Oncolytic activity of vesicular stomatitis virus in primary adult T-cell leukemia.

Treatments for hematological malignancies have improved considerably over the past decade, but the growing therapeutic arsenal has not benefited adult T-cell leukemia (ATL) patients. Oncolytic viruses such as vesicular stomatitis virus (VSV) have recently emerged as a potential treatment of solid tumors and leukemias in vitro and in vivo. In the current study, we investigated the ability of VSV to lyse primary human T-lymphotropic virus type 1 (HTLV-1)-infected T-lymphocytes from patients with ATL. Ex vivo primary ATL cells were permissive for VSV and underwent rapid oncolysis in a time-dependent manner. Importantly, VSV infection showed neither viral replication nor oncolysis in HTLV-1-infected, nonleukemic cells from patients with HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), and in naive CD4(+) T-lymphocytes from normal individuals or in ex vivo cell samples from patients with chronic lymphocytic leukemia (CLL). Interestingly, activation of primary CD4(+) T-lymphocytes with anti-CD3/CD28 monoclonal antibody, and specifically with anti-CD3, was sufficient to induce limited viral replication and oncolysis. However, at a similar level of T-cell activation, VSV replication was increased fourfold in ATL cells compared to activated CD4(+) T-lymphocytes, emphasizing the concept that VSV targets genetic defects unique to tumor cells to facilitate its replication. In conclusion, our findings provide the first essential information for the development of a VSV-based treatment for ATL.

Quantification of HTLV type I and HIV type I DNA load in coinfected patients: HIV type 1 infection does not alter HTLV type I proviral amount in the peripheral blood compartment.

Several reports suggest that HTLV-I/HIV coinfection may be associated with an increased risk of HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP). In HTLV-I-monoinfected patients, the occurrence of HAM/TSP is associated with high peripheral blood HTLV-I proviral load. Using a real-time quantitative PCR assay, we assessed the proviral DNA load in peripheral blood mononuclear cells (PBMCs) from 15 asymptomatic HTLV-I-monoinfected patients, 15 HTLV-I-monoinfected patients with HAM/TSP, and 25 HTLV-I/HIV-1 coinfected patients, including 4 with HAM/TSP. We also measured HIV-1 proviral DNA load in PBMCs from the coinfected patients. The median HTLV-I proviral loads were 6,800 and 4,100 copies per 10(6) PBMCs in the asymptomatic monoinfected and coinfected groups, and 58,800 and 43,300 copies per 10(6) PBMCs in the monoinfected and coinfected patients with HAM/TSP, respectively. The difference between HTLV-I proviral loads in HAM/TSP and asymptomatic monoinfected patients was statistically significant (p < 0.0001), but there was no difference between the HTLV-I-monoinfected and HTLV-I/HIV-1-coinfected groups. There was no correlation between HTLV-I and HIV-1 proviral load. HTLV-I proviral load did not correlate with the CD4+ T lymphocyte count. Among patients with no HTLV-I disease, the median copy number of HTLV-I per 10(6) circulating CD4+ T cells was 114,000 in the coinfected group and 16,700 in the monoinfected group, but the difference was not significant (p = 0.089). These data do not confirm the hypothesis in which HIV-1 coinfection would increase HTLV-I proviral burden in the PBMCs. However, depletion of the CD4+ T cell subset, the main target of HTLV-I, could be counterbalanced by an up-regulation of HTLV-I replication or by greater resistance of HTLV-I-infected cells to HIV-1-induced destruction.

Tissue inhibitor of metalloproteinase 3, matrix metalloproteinase 9, and neopterin in the cerebrospinal fluid: preferential presence in HTLV type I-infected neurologic patients versus healthy virus carriers.

