Current medico-psycho-social conditions of patients with West syndrome in Japan.

OBJECTIVE: To unveil current medical and psychosocial conditions of patients with West syndrome in Japan. METHODS: A cross-sectional analysis was performed in patients with West syndrome registered in the Rare Epilepsy Syndrome Registry (RES-R) of Japan. Furthermore, new-onset patients registered in the RES-R were observed prospectively and their outcomes after one and two years of follow-up were compared with data at onset. RESULTS: For the cross-sectional study, 303 patients with West syndrome were included. Seizures (such as spasms, tonic seizures and focal seizures) occurred daily in 69.3% of the patients at registration. Seizure frequency of less than one per year was observed in cases of unknown etiology (22.6%), genetic etiology (23.8%) and malformation of cortical development (MCD; 19.1%). Neurological findings were absent in 37.0%, but a high rate of abnormality was seen in patients with Aicardi syndrome, hypoxic-ischemic encephalopathy (HIE), genetic etiology and MCD other than focal cortical dysplasia, accompanied by a >50% rate of bedridden patients. Abnormal EEG was found in 96.7%, and CT/MRI was abnormal in 62.7%. Treatments included antiepileptic drug therapy (94.3%), hormonal therapy (72.6%), diet therapy (8.3%) and surgery (15.8%). Intellectual/developmental delay was present in 88.4%, and was more severe in patients with Aicardi syndrome, genetic etiology and HIE. Autism spectrum disorder was found in 13.5%. For the longitudinal study, 27 new-onset West syndrome patients were included. The follow-up study revealed improved seizure status after two years in 66.7%, but worsened developmental status in 55.6%, with overall improvement in 51.9%. SIGNIFICANCE: The study reveals the challenging neurological, physical and developmental aspects, as well as intractable seizures, in patients with West syndrome. More than a half of the children showed developmental delay after onset, even though seizures were reduced during the course of the disease.

Coffin-Siris syndrome with bilateral macular dysplasia caused by a novel exonic deletion in ARID1B.

Coffin-Siris syndrome (CSS) is a congenital anomaly syndrome characterized by developmental delay, coarse facial features, and hypoplasia of the fifth digit's nail or phalanges. Herein, we report a case of the 8-year-old female patient who showed developmental delay associated with dysplasia in the macular and large toe area. Comprehensive genomic analysis showed no possible candidate variants, but the subsequent genomic copy number analysis revealed a novel exonic deletion in the coding region of AT-rich interactive domain-containing protein 1B (ARID1B), a gene responsible for CSS. Genomic copy number analysis can aid in diagnosing CSS by confirming undiagnosed exonic deletions in ARID1B. Furthermore, this is the first report of CSS associated with bilateral macular dysplasia.

Further characterization of CAPOS/CAOS syndrome with the Glu818Lys mutation in the ATP1A3 gene: A case report.

A 38-year-old female patient experienced recurrent episodes of neurological deterioration during febrile illness at the age of 7 and 8 months, and 2, 4, and 37 years. Acute symptoms comprised unconsciousness, headache, abnormal ocular movements, flaccid paralysis with areflexia, ataxia, dysphagia, and movement disorders. Each episode of neurological deterioration was followed by partial recovery with residual symptoms of progressive disturbance of visual acuity with optic atrophy and hearing loss, moderate intellectual disability, strabismus, ophthalmoplegia, as well as fluctuating degree of gait ataxia, chorea, tremor, and myoclonus. In addition, electrocardiography revealed incomplete right bundle branch block. The genetic testing revealed a de novo heterozygous mutation of c.2452G > A (p.Glu818Lys) in the ATP1A3 gene, which was compatible with the clinical phenotype of CAPOS (cerebellar ataxia, areflexia, pes cavus, optic atrophy, and sensorineural hearing loss)/CAOS syndrome. Here we discuss the significance of clinical features of a patient, overlapping with those of alternating hemiplegia of childhood, along with a literature review.

Long-term follow up of an adult with alternating hemiplegia of childhood and a p.Gly755Ser mutation in the ATP1A3 gene.

