A new juvenile hexosaminidase deficiency disease presenting as cerebellar ataxia: Clinical and biochemical studies

Article abstract

A boy with mild hand tremor since age 2M was found at 4 to have cherry-red spots and mild truncal ataxia without seizures or dementia. Biochemically, he had striking hexosaminidase deficiency (serum: 4.6 percent of normal, 88.9 percent heat-labile; leukocyte: 2.2 percent of normal, 84.6 percent heat-labile; fibroblast 12.8 percent of normal, 93.1 percent heat-labile). The residual hexosaminidase activity migrated electrophoretically in two bands. The major band comigrated with hexosaminidase A, the minor with hexosaminidaseS. Hexosaminidase B was totally absent. The parents had partially reduced hexosaminidase with a decreased heat-stabile fraction. This disorder may result from a new mutation closely related to that causing Sandhoff-Jatzkewitzdisease. NEUROLOGY 27: 1012-1018, November 1977

A new juvenile hexosaminidase deficiency disease presenting as cerebellar ataxia Clinical and biochemical studies WILLIAM G. JOHNSON, M.D., ABE CHUTORIAN, M.D., and ARMAND MIRANDA, Ph.D.

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andhoff-Jatzkewitz d i s e a s e was described in 196S1in a patient with the onset at 5 months of a neurologic disease characterized by progressive mental, motor, and visual deterioration, tnyoclonic seizures, macular cherry red spots, and death at age 2Y2. Biochemically, there was nearly total deficiency of 8-D-N-acetylhexosaminidase (hexosaminidase). 2- At least 34 similar cases have been described subsequently. .5-36 The small amount of residual hexosaminidase activity differed from the two major normal hexosaminidase components, hexosaminidases A and B, by a lower isoelectric pointt3 and has been designated hexosaminidase S . 6 ~ 3 Sandhoff-Jatzkewitz disease represents, then, a combined deficiency of

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From the Departments of Neurology and Pathology, Columbia University College of Physicians and Surgeons, and Neurological Institute of New York This investigationwas supported by Clinical Center grantsfrom the National Instituteof Neurologicaland Communicative Disorders and Stroke (1 1766) and the Muscular Dystrophy Association, Inc, and by grant No 1-R01-CA-18172, awarded by the National Cancer Institute, DHEW Accepted for publication March 1977 Dr Johnson s address is Department of Neurology, College of Physicians and Surgeons, Columbia University, 710 West 168th Street, New York, NY 10032

1012 NEUROLOGY November 1977

hexosaminidases A and B with residual hexosaminidase S in an infant with a disorder that is clinically indistinguishable from classical Tay-Sachs disease. We now describe a patient with a unique and distinctive clinical syndrome, different from that of Sandhoff-Jatzkewitz disease, in whom combined deficiency of hexnsaminidase A and B was found in serum. In leukocytes and cultured fibroblasts, however, small amounts of hexosaminidase S-like and greater amounts of hexosaminidase A-like activity were found. Case report. This boy was the only child of healthy, unrelated parents. The family history was remarkable only for the presence of rnyasthenia gravis in a maternal cousin and Parkinson disease in a maternal uncle. The father was 27 years old and of Irish descent. The mother was 28 years old and of lrish and Italian descent. The patient’s perinatal history was unremarkable. His early growth and development were norrnal or even precocious. He sat at 5 months, walked at I0 months. and began to talk at I year ofage. He was toilet-trained before 2 years. However, he never learned to run and never could pedal a tricycle, which he pushed instead. At age 2’/2 he began to develop a slight intention tremor ofthe arms. This was most notable when he used the hands, as in holding a cup. Some days, the tremor was scarcely noticeable: other days, it was quite severe and his cup would drop out of his