The human retrovirus HTLV-I is responsible for the chronic progressive myelopathy, TSP/HAM, characterized by the presence of infiltrated T lymphocytes, cytokines, and matrix metalloproteinases (MMPs) within spinal cord lesions. MMPs have been associated with several neurological diseases, and we previously reported the specific presence of the extracellular matrix-degrading protease, MMP-9, in the cerebrospinal fluid of TSP/HAM patients. Nevertheless, previous studies have not yet shown whether the expression of MMP-9 is associated with HTLV-I infection per se, or with neurological symptoms following infection. In the present work, the presence of tissue inhibitors of metalloproteinases 1 and 3 (TIMP-1 and TIMP-3) and of MMP-9 in the CSF of HTLV-I-infected individuals was compared in TSP/HAM patients versus HTLV-I carriers without neurological symptoms. TIMP-3, a regulator of MMP activity and cell survival, was detected with a significantly higher frequency in the TSP/HAM group and paralleled the increased levels of MMP-9 and neopterin, a sensitive indicator of cellular immune activation. These data may reflect the intense cell remodeling that occurs intrathecally in inflamed tissue. Changes in MMP, TIMP, and neopterin expression were not related to age at onset of disease, grade of motor disability, progressor status, or duration of disease, presumably indicating that TSP/HAM patients are continuously subjected to viral and immunological pressure. All these observations suggest that TIMPs and MMPs may contribute to the pathogenesis of TSP/HAM, and hence a new therapeutic strategy targeting the MMP/TIMP balance is needed. These observations also suggest that MMP-9 and TIMP-3 in CSF may be useful markers in the follow-up of the efficacy of therapeutic trials in TSP/HAM patients.

Seroindeterminate patterns and seroconversions to human T-lymphotropic virus type I positivity in blood donors from Martinique, French West Indies.

BACKGROUND: Screening for human T-lymphotropic virus type I (HTLV-I) antibodies in volunteer blood donors has been systematic in the French West Indies since 1989. Western blot-indeterminate results are commonly obtained. The significance of these indeterminate serologic patterns in HTLV-I-endemic areas is still unclear. STUDY DESIGN AND METHODS: During a 2-year period, 9759 blood donors were tested for HTLV-I antibodies. The epidemiologic features of HTLV-I-seropositive, -seroindeterminate, and -seronegative donors were compared. A lookback investigation was performed for the HTLV-I-seropositive donors, and the HTLV-I-seroindeterminate individuals were followed up. RESULTS: Thirty-nine donors (0.4%) were HTLV-I seropositive and 49 (0.5%) were seroindeterminate. The age and sex ratio characteristics of the seroindeterminate donors are divergent from those of the HTLV-I-seropositive group and are more like those of the seronegative population. However, during the study period, three cases of seroconversion after an initial seroindeterminate profile were reported. Two cases were detected through follow-up of 38 HTLV-I-seroindeterminate donors over a mean of 8 months (2-24 months). The third seroconverter belonged to the HTLV-I-seropositive group and was identified through lookback investigation. This case is atypical, with p19 reactivity for several months before HTLV-I seropositivity. CONCLUSION: These findings indicate that, although HTLV-I-seroindeterminate donors mainly are HTLV-I-noninfected, the rate of seroconversion in a repeat blood donor population from an endemic region must be taken into consideration. Moreover, the case of delayed seroconversion observed in this study suggests the difficulty of counseling seroindeterminate blood donors in endemic regions.

Linkage disequilibrium at the SCA2 locus.

Spinocerebellar ataxia type 2 (SCA2) is caused by the expansion of an unstable CAG repeat encoding a polyglutamine tract. Repeats with 32 to 200 CAGs are associated with the disease, whereas normal chromosomes contain 13 to 33 repeats. We tested 220 families of different geographical origins for the SCA2 mutation. Thirty three were positive (15%). Twenty three families with at least two affected subjects were tested for linkage disequilibium (LD) between the SCA2 mutation and three microsatellite markers, two of which (D12S1332-D12S1333) closely flanked the mutation; the other (D12S1672) was intragenic. Many different haplotypes were observed, indicating the occurrence of several ancestral mutations. However, the same haplotype, not observed in controls, was detected in the German, the Serbian, and some of the French families, suggesting a founder effect or recurrent mutations on an at risk haplotype.

[A familial form of bilateral recurrent dislocation of the patella with major trochlea dysplasia]. / Une forme familiale de luxation récidivante bilatérale de la rotule avec dysplasie trochléenne majeure.

