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Among Ashkenazi Jewish individuals with mucolipidosis IV (ML IV), two mutations in the ML IV gene, IVS3-1A --G and delEX1-EX7, account for more than 95% of disease alleles. The reported method of genotyping for the delEX1-EX7 mutation involves a cumbersome multistep procedure. In the present study, a new simplified one-step procedure is described that detects this mutation in both patients and carriers. An improved procedure is also described for detection of the IVS3-1A --G mutation. Using these improved procedures, we have characterized the ML IV mutant alleles in 27 patients and 95 of their relatives from 22 families, and in 123 unrelated and unaffected Ashkenazi Jewish controls. Of the 27 ML IV patients, 16 patients (59.3%) were found to be homozygous for the IVS3-1A --G mutation and 1 patient (3.7%) homozygous for the delEX1-EX7 mutation. Additionally, 9 patients (33.3%) were compound heterozygotes for IVS3-1A --G/delEX1-EX7. Among the 123 Ashkenazi Jewish controls, two individuals were identified as heteroallelic with one IVS3-1A --G mutation (carrier frequency: approximately 1 in 61); none showed the delEX1-EX7 mutation. The modifications described here provide a more facile means of genotyping patients and carriers and expand the possibilities for screening at-risk populations.

An adenovirus, AdCDTK, expressing both bacterial cytosine deaminase (CD) and herpes simplex virus thymidine kinase (HSVTK) was constructed and introduced into glioma cells. AdCDTK selectively rendered glioma cells sensitive to both 5-fluorocytosine (5-FCyt) and ganciclovir (GCV) (termed AdCDTK/5-FCyt-GCV). AdCDTK/5-FCyt-GCV not only potently mediated apoptosis and the arrest of glioma cell growth in vitro, but also significantly increased the survival time of glioma-bearing rats as compared with controls. The 90-day survival time was observed in 50% of rats. Interferon-alpha (IFN-alpha) further enhanced the tumor cell killing of AdCDTK/5-FCyt-GCV. In the group of AdCDTK/5-FCyt-GCV/IFN-alpha, the average survival time was significantly increased, and the average tumor size was smaller than that in the group of AdCDTK/5-FCyt-GCV. Ninety-day survival increased from 50% in the group of AdCDTK/5-FCyt-GCV to 75% in the group of AdCDTK/5-FCyt-GCV/IFN-alpha. Complete tumor regression was observed in 50% of rats in the group of AdCDTK/5-FCyt-GCV/IFN-alpha. The data indicate that AdCDTK/5-FCyt-GCV induces glioma cell killing greater than that induced by either CD/5-FCyt or HSVTK/GCV alone. IFN-alpha synergistically enhances this effect by increasing apoptosis.

Whereas two mutations in the ASPA gene account for more than 98% of all mutant alleles causing Canavan disease in the Ashkenazi Jewish population, many different mutations can be found in non-Jewish individuals with Canavan disease. In our investigation of 40 non-Jewish patients with Canavan disease, we have found 24 novel mutations and one new polymorphism in the ASPA gene.

We describe two sisters with a mild-onset variant of Canavan's disease who presented at age 50 and 19 months with developmental delay but without macrocephaly, hypotonia, spasticity, or seizures. Remarkably, both patients had age-appropriate head control, gross motor development, and muscle tone. There were very mild deficits in fine motor skills, coordination, and gait. Both sisters had a history of strabismus, but otherwise vision was normal. The older child showed evidence of mild cognitive and social impairment, whereas language and behavior were normal for age in the infant. Both patients were found to be compound heterozygotes for C914A (A305E) and G212A (R71H) mutations in ASPA. Like all other known ASPA mutations, this previously unknown G212A mutation appears to have low absolute enzyme activity. Nevertheless, it is associated in these patients with an extremely benign phenotype that is highly atypical of Canavan's disease. Biochemical and clinical data were evaluated using a generalized linear mixed model generated from 25 other subjects with Canavan's disease. There were statistically significant differences in brain chemistry and clinical evaluations, supporting a distinct variant of Canavan's disease. Future studies of ASPA enzyme structure and gene regulation in these subjects could lead to a better understanding of Canavan's pathophysiology and improvements in ASPA gene therapy.

This work was supported by Comision Interministerial de Ciencia y Technologia (SAF 97-0074). We are grateful to Dr. E. Beutler for critical reading of the manuscript; to L. Gort, M. Martinez, and J. Armstrong for technical assistance; to R. Rycroft for revising the English; and to Dr. N. Espiro and Dr. J. Israel for helpful comments on Jewish history.