hand. At about the same time he lost control of urinary function. The tremor slowly worsened. Both arms were involved. At times the entire trunk seemed to shake when he lifted an object with both hands or was in an unsteady posture. The tremor was worse during an infectious illness or fever. There was no change or regression in mental functions, which continued to develop, no behavioral change, no obvious visual or ocular difficulty, no hearing loss, and no abnormality of speech or articulation. There was no resting tremor, weakness, wasting, twitching, sensory complaint, or seizure. At age 4 years, 4 months he was a cheerful, bright, cooperative child with normal facies and stature. Speech was fluent and articulation was normal. Head circumference was 51.5 c m . There was no skin lesion, cardiopulmonary abnormality, or other abnormality on general physical examination. Visual acuity was 20/100 or better bilaterally. The disks were pale and bilateral cherry red spots were present. Ocular motions were full, but both eyes showed slowing of voluntary and optokinetic saccadic movements in abduction. No nystagmus or dysarthria was seen. Other cranial nerve functions were normal. There was a mild truncal ataxia, and slight clumsiness of gait was evident. He had a mild intention tremor of the arms withclumsiness of voluntary movement, more marked on the left than on the right. There was no resting or sustention tremor, no lower extremity tremor, no weakness, wasting, or fasciculation. Sensation appeared intact. Tendon reflexes were present, normally active, and symmetrical. Abdominal reflexes were symmetrical. Plantar reflexes were flexor. Since that time the gait ataxia and intention tremor have become slightly worse. Laborarory csaminarions. Normal studies included hemogram, urinalysis, bone marrow, serum electrolytes, uric acid, and glutamic-oxaloacetic transaminase. Urine ferric chloride test, DNPH test, and nitroprusside test were normal. Cerebrospinal fluid was clear and colorless with no cells; glucose was 68 mg per deciliter, protein was 32 mg per deciliter with 2 . 5 percent gamma globulin. Roentgenograms of the skull and l o n g bones w e r e n o r m a l . E l e c t r o c a r d i o g r a m , electromyography , and nerve conduction velocities including distal sensory latencies were all normal. Electroencephalogram, awake and asleep, was normal for his age. Photic stimulation produced no significant driving. Electroretinogram was normal. Visual evoked responses were very monoform and extremely

high voltage in response, although the latencies were generally within usual limits. Computerized axial tomography was performed with and without contrast. A small focus of apparent enhancement was seen in the right temporal region in an otherwise normal study.

Materials and methods. Clotted blood and heparinized blood were collected by venipuncture. Leukocyte pellets were prepared as previously d e s ~ r i b e d . ) ~ Hexosaminid ase was deter mined as previously described39 using the artificial fluorogenic substrate 4 methylumbelliferyl - 2 - deoxy - 2 - acetamidog - D -glucopyranoside (4-MU-J-GlcNAc). which was obtained from Koch-Light, Ltd. Hexosaminidase A and hexosaminidase B were determined in serum4* and leukocytes and fibroblasts41 by the heat denaturation method at 49” C. Hexosaminidase isozymes were separated by horizontal starch gel e l e c t r o p h ~ r e s i sat~pH ~ 7.1 using a Tris-citrate-borate-LiOHor Tris-citrate buffer s y ~ t e m . ~ Electrophoresis ~*~’ was performed at I volt per centimeter at room temperature for 17 hours. The sliced gels were stained by overlaying a filter paper strip saturated with a solution of 1 mM 4-MU$’-GlcNAc in 0.2 M sodium citrate buffer pH 4.85. The staining was carried out for 24 hours at room temperature in a closed humid box. Spots were visualized for photography with a long ultraviolet light (Applied Science) and photographed in an otherwise dark room using a Kodak Wratten No. 3 filter, Polaroid 105 film, and exposures of up to 2.5 minutes. The gels were scanned using a Zeiss system with Xenon lamp, incident wavelength of 344 nm and 430 nm single-barrier filter for the emitted light. Protein was determined by the method of L0w1-y.~~ Cultured skin fibroblasts from a patient with Sandhoff-Jatzkewitz disease (GM 294) were obtained from the Institute for Medical Research in Camden, New Jersey. Skin biopsies were dbtained by the punch method. 1 to I ?hmm explants were grown in Eagle’s minimal essential medium and 15 percent fetal calf serum, supplemented with nonessential

Table 1. Hexosaminidase in serum, leukocytes, and fibroblasts

NEUROLOGY November 1977 1013

New juvenile hexosaminidase deficiency disease

amino acids and vitamins, penicillin, 60 units per milliliter, streptomycin, 60 units per milliliter and amphotericin B, 0.22 p g per milliliter (Grand Island Biological Company, Grand Island, NY) in cluster dishes (Costar, Cambridge, Massachusetts). After sufficient growth of fibroblasts (2 to 3 weeks) the explants were removed and the outgrowth trypsinized and transferred to 75 cm2 plastic culture flasks (Lux Scientific Company, Newbury, California). All cultures were used at early passage (six or less). They were fed every third day (last feeding 24 hours before harvesting). When the monolayer