Alternating hemiplegia of childhood (AHC) is a rare neurological disease mainly caused by mutations in the ATP1A3 gene and showing varied clinical severity according to genotype. Patients with a p.Gly755Ser (p.G755S) mutation, one of minor genotypes for AHC, were recently described as having a mild phenotype, although their long-term outcomes are still unclear due to the lack of long-term follow up. Here, we demonstrate the full clinical course of a 43-year-old female AHC patient with p.G755S mutation. Although her motor dysfunction had been relatively mild into her 30 s, she showed a subsequent severe aggravation of symptoms that left her bedridden, concomitant with a recent recurrence of seizure status. The seizures were refractory to anti-epileptic drugs, but administration of flunarizine improved seizures and the paralysis. Our case suggests that the phenotype of AHC with p.G755S mutation is not necessarily mild, despite such a presentation during the patient's younger years.

A de novo missense mutation of GABRB2 causes early myoclonic encephalopathy.

BACKGROUND: Early myoclonic encephalopathy (EME), a disease with a devastating prognosis, is characterised by neonatal onset of seizures and massive myoclonus accompanied by a continuous suppression-burst EEG pattern. Three genes are associated with EMEs that have metabolic features. Here, we report a pathogenic mutation of an ion channel as a cause of EME for the first time. METHODS: Sequencing was performed for 214 patients with epileptic seizures using a gene panel with 109 genes that are known or suspected to cause epileptic seizures. Functional assessments were demonstrated by using electrophysiological experiments and immunostaining for mutant γ-aminobutyric acid-A (GABAA) receptor subunits in HEK293T cells. RESULTS: We discovered a de novo heterozygous missense mutation (c.859A>C [p.Thr287Pro]) in the GABRB2-encoded ß2 subunit of the GABAA receptor in an infant with EME. No GABRB2 mutations were found in three other EME cases or in 166 patients with infantile spasms. GABAA receptors bearing the mutant ß2 subunit were poorly trafficked to the cell membrane and prevented γ2 subunits from trafficking to the cell surface. The peak amplitudes of currents from GABAA receptors containing only mutant ß2 subunits were smaller than that of those from receptors containing only wild-type ß2 subunits. The decrease in peak current amplitude (96.4% reduction) associated with the mutant GABAA receptor was greater than expected, based on the degree to which cell surface expression was reduced (66% reduction). CONCLUSION: This mutation has complex functional effects on GABAA receptors, including reduction of cell surface expression and attenuation of channel function, which would significantly perturb GABAergic inhibition in the brain.

Treatment with Oral ATP decreases alternating hemiplegia of childhood with de novo ATP1A3 Mutation.

BACKGROUND: Alternating hemiplegia of childhood is an intractable neurological disorder characterized by recurrent episodes of alternating hemiplegia accompanied by other paroxysmal symptoms. Recent research has identified mutations in the ATP1A3 gene as the underlying cause. Adenosine-5'-triphosphate has a vasodilatory effect, can enhance muscle strength and physical performance, and was hypothesized to improve the symptoms of paroxysmal hemiplegia. METHODS: A 7-year-old boy with alternating hemiplegia of childhood who was positive for a de novo ATP1A3 mutation was treated with adenosine- 5'- triphosphate supplementation orally as an innovative therapy for 2 years. Outcome was evaluated through the follow-up of improvement of hemiplegic episodes and psychomotor development. Side effects and safety were monitored in regularity. RESULTS: With the dosage of adenosine-5'-triphosphate administration increased, the patient showed significantly less frequency and shorter duration of hemiplegic episodes. Treatment with adenosine-5'-triphosphate was correlated with a marked amelioration of alternating hemiplegia of childhood episodes, and psychomotor development has improved. The maximum dose of oral administration of adenosine-5'-triphosphate reached 25 mg/kg per day. Adenosine-5'-triphosphate therapy was well tolerated without complaint of discomfort and side effects. CONCLUSIONS: The 2-year follow-up outcome of adenosine-5'-triphosphate therapy for alternating hemiplegia of childhood was successful.

Clinical and genetic features of acute encephalopathy in children taking theophylline.

BACKGROUND: Theophylline has recently been suspected as a risk factor of acute encephalopathy with biphasic seizures and late reduced diffusion (AESD), although there has been no systematic study on the relationship between acute encephalopathy in children taking theophylline (AET) and AESD. METHODS: We recruited 16 Japanese patients (11 male and 5 female, median age of 2 years and 7 months) with AET from 2008 to 2013. We evaluated their clinical features, such as the duration of first seizure, biphasic clinical course and cranial CT/MRI imaging and compared them with those of AESD. We analyzed the polymorphisms or mutations of genes which are associated with AESD. RESULTS: Clinically, 12 patients had neurological and/or radiological features of AESD. Only one patient died, whereas all 15 surviving patients were left with motor and/or intellectual deficits. Genetically, 14 patients had at least one of the following polymorphisms or mutations associated with AESD: thermolabile variation of the carnitine palmitoyltransferase 2 (CPT2) gene, polymorphism causing high expression of the adenosine receptor A2A (ADORA2A) gene, and heterozygous missense mutation of the voltage gated sodium channel 1A (SCN1A) and 2A (SCN2A) gene. CONCLUSIONS: Our results demonstrate that AET overlaps with AESD, and that AET is a multifactorial disorder sharing a genetic background with AESD.

