J Med Genet 2000;37:321–335 321

Review article J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from

The Smith-Lemli-Opitz syndrome

Richard I Kelley, Raoul C M Hennekam

Abstract genitalia. In all informative families, segregation The Smith-Lemli-Opitz syndrome (SLOS) of either form of SLOS was consistent with is one of the archetypical multiple congeni- autosomal recessive inheritance.212The genetic tal malformation syndromes. The recent cause of the disorder was not suspected for many discovery of the biochemical cause of years, although by the mid 1980s a number of SLOS and the subsequent redefinition of abnormalities of metabolism in SLOS SLOS as an inborn error of had been reported, including enlarged, lipid metabolism have led to important new depleted adrenal glands17 and aberrant patterns treatment possibilities for aVected pa- of steroid sulphates in plasma and urine.12 17 18 tients. Moreover, the recent recognition of Nevertheless the primary defect remained un- the important role of cholesterol in verte- known until Natowicz and Evans19 found that a brate embryogenesis, especially with re- patient with SLOS had essentially undetectable gard to the hedgehog embryonic signalling levels of normal urinary bile acids. An analysis of pathway and its eVects on the expression of that patient’s plasma sterols led to the homeobox genes, has provided an explana- discovery20 that the patient had a more than tion for the abnormal morphogenesis in 1000-fold increase in the level of the syndrome. The well known role of chol- 7-dehydrocholesterol (cholesta-5,7-dien-3beta- esterol in the formation of steroid hor- ol; 7DHC), suggesting a deficiency of mones has also provided a possible 7-dehydrocholesterol reductase (DHCR7), the explanation for the abnormal behavioural final step in the Kandutsch-Russell cholesterol characteristics of SLOS. biosynthetic pathway.21 The same sterol pattern (J Med Genet 2000;37:321–335) has subsequently been found in most patients with either type of SLOS, as well as in patients http://jmg.bmj.com/ Keywords: Smith-Lemli-Opitz syndrome; cholesterol with variant syndromes that could not be metabolism; 7-dehydrocholesterol reductase; clinical history; management assigned the diagnosis of SLOS on clinical grounds alone.22 23 Although initial evidence suggested that the human gene for SLOS was History located at 7q32.1, the human DHCR7 gene was The Smith-Lemli-Opitz syndrome was first later cloned and localised to chromosome described in 1964 by the late David Smith, the 11q12-13 by Moebius et al.24 Shortly afterwards, on September 30, 2021 by guest. Protected copyright. Belgian paediatrician Luc Lemli, and John three groups independently reported apparently 1 Opitz in a report of three patients who had in disabling mutations of DHCR7 in patients with common a distinctive facial appearance, micro- SLOS.24–26 cephaly, broad alveolar ridges, hypospadias, a characteristic dermatoglyphic pattern, severe feeding disorder, and global developmental Table 1 Findings in 167 clinically diagnosed cases of The Johns Hopkins Smith-Lemli-Opitz syndrome compared with 164 University, Kennedy delay. A more complete delineation of SLOS biochemically confirmed cases Krieger Institute, 707 was presented in 1969 as the “RSH syndrome”, North Broadway, a non-descriptive acronym of the first letters of Clinically Biochemically Baltimore, Maryland 2 diagnosed confirmed the original patients’ surnames. The description Finding (%) (%) 21205, USA of many new cases of SLOS over the next 20 R I Kelley years expanded the known characteristics of the Mental retardation 97 95 Postnatal growth retardation 85 82 Institute for Human syndrome, especially in the recognition of multi- Microcephaly 80 84 2–11 Genetics and ple internal anomalies (table 1). Many Structural brain anomalies 60 37 Department of aVected children died in the first year from fail- Ptosis 69 70 23 22 Paediatrics, Academic ure to thrive and infections, but many others Anteverted nares 90 78 Medical Centre, survived to adulthood. Somewhat later, several Cleft palate* 51 47 University of authors described patients with a lethal syn- Congenital heart defect 50 54 Amsterdam, Abnormal lung lobation 40 45 Meibergreef 15, 1105 drome that resembled SLOS, and which they Pyloric stenosis 15 14 12–16 AZ Amsterdam, designated “type II SLOS”. These children Colonic aganglionosis 12 16 Renal anomalies 40 43 The Netherlands had many of the anomalies found in SLOS, but Genital anomalies 74 65 R C M Hennekam died in the newborn period from internal 2/3 toe syndactyly 85 97 malformations. Moreover, most 46,XY “males” Polydactyly† 52 48 Correspondence to: Dr Hennekam, with so called type II SLOS had severe *Includes cleft soft palate, submucous cleft, and cleft uvula. [email protected] hypogenitalism or female appearing external †Includes postaxial polydactyly of hand(s)/foot. 322 Kelley, Hennekam

