in he lanking in onic egions ± 20bp was pe o med by Cen ogene (Ros ock, Ge many) using he Cen oXome Gold p o ocol and gua an eeing a dep h o eads o 100X. Typically, 97–98% o he a ge bases a e > 10 × co e ed. Nex , a p op ie a y bioin o ma - ics me hod was applied ha includes anno a ion o bases, alignmen o he hg19 e e ence human genome (Genome Re e ence Conso ium GRCh37), il e ing o low-quali y eads and possible a i ac s, and anno a ion o a ian s (Cen ogene, Ros ock, Ge many). Fo he med- ical e alua ion, we conside ed all he pa hogenic a ian s epo ed in The Human Gene Da abase (HGMD®) and ClinVa and all a ian s wi h minimal allelic equency (MAF) below 1% in Genome Agg ega ion Da abase (gno- mAD). All possible inhe i ance pa e ns we e consid- e ed based on he clinical in o ma ion con ibu ed. Only a ian s ela ed o he pheno ype o he pa ien we e epo ed. The pa ien ’s clinical in o ma ion and amily his o y we e used o e alua e he iden i ied a ian s. All iden i ied a ian s we e e alua ed based on hei pa ho- genici y and causali y, and hose associa ed wi h he phe- no ype o he pa ien ha we e no benign o p obably benign we e epo ed. Consis en wi h he gene ic analysis, espi a o y chain ac i i y was dec eased in he CII and CI + III complexes and PDC ac i i y was dec eased [9]. The esul s o he muscle biopsy s udy showed a de ici o mi ochond ial espi a o y chain complex I (le el 8.03, pa hological alue < 10). Addi ionally, signs o mi ochond ial p oli e a- ion, such as a ci a e syn hase alue o 1353 ( e e ence alue 200–900), we e obse ed. We only ha e quali a i e da a o PDC ac i i y. Page 4 o 10 Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 Physical examina ion andaddi ional es s A e he con i ma ion o he diagnosis and jus be o e s a ing ea men , he physical examina ion showed he p og essi e de elopmen o scanning speech, se e e dys onia, a axia, hypo onia wi h hype lo dosis and mode a e o se e e kyphoscoliosis, and he ex emi- ies we e in in e nal o a ion wi h g ea di icul y walk- ing. No abno mal eye mo emen s we e obse ed. The Bu ke-Fahn-Ma sden Dys onia Ra ing Scale (BFMDRS) showed an 18/30 disabili y sco e. The G oss Mo o Func ion Classi ica ion Sys- em (GMFCS) o unc ional mo o impai men , psychomo o delay, and in ellec ual disabili y was de e - mined o be le el IV by p incipal clinical judgmen s. Magne ic esonance imaging (MRI) pe o med in 2016 showed T2 hype in ensi y in bo h he cauda e and pu a- men nuclei, wi h mo e ma ked hype in ensi y in he pos- e io and medial le pu amen, sugges ing ca i a ion. No b ain a ophy o whi e ma e in ol emen was obse ed (Fig.2, Panels A and B). Bila e al op ic a ophy was de ec ed a e oph hal- mic e alua ion. Conce ning he biochemical ea u es, he enous acid–base analysis showed a base excess o 2.80mmol/L, HCO3 27.20mmol/L, pH 7.44 and lac a e 1.10mmol/L. Addi ionally, he ce eb ospinal luid (CSF) Fig. 2 The axial T2 sequence in ce eb al magne ic esonance a diagnosis (Panel A) and a e 3 yea s o die a y in e en ion (Panel C) and he co esponding luid‑a enua ed in e sion eco e y (FLAIR) images (Panel B, D) Page 5 o 10 Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 s udy, including amino acid and hiamine de e mina ion, was no mal, and he amino acid and acylca ni ine p o- iles measu ed in he plasma and o ganic acids in u ine we e also no mal. In pa icula , no mal u ina y exc e ion o 2-me hyl-2,3-hyd oxybu y ic acid was ound. T ea men A e gene ic diagnosis and based on he po en ially a ec ed me abolic pa hways, he die o his pa ien was modi ied. Valine and a , pa icula ly dai y a , we e es ic ed. This p o ein- and a - es ic ed die was ini i- a ed wi h adequa e ole ance