Case of Desbuquois dysplasia type 1: potentially lethal skeletal dysplasia.

We report a boy with Desbuquois dysplasia type 1. He had the typical skeletal changes: a "Swedish key" appearance of the proximal femora; advanced carpal ossification and other distinctive features of the hand, including an extra-ossification center at the base of the proximal phalanx of the index and middle fingers; dislocation of the metacarpophalangeal joint of the index finger; and bifid distal phalanx of the thumb. In addition, he presented with very severe prenatal growth failure, respiratory distress as a neonate, subsequent failure to thrive and susceptibility to airway infection, and sudden death in early childhood. Molecular analysis identified homozygous 1 bp deletion in the Calcium-Activated Nucleotidase 1 gene (CANT1). To our knowledge, this is the first report of Desbuquois dysplasia type 1 in Japan. Our experience suggests potential lethality in the disorder.

Genotype-phenotype correlations in alternating hemiplegia of childhood.

OBJECTIVE: Clinical severity of alternating hemiplegia of childhood (AHC) is extremely variable. To investigate genotype-phenotype correlations in AHC, we analyzed the clinical information and ATP1A3 mutations in patients with AHC. METHODS: Thirty-five Japanese patients who were clinically diagnosed with AHC participated in this study. ATP1A3 mutations were analyzed using Sanger sequencing. Detailed clinical information was collected from family members of patients with AHC and clinicians responsible for their care. RESULTS: Gene analysis revealed 33 patients with de novo heterozygous missense mutations of ATP1A3: Glu815Lys in 12 cases (36%), Asp801Asn in 10 cases (30%), and other missense mutations in 11 cases. Clinical information was compared among the Glu815Lys, Asp801Asn, and other mutation groups. Statistical analysis revealed significant differences in the history of neonatal onset, gross motor level, status epilepticus, and respiratory paralysis in the Glu815Lys group compared with the other groups. In addition, 8 patients who did not receive flunarizine had severe motor deteriorations. CONCLUSIONS: The Glu815Lys genotype appears to be associated with the most severe AHC phenotype. Although AHC is not generally seen as a progressive disorder, it should be considered a disorder that deteriorates abruptly or in a stepwise fashion, particularly in patients with the Glu815Lys mutation.

A recurrent KCNT1 mutation in two sporadic cases with malignant migrating partial seizures in infancy.

We performed analysis of KCNT1 in two unrelated patients with malignant migrating partial seizures in infancy. Both patients had intractable focal seizures since two months of age. Their seizures were characterized by a shift of epileptic focus during a single seizure and were resistant to most antiepileptic drugs but responded to vagus nerve stimulation in one and clorazepate in the other. Bidirectional sequencing for KCNT1 was analyzed by standard Sanger sequencing method. A de novo c.862G>A (p.Gly288Ser) missense mutation was identified at the pore region of KCNT1 channel in both patients, whereas all KCNT1 mutations in the previous reports were identified mostly in the intracellular C-terminal region. Computational analysis suggested possible changes in the molecular structure and the ion channel property induced by the Gly288Ser mutation. Because the G-to-A transition was located at CG dinucleotide sequences as previously reported for KCNT1 mutations, the recurrent occurrence of de novo KCNT1 mutations indicated the hot spots of these locations.

Newborn screening for Fabry disease in Japan: prevalence and genotypes of Fabry disease in a pilot study.