Clinical overview lar genetic studies confirmed that the diVer-

GENERAL ences in severity between types I and II SLOS J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from Although most published reports of SLOS are explained by the severity of the mutations describe only a single patient or a small number responsible. However, these results do not pre- of patients, there are a few exceptions in which clude the existence of genetic heterogeneity in 10 or more cases are described.2 9 12 23 27–29 Fur- SLOS; recently two mildly aVected sibs were thermore, several excellent older reviews found to have probably a related but biochemi- exist.10 15 30 A tabulation of the major clinical cally diVerent disorder of sterol metabolism.33 features of patients described in suYcient detail is provided in table 1. A comparison of INCIDENCE the most common characteristics of patients The earliest estimate of the incidence of SLOS ascertained by clinical v biochemical diagnosis was made by Lowry and Yong,34 who found an shows similar frequencies of the physical incidence in British Columbia of 1/40 000 anomalies. Only structural brain anomalies and births and a carrier frequency of 1%. The inci- anomalies of the genitalia are somewhat more dence in that survey increased to 1/20 000 common in the clinically diagnosed group. births when suspected but less definite cases Before the recognition of the biochemical cause were included. The incidence in a completely of SLOS, several authors suggested division of ascertained newborn population in Middle Bohemia (Czech Republic) was estimated to be SLOS into the relatively less expressed type I 35 form and the more severe type II form.12 13 31 greater than 1 in 10 000. However, in contrast Curry et al12 cautiously suggested that type I to these relatively high incidences based exclu- and type II might diVer genetically, whereas sively on clinical diagnosis, the incidence of others thought the subdivision was not SLOS diagnosed biochemically in similar justified.32 To address this question, Bialer et populations appears to be much lower. For example, between 1995 and 1998, when al15 devised a scoring system to evaluate the knowledge of the biochemical defect was wide- clinical severity of SLOS. Upon scoring 122 spread, the two laboratories that perform at published cases, they found a unimodal least 80% of biochemical testing for SLOS in frequency distribution of the scores for various the United States identified only about 40 new anomalies, which was interpreted as evidence cases per year, or an estimated incidence of less for a continuum of severity in SLOS and than 1 in 60 000 births (Kelley and Tint, against a genetic distinction between types I unpublished data). Similarly, estimates of only and II SLOS. However, because length of sur- 1 in 60 000 newborns in the United vival was part of the original scoring in addition Kingdom,27 1 in 80 000-100 000 births in The to separate scores for individual visceral Netherlands (Waterham et al, unpublished anomalies, the original scoring system of Bialer 36 15 data), and an even lower incidence in Japan et al was overweighted for internal anomalies. have been reported. The number of cases with 23 37 38 39 40 Therefore, we (Kelley and Hennekam, unpub- African, Asian, or South American http://jmg.bmj.com/ lished data) modified the Bialer score, to ancestry is also low. Although there remains weight embryologically separate organ systems some uncertainty about the absolute incidence equally. The revised scoring system (table 2) of SLOS in some countries, there clearly are has been used in two series of biochemically strikingly diVerent incidences among various 23 27 confirmed cases and showed a continuum ethnic groups. Both heterozygote advantage of severity and strong correlations with various and founder eVect have been suggested to biochemical parameters. Subsequent molecu- explain the relatively higher incidence of SLOS on September 30, 2021 by guest. Protected copyright. Table 2 Adapted (from ref 15) severity score for anatomical abnormalities in among those of European descent. The per- Smith-Lemli-Opitz syndrome centage of patients born to consanguineous parents is relatively low for an autosomal reces- Organ Score Criteria sive entity,23 27 35 which suggests persistence in Brain 1 Seizures; qualitative MRI abnormality the population of multiple mutant alleles 2 Major CNS malformations; gyral defects through heterozygote advantage. Oral 1 Bifid uvula or submucous cleft 2 Cleft hard palate or median cleft lip Acral 0 Non-Y shaped minimal toe syndactyly CRANIOFACIAL CHARACTERISTICS 1 Y shaped 2/3 toe syndactyly; club foot; upper or lower In the following listing of cases with various polydactyly; other syndactyly 2 Any two of the above features, no indication is made of whether a Eye 2 Cataract; frank microphthalmia case was biochemically confirmed or not. In Heart 0 Functional defects general, cases published after 1994 will have 1 Single chamber or vessel defect 2 Complex cardiac malformation been biochemically confirmed, but in cases Kidney 0 Functional defect from earlier papers this remains unknown. The 1 Simple cystic kidney disease “SLOS face” is highly characteristic of the syn- 2 Renal agenesis; clinically important cystic disease Liver 0 Induced hepatic abnormality drome and easily recognised in most patients, 1 Simple structural abnormality but may be very subtle in some.27 41 The most 2 Progressive liver disease salient features are microcephaly, bitemporal Lung 0 Functional pulmonary disease 1 Abnormal lobation; hypoplasia narrowing, ptosis, a short nasal root, an- 2 Pulmonary cysts; other major malformations teverted nares, and a small chin (fig 1). Bowel 0 Functional GI disease Congenital microcephaly is very common 1 Pyloric stenosis 2 Hirschsprung disease (for exact percentages of this and other symp- Genitalia 1 Simple hypospadias toms see table 1). Although a prominent ridge 2 Ambiguous or female genitalia in 46,XY; frank genital along the metopic suture can often be palpated, malformation in 46,XX true craniosynostosis is uncommon.212 More The Smith-Lemli-Opitz syndrome 323 J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from

Figure 1 Patients with Smith-Lemli-Opitz syndrome. than half of the patients have ptosis, often sequence has been found in asymmetrical or unilateral. The palpebral patients with abnormal cholesterol metabo- fissures are usually straight, but both upward lism.58 In some patients, the mouth is large, and downward slanting occurs. Other less which, combined with frequent micrognathia, frequent external eye anomalies include hyper- gives a distinctive appearance.59 More severe telorism, epicanthic folds, absence of lacrimal micrognathia, including the classical Pierre puncta,42 and unusually long cilia.43 Mild Robin sequence, is not rare in SLOS. exophthalmos has not been reported often,2 The intraoral anomalies of SLOS are diag- but is relatively commonly seen in figures in nostically important. The palate is usually

publications. Ocular defects include mainly highly arched, often with a midline cleft of the http://jmg.bmj.com/ congenital and occasionally postnatal cata- uvula, soft palate, or hard palate. In addition, racts, strabismus, and nystagmus.11 44–46 Vision the alveolar ridges typically are abnormally is usually normal. Less common symptoms are broad and conspicuously ridged. The tongue sclerocornea,13 43 47 heterochromia iridis,48 colo- can be small with redundant sublingual tissue12 boma of the iris,16 42 posterior synechiae,64549 or sublingual cysts13 31 in the more severely glaucoma,212retinal hyperpigmentation,2 optic aVected children. More rarely, the tongue is atrophy,21245 optic pits,42 microphthalmia,523 bifid.27 Crowded teeth and widely spaced inci- 27 44 52 and abnormalities of eye movements.44 Aniridia sors are not uncommon, and oligodontia on September 30, 2021 by guest. Protected copyright. was reported by Gracia et al,50 but the diagno- or polydontia,44 unusually large central upper sis in this mentally normal child with bilateral incisors,27 59 enamel hypoplasia,10 and prema- gonadoblastoma may be questioned. ture tooth eruption60 have been reported. A A hallmark of the syndrome is the shape of detailed dental study in SLOS, however, has the nose; usually the nasal bridge and base are not been published. Pharyngeal abnormalities 13 broad, the nasal root short, and the nares have included a small larynx and small vocal anteverted. The nasal bridge can be flat or cords with a subglottal shelf of excess fibrocar- 12 high, often with a striking capillary haemangi- tilaginous tissue. oma extending across the glabella. The degree The neck can appear short and excessive of anteversion of the nares decreases with age, skin folds or nuchal oedema are common, 61 but remains distinct in many adults.27 48 59 Nar- especially prenatally. row or atretic choanae may occur.51 The ears often appear low set and posteriorly CENTRAL NERVOUS SYSTEM rotated, but are otherwise unremarkable. The structural brain abnormalities of SLOS Rarely, the ear canals have been reported to be have been reviewed by Garcia et al,62 Cherstvoy very small.852Congenital sensorineural hearing et al,63 and Marion et al.64 In addition to micro- deficits may aVect as many as 10% of patients, cephaly, which is almost universal in SLOS, but many more severely aVected children are common abnormalities include enlarged not tested. Anomalies of the inner ear shown by ventricles,21252636566 hypoplastic or absent radiography or found at necropsy have been corpus callosum,12 27 57 63 66 67 hypoplastic fron- described.53 54 The philtrum is long, as can be tal lobes,39 and pituitary lipoma.68 Cerebellar expected in shortening of the nose. Although hypoplasia, sometimes with severe hypoplasia cleft lip has been reported in SLOS,13 55–57 only or aplasia of the vermis, is also not the “midline” type of cleft associated with the uncommon.25927445253626467Various forms of 324 Kelley, Hennekam