PURPOSE OF THE STUDY: We report four cases of bilateral recurrent dislocation of the patella with major trochlear dysplasia, in the same family. MATERIAL AND METHODS: Details of clinical examination of all members of this family and measurements on knee radiographs are reported. RESULTS: In all cases a severe proximal dysplasia of the trochlea was described on lateral views. The patella and the trochlea had a normal shape on the axial view. DISCUSSION: In some recurrent dislocation, with no associated disease, possibility of a genetic transmission have been suggested in some publications. Cases involving the same family have never been reported to confirm a genetic transmission of a bilateral and major trochlear dysplasia. CONCLUSION: This report points out a genetic origin of severe trochlear dysplasia. To know more about transmission and chromosomic localisation, careful investigations on others families of bilateral dislocations with trochlear dysplasia must be done.

Molecular and clinical correlations in spinocerebellar ataxia 2: a study of 32 families.

Spinocerebellar ataxia 2 (SCA2) is caused by the expansion of an unstable CAG repeat encoding a polyglutamine tract. One hundred and eighty four index patients with autosomal dominant cerebellar ataxia type I were screened for this mutation. We found expansion in 109 patients from 30 families of different geographical origins (15%) and in two isolated cases with no known family histories (2%). The SCA2 chromosomes contained from 34 to 57 repeats and consisted of a pure stretch of CAG, whereas all tested normal chromosomes (14-31 repeats), except one with 14 repeats, were interrupted by 1-3 repeats of CAA. As in other diseases caused by unstable mutations, a strong negative correlation was observed between the age at onset and the size of the CAG repeat (r = -0.81). The frequency of several clinical signs such as myoclonus, dystonia and myokymia increased with the number of CAG repeats whereas the frequency of others was related to disease duration. The CAG repeat was highly unstable during transmission with variations ranging from -8 to +12, and a mean increase of +2.2, but there was no significant difference according to the parental sex. This instability was confirmed by the high degree of gonadal mosaicism observed in sperm DNA of one patient.

Autosomal dominant cerebellar ataxia type I in Martinique (French West Indies): genetic analysis of three unrelated SCA2 families.

Autosomal dominant cerebellar ataxias (ADCAs) are a group of neurodegenerative disorders that are clinically and genetically heterogeneous. We report here a genetic linkage study, with five chromosome 12q markers, of three Martinican families with ADCA type 1, for which the spinocerebellar ataxia 1 (SCA1) locus was excluded. Linkage to the SCA2 locus was demonstrated with a maximal lead score of 6.64 at theta = 0.00 with marker D12S354. Recombinational events observed by haplotype reconstruction demonstrated that the SCA2 locus is located in an approximately 7-cM interval flanked by D12S105 and D12S79. Using the z(max)-1 method, multipoint analysis further reduced the candidate interval for SCA2 to a region of 5 cM. Two families shared a common haplotype at loci spanning 7 cM, which suggests a founder effect, whereas a different haplotype segregated with the disease in the third family. Finally, a mean anticipation of 12+/-14 years was found in parent-child couples, with no parental sex effect, suggesting that the disease might be caused by an expanded and unstable triplet repeat.

Autosomal dominant cerebellar ataxia type I in Martinique (French West Indies). Clinical and neuropathological analysis of 53 patients from three unrelated SCA2 families.

Autosomal dominant cerebellar ataxia type I was diagnosed in three unrelated families from Martinique (French West Indies), and linkage to the locus for spinocerebellar ataxia 2 (SCA2) was established. Neuropathological findings in two patients were those of olivopontocerebellar atrophy without oligodendroglial cytoplasmic inclusions. Cerebellar ataxia was associated with hyporeflexia in 68% of 31 examined patients, with slowed and/or limited eye movements in 65% and with dementia in 29%. No patients had optic atrophy, pigmentary retinal degeneration, spasticity or parkinsonism. Mean age at onset was 33 +/- 16 years, and onset before the age of 20 years was correlated with a more rapid and severe course of the disease. Movement disorders, oculomotor disturbances, sphincter disturbances and cognitive impairment were significantly more frequent in early than in late onset patients. This explains why the phenotype was strikingly different in one family, in which mean age at onset was much earlier. Comparison with previously described SCA2 families indicated similarities, such as reduced saccade velocity, supranuclear ophthalmoplegia and decreased reflexes, although phenotypic heterogeneity remains the outstanding feature of this disorder.