Fabry disease (FD) is an X-linked lysosomal storage disorder caused by a deficiency of α-galactosidase A (GLA) activity. Enzyme replacement therapy (ERT) for FD is available, and newborn mass screening for FD is being implemented. Here, we undertook a pilot study of newborn mass screening for FD in Japan. GLA activity in dried blood spots was measured using a fluorescence assay and confirmed by measurement of GLA activity in white blood cells (WBCs) in infants with abnormally low GLA activity. This was followed up by genetic testing. A total of 21 170 neonates were enrolled in the study. Of these, seven (five boys, two girls) had low GLA activities, which were verified by the WBC GLA activity assay. Thus, the initial fluorescence assay was suitable for newborn mass screening for FD. Pathogenic mutations of the GLA gene, that is, V199M and IVS4+919G>A, were found in two boys and one boy, respectively. Functional mutations, E66Q and c.-10C>T: g.1170C>T, were found in two boys and one girl, respectively. The prevalence of test-positive newborns was 1/3024, while that of those with a pathogenic mutation was 1/7057. The numbers are higher than those previously anticipated. Standardized management for FD found during newborn mass screening, including an ERT regimen, remains to be established.

A Pancreatic Solid Pseudo-Papillary Tumor Detected After Abdominal Injury.

Solid pseudo-papillary tumor (SPT) of the pancreas is a relatively benign tumor that is more frequently reported in females. Most patients usually present with abdominal pain or mass. We experienced the girl who identified SPT with the injury. We diagnosed SPT in a previously healthy 14-year-old Asian girl after abdominal injury. She experienced upper abdominal pain and vomiting after being hit by a basketball. Blood examination revealed a high serum amylase level. Abdominal radiography indicated abnormal bowel gases. Contrast-enhanced computed tomography revealed a smooth, peripheral and unilocular mass approximately 55 mm in diameter in the pancreatic tail. Based on these observations, acute pancreatitis complicated by a pancreatic mass was initially diagnosed. Therapy for acute pancreatitis was instituted, while we simultaneously investigated the mass. Levels of tumor markers were not profoundly elevated in serum. Dynamic contrast-enhanced magnetic resonance imaging (MRI) revealed moderate and gradual increase in contrast-enhanced imaging, consistent with findings of SPT of the pancreas. We thus elected surgical resection for her. Pathological examination of the surgical specimen confirmed our diagnosis of SPT. SPT of the pancreas should be considered as a differential diagnosis of acute abdomen disorders, especially in instances after minor abdominal injuries in young women, and diagnoses must be confirmed with MRIs.

Identification of ATP1A3 mutations by exome sequencing as the cause of alternating hemiplegia of childhood in Japanese patients.

BACKGROUND: Alternating hemiplegia of childhood (AHC) is a rare disorder characterized by transient repeated attacks of paresis and cognitive impairment. Recent studies from the U.S. and Europe have described ATP1A3 mutations in AHC. However, the genotype-phenotype relationship remains unclear. The purpose of this study was to identify the genetic abnormality in a Japanese cohort of AHC using exome analysis. PRINCIPAL FINDINGS: A total of 712,558 genetic single nucleotide variations in 8 patients with sporadic AHC were found. After a series of exclusions, mutations of three genes were regarded as candidate causes of AHC. Each patient harbored a heterozygous missense mutation of ATP1A3, which included G755C, E815K, C927Y and D801N. All mutations were at highly conserved amino acid residues and deduced to affect ATPase activity of the corresponding ATP pump, the product of ATP1A3. They were de novo mutations and not identified in 96 healthy volunteers. Using Sanger sequencing, E815K was found in two other sporadic cases of AHC. In this study, E815K was found in 5 of 10 patients (50%), a prevalence higher than that reported in two recent studies [19 of 82 (23%) and 7 of 24 (29%)]. Furthermore, the clinical data of the affected individuals indicated that E815K resulted in a severer phenotype compared with other ATP1A3 mutations. INTERPRETATION: Heterozygous de novo mutations of ATP1A3 were identified in all Japanese patients with AHC examined in this study, confirming that ATP1A3 mutation is the cause of AHC.

Detection of human adenovirus hexon antigen using carbon nanotube sensors.

Human adenoviruses (HAdVs) have been implicated in a wide range of diseases affecting primarily the respiratory, ocular and gastrointestinal systems. A rapid and efficient method for the detection of HAdV hexon antigen is described using carbon nanotube (CNT) sensors. Anti-HAdV antibody was immobilised on the reverse surface of a CNT sensor. As a control, non-specific mouse IgG was immobilised on another CNT sensor. I-V(gate) curves were measured after incubation of various concentrations of recombinant HAdVs hexon antigen with anti-HAdVs antibody-immobilised or non-specific mouse IgG-immobilised sensors. The curves showed a positive shift that was dependent on the hexon antigen concentration in the anti-HAdV antibody-immobilised sensor, whereas no such shift was observed in the non-specific mouse IgG-immobilised sensor. The sensitivity of the CNT sensor method was greater than that of enzyme-linked immunosorbent assay. Hence, this method offers a new tool for HAdV detection by analysing antigen-antibody interactions.