and adhe ence. The nu i ional plan included he consump ion o a con enien , educed olume, eady- o-d ink p o ein sub- s i u e wice a day om 3yea s o age. This subs i u e is also used o o he congeni al inbo n e o s o me abo- lism, i.e., me hylmalonic and p opionic acidemias. The p oduc (MMA/PA PA coole ™) was supplied by Vi a- lo®, Spain, a Nes lé Heal h Science company, in a 130ml pouch (h ps:// www. nes l eheal hsci ence. es/ i a l o). Table1 shows he nu i ional composi ion o he sup- plemen . Indeed, he p o ein subs i u e p o ided all o he amino acids ound in die a y p o ein excep me hio- nine, h eonine, and aline and con ained only a small amoun o isoleucine. This p o ein subs i u e was used in combina ion wi h he amoun o p o ein ole a ed om he indi idual’s usual die . The oods in his die a ied. Howe e , he dai y p oduc s we e om skimmed milk, and he die was low in a y oods. This die also con- ained ela i ely high amoun s o complex ca bohyd a es a each eeding, especially h ough he in ake o b ead and pas a. The na u al p o eins o his die we e p o ided by small quan i ies o dai y p oduc s, ce ain ce eals, legumes (in a con olled way), gela in, and a minimum quan i y o ish. The maximum daily amoun o hese die- a y p o eins (in g ams) we e gi en o he amily. In addi- ion, ee ui and ege ables we e gi en. A quan i a i e 24h die a y eco d was e alua ed du - ing e e y isi . On a e age, he pa ien had a daily ene gy in ake o 1312kcal, o which 16.7% was de i ed om die a y p o ein (54.8g/d o al p o ein, o which 29.9g o p o ein equi alen was ob ained om he amino acid supplemen ), 70.4% (231g/d) was de i ed om ca bohy- d a es, and 12.9% (18.7g/d) was de i ed om lipids. Resul s A e wo yea s o his ea men , he child exhibi ed be e s abili y. He equi ed minimal suppo o walk o s and, which enhanced his au onomy a school and imp o ed his hygiene and pe sonal ca e ou ines. He had no ad e se e ec s and had a good nu i ional p o ile and g ow h. A e h ee yea s o ea men , his cu en weigh is 29.5kg (pe cen ile 4), his heigh is 144cm (pe cen ile 10), his body mass index (BMI) is 14.04 wi h a p epu- be al s a us. The amily con inues o pe o m die a y calcula ions as expec ed, complying wi h wha was p e- sc ibed. Clinical e alua ion and neu ological es s a e ca ied ou e e y 6mon hs. G oss mo o unc ion has imp o ed, and his GMFCS classi ica ion was educed om le el IV o le el III. The child now shows g ea e s abili y. He equi es less suppo when walking o s anding, his language speed has inc eased, and he has mo e au onomy (i.e., he is now able o ge d essed and showe alone). Mino hype lo dosis and kyphoscoliosis and ewe dys onic posi ions we e obse ed, imp o ing he abili y o he pa ien o walk. He is e en able o walk Table 1 Composi ion o he eady‑ o‑d ink p o ein subs i u e (1 pouch o 130 ml) used o he die a y managemen o he ECHS1 case EPA, eicosapen aenoic acid; DHA, docosahexaenoic acid a Suc ose, glucose sy up, and modi ied co n s a ch Ene gy (kcal) 103 Mine als To al amino acids (g) 18.2 Sodium (mg) 127 P o ein equi alen (g) 15.0 Po assium (mg) 235 Ca bohyd a es (g)17.0 Chlo e (mg) 182 Suga s (g) 5.9 Calcium (mg) 299 Lipids (g) 1.7 Phospho us (mg) 267 Sa u a ed a (g) 0.33 Magnesium 94 DHA (mg) 100 I on (mg) 5.4 EPA (mg) 23 Coppe (mg) 0.55 Amino acids (g) Zinc (mg) 5.4 Alanine 1.68 Manganese (mg) 0.8 A ginine 1.73 Iodine (μg) 63 Aspa ic acid 3.0 Molybdenum (μg) 36 Cys ine 0.73 Selenium (μg) 22 Glu amine 0.00 Ch omium (μg) 22 Glycine 0.79 Vi amins His idine 1.13 A (μg) 208 Isoleucine 0.05 D (μg) 7.5 Leucine 2.37 E (mg) 3.9 Lysine 1.85 K (μg) 25 Me hionine 0.00 C (mg) 27 Phenylalanine 1.13 Thiamine (B1) (mg) 0.50 P oline 0.83 Ribo