the holoprosencephaly sequence occur in has been found in only one biochemically con- 513172358 87 about 5% of patients. On histological firmed patient, who also had a de novo J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from examination of the brain, the most important balanced chromosome translocation. findings are disturbed cerebral neuronal 12 27 64 67 69 72 73 12 migration, extensive gliosis, dys- GENITAL ANOMALIES plasia of the medial olivary nuclei, and ectopic The genitalia in male SLOS patients range Purkinje cells.944 Several authors27 44 62 67 70–72 74 from normal to the appearance of complete have reported maturational abnormalities of sex reversal.12–15 60 65 Classically, hypospadias the white matter; however, most cranial MRI varies from coronal to perineoscrotal hypospa- studies do not show white matter abnormali- dias, although the latter is uncommon except ties. Although seizures are not uncommonly in the biochemically most severely aVected reported in SLOS, including infantile cases. Maldescent of the testes is common, spasms,32 75 they are uncommon in biochemi- but, even with severely malformed genitalia, cally proven cases of SLOS and may not be the testes are often easily palpated in the scro- substantially more frequent than in children tum, which is sometimes bifid.72 88 Mullerian without SLOS.237274852626976The same reser- duct derivatives are usually absent in 46,XY vation must be applied to a number of central males, as expected, but blind ending vagina, nervous system malformations reported in rudimentary or bicornuate uterus, and persist- older SLOS publications before biochemical ent cloaca have been described.12 13 15 31 51 89 confirmation was available. The gonads vary from normal testes to ovotes- tes to normal ovaries, or may be missing.5 In SKELETAL ANOMALIES females, the external genitalia may appear The skeletal anomalies have been reviewed in normal or there may be distinct hypoplasia of detail.10 15 23 27 63 Bilateral or unilateral postaxial the labia majora and minora. There are also polydactyly can be present in the hands or, less single reports of premature thelarche and high commonly, the feet, or both. Preaxial polydac- serum prolactin levels in a 15 month old girl tyly has not been reported in a biochemically with SLOS,90 and a malignant germ cell proven case. The thumb is generally short and tumour with a contralateral streak gonad in proximally placed and the first metacarpals and another female.91 Menstrual function is often thenar eminences are typically hypo- irregular but otherwise normal in most SLOS plastic.2 12 77–79 Other unusual digital abnor- adolescent females and adults, although me- malities include ectrodactyly,56 70 80 monodac- narche is often delayed. One adolescent girl tyly,80 oligodactyly,56 81 82 radial agenesis,80 with (biochemically unproven) SLOS and brachydactyly, absent middle phalanx of the borderline intelligence gave birth to an appar- second finger, radial or ulnar deviation of the ently normal daughter.34 fingers, clinodactyly, camptodactyly, and vari- 2101227567080 ous syndactylies. Rhizomelic and CARDIOVASCULAR ANOMALIES

mesomelic limb shortness and, more rarely, The cardiac anomalies of SLOS have been http://jmg.bmj.com/ “chondrodysplasia punctata” occur in SLOS, reviewed by Robinson et al,7 Johnson,10 and, but a true chondrodystrophy is not most extensively, by Lin et al.92 Almost half of found.12 13 16 23 27 29 63 70 Dermatoglyphics in SLOS patients have a congenital heart defect, SLOS have been reported to be distinctive with although if only biochemically confirmed an increased proportion of whorls and de- patients are taken into consideration, this creased proportion of ulnar loops on the finger percentage is somewhat lower.23 27 92 There is a tips in most series,2101252 but not all.83 The strong predominance of endocardial cushion presence of whorls may point to a pathogenesis defects and the hypoplastic left heart sequence, on September 30, 2021 by guest. Protected copyright. through puVy finger tip pads or oedema during whereas conotruncal defects are uncommon. early fetal development. There is no study as Almost every known cardiac defect has been yet of dermatoglyphic findings in a group of described at least once. The five most prevalent biochemically proven cases, but whorls appear defects found in a study of 95 biochemically to predominate as commonly reported in the confirmed cases of SLOS were atrioventricular early case reports. canal (25%), primum atrial septal defect One of the most consistently present anoma- (20%), patent ductus arteriosus at term (18%), lies in SLOS is the distinctive “Y shaped” cuta- and membranous ventricular septal defect neous syndactyly of the second and third toes, (10%).92 Lin et al92 hypothesised that the which has been reported in up to 99% of abnormal development of the extracellular biochemically proven cases.23 27 Postaxial poly- matrix may be the cause of both the cardiac dactyly of the feet is common in severe SLOS, defects and the absence of ganglion cells in the and sometimes takes the form of polysyndac- bowel (Hirschsprung disease) because of al- tyly with a “windswept”” foot deform- tered cell membranes and cell to cell interac- ity.12 13 15 16 27 45 48 63 72 84 Other lower limb abnor- tions. However, abnormal migration or prolif- malities include club foot, varus or valgus foot eration of neural crest derived cells, which deformities, short first toes, and hip contributes substantially to the endocardial dislocations.9122731Occasionally reported skel- cushions, could also explain both the cardiac etal abnormalities include dense base of the defects and abnormal intestinal ganglion cells. skull,31 scoliosis,34 52 69 78 kyphosis,35 71 78 ovoid In addition to structural heart defects, there vertebrae,68 cervical ribs,54 78 82 thin ribs,57 68 85 is a substantially increased frequency of and missing ribs.71 Although epiphyseal stip- pulmonary hypertension in the newborn pling (“chondrodysplasia punctata”) has been period and persistent hypertension postnatally, reported in a few cases,132576886such stippling but limited largely to patients with especially The Smith-Lemli-Opitz syndrome 325

low cholesterol levels, possibly related to the A small number of SLOS patients have had

abnormal steroid metabolism of these the unusual finding of dysplasia or aplasia of J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from patients.69 71 72 93 the gall bladder12 27 or gallstones in infancy or later childhood.12 69 More common, however, is RENAL AND ADRENAL ANOMALIES transient or, more rarely, lethal cholestatic liver About one quarter of patients with biochemi- disease.12 27 67 100 Histologically, iron pigment in cally confirmed SLOS have renal liver cells has been found to be increased,51 67 as anomalies,13 27 94 most commonly renal hypopla- is also seen in peroxisomal disorders. Although sia or aplasia,10 12 13 27 55 94 95 renal cortical lipidosis is not commonly associated with cysts,912274457 hydronephrosis,9102744606772 SLOS, diVuse lipid storage was reported by renal ectopia,1 9–31 44 57 67 72 79 ureteral duplica- Parnes et al,74 and Porter et al101 recently tion,31 60 and persistent fetal lobation.15 27 31 79 A described impaired intracellular trafficking of number of cases with the oligohydramnios LDL in SLOS fibroblasts. sequence caused by bilateral renal aplasia or Pancreatic islet cell hyperplasia has been other renal causes of severely diminished urinary reported frequently in severe SLOS.912173165 output have been described.12 13 55 82 95 The blad- The histology in these cases features nesidio- der and ureters may be hypoplastic, probably blastosis and reduced quantities of somatosta- secondary to renal hypoplasia or aplasia. tin.17 102 Other abdominal malformations less Both adrenal hyperplasia12 17 and adrenal frequently reported include intestinal malrota- hypoplasia13 have been reported in SLOS, but tions,912274584 absence of the diaphragm or the growth and shape of the adrenals appear to diaphragmatic hernia,955 polysplenia and be normal in most.13 29 Histological studies of asplenia,9 anal stenosis or atresia,9135265103and hyperplastic SLOS adrenal glands17 typically Meckel’s diverticulum.27 show deficient cortical lipid, where normally fetal adrenals contain much cholesterol. Post- OTHER ANOMALIES AND CLINICAL PROBLEMS natal studies of adrenal function in children Involuted67 and hypoplastic99 thymus and with SLOS have shown either normal absent parathyroids12 have been found. Among 73 function or, in several biochemically severely the more common dermatological and hair 96 aVected children, decreased steroid synthesis. abnormalities are hypopigmented hair,278mild to extreme skin photosensitivity in more than PULMONARY ANOMALIES half of patients,27 hyperhidrosis of the palms,104 Abnormal pulmonary lobation and pulmonary marked cutis marmorata,99 and eczema.27 After hypoplasia are common in the more severely infancy, many children have mild to marked 9121331515382 aVected cases of SLOS. As ex- acrocyanosis that appears to be autonomic in pected, pulmonary hypoplasia is also common nature and often resolves after cholesterol in SLOS patients with the oligohydramnios therapy. Widely spaced nipples are mentioned 12 13 55 82 sequence secondary to renal aplasia. repeatedly, but measurements are rarely pro- Accessory pulmonary arteries have also been vided. There are two reports of excessive mus- http://jmg.bmj.com/ 45 described. Anomalies of the laryngeal and cle rigidity after halothane anaesthesia, but tracheal cartilages are common even among diagnostically raised creatine kinase levels were patients with mild forms of SLOS and may not found.105 106 cause obstructive sleep apnoea. Serious com- plications have resulted from diYculties with emergent and even elective intubation because Natural history of marked tracheal narrowing and other abnor- The neonatal period and infancy of SLOS malities of the laryngeal and tracheal patients are almost invariably disturbed by on September 30, 2021 by guest. Protected copyright. structures.69 feeding problems, including poor or abnormal suck, swallowing diYculties, vomiting, and lack GASTROINTESTINAL ANOMALIES of interest in feeds. Oral tactile defensiveness Pyloric stenosis is a prominent clinical problem and failure of progression to textured foods in noted in the original description of SLOS and later months is also characteristic of the SLOS in many subsequent case reports.12797Pyloric infant. As a result, more than 50% of patients stenosis in SLOS has the same clinical and require nasogastric tube feedings, often pro- anatomical characteristics as in otherwise nor- gressing to gastrostomy feeding for several mal children, but vomiting and other feeding years. As expected, patients with a cleft palate problems commonly persist after surgical and a small mandible () repair, in part because of apparent intrinsic have the most severe feeding problems. Al- abnormalities of intestinal motility. In the more though gastro-oesophageal reflux is a common severe cases, intestinal aganglionosis occurs, problem in patients with SLOS, such reflux is both short segment and more extensive in- often caused by a failure to recognise that the volvement of the upper and lower intestinal children have congenitally small stomachs and tract.12 87 98 99 Even among SLOS patients lack- intestinal dysmotility. In addition, gastro- ing histological evidence of intestinal aganglio- oesophageal reflux secondary to milk or soy nosis, intestinal dysmotility is common, espe- protein allergy seems to be unusually common cially in the first year. However, whereas pyloric in children with SLOS. Pyloric stenosis is also stenosis historically has been reported in at frequent in SLOS as shown by Lin et al,92 who least 10% of SLOS patients, it is now uncom- found that pyloric stenosis occurred in at least mon in SLOS patients treated with supplemen- 7% of their biochemically confirmed cases and tary cholesterol starting shortly after birth 11% of published cases. With early recognition (Kelley, unpublished data). of these causes of gastro-oesophageal reflux 326 Kelley, Hennekam