Figure 1. Starch gel electrophoresis o f serum hexosaminidase(anode at the top, cathode at the bottom). 30 pl serum was applied to each lane. Lane 1: normal control; lane 2: patient; lane 3: father of patient; lane 4: Tay-Sachs disease; lane 5 : mother of patient. Fine arrows: origin; short thick arrows: hexosaminidase B; long thick arrows: hexosaminidase A.

Figure 2. Starch gel electrophoresis of leukocyte hexosaminidase(anode at the top, cathode at the bottom). Leukocyte pellets from 10 ml blood were sonicated (15 sec x2) in 2 ml .05 M sodium citrate buffer pH 5.0 and the suspension centrifuged at 10,000 X g for 30 minutes. 30 pl of the supernate was applied to each lane. Lane 1: mother of patient (35.4 p g protein); lane 2: patient (68.4 p g protein); lane 3: father of patient (42.0 p g protein); lane 4: Tay-Sachs heterozygote (6.0 p g protein); lane 5 : normal control (10.8 p g protein). Fine arrows: origin, hexosaminidase B; thick arrows: hexosaminidase A.

1014 NEUROLOGY November 1977

approached confluency , cells were harvested by trypsinization. Trypsin activity was abolished by brief addition of 5 ml complete medium at 4” C . Cells were pelleted by centrifugation, resuspended, and washed three times with phosphate-buffered saline (pH 7.2).

Results. Serum hexosaminidase was strikingly decreased in the patient and only slightly decreased in both parents (table 1). The patient’s value was far lower (4.6 percent of mean control hexosaminidase) than any control value but the parents’ values were in the lower control range. In leukocytes, the patient’s total hexosaminidase activity was strikingly decreased (2.2 percent of mean control hexosaminidase) and the parent’s levels were partially decreased. The patient’s value and the parents’ values were lower than any control. In fibroblasts, the patient’s hexosaminidase level was strikingly decreased, (12.8 percent of mean control hexosaminidase) and the parent’s values were partially decreased. All three were lower than any control. About 90 percent of the p a tie n t’ s residual hexosaminidase in serum, leukocytes, and fibroblasts (table 1) was heat-labile, a far higher fraction than in controls. One parent had a higher fraction of heat-labile hexosaminidase than the average for controls; the other was similar to controls in serum but had a higher fraction in leukocytes and fibroblasts. Starch gel electrophoresis failed to show any residual hexosaminidase in the patient’s serum (figure 1). However, in leukocyte and fibroblast preparations from the patient, significant residual hexosaminidase was seen (figures 2, 3, ahd 4). All of the residual enzyme migrated

Figure 3. Starch gel electrophoresis of fibroblast hexosaminidase(anode at the top, cathode at the bottom). Fibroblast pellet, twice washed with normal saline, was sonicated (15 sec x2) in 1 ml .05 M sodium citrate buffer pH 5.0 and the suspension centrifuged at 10,000 x g for 30 minutes. 30 pi of the supernate was applied to each lane. Lane 1: control #1 (16.2 p g protein); lane 2: patient (12.0 p g protein); lane 3: father of patient (8.4 p g protein): lane 4: control #2 (11.4 p g protein): lane 5: mother of patient (18.3 p g protein). Gels were sliced at 90 degrees to the direction of electrophoresis; a slicing artifact is seen in lane 4. Fine arrows: origin, hexosaminidase B; long thick arrows: hexosaminidase A; short arrow: hexosaminidase S.