la in (B2) (mg) 0.57 Se ine 1.25 Niacin (mg) (B5) 6.2 Th eonine 0.00 Py idoxamine (B6) (mg) 0.70 T yp ophan 0.47 Folic acid (B9) (μg) 100 Ty osine 1.13 B12 (μg) 1.2 Valine 0.00 Bio in (B7) 47 Ca ni ine (mg) 17 Pan o henic acid (B3)(mg) 2.0 Tau ine (mg) 33 Choline (mg) 150 Page 6 o 10 Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 up igh o 1h a day. He u ns wi h less di icul y, and he now has be e ine mo o skills and mino emo s. Fo example, he is now able o ea a yogu . He has a be e unde s anding and is conscien ious o he con- cep s o his in e es . His amily seeks social in eg a ion. He is adap ing o his high-school class, whe e he usu- ally mo es au onomously wi h a icycle. On he BFM- DRS, he disabili y sco e a e 3yea s imp o ed om 18/30 o 9/30. His quali y o li e and mo i a ion o con- inue his ea men ha e g ea ly inc eased. A e ea - men , he e was no signi ican p og ession as e alua ed by neu oimaging. In ac , MRI showed mo e ma ked hype in ensi ies in bo h he cauda e and pu amen nuclei, sugges ing inc eased ca i a ion (Fig.2, Panels C and D). A ideo wi h images o he pa ien immedia ely a e diagnosis and 3yea s a e ea men is included in he Addi ional ile1: Video. Rega ding he biochemical ea u es, Table 2 shows he plasma concen a ions o aline, leucine, and isole- ucine a he ime o diagnosis and a e 1.5 and 3yea s o ollow-up. Valine was dec eased du ing he clini- cal cou se compa ed wi h he baseline le el, while iso- leucine and leucine emained ai ly cons an . In he las analysis, he exc e ion o 2-me hyl-2,3-hyd oxybu y ic acid was 1 nmol/mol c ea inine, emaining wi hin he no mal alues (< 2 nmol/mol c ea inine). Addi ionally, no u ina y me hac ylyl-glycine was de ec ed. In addi- ion, he S-(2-ca boxyp opyl) cys eine concen a ion was 0.18mmol/mol c ea inine (no mal alue < 0.39) and ha o S-(2-ca boxye hyl)-cys eine was 0.23mmol/mol c ea inine (no mal alue < 0.15). Howe e , N-ace yl-S- (2-ca boxye hyl)-cys eine, N-ace yl-S-(2-ca boxyp opyl)- cys eine and S-(2-ca boxyp opyl)-cys eine ca ni ine we e unde ec able. In addi ion, he plasma le els o acylca ni- ines we e wi hin he ange o no mali y, and he le els o C4-OH [3-OH-bu y yl/3-OH-isobu y yl-ca ni ine] (0.03µmol/L) we e also wi hin he ange o no mali y (0.01–0.06µmol/L). Discussion The main inding o he p esen s udy was ha ea men o male child wi h an ECHS1 de iciency wi h a aline- and a - es ic ed die esul ed in an imp o emen in his quali y o li e and he amelio a ion o clinical ea u es. In ac , a e h ee yea s o ea men , he pa ien showed g ea e s abili y, equi ed less suppo when walking o s anding, had imp o ed language skills, and was mo e au onomous. In ou pa ien , a he ime o diagnosis, MRI de ec ed T2 hype in ensi y in he cauda e and pu amen nuclei. Howe e , no b ain a ophy o whi e ma e in ol emen was obse ed. In addi ion, ou pa ien exhibi ed bila - e al op ic a ophy soon a e he diagnosis. In ECHS1 de iciency pa ien s, mic ocephaly and impai ed ision a e usually de ec ed by physical examina ion [9]. These pa ien s also ha e di icul y ixing o ollowing objec s, nys agmus, and op ic ne e a ophy, which could explain ision p oblems [26]. In addi ion, hey show di icul ies in speech. Howe e , hey a e awa e o hei su oundings and communica e in o he ways, such as by sounds o smiling. These pa ien s ha e poo head con ol, so hey a e unable o si up o ambula e. This symp om occu ed in ou pa ien . Al hough clinical and biochemical ea u es could sug- ges ECHS1 de iciency, he diagnosis o his condi ion is mos ly es ablished by he iden i ica ion o biallelic pa ho- genic a ian s in he ECHS1 gene by molecula gene ic es ing. In he p esen case epo , he i s a ian in he ECHS1 gene, c.123_124del p. (Gly42Glu s*3), al e s he eading ame a codon 42. The new eading ame ends 2 posi ions la e in a s op codon. I is classi ied as likely pa hogenic (class 2) acco ding o he Ame ican Col- lege o Medical Gene ics and Genomics (ACMG) ec- ommenda ions. The second a ian in he ECHS1 gene, c.830C > T p.