and vomiting in SLOS, fundoplications and described as severely mentally retarded (IQ 20

other surgical interventions can be minimised. to 40), such apparently poor development in J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from Severe hypotonia is almost universal in part reflected diYculties in testing. In general, SLOS during infancy. Although the hypotonia SLOS children are very sociable, have much is partly central in origin, congenital muscle better receptive than expressive language, and hypoplasia also contributes to the hypotonia. may be surprisingly mechanically adept for However, during the second year, muscle mass their apparent degree of cognitive impairment. and tone often improve, and muscle strength Because of their poor expressive language and and tone are typically normal in older children hyperactivity, routine developmental testing with SLOS. In later childhood, increased mus- often underestimates their cognitive abilities. cle tone may occur and can lead to joint and Gross motor development is typically more skeletal problems in non-ambulatory children. severely delayed than fine motor development, Such hypertonia appears to be extrapyramidal but most children with classical SLOS learn to rather than spastic. walk between 2 and 4 years. With the availabil- “Failure to thrive” is also almost universally ity of biochemical testing and the more diagnosed in children with SLOS, but the frequent recognition of more mildly aVected diagnosis more often than not is incorrect. SLOS children, the known developmental Infants with SLOS are small for gestational age spectrum of SLOS has widened substantially. and most continue to grow below the 3rd cen- Approximately 10% of children with biochemi- tile despite adequate caloric intake, indicating a cally diagnosed SLOS have development in the fundamental, genetic hypotrophy as the basis mildly retarded range (IQ 50 to 70). A few of the growth retardation. Weight gain can be patients with normal or borderline normal even poorer in the first two years because of development have been described,27 34 and it is feeding and GI motility problems. Although likely that the proportion of recognised patients even well nourished infants will experience a with borderline and normal intelligence will fall oV from their birth centiles over the first increase. year, most children with SLOS have normal In the newborn period, excessive sleeping growth velocities for length and weight in later and poor responsiveness are common. How- years. Similarly, although head circumference ever, hours of shrill screeching or inconsolable is proportionately the smallest measurement at screaming, especially at night and in the early birth and postnatally, head circumference also morning hours, is a major behavioural charac- usually remains proportionate with other teristic of untreated SLOS later in infancy.10 27 measurements. Most measurements for classi- Others appear hypersensitive to all visual and cal SLOS fall between −1 and −5 SD below auditory stimuli and must be kept in quiet, normal, but measurements as low as −8 to −10 dark rooms. Ryan et al27 drew attention to the SD occur in the more severely aVected strikingly diminished amount of sleep in early patients. With a few exceptions at both childhood, which they found in 70% of their

extremes, final adult height and head circum- patients, some of whom slept for only two or http://jmg.bmj.com/ ference are between 2 and 5 SD below three hours at night. Although this abnormal normal.12 13 27 107 In several published series, sleeping pattern may improve with age, adult final height in adults with “type I” SLOS was SLOS patients with similar sleep problems are between 143 and 170 cm.27 48 59 Size at birth known. Many patients, even those with very and growth of the biochemically more severely mild clinical disease, may show self-injurious aVected patients is substantially less. and aggressive behaviour. Most characteristic During both infancy and childhood, children among these behaviours over the age of 3 years with SLOS appear to have an increased are forceful hyperarching (what we have called on September 30, 2021 by guest. Protected copyright. number of infections. Although many of the “opisthokinesis”), with or without head bang- infections are otitis media, skin infections and ing, and arm and hand biting. Marked tactile pneumonias also seem to occur more often. hypersensitivity of the hands and feet is also Despite the frequency of reflux and hypotonia seen in more than 50% of patients. Behavioural in children with SLOS, aspiration pneumonia characteristics of autism, such as hand flap- is surprisingly uncommon, most probably ping, abnormal obsessions, insistence on rou- because of the children’s exaggerated gag tine, and poor visual contact, are common in reflex. Except for a single report of abnormal children with SLOS. Despite these many monocyte oxidative metabolism,108 no specific behavioural problems, most parents describe primary immune disturbances have been de- their children often as loving, aVectionate, and scribed in SLOS. However, death from sudden happy.227 overwhelming infections is not rare in SLOS and suggests a fundamental abnormality in LIFE EXPECTANCY immune defenses or, possibly, adrenal Some investigators have suggested that there is function.96 Apart from the high frequency of an increased rate of spontaneous abortion in milk and soya protein allergy71 and, possibly, an families with SLOS, but Ryan et al,27 in the only increased frequency of reactive airway disease, systematic study of prenatal losses, found 39 other primary immunological diseases do not probands, 51 healthy sibs, 16 spontaneous seem to be common in SLOS. abortions, and seven elective terminations among 43 sibships with 113 known concep- 95 MENTAL DEVELOPMENT AND BEHAVIOUR tions. Furthermore, Kratz and Kelley found With rare exceptions, global psychomotor no increased recognised rate and retardation is characteristic of SLOS. Al- an expected segregation ratio of 25% in a though, historically, most patients have been cohort of prospectively monitored pregnancies. The Smith-Lemli-Opitz syndrome 327