Figure 4. Starch gel electrophoresis of fibroblast hexosaminidase(anode at the top, cathode at the bottom). Fibroblast pellet, twice washed with normal saline, was sonicated (15 sec x 2) in 0.2 to 1.O ml.05 M sodium citrate buffer pH 5.0 and the suspensioncentrifuged at 10,000 x g for 30 minutes. 30 pI of the supernate was applied to each lane. Lane 1 : control (16.2 p g protein); lane 2: patient (12.0 pg protein); lane 3: Sandhoff-Jatzkewitzdisease, GM 294 (18.9 p g protein); lane 4: mother of patient (18.3 pg protein); lane 5: father of patient (8.4 p g protein). Fine arrows: origin, hexosaminidase B; long thick arrows: hexosaminidase A; short arrow: hexosaminidase S. The photograph has been overexposed to bring out the faint spot in lane 3, thus blurring the double spot in lane 2.

in the region of hexosaminidase A or S. No residual hexosaminidase B was detected by electrophoresis in the patient’s serum, leukocytes, or fibroblasts. These findings were confirmed by gel scanning.

Discussion. This patient presents a unique clinical syndrome. He had neurologic symptoms for 1% years, but no dementia and no seizures and the EEG was normal. He had no evidence of any hemispheric disorder except perhaps the abnormality of visual evoked response. Despite the bilateral cherry red spots, there was no noticeable visual defect. The only symptoms were limb ataxia (worse in the arms) and mild truncal ataxia, which progressed only slightly over 2 years. The enzymatic picture also was unique. The heat denaturation assay showed an enzyme deficiency pattern r e s e m b l i n g that of S a n d h o f f d i s e a s e : T o t a l hexosaminidase depressed to less than 10 percent of control values, with most of the residual fraction being heat-labile. However, starch gel electrophoresis at pH 7.1 showed residual activity in two bands, the major band m i g r a t i n g w i t h t h e m o r e a n o d a l p o r t i o n of hexosaminidase A and the minor band migrating with hexosaminidase S. Hexosaminidase B was totally absent. At least 29 cases of combined hexosaminidase A and B deficiency have been reported in sufficient detail for comparison with this case (table 2 ) . The clinical picture has been uniform, with onset of symptoms around 6 months of age, survival to the second or third year of life, and neurologic deterioration characterized by seizures,

progressive loss of mental and motor function, progressive loss of vision, and cherry red spot in the fundus. The cases of Van Hoof, Evrard, and Hersz0 and Spence and associates33were atypical clinically because of onset in the second year of life and somewhat slower progression. The case of Van Hoopo differs from the present case, however, because of more rapid progression, seizures, intellectual deterioration, severe ataxia, spastic quadriparesis, decreased nerve conduction velocities, and lack of ocular cherry red spot. The case of S p e n ~ ediffers ~ ~ from the present one because of more rapid progression, seizures, intellectual deterioration, and hypotonia. Similarly affected cousins of that patient died between ages 3 and 4 years. Two other cases of combined hexosaminidase deficiency had unusual clinical findings. One of these45 began at age 4 and was very slowly progressive but differs from the present patient because of prominent pyramidal and anterior horn cell disease, dystonia, and lack of cherry red spot. The other46was an asymptomatic adult with combined hexosaminidase deficiency. The present case appears to be clinically unique. The cases of Clausen, Melchior, and Paerregaardls and of Spence et a133 were biochemically atypical for Sandhoff disease and unlike the present case. The case of Clausen et a l l s had large amounts of residual hexosaminidase but only 20 percent of the residual activity was heat-labile. Other methods of fractionating hexosaminidase were not employed. The case of Spence et a133had a large amount of residual hexosaminidase in serum, less in leukocytes and fibroblasts, and very little in liver. The residual enzyme was largely heat-labile, but some hexosaminidase B was present on electrophoresis, unlike Sandhoffs disease or the present case. The biochemical findings in the present case of combined hexosaminidase deficiency appear unique. Sandhoff disease is sometimes called “total hexosaminidase deficiency’ ’ but is defined biochemically by total hexosaminidase activity less than 10 percent of normal. Obviously, these two statements are inconsistent: 90 percent deficiency is not “total deficiency.” Hexosaminidase A and B are totally deficient in typical Sandhoff disease when isoelectric f o c u ~ i n g , ~ ion J~ exchange c h r o m a t ~ g r a p h y , ~or, ~electroph~resis’~ ~,~~ is used to fractionate the hexosaminidase. The residual hexosaminidase in Sandhoff disease appears to be a more acidic protein, different from either hexosaminidase A or For this reason, we prefer the hexosaminidase B. term “combined hexosaminidase deficiency.” In most cases, however, the residual hexosaminidase has been fractionated by heat denaturation. The heat-labile fraction is called “hexosaminidase A”; the heat-stabile fraction, “hexosaminidase B.” Such statements are misleading. The residual heat-labile hexosaminidase activity in Sandhoff disease does not correspond to hexosaminidase A, nor does the heat-stabile residual activity correspond to hexosaminidase B. Whenever electrophoresis or isoelectric focusing was used, typical Sandhoff disease was characterized by absence of both hexosaminidases A 13736737