(Th 277Ile), causes an amino acid change om h eonine o isoleucine a posi ion 277. In ClinVa , his a ian is lis ed as likely pa hogenic bu does no p o- ide e idence suppo ing i s pa hogenici y (clinical s ud- ies, ID: 397,546). The e o e, i is classi ied as a a ian o unce ain signi icance (class 3) acco ding o ACMG ecommenda ions. U ina y exc e ion o 2-me hyl-2,3-dihyd oxybu y ic acid and o he me aboli es, such as S-(2-ca boxyp opyl)- cys eine, S-(2-ca boxyp opyl)-cys eamine, and N-ace yl- S-(2-ca boxyp opyl)-cys eine, is use ul o diagnose ECHS1 de iciency. These compounds ha e been p oposed as help ul bioma ke s o ini ial sc eening p ocesses [6]. Table 2 The plasma concen a ions o b anched‑chain amino acids in a case o ECHS1 de iciency ea ed wi h a aline‑ and a ‑ es ic ed die Amino acid (µmol/L) Diagnosis 1.5 y a e diagnosis 3 y a e diagnosis Range o no mali y Valine 158 95 104 219 ± 47 Isoleucine 39 40 36 58 ± 15 Leucine 88 92 93 115 ± 26 Page 7 o 10 Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 Despi e hese obse a ions, nei he 2-me hyl-2,3-dihy- d oxybu y ic acid no abno mal a y acylca ni ines and o ganic acids in he u ine we e de ec ed in ou pa ien . Mo eo e , lac a e le els in he plasma we e no signi i- can ly ele a ed (1.10 mmol/L), adding e idence o he ela i ely mild ac i i y o he PDC. A de ini i e cu e o he ECHS1 diso de has no ye been disco e ed. Despi e he encou aging esul s ha some s udies ha e epo ed abou clinical imp o emen in adap ing pa ien s’ die s, he e is sca ce li e a u e abou die a y ea men . Loupa y e al. [27] i s sugges ed ha ea ing pa ien s wi h equen ca bohyd a e- ich meals, in which he main sou ce o ene gy is ca bohyd a es and p o ein in ake is es ic ed, could be a good he apeu ic op ion o 3-hyd oxyisobu y yl-CoA hyd olase (HIBCH) de iciency. HIBCH de iciency is a mi ochond ial disease ha is simila o ECHS1 de iciency. La e , Sole -Al onso e al. [28] showed ha a HIBCH-de icien child clinically esponded o a aline- es ic ed die using a a ge ed o mula o maple sy up u ine disease. Gi en he clini- cal and pa hophysiological simila i ies be ween HIBCH and ECHS1 de iciencies, i was hough ha he same es ic ed die a y ea men could be po en ially ben- e icial in ECHS1 subjec s. Acco ding o wha had been assumed, some s udies demons a ed imp o emen in clinical ou comes a e a aline- es ic ed die wi h N-ace ylcys eine supplemen a ion [29, 30]. Recen ly, Abdenu e al. [19] showed enhancemen in bo h clinical and biochemical mani es a ions, wi h a dec ease in S-(2- ca boxyp opyl)-cys eine in pa ien s wi h ei he HIBCH o ECHS1 de iciencies. In he p esen case, die a y ea men was ini ia ed a e se e al ole ance es s using an a ailable o mula ha was designed o pa ien s wi h me hylmalonic and p opionic acidemia (MMA/PA). This MMA/PA coole 15 is a eady- o-d ink me hionine- ee, h eonine- ee, aline- ee and low isoleucine medical ood, and i was gi en in di e en amoun s du ing he day. In addi ion, he pa ien ’s die was es ic ed o animal p o eins, leg- umes, and de a ed o skimmed p oduc s. Howe e , he in ake o na u al p o eins and ca bohyd a es had o be ensu ed. We se he consump ion limi o high-biologi- cal alue p o eins a 1g/kg/day. Plasma amino acids we e moni o ed du ing ea men , and he amily o was asked o adjus he die when low le els we e de ec ed. In ou case, he aim o speci