Although these data do not support the (fig 2). The markedly increased levels of 7DHC

hypothesis of an increased miscarriage with almost no 7-dehydrodesmosterol in pa- J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from rate,10 109 110 it may still occur in families with tients with SLOS suggests that the principal more severely aVected children, since severe route of cholesterol biosynthesis in man may be cardiac or renal abnormalities have been the Kandutsch-Russell pathway.21 Never- known to lead to mid-trimester intrauterine theless, the relative abundance of death of SLOS fetuses. As supported by the in neuronal tissues, the testes, and breast data of CunniV et al, the low 17% segregation milk125–127 suggests that desmosterol may be the ratio found in one large series10 could be penultimate sterol in some tissues, or have spe- explained by the inclusion of a proportion of cific functions in the tissue where it is genetically diVerent disorders at a time when abundant. biochemical confirmation of the diagnosis was Before discovery as the marker metabolite of not possible. SLOS, 7DHC was known as a minor constitu- There are no recent figures for life expect- ent of plasma and solid tissues. It was found to ancy in SLOS. Johnson10 found that 27% of be increased up to three-fold in conditions cases in her series died before 2 years of age. associated with increased loss of bile acids, for However, an analysis of the causes of death was example, ileal resection, and, therefore, in- not provided. As ascertainment of SLOS, even creased cholesterol synthesis. 7DHC has spe- in the era of biochemical diagnosis, has been cial physiological importance as the precursor

incomplete, the exact percentage of cases with of vitamin D3 via photic conversion of 7DHC

early lethality remains uncertain. Clearly, how- to pre-vitamin D3 in skin. ever, life expectancy in SLOS is determined Whereas most of the early steps of choles- largely by the severity of the internal malforma- terol biosynthesis are well characterised at the tions and the quality of general supportive care DNA, RNA, and protein levels, the individual and not by an intrinsic degenerative process or enzymes, cofactors, carrier proteins, and intra- biochemical toxicity per se. cellular transport steps involved in the conver- sion of lanosterol to cholesterol are less well DiVerential diagnosis understood, as is the complex intracellular Many papers describe patients with a clinical traYcking of cholesterol. A genetic disruption phenotype resembling SLOS, especially before of one of these transport pathways appears to the era of biochemical diagnosis. In German be the cause of the lipid storage in type C publications the designations “Ullrich- Niemann-Pick disease.128 Understanding intra- Feichtiger syndrome”111 or “Typus Rostockien- cellular traYcking and regulation of cholesterol sis”,112 a multiple congenital anomalies syn- uptake as well as de novo cholesterol synthesis drome with facial anomalies, polydactyly, and will be important to understanding the cellular hypospadias, probably describe severe forms of pathology of SLOS. SLOS.113 Among clinical diagnoses that have In contrast to many other small metabolites,

been proven or suspected to be cases of SLOS very little cholesterol is transported after the http://jmg.bmj.com/ are the acrodysgenital syndrome,31 85 114 first trimester from the mother to the fetus. Gardner-Silengo-Wachtel syndrome (OMIM Instead, most cholesterol must be synthesised 231060),115 116 and holoprosencephaly- by the fetus.129 130 Both in later fetal life and polydactyly (“pseudotrisomy 13”) syndrome after birth, most if not all cholesterol in the (OMIM 264480).117 118 However, the predomi- brain is synthesised locally, not transported nance in the Gardner-Silengo-Wachtel syn- from blood lipoproteins.130–133 However, recent drome of conotruncal malformations and in molecular biological studies have delineated a holoprosencephaly-polydactyly syndrome of go- system for the delivery of maternal LDL to on September 30, 2021 by guest. Protected copyright. nadal dysgenesis, both uncommon abnormali- LDL receptors in the embryonic neuroepithe- ties in SLOS, suggests that some of these lium in the first trimester.134 135 The fundamen- biochemically untested patients do not have tal importance of this system is supported by SLOS. Among diagnoses that have been incor- the discovery135 that transgenic mice lacking rectly assigned to patients with SLOS are Noo- megalin, the LDL receptor in the embryonic nan syndrome (OMIM 163950), Opitz syn- neuroepithelium, develop holoprosencephaly, drome (OMIM 145410, 300000), and which has been linked to abnormal cholesterol (OMIM 214100). Con- metabolism at several levels.58 136 137 Thus, versely, several patients with á thalassaemia- whereas delivery of cholesterol to the fetus after mental retardation syndrome (OMIM 301040) development of the placenta may be minimal, have been given the diagnosis of SLOS. Other critical aspects of embryonic tissue diVerentia- disorders that resemble SLOS, but less so, are tion may be sensitive to maternal blood choles- Meckel syndrome (OMIM 249000), hydrole- terol and LDL levels. thalus syndrome (OMIM 236680), Pallister- Although most cholesterol in tissues serves Hall syndrome (OMIM 146510), orofaciodig- as a major structural lipid of membranes, some ital syndrome type VI (OMIM cholesterol also enters pathways for bile acid 277170),12 13 112 118 119 and a number of individual metabolism and the synthesis of steroid case reports.74 120–124 hormones. Another unexpected and recently discovered role of cholesterol is as a covalently Sterol biosynthesis bound element of active hedgehog proteins, a Cholesterol is a 27-carbon, mono-unsaturated family of embryonic signalling proteins.58 137 138 sterol, synthesised from lanosterol by a series of Already in 1966, Roux and Aubry139 oxidations, reductions, and demethylations, had shown that inhibitors of enzymes of the mostly limited to the endoplasmic reticulum distal cholesterol biosynthetic pathway caused 328 Kelley, Hennekam

holoprosencephaly, microcephaly, pituitary presence of cholesterol cleaves itself into a

agenesis, limb defects, and genital anomalies in non-signalling COOH-terminal half and a J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from the pups of exposed pregnant rats or mice. The mature, cholesterol substituted, N-terminal same group also showed that feeding choles- half, “hh-N”. The processed amino half terol to the pregnant, treated rats substantially appeared to possess all hh signalling activity, limited or blocked the teratogenic eVects of which in vertebrates includes patterning of AY-9944, an inhibitor of DHCR7.140 Before development in the ventral forebrain and limb 1993, holoprosencephaly had been reported in buds. The hh-N fragment appears to have a only one SLOS patient.17 However, using an role in the attachment and localisation of hh-N increased 7DHC level as a diagnostic marker to cell membranes.137 The proteins in verte- for SLOS, holoprosencephaly of variable sever- brates homologous to Shh, Desert hedgehog ity has been found to occur in about 5% of and Indian hedgehog, appear to have similar patients with SLOS, most of whom have also roles in, respectively, genital and skeletal devel- been found to have mutations in DHCR7.23 58 opment, both important aspects of malforma- This association was soon complemented by tion in SLOS.142 143 Another link between these the discovery at about the same time138 that malformations and hedgehog proteins was targeted disruption in mice of Sonic hedgehog made by Roessler et al,136 who found that (Shh) causes not only holoprosencephaly, but heterozygosity for mutations in SHH, the 7q36 also distal limb defects and other skeletal linked gene encoding SHH in humans, causes anomalies. The same group showed that cova- autosomal dominant holoprosencephaly. lent addition of cholesterol to the equivalent This convergence of holoprosencephaly, hedgehog protein in Drosophila, hh, was an SLOS, Shh, and cholesterol metabolism fo- essential part of the “autoprocessing” of cused attention on the possibility that the hh,137 141 wherein precursor Shh protein in the covalent attachment of cholesterol to Shh-N

24 25 9 14 8 3 HO 5 7 6 Lanosterol

4 2 http://jmg.bmj.com/ ?