Polyion complex stability and gene silencing efficiency with a siRNA-grafted polymer delivery system.

An siRNA-grafted polymer through disulfide linkage was prepared to improve the physicochemical properties and transfection efficacies of the polyion complexes (PICs) as a nanocarrier of siRNA. The siRNA-grafted polymer formed stable PICs due to its larger numbers and higher density of anionic charges compared with monomeric siRNA, leading to effective internalization by cultured cells. Following the endosomal escape of the PIC, the disulfide linkage of the siRNA-grafted polymer allowed efficient siRNA release from the PIC under intracellular reductive conditions. Consequently, the PIC from the siRNA-grafted polymer showed a potent gene silencing effect without cytotoxicity or immunogenicity, demonstrating a promising feature of the siRNA-grafted polymer to construct the PIC-based nanocarrier for in vivo siRNA delivery.

Introduction of stearoyl moieties into a biocompatible cationic polyaspartamide derivative, PAsp(DET), with endosomal escaping function for enhanced siRNA-mediated gene knockdown.

Applications of siRNA for cancer therapy have been spotlighted in recent years, but the rational design of efficient siRNA delivery carriers is still controversial, especially because of possible toxicity of the carrier components. Previously, a cationic polyaspartamide derivative, poly{N-[N-(2-aminoethyl)-2-aminoethyl]aspartamide} (PAsp(DET)), was reported to exert high transfection efficacy for plasmid DNA with negligible cytotoxicity. However, its direct application for siRNA delivery was fairly limited due to the unstable polymer/siRNA complex formation. In this study, to overcome such instability, stearic acid as a hydrophobic moiety was conjugated to the side chain of PAsp(DET) with various substitution degrees. The stearoyl introduction contributed not only to siRNA complex formation with higher association numbers but also to complex stabilization. The obtained stearoyl PAsp(DET)/siRNA complex significantly accomplished more efficient endogenous gene (BCL-2 and VEGF) knockdown in vitro against the human pancreatic adenocarcinoma (Panc-1) cells than did the unmodified PAsp(DET) complex and commercially available reagents, probably due to the facilitated cellular internalization. This finding suggests that the hydrophobic PAsp(DET)-mediated siRNA delivery is a promising platform for in vivo siRNA delivery.

GM3 synthase gene is a novel biomarker for histological classification and drug sensitivity against epidermal growth factor receptor tyrosine kinase inhibitors in non-small cell lung cancer.

Expression of gangliosides and alterations in their composition have been observed during cell proliferation and differentiation and in certain cell cycle phases, brain development and cancer malignancy. To investigate the characteristics of GM3 synthase, SAT-I mRNA and ganglioside GM3 expression levels in lung cancer, we examined the expression levels of SAT-I mRNA as well as GM3 in 40 tumor tissues surgically removed from non-small cell lung cancer patients. Adenocarcinoma tissues expressed SAT-I mRNA levels that were significantly higher than those of squamous and other carcinomas (P < 0.0001). Moreover, the SAT-I mRNA levels were high in the bronchioalveolar carcinoma subtype and low in the solid and mucin subtypes of adenocarcinomas (P = 0.049, 0.049 and 0.013, respectively). To clarify the relationship between SAT-I mRNA and epidermal growth factor receptor (EGFR)-tyrosine kinase (TK) inhibitor sensitivity, we carried out drug sensitivity tests for the EGFR-TK inhibitors gefitinib and AG1478 using eight adenocarcinoma cell lines expressing no EGFR mutations. The IC(50) values for gefitinib and AG1478 decreased dramatically with increasing SAT-I mRNA levels (R(2) = 0.81 and 0.59, respectively), representing a wide range of drug sensitivities among adenocarcinoma cell lines. To explore a possible mechanism of how GM3 could enhance the sensitivity to EGFR-TK inhibitors, the SAT-I gene was introduced stably into a GM3-negative clone of murine 3LL lung cancer cells to produce GM3-reconstituted clones. We found an increase in EGFR protein levels and gefitinib sensitivity in GM3-reconstituted cells, suggesting the involvement of GM3 in the turnover of EGFR protein. Therefore, it is highly expected that, by measuring the expression levels of SAT-I mRNA in lung biopsy samples from non-small cell lung cancer patients, enhanced pathological identification and individualized chemotherapeutic strategies can be established for the appropriate use of EGFR-TK inhibitors.