ically es ic ing aline was ocused on educing neu o oxici y due o enzyma ic blockade and he accumula ion o me aboli es in he ca - abolic pa hway. Mi ochond ia play a key ole in he consump ion o ene gy in euka yo ic cells by oxidizing a y acids and sug- a s o gene a e ATP. Mi ochond ial a y acid β-oxida ion (FAO) and oxida i e phospho yla ion (OXPHOS) a e wo key pa hways in ol ed in his p ocess, and ECHS1 is a key enzyme in ol ed in mi ochond ial FAO. Fo he e y-long-chain and long-chain a y acid (C14–24) a y acyl-CoA es e s, he second, hi d and ou h s eps o β-FAO a e pe o med by he mul idomain mi ochond ial i unc ional p o ein (MTP). Howe e , medium-(C8– C12) and sho -chain (C4–C6) a y acyl-CoA es e s a e me abolized ia a di e en se o enzymes ha pe o m hyd a ion (s ep 2; ECHS1). Dis up ion o FAO can cause human disease. Howe e , pa ien s wi h de iciencies in he FAO enzyme ECHS1 a e ypically diagnosed wi h Leigh synd ome, which is no - mally associa ed wi h OXPHOS dys unc ion [31]. In ou pa ien , he esul o he muscle biopsy s udy showed a de ici o he mi ochond ial espi a o y chain complex I (le el 8.03, pa hological alue < 10) oge he wi h signs o mi ochond ial p oli e a ion, such as a ci a e syn hase alue o 1352.98 ( e e ence alue 200–900). Fo PDC ac i i y, we only ha e quali a i e da a ha suppo s mild ac i i y. In e es ingly, he e may be a co ela ion be ween he ECHS1 pheno ypic se e i y and PDC ac i i y [31]. Pa ien s wi h low PDC ac i i y also ha e high lac a e le - els. Se e al o hese pa ien s p esen wi h a mo e se e e p ognosis [4, 7, 32], including dea h wi hin 48h o bi h [2, 33]. In con as , milde cases o ECHS1 de iciency do no show educed PDC ac i i y o lac ic acidosis [6, 12, 34, 35]. Since ECHS1 de iciency is associa ed wi h se e e aci- dosis and low ATP om impai ed ae obic oxida ion usually occu s, we hypo hesized ha a die wi h a ela- i ely low con en o a and a pa icula ly low con en o medium- and sho -chain a y acids migh be bene icial o pa ien s wi h impai ed OXPHOS. Res ic ing he consump ion o speci ic a y acids has been es ed in se e al FAO diso de s [22], aiming o educe he syn hesis o oxic in e media es [23]. In addi- ion, al hough high- a , educed-ca bohyd a e ke ogenic die s ha e been used in some cases o mode a e ECHS1 de iciency, hey ha e been ine ec i e in imp o ing he symp oms o he disease in se e e cases [24, 33]. Bo ine milk lipids con ain app oxima ely 12 a y acids in amoun s g ea e han 1%. Bu y ic acid (4:0) and cap oic acid (6:0) a e he mos impo an subs a es o ECHS1, and hey ep esen 11.8% and 4.6%, espec- i ely, o he o al iacylglyce ol a y acids. In addi ion, all o he 4:0, 83% o 6:0 and 63% o 8:0 a e es e i ied a he sn-3 posi ion and a e easily diges ed by gas ic and panc ea ic lipases [36, 37]. These sho and medium- chain a y acids each he sys emic ci cula ion and eely di use ac oss he plasma memb ane. They en e di ec ly in o mi ochond ia wi hou he assis ance o he ca ni ine shu le sys em [38]. They a e apidly oxi- dized in he li e , muscle, and o he pe iphe al issues. Page 8 o 10 Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 Hence, educing he amoun o o al a , pa icula ly dai y a , in he die could educe he impac o FAO on he symp oms o mi ochond ial ECHS1 de iciency. A limi a ion o his s udy is ha impo an biochemi- cal ma ke s, such as S-(2-ca boxyp opyl)-cys eine, S-(2-ca boxye hyl)-cys eine, N-ace yl-S-(2-ca boxye hyl)- cys eine, N-ace yl-S-(2-ca boxyp opil)-cys eine, and S-(ca boxyp opyl)-cys eine ca ni ine, we e no meas- u ed a he beginning o die a y ea men , so we canno compa e hem wi h cu en le els. Ano he limi a ion o ou s udy is ha he pa ien has a mild pheno