NEUROLOGY November 1977 1015

New juvenile hexosaminidase deficiency disease Table 2. Clinical and biochemical data-Cases of Sandhoff-Jatzkewitz disease

and B, and the residual hexosaminidase activity was distinct from either. This residual activity migrates more rapidly toward the anode than hexosaminidase A in conventional starch gel electrophoresis systems (figure 4), and has a lower isoelectric point than hexosaminidase A by isoelectric focusing.13 The present case is biochemically unique because the 1016 NEUROLOGY November 1977

electrophoretic pattern of residual hexosaminidase consisted of two bands. The major band migrated with hexosaminidase A and a minor band migrated more anodally. This was most clearly demonstrated in the fibroblast extracts, where a distinct two-banded pattern was seen (figure 3). Residual activity in the patient’s leukocytes migrated with hexosaminidase A and extended

more anodally, but no two-banded pattern was seen (figure 2 ) . No residual activity was seen on electrophoresis of the patient’s serum (figure 1). Like Sandhoff disease, however, the present case showed a total deficiency of hexosaminidase B in serum, leukocytes, and fibroblasts (figure 1-4). The identity of this patient’s residual hexosaminidase was not clear. The major band on electrophoresis resembled hexosaminidase A in mobility and the minor band, hexosaminidase S (figures 3 and 4). The residual hexosaminidases were heat-labile (table 1 ) like hexosaminidase A and hexosaminidase S.36 The mild clinical course, quite unlike that associated with Sandhoff disease, suggests that the residual hexosaminidases may function to some extent in vivo on natural substrates. If the fast-moving hexosaminidase S-like band in fact represents the same hexosaminidase S found in classical Sandhoff disease, then the mild clinical course in this patient might be related to the additional presence of the major, hexosaminidase A-like band. This major residual band may be quite unrelated to hexosaminidase A; it may represent a small amount of normal or mutationally altered hexosaminidase A, or it may represent a normal or mutationally altered subcomponent of hexosaminidase A. The latter possibility is consistent with the observation of Bach and Suzuki4’ that an acidic subcomponent of hexosaminidase A appears to be responsible for G M ~ cleavage. In addition, this major residual band may be related to hexosaminidase S’ found in trace amounts in Sandhoff-Jatzkewitz disease fibroblast^^^ as a minor component of the residual hexosaminidase. Further studies are in progress to clarify the nature of these residual he xosaminidases. The clinically normal parents of the patients both showed partial hexosaminidase deficiency in leukocytes and fibroblasts (table 1). This is best explained as representing the carrier state of an autosomal recessive disorder. The parents’ hexosaminidase levels were intermediate between the patient and controls. The parents were not identical, however. This may represent variability between individuals carrying the same mutation. Alternatively, the parents may carry different, perhaps allelic, mutations, and the patient could be a genetic compound. Family studies are in progress to answer this question.

Acknowledgments We wish to thank Andrew Hiatt, Julie Ruben, and Richard Goldenberg for skilled technical assistance. We wish to thank Dr. Lewis P. Rowland and Dr. Virginia Tennyson for helpful comments on the manuscript.

REFERENCES

1. Pilz H, Muller D, Sandhoff K, et al: Tay-Sachssche Krankheit mit Hexosaminidase-Defekt. Deutsch Med Wochenschr 93:1833-1839, 1968 2. Sandhoff K, Andreae U, Jatzkewitz H: Deficient hexosaminidase activity in an exceptional case of Tay-Sachs disease with additional storage of kidney globoside in visceral organs. Life Sci 7:283-288, 1968