Role for up-regulated ganglioside biosynthesis and association of Src family kinases with microdomains in retinoic acid-induced differentiation of F9 embryonal carcinoma cells.

Mouse F9 embryonal carcinoma cells have been widely used as a model for studying the mechanism of embryonic differentiation, because they are similar to the inner cell mass of early mouse embryos and can differentiate into primitive endoderm (PrE) following retinoic acid (RA) treatment. During F9 cell differentiation, the carbohydrate chains of glycoproteins and their corresponding glycosyltransferases are known to undergo rapid changes. However, there have been no corresponding reports on the expression of gangliosides. We have developed a custom cDNA array that is highly sensitive for the genes responsible for sphingolipid (SL) biosynthesis and metabolism. Using this, we found that, of the 28 selected genes, 26 exhibited increased expression during F9 differentiation into PrE. Although neutral glycosphingolipids (GSLs) were expressed at similar levels before and after differentiation, a greater than 20-fold increase in total ganglioside content was evident in PrE. Glucosylceramide synthase inhibitors (d-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol [d-PDMP] and its analog) depleted gangliosides and this resulted in delayed expression of Disabled-2 (Dab-2), suggesting the involvement of gangliosides in F9 cell differentiation. Disruption of cholesterol-enriched membrane microdomains by methyl-beta-cyclodextrin (MbetaCD) also delayed differentiation. Both MbetaCD and d-PDMP blocked the accumulation of Src family kinases (SFKs) to microdomains. However, d-PDMP did not block flotillin accumulation, yet MbetaCD did. Additionally, confocal laser microscopy revealed the formation of distinct functional microdomains integrating SFKs with gangliosides and cholesterol during PrE differentiation. Thus, we demonstrate the outstanding up-regulation of ganglioside biosynthesis and its importance in the formation of distinct microdomains incorporating SFKs with gangliosides during RA-induced differentiation of F9 cells.

Ganglioside G(M3) overexpression induces apoptosis and reduces malignant potential in murine bladder cancer.

We demonstrated previously (S. Kawamura et al., Int. J. Cancer, 94: 343-347, 2001) that large amounts of ganglioside G(M3) accumulate in superficial bladder tumor, compared with invasive bladder tumors and that exogenous G(M3) inhibits the invasive potential of bladder tumor cells. To apply the G(M3) overexpression system to bladder tumor therapy, direct evidence for the important role of G(M3) in bladder tumor invasion must be obtained through transfer of the gene responsible for G(M3) overexpression. To determine the most appropriate cancer cell line for elucidating the antitumor effect of ganglioside G(M3) overexpression, the present study examined glycolipid composition, enzyme activity, and mRNA expression of the glycosyltransferases responsible for G(M3) synthesis in the bladder tumor cell lines KK-47, J82, MGH-UI, YTS-1, and MBT-2. A murine bladder carcinoma cell line (MBT-2) was transfected with a G(M3) synthase [(lactosylceramide alpha2,3-N-acetyl sialic acid transferase); sialyltransferase-I; SAT-I] cDNA, because this line does not naturally express G(M3). Stable transfectants (MBT-2-SAT-I) that overexpressed G(M3) were characterized by a reduced potential for cell proliferation, motility, invasion, and xenograft tumor growth, and an increase in the number of apoptotic cells. In the proportion of synthetic S phase, cells did not differ between MBT-2-SAT-I and mock-transfectant cells. These results suggest that the decreased proliferative potential related to G(M3) overexpression was attributable to the increased number of apoptotic cells. Although details of the mechanism of apoptosis remain unclear, the overexpression of G(M3) by gene transfer of SAT-I may present a novel therapeutic modality.

DNA cleavage potency, cytotoxicity, and mechanism of action of a novel class of enediyne prodrugs.

We have discovered a novel class of (E)-3-acyloxy-4-(arylmethylidene)cyclodeca-1,5-diynes which exhibit promising enediyne-like DNA cleavage and cytotoxic activities. LC-MS analysis of the incubation mixture (pH 8.5, 37 degrees C) confirmed formation of 10-membered ring enediyne presumably via an allylic cation and suggested that the 1,4-benzenoid diradical might be one of the active species for DNA damage and cytotoxicity.