ype, as e i- denced by low le els o al e ed me aboli es as well as an imp o ed clinical cou se. In only wo yea s o ea men , he pa ien has p og essed om g ea di icul y walking, se e e dys onia and hypo onia wi h se e e hype lo dosis and kyphoscoliosis o showing be e s abili y, less dys- onic posi ions, and much mo e au onomy in walking and daily ou ine. He has also demons a ed g ea social in e- g a ion in school due o mo e luen language and be e unde s anding. In his a e disease, clinical esul s should be consid- e ed wi h cau ion because he e is no li e a u e abou how o pe o m a clinical ollow-up and wha es s could be pe o med o bes ca y ou a speci ic e alua ion o con ol i s p og ession o imp o emen . The alida ed in e na ional es s used, BFMDRS and GMFCS, we e applied o pe o m a neu ological e alua ion based on mo o unc ion and disabili y. The BFMDRS gi es a dis- abili y sco e, and i is a uni e sally applied ins umen o he quan i a i e assessmen o dys onia in bo h chil- d en and adul s. I should be used longi udinally, as i was used o his pa ien , e e y 6mon hs, and he age o he pa ien should be conside ed. The GMFCS meas- u es he changes in g oss mo o unc ion in child en wi h ce eb al palsy o simila symp oms. The e o e, i is used bo h o desc ibe a child’s cu en abili ies and o quan i y changes in unc ion o e ime as a esul o de elopmen , he apy, o aining. In addi ion, ex ensi e biochemical e alua ion, pa icu- la ly hose ha a e ela ed o he me aboli es associa ed wi h ECHS1 de iciency, was no pe o med by clinicians be o e ea men . This is because o di icul ies in he hos- pi al in ca ying ou speci ic biochemical analyses bu also because o sca ce knowledge abou he biochemical pa h- ways ela ed o his ul a a e disease. Mo eo e , he di ec e ec o a low- aline and low- a die on hose me abo- li es in he b ain canno be di ec ly known, and pe haps hese me aboli es a e ound in di e en concen a ions in he b ain han in blood. The e o e, whe he he die esul ed in biochemical amelio a ion, as i seems o ha e done o an ex en , canno be p o en in he p esen case. Indeed, he clinical imp o emen s a e 3yea s on his low- aline, low- a die could be a o ui ous esul , as we did no ob ain clea biochemical e idence o imp o e- men . Howe e , he p og ession o he pa ien , p io o he gene ic diagnosis, was e iden , and clinical s abiliza- ion wi h some imp o emen s would seem o be associ- a ed wi h ou easonable assump ion o a di ec e ec o he die . In he u u e, we hope o be able o p o ide new e idence ela ed o he con inui y o he die in he long e m since i does no cause ad e se e ec s. Th ough hese u u e s udies, i s e ec can be con i med. Conclusion The e is s ill a subs an ial lack o knowledge abou his a e diso de . especially knowledge o u u e e ec i e ea men s. The die a y app oach o ECHS1 has shown bene i s in mul iple pa ien s. Howe e , he co ec o - mula ion be ween a aline- es ic ed die o a combined aline- a es ic ion will need addi ional pa ien in o - ma ion. Indeed, ea ly diagnosis and he he apeu ic use o a aline- and a - es ic ed die suppo ed by a speci ic and longi udinal clinical e alua ion and me aboli e meas- u emen s could help o con i m i s e ec s on clinical dis- ease p og ession in pa ien s wi h ECHS1 de iciency. Abb e ia ions ACMG: Ame ican College o Medical Gene ics and Genomics; BFMDRS: Bu ke‑ Fahn‑Ma sden Dys onia Ra ing Scale; BMI: Body Mass Index; CFS: Ce eb ospi‑ nal luid; CoA: Coenzyme A; C4OH: 3‑OH‑isobu y yl ca ni ine; ECHS1: Enoyl‑ CoA hyd a ase, sho ‑chain 1; FAO: Fa y acid oxida ion; GMFCS: G oss mo o unc ion classi ica ion sys em; GnomAD: Genome agg ega ion da abase; HGMD®: The human gene da abase; HIBCH: 3‑Hyd oxyisobu y yl‑CoA hyd o‑ lase; MAF: Minimal allelic equency; MMA/PA: Me hylmalonic and p opionic acidemia; MRI: Magne ic esonance imaging; MS: Mass spec oscopy; MTP: Mi ochond ial i unc ional p o ein; OXPHOS: Oxida i e phospho yla ion; WES: Whole exome sequencing. Supplemen a y In o ma ion The online e sion con ains supplemen a y ma e ial a ailable a h ps:// doi. o g/ 10. 