[Involvements of the peripheral nervous system and skeletal muscles in HTLV1-related paraplegia. Study of 70 cases seen in Martinique]. / Atteintes du système nerveux périphérique et du muscle squelettique au cours des paraplégies associées au virus HTLV1. Etude de 70 cas observés en Martinique.

The main neurological manifestation due to Human T-Cell Lymphotropic Virus type (HTLV1) infection is a chronic spastic paraparesis called HTLV1-associated paraparesis (HAP). Since 1985, we observed in Martinique (French West Indies) 276 cases of HAP. Among them, 70 patients fulfilled clinical, electrophysiological or histological criteria of associated peripheral nervous system involvement (42/70), myositis (8/70), or both (20/70). Muscle biopsy revealed neurogenic atrophy of muscle fibers or myositic changes in 41/70. Neuromuscular involvement was only mild in 19/70. On the other hand, 25 patients presented with a syndrome mimicking amyotrophic lateral sclerosis, and 7 other patients with features of polymyositis. This study shows that neurological manifestations associated to HTLV1 infection may be more complex than the well-known chronic spastic paraparesis, since 25.4% of the HAP Martinican patients exhibit neuropathic or myositic features.

Autosomal dominant cerebellar ataxia type I linked to chromosome 12q (SCA2: spinocerebellar ataxia type 2).

Spinocerebellar ataxia 2 (SCA2) is one of the loci for the clinically and genetically heterogeneous group of autosomal dominant type I cerebellar ataxias. After initial linkage to chromosome 12q in Cuban families, SCA2 was shown to be the gene responsible for the disease in Italian, Tunisian, French-Canadian, Austrian-Canadian and Martinican kindreds with dominant ataxia, and the candidate interval was reduced to 6.4 cM between markers D12S84 and D12S79. Comparison of patients from families of different geographical origins clearly demonstrates the clinical interfamilial variability of the clinical signs which reaches statistical significance for the frequency of extrapyramidal rigidity, postural tremor and dementia. The most striking difference between the 29 Martinican SCA2 patients and those with SCA1 on chromosome 6p or SCA3/MJD on chromosome 14q is the greater frequency of hyporeflexia in the former. A mean 12.5 year anticipation is observed, with a more rapid clinical course of the disease in successive generations, indicating that an expanded trinucleotide repeat probably constitutes the underlying molecular mechanism.

Prosomatostatin is processed in the Golgi apparatus of rat neural cells.

Proteolytic processing of somatostatin precursor produces several peptides including somatostatin-14 (S-14), somatostatin-28 (S-28), and somatostatin-28 (1-12) (S-28(1-12)). The subcellular sites at which these cleavages occur were identified by quantitative evaluation of these products in enriched fractions of the biosynthetic secretory apparatus of rat cortical or hypothalamic cells. Each of the major cellular compartments was obtained by discontinuous gradient centrifugation and was characterized both by specific enzyme markers and electron microscopy. The prosomatostatin-derived fragments were measured by radioimmunoassay after chromatographic separation. Two specific antibodies were used, allowing the identification of either S-28(1-12) or S-14 which results from peptide bond hydrolysis at a monobasic (arginine) and a dibasic (Arg-Lys) cleavage site, respectively. These antibodies also revealed prosomatostatin-derived forms containing at their COOH terminus the corresponding dodeca- and tetradecapeptide sequences. Whereas the reticulum-enriched fractions contained the highest levels of prosomatostatin, the proportion of precursor was significantly lower in the Golgi apparatus. In the latter fraction, other processed forms were also present, i.e. S-14 and S-28(1-12) together with the NH2-terminal domain (1-76) of prosomatostatin (pro-S(1-76). Inhibition of the intracellular transport either by monensin or by preincubation at reduced temperature resulted in an increase of prosomatostatin-derived peptides in the Golgi-enriched fractions. Finally, immunogold labeling using antibodies raised against S-28(1-12) and S-14 epitopes revealed the presence of these forms almost exclusively in the Golgi-enriched fraction mainly at the surface of saccules and vesicles. Together these data demonstrate that in rat neural cells, prosomatostatin proteolytic processing at both monobasic and dibasic sites is initiated at the level of the Golgi apparatus.