HO Zymosterol HO Cholest-8(9)-en-3β-ol HO 8-Dehydrocholesterol

1 1 1

4 on September 30, 2021 by guest. Protected copyright.

HO Cholesta-7,24-dien-3β-ol HO Lathosterol

2 2

4

HO 7-Dehydrodesmosterol HO 7-Dehydrocholesterol

AY-9944 3 – 3 BM 15,766

4 –

HO Desmosterol HO Cholesterol Figure 2 Enzymatic steps and major sterol intermediates comprising the pathway of cholesterol from the first sterol, lanosterol. The denoted enzymatic steps are (1) 3beta-hydroxysteroid-delta8,delta7-isomerase (EC 5.3.3.5); (2) 3beta-hydroxysteroid-delta5-desaturase (lathosterol dehydrogenase, EC 1.3.3.2); (3) 3beta-hydroxysteroid-delta7-reductase (DHCR7,EC 1.3.1.21); and (4) 3beta-hydroxysteroid-delta24-reductase (desmosterol reductase). The Smith-Lemli-Opitz syndrome 329

and related hedgehog proteins is the link much lower total sterol levels and higher

between the abnormal cholesterol metabolism 7DHC/sterol ratios than surviving SLOS J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from of SLOS and abnormal morphogenesis in patients.23 152 Although it was suggested153 that SLOS. However, Cooper et al144 showed that essentially all SLOS patients with cholesterol autoprocessing of hedgehog is not impaired levels lower than 10 mg/dl die at birth or in the when cholesterol in the reaction medium is first few months,152 death in SLOS is caused replaced with 7DHC or any of many other largely by specific visceral malformations 27-carbon sterols. In vitro studies with AY- rather than some biochemical consequences of 9944 and other teratogens that cause SLOS- the abnormal sterol levels. Patients with levels like malformations in rats strongly suggested less than 10 mg/dl at diagnosis who are long that the defect in Shh signalling resides in sterol survivors are not rare and are usually dis- mediated changes in the target tissue and not tinguished by a lack of major internal anoma- an abnormality of the Shh-N signal itself.144 lies. The genetic basis of the diVerences in bio- There are other possible levels at which the chemical and clinical severity is supported by sterol defect of SLOS may cause impaired sig- the finding that sibs with SLOS usually have nalling of Shh, such as the sterol sensing similar plasma levels of 7DHC and cholesterol domain in Patched,145 and the lamin B and similar degrees of clinical severity.23 receptor, which has both sterol-14-reductase Indeed, subsequent studies of DHCR7 muta- and a binding site structure similar to that of tions have shown a strong correlation between DHCR7.24 146 Shh is just one of several similarly the predicted enzymatic eVect of a patient’s functioning hedgehog proteins, all of which particular mutations and the clinical may interact with Patched in a sterol sensitive phenotype.163 manner. Thus, it is possible that impaired As expected for an autosomal recessive signalling activities of Desert hedgehog142 and disorder, the parents of SLOS patients have no Indian hedgehog143 also play a role in SLOS in or minimal physiological signs of abnormal the abnormal morphogenesis of tissues that are sterol metabolism. About half of the parents not special targets of Shh signalling. are found to have mildly increased plasma Other less specific disturbances may also 7DHC levels, with a mean level for all parents contribute to the abnormal morphogenesis of that is slightly higher than normal.153 However, SLOS. Cholesterol is severely deficient in all more than 90% of SLOS parents have tissues, and such a severe disturbance in mem- abnormally increased 7DHC to cholesterol brane sterol composition may aVect develop- ratios when their lymphoblasts are cultured in mental processes that involve cell-cell interac- cholesterol depleted medium.33 Although there tions, as suggested by DeHart et al.147 are no obvious adverse clinical consequences of Interestingly, in recent studies with the SLOS mildly abnormal 7DHC metabolism in hetero- mimicking teratogen BM 15 766, Lanoue et zygotes, no systematic survey for associated al148 showed that more severe and more charac- minor anomalies has been performed.

teristic SLOS malformations were produced Apart from occasional case studies reporting http://jmg.bmj.com/ when BM 15 766 was given to mice with a adrenal or gonadal steroid levels,17 72 95 154 there genetic deficiency of LDL receptors, suggest- have been few studies of steroid metabolism in ing that, in embryonic tissue, a deficiency of SLOS. Reports of DHEA levels have shown cholesterol itself does indeed have a contribu- both abnormally low and high levels, and tory role in the abnormal embryologic signal- testosterone levels were found to be either low ling of SLOS. or normal, with normal to mildly increased Despite the apparent negligible transfer of levels of FSH and LH in some. However, the cholesterol from the mother to the human anecdotal and usually uncontrolled nature of on September 30, 2021 by guest. Protected copyright. fetus, the most severely aVected SLOS patients these studies precludes an informed under- have measurable cholesterol levels at birth, standing of sex steroid metabolism in SLOS. typically 5 to 20 mg/dl.23 131 Moreover, initial Moreover, the high frequency of hypogenital- rates of cholesterol synthesis in cultured fibro- ism in SLOS is not suYcient evidence to blasts from patients predicted on the basis of implicate inadequate steroid production in their mutations to have no DHCR7 activity utero, as the persistence of Mullerian remnants may be as high as 50% of all sterols (Kelley, in some SLOS patients15 suggests defective unpublished observations). Thus, there may be genital morphogenesis unrelated to steroid another genetic source of DHCR7 activity or a hormone levels, possibly mediated through pathway of cholesterol synthesis not requiring sterol related dysfunction of Desert hedgehog DHCR7. The most likely source of such activ- in genital tissues.155 ity is the peroxisome, which has been shown to have an essential role in the early steps of chol- Molecular genetics esterol biosynthesis.149–151 The finding of hypocholesterolaemia and Clinical severity correlates best with the level markedly increased levels of 7DHC in patients of cholesterol (inversely) or with 7DHC (or the with SLOS immediately focused attention on sum of all dehydrosterols) expressed as a frac- 7-dehydrocholesterol reductase (DHCR7, EC tion of total sterols.23 The dehydrosterol 1.3.1.21) as a candidate deficient enzyme fraction better expresses the systemic sterol causing the disorder. Additional evidence came abnormality than absolute blood sterol levels, from drugs that inhibit distal steps of choles- which are subject to wide physiological varia- terol biosynthesis produced SLOS-like malfor- tions independent of the rate of sterol synthe- mations in mice and rats, suggesting that sis. SLOS patients with multiple internal the cholesterol lowering action of these drugs anomalies and early death have had, in general, may rather be the principal teratogenic 330 Kelley, Hennekam

factor.147 156–158 Before the renewed interest in IVS8-1G>C mutation constituted 29% of