1186/ s13023‑ 022‑ 02468‑6. Addi ional ile1. This ideo shows he clinical e olu ion o he pa ien wi h ECHS1 de iciency be o e and 3 yea s a e ea men wi h a low‑ aline and low‑ a die . One can obse e how a e ea men he shows be e s abili y, less walking and u ning di icul ies, an imp o emen in ine mo o skills and emo s. Acknowledgemen s The esea che s hank he pa ien and his amily and Raquel Muñoz Sánchez, a adiologis a he Reina So ía Uni e si y Hospi al, o hei pa icipa ion in he p esen s udy. Au ho con ibu ions AG designed he manusc ip and con ibu ed o he biochemical explana‑ ion o he ela ionship be ween biochemical e en s and die a y esponses. S.P. and MG‑C ob ained he clinical esul s and w o e he manusc ip . KF‑R included he in o ma ion ela ed o he die . EL‑L inco po a ed neu ologic e alua ions and da a. LM‑S, HB‑M, and BP‑D we e esponsible o he Page 9 o 10 Pa ae al. O phane Jou nal o Ra e Diseases (2022) 17:340 biochemical and molecula analyses. All he au ho s ha e e iewed and app o ed he inal e sion o his manusc ip . Funding The biochemical and gene ic analyses we e unded by Ins i u o de Salud Ca los III, G an /Awa d Numbe : PI18/01319, CERCA P og amme/Gene ali a de Ca alunya, Agència de Ges ió d’Aju s Uni e si a is i de Rece ca (AGAUR), Cen o de In es igación Biomédica en Red de En e medades Ra as (CIBERER), and Ins i u o de Salud Ca los III (ISCIII), G an /Awa d Numbe s: PI19/01310, PI16/01048. A ailabili y o da a and ma e ials All da a gene a ed o analyzed du ing his s udy a e included in his published a icle and i s Addi ional iles. Decla a ions E hics app o al and consen o pa icipa e W i en in o med consen was ob ained om he pa ien ’s pa en s o pa icipa ion in he s udy. All p ocedu es we e pe o med unde he e hical s anda ds o good clinical p ac ice and we e in acco dance wi h he Decla a‑ ion o Helsinki. This s udy was app o ed by he Resea ch E hics Commi ee o Có doba. Consen o publica ion W i en in o med consen was ob ained om he pa en s o ou pa ien o he publica ion o his s udy and he use o he ideo. Compe ing in e es s The au ho s decla e ha hey ha e no compe ing in e es s. Au ho de ails 1 Pedia ic Resea ch and Me abolism Uni , Reina So ia Uni e si y Hospi al, Uni‑ e si y o Có doba, 14010 Có doba, Spain. 2 Maimónides Ins i u e o Biomedi‑ cal Resea ch o Có doba (IMIBIC), Có doba, Spain. 3 Depa men o Biochemis‑ y and Molecula Biology II, Ins i u e o Nu i ion and Food Technology “José Ma aix,” Biomedical Resea ch Cen e , Pa que Tecnológico de la Salud, Uni e ‑ si y o G anada, A enida del Conocimien o s/n, A milla, 18100 G anada, Spain. 4 Ins i u o de In es igación Biosani a ia IBS.GRANADA, A milla, 18100 G anada, Spain. 5 CIBEROBN (Physiopa hology o Obesi y and Nu i ion), Ins i u o de Salud Ca los III (ISCIII), 28029 Mad id, Spain. 6 Pedia ic Neu ology Uni , Reina So ia Uni e si y Hospi al, 14010 Có doba, Spain. 7 CIBERER (Ra e Diseases), Ins i u o de Salud Ca los III (ISCIII), 28029 Mad id, Spain. 8 Depa men o Clini‑ cal Biochemis y, Ins i u de Rece ca San Joan de Déu, Ba celona, Spain. 9 Uni e si a de Ba celona, Ba celona, Spain. 10 Pedia ic Neu ology Resea ch G oup, Hospi al Vall d’Heb ón, Ba celona, Spain. 11 Uni e si a Au ònoma de Ba celona, Ba celona, Spain. 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