Dichloroacetate causes toxic neuropathy in MELAS: a randomized, controlled clinical trial.

OBJECTIVE: To evaluate the efficacy of dichloroacetate (DCA) in the treatment of mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS). BACKGROUND: High levels of ventricular lactate, the brain spectroscopic signature of MELAS, correlate with more severe neurologic impairment. The authors hypothesized that chronic cerebral lactic acidosis exacerbates neuronal injury in MELAS and therefore, investigated DCA, a potent lactate-lowering agent, as potential treatment for MELAS. METHODS: The authors conducted a double-blind, placebo-controlled, randomized, 3-year cross-over trial of DCA (25 mg/kg/day) in 30 patients (aged 10 to 60 years) with MELAS and the A3243G mutation. Primary outcome measure was a Global Assessment of Treatment Efficacy (GATE) score based on a health-related event inventory, and on neurologic, neuropsychological, and daily living functioning. Biologic outcome measures included venous, CSF, and 1H MRSI-estimated brain lactate. Blood tests and nerve conduction studies were performed to monitor safety. RESULTS: During the initial 24-month treatment period, 15 of 15 patients randomized to DCA were taken off study medication, compared to 4 of 15 patients randomized to placebo. Study medication was discontinued in 17 of 19 patients because of onset or worsening of peripheral neuropathy. The clinical trial was terminated early because of peripheral nerve toxicity. The mean GATE score was not significantly different between treatment arms. CONCLUSION: DCA at 25 mg/kg/day is associated with peripheral nerve toxicity resulting in a high rate of medication discontinuation and early study termination. Under these experimental conditions, the authors were unable to detect any beneficial effect. The findings show that DCA-associated neuropathy overshadows the assessment of any potential benefit in MELAS.

Cerebral lactic acidosis correlates with neurological impairment in MELAS.

OBJECTIVE: To evaluate the role of chronic cerebral lactic acidosis in mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes (MELAS). METHODS: The authors studied 91 individuals from 34 families with MELAS and the A3243G point mutation and 15 individuals from two families with myoclonus epilepsy and ragged red fibers (MERRF) and the A8344G mutation. Subjects were divided into four groups. Paternal relatives were studied as controls (Group 1). The maternally related subjects were divided clinically into three groups: asymptomatic (no clinical evidence of neurologic disease) (Group 2), oligosymptomatic (neurologic symptoms but without the full clinical picture of MELAS or MERRF) (Group 3), and symptomatic (fulfilling MELAS or MERRF criteria) (Group 4). The authors performed a standardized neurologic examination, neuropsychological testing, MRS, and leukocyte DNA analysis in all subjects. RESULTS: The symptomatic and oligosymptomatic MELAS subjects had significantly higher ventricular lactate than the other groups. There was a significant correlation between degree of neuropsychological and neurologic impairment and cerebral lactic acidosis as estimated by ventricular MRS lactate levels. CONCLUSIONS: High levels of ventricular lactate, the brain spectroscopic signature of MELAS, are associated with more severe neurologic impairment.

Long survival and therapeutic responses in patients with histologically disparate high-grade gliomas demonstrating chromosome 1p loss.

OBJECT: Allelic loss of chromosome 1p is a powerful predictor of tumor chemosensitivity and prolonged survival in patients with anaplastic oligodendrogliomas. Chromosome 1p loss also occurs in astrocytic and oligoastrocytic gliomas, although less commonly than in pure oligodendroglial tumors. This observation raises the possibility investigated in this study that chromosome 1p loss might also provide prognostic information for patients with high-grade gliomas with astrocytic components. METHODS: The authors report on seven patients with high-grade gliomas composed of either pure astrocytic or mixed astrocytic-oligodendroglial phenotypes, who had remarkable neuroradiological responses to therapy or unexpectedly long survivals. All of the tumors from these seven patients demonstrated chromosome 1p loss, whereas other genetic alterations characteristic of high-grade gliomas (p53 gene mutations, EGFR gene amplification, chromosome 10 loss, chromosome 19q loss, or CDKN2A/p16 deletions) were only found in occasional cases. The authors also assessed the frequency of chromosome 1p loss in a series of anonymous high-grade astrocytoma samples obtained from a tumor bank and demonstrate that this genetic change is uncommon, occurring in only 10% of cases. CONCLUSIONS: Although any prognostic importance of chromosome 1p loss in astrocytic or mixed astrocytic-oligodendroglial gliomas can only be determined in larger and prospective series, these findings raise the possibility that some high-grade gliomas with chromosome 1p loss, in addition to pure anaplastic oligodendrogliomas, may follow a more favorable clinical course.

Complement depletion facilitates the infection of multiple brain tumors by an intravascular, replication-conditional herpes simplex virus mutant.