3. Sandhoff K, Andreae U, Jatzkewitz H: Deficient hexosaminidaseactivity in an exceptional case of Tay-Sachs disease with additional storage of kidney globoside in visceral organs. Pathol Eur 3:278-285, 1968 4. Sandhoff K, Jatzkewitz H, Peters G: Die infantile amaurotische ldiotie und verwandte Formen als Gangliosid-Speicherkrankheiten. Naturwissenschaften 56:356-362. 1969 5. Sandhoff K: Variation of Jl-N-acetylhexosaminidase pattern in Tay-Sachs disease. FEBS Lett 4:351-354, 1969 6. OBrien JS: Five gangliosidoses. Lancet 2:805, 1969 7. Young EP, Ellis RB, Lake BD, et al: Tay-Sachs disease and related disorders: Fractionation of brain N-acetyl$?-hexosaminidase on DEAE-cellulose. FEES Lett 9:l-4, 1970 8. Fontaine G, Maillard E, Farriaux J-P, et al: Un cas de gangliosidose d GMZ.Presse Med 78:603, 1970 9. Rdsibois A, Tondeur M: Ultrastructure of Sandhoffs disease. Vle Congres International de Neuropathologie, Paris, Masson et Cie, 1970, pp 1047-1048 10. Strecker G, Montreuil J: Description dune oligosaccharidosurie accompagnant une gangliosidose GMZ a deficit total en N-acetyl-hexosaminidases.Clin Chim Acta 33:395-401, 1971 11. Fontaine G, Rdsibois A, Tondeur M, et al: Gangliosidosis with total hexosaminidase deficiency: Clinical, biochemical, and ultrastructural studies and comparisonwith conventional cases of Tay-Sachs disease. Acta Neuropathol (Berl) 23:118-132, 1973 12. Suzuki Y, Jacob JC, Suzuki K, et al: GMz-gangliosidosis with total hexosaminidase deficiency. Neurology (Minneap) 2313-32W,et al:Enzyme alterations and ipid storage in three variants of Tay-Sachs disease. J Neurochem 18:2469-2489, 1971 14. Harzer K, Sandhoff K, Schagern einer Variante der Tay-Sachsschen Erkrankung (Variante 0). Klin Wochenschr 493189-1191, 1971 15.Krivit W, Desnick RJ, Lee J, et al: Generalized accumulation of neutral glycosphingolipidswith GMZganglioside accumulation in the brain. Am J Med 52:763-769. 1972 6. Snyder PD, Krivit W, Sweeley CC: Generalized accumulationof neutral glycosphingolipids with GM2 ganglioside accumulation in the brain. J Lipid Res 13328-136, 1972 7.Okada S, McCrea M, O’Brien JS: Sandhoff’s disease (GM2 gangliosidosis type 2): Clinical, chemical and enzyme studies in five patients. Pediatr Res 6:606-615, 1972 8. Clausen J, Melchior JC, Paerregaard P: Enzymic differentiation between different types of Tay-Sachs disease of similar clinical appearance. Eur Neurol 756-64, 1972 9. Kolodny EH: Sandhoffs disease: Studies on the enzyme defect in homozygotes and detection of heterozygotes. In Volk BW, Aronson SM (Editors): Sphingolipids, Sphingolipidoses and Allied Diseases. New York, Plenum Press, 1972, pp 321-341 20. Van Hoof F, Evrard P, Hers HG: An unusual case of Gun-gangliosidosis with deficiency of hexosaminidase A and B. /bid,’9 pp 343-350 21. Bain AD, Tateson R, Anderson JM, et al: Sandhoff’s disease (GM2 gangliosidosis, type 2) in a Scottish family. J Ment Defic Res 16:119-127, 1972 22. Garner A: Ocular pathology of GM2gangliosidosis-type 2 (Sandhoff’s disease). Br J Ophthalmol 57514-520. 1973 23. Soeda S, Suzuki H, Tanimura R, et al: A case of Tay-Sachs disease (Sandhoff type) with total hexosaminidase deficiency. J Tokyo Women’s Med Coll 42:1030-1035, 1972 24. Sandhoff K, Harzer K: Tay-Sachs disease (variant 0).In Hers HG, Van Hoof F (Editors): Lysosomes and Storage Diseases. New York, Academic Press, 1973, pp 345-356 25. Dolman CL, Chang E, Duke RJ: Pathologic findings in Sandhoff disease. Arch Pathol 96:272-275, 1973 26. Vidailhet M, Neimann N, Grignon G, et al: Maladie de Sandhoff (gangliosidose a GMZ,de type 2). Arch Franc Pdd 30:45-60, 1973 27. Juif JG, Luckel JC, Nussbaum JL, Stoebner R, Kapps R: La gangliosidose GM2 avec deficit complet en@-N-acetyl-hexosaminidase ou maladie de Sandhoff. Arch Fr Pediatr 3029-43,1973 28. Desnick RJ, Krivit W, Sharp HL: In utero diagnosis of Sandhoffs disease. Biochem Biophys Res Commun 51:20-26. 1973 29. Elieden LC, Desnick RJ, Carter JB, et al: Cardiac involvement in Sandhoffs disease. Am J Cardiol 34:83-88, 1974 30. Desnick RJ, Snyder PD, Desnick SJ, et al: Sandhoff‘s disease: Ultrastructural and biochemical studies. /bid,1s pp 351-371 31. Tremblay M, Szots F: GMz Type 2-gangliosidosis (Sandhoff’s disease). Ocular and pathological manifestations. Can J Ophthalmol9:338-346, 1974 32. Melancon SB, Potier M, Geoffroy G, et al: Maladie de Sandhoff: Etude clinique et gdnetique chez une enfant canadienne-francaise. Union Med Can 103:121&1222, 1974 33. Spence MW, Ripley BA, Embil JA, et al: A new variant of Sandhoffs disease. Pediatr Res 8:628-637, 1974 34. Tomasi LG, Fukushima DK, Kolodny EH: Steroid hexosaminidase activity in Tay-Sachs and Sandhoff-Jatzkewitz diseases. Neurology