To the Editor:We have recently described a common origin for the most prevalent mutation (N370S) observed among Gaucher disease (GD) patients of Ashkenazi Jewish (AJ) and Spanish descent (Diaz et al. 1999xGaucher disease: the N370S mutation in Ashkenazi Jewish and Spanish patients has a common origin and arose several thousand years ago. Diaz, A, Montfort, M, Cormand, B, Zeng, BJ, Pastores, GM, Chabas, A, Vilageliu, L et al. Am J Hum Genet. 1999; 64: 1233–1238Abstract | Full Text | Full Text PDF | PubMed | Scopus (15)See all References1999). We also estimated the age of this mutation, using a formula described by Risch et al. (1995bxGenetic analysis of idiopathic torsion dystonia in Ashkenazi Jews and their recent descent from a small founder population. Risch, N, DeLeon, D, Ozelius, L, Kramer, P, Almasy, L, Singer, B, Fahn, S et al. Nat Genet. 1995b; 9: 152–159Crossref | PubMed | Scopus (299)See all References1995b). Unfortunately, as R. Colombo pointed out in a recent report (Colombo 2000xAge estimate of the N370S mutation causing Gaucher disease in Ashkenazi Jews and European populations: a reappraisal of haplotype data. Colombo, R. Am J Hum Genet. 2000; 66: 692–697Abstract | Full Text | Full Text PDF | PubMed | Scopus (22)See all References2000), there was an error in the formula presented in the original publication that was never rectified. In a reply (Risch et al. 1995axITD in Ashkenezi Jews—genetic drift or selection? In reply. Risch, N, DeLeon, D, Fahan, S, Ozelius, L, Breakfield, X, Kramer, P, Almasy, L et al. Nat Genet. 1995a; 11: 14–15CrossrefSee all References1995a) to criticisms raised by Zoossmann-Diskin (1995xITD in Ashkenazi Jews—genetic drift or selection?. Zoossmann-Diskin, A. Nat Genet. 1995; 11: 13–14Crossref | PubMed | Scopus (4)See all References1995), Risch et al. made no mention of an error in the formula. The continued application of the formula by researchers who may not be well versed in the field may lead to repeated mistakes if the error remains uncorrected. We apologize for our failure to recognize the error, but we are pleased that it has been identified by Colombo, who had no difficulty in re-estimating the age of the mutation, using our data. This reappraisal indicates that the N370S mutation may have occurred between the 11th and 13th centuries or even in the 10th century, considering 30 years per generation as suggested recently by Tremblay and Vezina (2000xNew estimates of intergenerational time intervals for the calculation of age and origin of mutations. Tremblay, M and Vezina, H. Am J Hum Genet. 2000; 66: 651–658Abstract | Full Text | Full Text PDF | PubMed | Scopus (117)See all References2000). This new estimated date for the mutation suggests that it occurred (or entered) the AJ population after the separation of the Ashkenazi and Sephardic Jewish traditions, which would be consistent with the apparent absence of this mutation among Sephardic patients. Using a totally different approach based on mutation detection in healthy Roman Jews, Oddoux et al. (1999xMendelian diseases among Roman Jews: implications for the origins of disease alleles. Oddoux, C, Guillen-Navarro, E, Ditivoli, C, Dicave, E, Cilio, MR, Clayton, CM, Nelson, H et al. J Clin Endocrinol Metab. 1999; 84: 4405–4409Crossref | PubMedSee all References1999) also got to the conclusion that the N370S is an old mutation. Further information on the origin of a mutation could be provided from the length of the chromosomal region noted with linkage disequilibrium (LD) and the strength of the LD. In this case, the data can be used to determine whether the N370S mutation had a Jewish or non-Jewish origin. The 3.2-cM region in LD in AJ (Diaz et al. 1999xGaucher disease: the N370S mutation in Ashkenazi Jewish and Spanish patients has a common origin and arose several thousand years ago. Diaz, A, Montfort, M, Cormand, B, Zeng, BJ, Pastores, GM, Chabas, A, Vilageliu, L et al. Am J Hum Genet. 1999; 64: 1233–1238Abstract | Full Text | Full Text PDF | PubMed | Scopus (15)See all References1999) seems to be shorter in Spanish chromosomes, as the flanking marker D1S2624 is in LD among AJ, whereas it is not in Spanish GD patients. Moreover, LD values are stronger in AJ than in Spanish chromosomes. These observations suggest that the mutation was introduced later in the AJ population, a statement that is independent of the actual age of the mutation. However, these results should be taken with caution because of the small number of Spanish chromosomes included in the study. Further work would be necessary to settle this issue which, to date, remains an open question.