Intravascular routes of administration can provide a means to target gene- and virus-based therapies to multiple tumor foci located within an organ, such as the brain. However, we demonstrate here that rodent plasma inhibits cell transduction by replication-conditional (oncolytic) herpes simplex viruses (HSV), replication-defective HSV, and adenovirus vectors. In vitro depletion of complement with mild heat treatment or in vivo depletion by treatment of athymic rats with cobra venom factor (CVF) partially reverses this effect. Without CVF, inhibition of cell infection by HSV is observed at plasma dilution as high as 1:32, while plasma from CVF-treated animals displays anti-HSV activity at lower dilutions (1:8). When applied to the therapy of intracerebral brain tumors, in vivo complement depletion facilitates the initial infection (assayed at the 2-day time point) by an intra-arterial replication-conditional HSV of tumor cells, located within three separate and distinct human glioma masses. However, at the 4-day time point, no propagation of HSV from initially infected tumor cells could be observed. Previously, we have shown that the immunosuppressive agent, cyclophosphamide (CPA), facilitates the in vivo propagation of an oncolytic HSV, delivered intravascularly, within infected multiple intracerebral masses, by inhibition of both innate and elicited anti-HSV neutralizing antibody response (K. Ikeda et al., Nat. Med. 5:881-889, 1999). In this study, we thus show that the addition of CPA to the CVF treatment results in a significant increase in viral propagation within infected tumors, measured at the 4-day time period. The concerted action of CVF and CPA significantly increases the life span of athymic rodents harboring three separate and large glioma xenografts after treatment with intravascular, oncolytic HSV. Southern analysis of viral genomes analyzed by PCR reveals the presence of the oncolytic virus in the brains, livers, spleens, kidneys, and intestine of treated animals, although none of these tissues displays evidence of HSV-mediated gene expression. In light of clinical trials of oncolytic HSV for malignant brain tumors, these findings suggest that antitumor efficacy may be limited by the host innate and elicited humoral responses.

Molecular genetic correlates of p16, cdk4, and pRb immunohistochemistry in glioblastomas.

The vast majority of glioblastomas have CDKN2A, CDK4, or RB gene alterations that perturb the p16-cdk4-pRb cell cycle regulatory cascade. To explore whether immunohistochemical methods provide an alternative means of assessing this pathway, we studied 25 glioblastomas using a combination of molecular genetic and immunohistochemical assays. Homozygous deletion of the CDKN2A gene was detected in 12 of 25 (48%) cases, CDK4 amplification in 4 of 25 (16%) tumors, and loss of heterozygosity at the RB gene in 8 of 22 (36%) informative cases. Five of 25 (20%) glioblastomas had diffuse p16 immunohistochemical positivity. Significantly, all of these had either CDK4 amplification or RB LOH, suggesting that p16 immunopositivity only occurs in those tumors with alterations of another component in the pathway. Nineteen (76%) cases were uniformly immunonegative for p16, and 12 (48%) had CDKN2A homozygous deletions, but the remaining 7 cases lacked CDKN2A deletions, mutations and promoter methylation. All glioblastomas stained diffusely for cdk4, irrespective of CDK4 gene amplification status. Extensive pRb staining was present in most cases that maintained both RB alleles, and absent in most cases with RB loss, but there were notable discrepancies. Thus, p16 and pRb immunohistochemistry cannot replace molecular genetic analysis of this critical regulatory cascade; instead, the combined results hint at complex regulation of this cell cycle checkpoint. From a practical point of view, although p16 immunonegativity does not necessarily indicate CDKN2A deletion, diffuse positive p16 immunostaining strongly suggests either CDK4 amplification or RB loss and excludes CDKN2A deletion.

Loss of heterozygosity at chromosome segment Xq25-26.1 in advanced human ovarian carcinomas.

To determine whether a tumor suppressor gene of importance to epithelial ovarian cancer resides on the X chromosome, we examined loss of heterozygosity (LOH) in 123 epithelial ovarian cancer cases. In 54 such cases, we examined LOH at 26 loci on the human X chromosome. In eight cases, we examined LOH in 14 loci and in 61 cases we examined LOH in 13 loci. Matched DNA samples from tumors and corresponding normal tissues were analyzed by polymerase chain reaction (PCR) amplification of microsatellite markers. Frequent losses were found in epithelial carcinomas at the Xq25-26.l region, including DXS1206 (34.5% loss in informative cases), DXS1047 (27.7%), HPRT (24.1%), and DXS1062 (33.3%). The minimum overlapping region of LOH was approximately 5 megabases (Mb), flanked by DXS1206 (Xq25) and HPRT (Xq26.1). The methylation status of the remaining allele of the androgen receptor gene in the tumors exhibiting LOH at the Xq25-26.1 region suggested that the loss was exclusively in the inactive X chromosome. We next determined whether a significant relationship exists between Xq LOH and other parameters, including histologic grade and/or clinical stage of the tumors and LOH at TP53. The Xq LOH had a significant association with grade 2 to 3 tumors at stages II to IV. Sixteen of 18 cases that showed Xq LOH revealed LOH at the TP53 locus, and 45% of tumors exhibiting LOH at TP53 showed Xq LOH. These results suggest that there may be a tumor suppressor gene or genes which escape inactivation of the X chromosome at Xq25-26.1, and that the loss of the gene(s) at Xq25-26.1 is frequently accompanied by loss of the TP53 or loss of another gene on chromosome 17. These losses may contribute to the progression from a well-differentiated to a more poorly differentiated state or to metastatic aggressiveness.