NEUROLOGY November 1977 1017

New juvenile hexosaminidase deficiency disease (Minneap) 24:1158-1165, 1974 35. Harzer K, Stengel-RutkowskiS, Gley E-0, etal: Pranatale Diagnoseder GMn-gangliosidoseTyp 2. Dtsch Med Wochenschr 100:106-106,1975 36. Beutler E, Kuhl W, Comings D: Hexosaminidaseisozyme in type0 GM2 gangliosidosis (Sandhoff-Jatzkewitz disease). Am J Hum Genet 27:628-638, 1975 37. lkonne JU, Rattaui MC. Desnick RJ: Characterization of Hex S, the major residual$-hexosaminidaseactivity in type 0 GM2 gangliosidosis (Sandhoff-Jatzkewitz disease). Am J Hum Genet 27:639-650, 1975 38. Brady RO, Johnson WG, Uhlendorf BW: Identification of heterozygous carriers of lipid storage diseases. Am J Med 51:423-431, 1971 39. Johnson WG, Mook G, Brady RO:@-HexosaminidaseA from human placenta. Methods Enzyrnol 28:857-861, 1972 40. OBrien JS, Okada S, Chen A, et al: Tay-Sachs disease: Detection of heterozygotes and homozygotes by serum hexosaminidase assay. N Engl J Med 283:15-20, 1970 41. Padeh B, Navon R: Diagnosis of Tay-Sachs disease by

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hexosaminidase activity in leukocytes and amniotic fluid cells. Isr J Med Sci 7:259-263. ~ . . ,1971 . 42. Swallow DM, Stokes DC, Corney G, et al: Differences between the N-acetyl-hexosaminidase isozymes in serum and tissues. Ann Hum Genet 37267-302, 1974 43. Hopkinson DA, Mestriner MA, Cortner J, et al: Esterase D, a new human polymorphism. Ann Hum Genet 37:119-137, 1973 44. Lowry OH, Rosebrough NJ, Farr AL, et al: Protein measurement with the Fohn phenol reagent. J Biol Chem 193265-275, 1951 45. Goldie W, Holtzman D, Suzuki K: Chronic total hexosaminidase deficiency-a new disease. Child Neurology Society, 5th National Meeting, Monterey, California, Oct 28-30, 1976, Abstract No. 16 46. DreyfusJ-C, Poenaru L, Svennerholm L:Absenceof hexosaminidaseA and B in a normal adult. N Engl J Med 29261-63, 1975 47. B a c h G, S u z u k i K : H e t e r o g e n e i t y of h u m a n h e p a t i c N - ace t y I-J- D - h exo Sam in id ase A activity toward natural glycosphingolipid substrates. J Biol Chem 250:1328-1332, 1975