DHCR7 caused by SLOS, DHCR7 had been DHCR7 mutant alleles, and that several other J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from little studied at a structural or DNA level, and alleles, R404C (11%), V326L (7%), W151X then mostly in lower organisms.159 At the time (8%), and T93M (8%), also occurred at of the discovery of apparent DHCR7 defi- relatively high frequencies.163 Collectively, the ciency in SLOS, the chromosomal location of five most common mutations from this study DHCR7 was unknown, but 7q32.1 was thought and the Dutch study group (Waterham et al, to be the best candidate, because two unrelated unpublished data) accounted for two-thirds of SLOS patients were known to have de novo the mutant alleles (fig 3). Although these high balanced translocations at that position. Fur- frequencies may have evolved because of a spe- ther genetic study of one of the biochemically cial heterozygote advantage aVorded by the confirmed translocation patients160 showed that particular mutations, more likely is a combina- the 7q32.1 breakpoint interrupted not a gene tion of founder eVect and a heterozygote involved in sterol metabolism but the human advantage that encouraged the persistence of metabotropic glutamate receptor 8 (GRM8), these several mutations and of diverse missense which is not known to have a role in cholesterol mutations. As would be predicted in a model of biosynthesis. However, Moebius et al24 shortly heterozygote advantage, such as increased syn- afterwards reported the cloning and mapping thesis of vitamin D by heterozygotes, the most of a human microsomal DHCR7 gene to chro- common mutations cause more severe enzy- mosomal location 11q12-13. Interestingly, matic defects than the less common mutations. DHCR7 was first cloned not as a gene for a There is as yet no correlation between sterol metabolising enzyme, but as a candidate individual mutations and their eVects on the gene for a “sigma type” drug binding protein.161 sterol or vitamin D metabolism in heterozy- Only after complete sequencing was the gene gotes. Such information, however, may provide found to have strong homology with DHCR7 evidence for increased vitamin D levels as a from Arabidopsis thaliana. Within months, possible heterozygote advantage, since Euro- three groups identified disabling mutations in pean palaeolithic studies have indicated that DHCR7 in SLOS patients.25 26 162 rickets was a common paediatric disease. As The 2957 bp cDNA for the human DHCR7 anticipated from biochemical studies, the most gene has an open reading frame of 1425 bp, severely aVected patients are either homozy- which codes for a protein with 475 amino acid gotes or compound heterozygotes for muta- residues and a calculated mass of 54.5 kDa.24 tions shown or predicted to have no or severely The gene, which spans 14 kb of genomic DNA, reduced DHCR7 activity.163 In contrast, most is organised into nine exons and, by hydropathy classical “type I” SLOS patients are compound plot, between six and nine transmembrane heterozygotes for a severe, truncating mutation alpha helices. In addition to having a high and a second missense mutation associated degree of homology with DHCR7 of Arabidop- with residual enzyme activity. Most patients

sis thaliana, the human DHCR7 has substantial with very mild clinical and biochemical pheno- http://jmg.bmj.com/ homology with genes encoding sterol-delta14- types are compound heterozygotes for two reductases across phyla and with the gene unique or uncommon missense mutations.163 encoding the human lamin B receptor, a nuclear inner membrane protein with intrinsic Biochemical diagnosis sterol-delta14-reductase activity but unknown DIAGNOSTIC METHODS nuclear function or physiological Because approximately 10% of patients with significance.24 146 Among the 26 alleles studied SLOS have normal serum cholesterol levels at by Fitzky et al,162 only one example of any age, including at birth, a blood cholesterol on September 30, 2021 by guest. Protected copyright. IVS8-1G>C, a splice site mutation that creates level is not a reliable screening test for SLOS. a 134 bp insertion between exons 8 and 9, was Moreover, because 8DHC and 7DHC react as found. However, study of a cohort of mostly cholesterol in cholesterol oxidase assay meth- North American patients with largely Euro- ods, hospital laboratories may report a normal pean and Hispanic ancestry showed that the “cholesterol” level even when 7DHC and

Exon 3 Exon 4 Exon 5 Exon 6 Exon 7 Exon 8 Exon 9

Missense mutation Splice site mutation Stop mutation Deletion

Figure 3 Distribution of 122 SLOS mutations of 3beta-hydroxysteroid-delta7-reductase (DHCR7) in 61 patients with Smith-Lemli-Opitz syndrome. The Smith-Lemli-Opitz syndrome 331

8DHC constitute more than half of the sterols dotrophin and á-fetoprotein) are also mildly

measured. In a few aVected children, the level depressed in some SLOS pregnancies. A J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from of 7DHC may be normal or equivocally number of aVected fetuses have been identified increased, but the 8DHC level is usually mildly by the discovery of suggestive fetal abnormali- but distinctly abnormal. In rare cases where the ties, such as nuchal oedema, microcephaly, levels of 7DHC and 8DHC are normal or fall cleft palate, polydactyly, cystic kidneys, am- in the heterozygote range, either DHCR7 biguous genitalia, or a 46,XY karyotype in a mutational testing, DHCR7 enzymatic assay, phenotypically female fetus. Determination of or analysis of sterol biosynthesis in cultured the level of 7DHC in the amniotic fluid is pos- cells must be undertaken.153 164 165 Under condi- sible, with typically a more than 500-fold tions of maximal growth stimulation in choles- increase in aVected pregancies.95 In some preg- terol depleted cultured medium, lymphoblasts nancies, the amniotic fluid level of cholesterol from the mild biochemical variants of SLOS is abnormally low, whereas in others the level is still develop markedly increased levels of surprisingly normal. Direct analysis of the 7DHC. sterol composition of chorionic villi at 10 weeks Not uncommonly, there is a need to make a is also a reliable method for diagnosis of SLOS, retrospective biochemical diagnosis of SLOS in although the relative increase in the 7DHC/ a child for whom there may be remaining cholesterol ratio is not as great as it is in amni- stored tissue or laboratory samples. Samples otic fluid.95 The initial experience with more commonly used for retrospective biochemical than 100 pregnancies at risk for SLOS has diagnosis of SLOS include archived plasma yielded no false negatives and no false and serum samples, Guthrie newborn screen- positives.95 Despite the feasibility of prenatal ing cards, frozen or formalin preserved diagnosis by molecular testing of villus tissue or necropsy tissues, and amniotic fluids. Recover- amniocytes, the simplicity and accuracy of bio- ing suYcient sterol for diagnosis from formalin chemical testing obviates the need for molecu- preserved tissue stored more than a few lar analysis except, perhaps, in a rare case with months is not assured, but can still be success- equivocal biochemical results. ful. In a 130 year old case from an Anatomical Museum the lowered cholesterol level could be Management proven.166 Because 7DHC and 8DHC are oxy- Until recently, SLOS could only be treated gen sensitive, diagnostic levels of 7DHC and supportively. However, with the recognition 8DHC may be lost with storage, but diagnoses and treatment of the deficiency of cholesterol using refrigerated Guthrie cards as old as five biosynthesis in SLOS, the care of patients has years have been made. changed substantially. The estimated daily synthetic need for cholesterol during infancy is OTHER CAUSES OF INCREASED LEVELS OF 7DHC 30 to 40 mg/kg/day, which decreases to 7DHC exists in normal human tissues at a approximately 10 mg/kg/day in adults.167 168

relatively constant ratio to cholesterol. Thus, in This daily requirement is usually met by a http://jmg.bmj.com/ common forms of hypercholesterolaemia, such combination of dietary cholesterol and de novo as familial hypercholesterolaemia resulting synthesis, balanced in a highly regulated man- from abnormalities of LDL metabolism, the ner. If basic regulation of cholesterol biosyn- absolute level of 7DHC may be increased, but thesis is normal in SLOS children, children the ratio relative to cholesterol is usually with SLOS may be able to absorb suYcient normal. In some other conditions where there dietary cholesterol to downregulate endo- is marked upregulation of cholesterol biosyn- genous sterol synthesis and therefore limit sig- thesis, such as cerebrotendinous xanthomato- nificantly the de novo production of 7DHC. on September 30, 2021 by guest. Protected copyright. sis, 7DHC may be increased 10 to 20-fold, but For this reason, children with SLOS routinely the presence of similarly increased levels of have been supplemented with dietary choles- other sterol precursors and cholestanol clearly terol with the expectation that blood choles- establish the correct diagnosis. Another cause terol levels will rise to normal at the same time of mildly increased levels of 7DHC is treat- that the abnormal accumulation of 7DHC is ment with haloperidol (Kelley, unpublished eliminated. A complete biochemical “cure” by observations), which has a high aYnity for the such therapy would require that dietary DHCR7 substrate binding site.24 Because cholesterol be distributed to all cellular and tis- DHCR7 is a member of the “sigma” class of sue compartments. This does not seem to drug binding proteins,24 it is likely that other occur, since cholesterol in the central nervous drugs of this class will cause increased levels of system appears to be synthesised locally and 7DHC through inhibition of DHCR7. More cannot be transported across the blood-brain importantly, however, such drugs may have barrier.133 Even if cholesterol could reach brain particularly adverse eVects on patients with cells and myelin, cognitive performance might SLOS, who have already markedly diminished not improve, since the mental retardation of activities of DHCR7. SLOS appears to reflect more the embryologi- cal micrencephaly and other abnormalities of PRENATAL DIAGNOSIS cerebral neuronal development than an exist- Prenatal diagnosis of SLOS has been accom- ing deficit of cholesterol or toxic eVect of plished in many pregnancies. As reported by 7DHC. several authors,13 95 154 one of the early signs of Initial experimental treatment protocols for an SLOS fetus is an abnormally low maternal SLOS provided 50 mg/kg/day cholesterol, and serum level of unconjugated oestriol. Other higher doses up to 300 mg/kg/day, either in elements of the triple screen (chorionic gona- natural form (eggs, cream, liver, meats, and 332 Kelley, Hennekam

meat based formulas) or as purified food grade terol supplements. After infancy, many SLOS

cholesterol, with or without supplementary bile patients will develop behaviours such as J Med Genet: first published as 10.1136/jmg.37.5.321 on 1 May 2000. Downloaded from acids.169–171 For substantially growth retarded rocking, finger flicking, object twirling, gaze children, treatment with cholesterol is some- avoidance, and various obsessions, which meet times followed by striking increases in the rate criteria for the diagnosis of autistic disorder of growth for several months until more appro- and which typically improve or resolve with priate weight and height centiles are reached. cholesterol treatment. Paradoxically, when SLOS children enter a A large proportion of patients may require growth spurt from better nutrition, cholesterol gavage or gastrostomy feeding for many levels typically fall and 7DHC levels rise months or years. Fundoplications are fre- substantially despite their marked clinical quently unsuccessful in treating gastro- improvement. Although the changes in plasma oesophageal reflux. In most instances, the sterol levels early in a treatment programme reflux is caused not by intrinsically abnormal usually do not reflect the obvious clinical ben- gastro-oesophageal function, but by protein efits of cholesterol supplementation, and nor- allergy or by simple overfeeding from ill malisation of the cholesterol level should not be advised attempts to make SLOS children with the ultimate goal in itself, over a period of years primordial dwarfism grow faster. The combi- all but the most severely aVected children usu- nation of gastrointestinal protein allergy, intrin- ally show a substantial increase in the level of sic intestinal dysmotility, and microgastria cholesterol and a corresponding fall in the lev- often make feeding management extremely els of 8DHC and 7DHC. More striking and diYcult; feeding with elemental formulas may rapid changes in blood sterol levels were be required for many months. reported in a rat model of SLOS created by An uncommon but sometimes severe nutri- treating rats with BM 15 766, a relatively tional problem in SLOS is idiopathic hyperme- specific inhibitor of DHCR7.172 The study tabolism, which most often occurs between the examined the eVects of cholic acid and lovasta- ages of 1 and 3 years. SLOS children who tin, an HMG-CoA reductase inhibitor, on the develop this unexplained hypermetabolism blood sterol levels and found that cholic acid may require up to 200 kcal/kg/day to maintain had little eVect. However, when the rats were body weight. Because of the complicating given lovastatin plus cholesterol, the antici- factors of microgastria and intestinal dysmotil- pated fall in 7DHC levels was prevented rather ity, such children often do not gain weight for than enhanced. Thus, because of the lack of many months, but can otherwise remain definitive evidence that 7DHC is “toxic”, and healthy. Tests for endocrine abnormalities or because of the concern that HMG-CoA intestinal malabsorption have been normal in reductase inhibitors might limit the synthesis of such cases, but signs of hypermetabolism such other critical isoprenoid compounds, HMG- as warm skin and tachycardia are often noted. CoA reductase inhibitors are not currently Although many SLOS patients have a severe

used for routine treatment of SLOS. Bile acid deficiency of normal bile acids, fat malabsorp- http://jmg.bmj.com/ supplements have not been included in more tion and deficiencies of fat soluble vitamins are recent SLOS treatment protocols. Indeed, not common in SLOS, and also clinical because certain bile acids may downregulate evidence of steroid hormone deficiency is sur- tissue levels of LDL receptors, the more rapid prisingly uncommon.96 Among these children, rise in plasma cholesterol levels and the the most common finding is a mild to moderate persistently high levels of 7DHC in bile acid deficiency of synthesis, usually supplemented patients may reflect impaired manifest as hyponatraemia and hyperkalaemia tissue uptake of sterols rather than enhanced during the added hormonal stress of an on September 30, 2021 by guest. Protected copyright. intestinal absorption of cholesterol.172 When infection. Glucocorticoid response to infection SLOS children are hospitalised for surgery or or to administered ACTH can also be acute medical problems and cholesterol cannot subnormal.96 However, for most SLOS chil- be given enterally, cholesterol in the form of dren glucocorticoid production appears to be LDL containing fresh frozen plasma can have normal and supplements during infections or striking beneficial eVects, especially for treat- perioperatively are probably not needed. ment of acute infections and poor wound heal- Nevertheless, the potential for developing min- ing. eralocorticoid and glucocorticoid deficiency In addition to the biochemical and physical postoperatively or during an acute illness must changes that follow the initiation of cholesterol be remembered. replacement therapy, there have also been a In conclusion, SLOS may now be considered number of striking behavioural changes. Upon the example par excellence of metabolic treatment, irritability and sleep disorders may dysmorphology. It is well defined biochemi- improve in young children with SLOS within cally, enzymatically, and molecularly. However, days or weeks, whereas tactile hypersensitivi- many parts of the pathogenesis, that is, the ties, especially of the hands and feet, take developmental pathways involved, still remain longer to ameliorate. Typically, all of these to be elucidated, and much work has to be behaviours return when cholesterol supple- done to reach optimal therapeutic regimens. mentation is stopped. The behavioural im- SLOS was the first malformation syndrome in provement does not appear to correlate with which a cholesterol metabolism disturbance any specific change in the plasma sterol profile, was found. There are now a number of and even children with normal or near normal other entities with such a defect, including plasma cholesterol levels may show substantial Conradi-Hunermann syndrome, CHILD syn- behavioural improvement when given choles- drome, desmosterolosis, Greenberg dysplasia, The Smith-Lemli-Opitz syndrome 333

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