Neuroprotective actions of 2-amino-N-(1,2-diphenylethyl)-acetamide hydrochloride (FPL 13950) in animal models of hypoxia and global ischemia.

2-Amino-N-(1,2-diphenylethyl)-acetamide hydrochloride (FPL 13950) has been demonstrated to have good anticonvulsant efficacy and a relative lack of acute side effects in rodents. Similar in structure to remacemide hydrochloride, it was also shown to possess weak potency as an uncompetitive antagonist of N-methyl-D-aspartic acid (NMDA) receptors. In our study FPL 13950 was profiled preclinically as a potential neuroprotective agent with respect to lengthening the survival time of rodents exposed to hypoxia, as well as for ability to protect the vulnerable CA1 pyramidal neurons of the rat and dog from the consequences of global ischemia. Under conditions of hypoxia, pretreatment of rodents with FPL 13950 resulted in an extension in the time to loss of the righting reflex (rats) and mortality (rats and mice). This occurred whether the rats were maintained at ambient body temperature or made hyperthermic. When administered after 30 min of four-vessel occlusion global ischemia for periods of either 7 (b.i.d.) or 14 days (s.i.d.), FPL 13950 exhibited protection of the vulnerable CA1 hippocampal neurons in rat. In the 14-day treatment study the CA3 neurons were evaluated and FPL 13950 was found to prevent the lesser degree of ischemic-induced damage to this hippocampal region. In ischemic rats treated with FPL 13950 for 7 days, electrophysiological responses of CA1 neurons (orthodromic and antidromic population spikes, in vitro) were also preserved after FPL 13950 treatment. FPL 13950 was administered i.v. at 30 min after 8 min of clamping the ascending aorta in dogs, followed by a b.i.d./s.i.d. dosing regimen for 1 wk. Neuronal damage to CA1 was considerable in the saline-treated ischemic animals but significantly protected in dogs receiving FPL 13950. FPL 13950 continues to serve as a potential backup candidate for remacemide HCl which is currently in clinical trials for patients with stroke and epilepsy.

Fluorescent in situ hybridization for assessing the proportion of cells with trisomy 4 in a patient with acute non-lymphoblastic leukemia.

A case of acute non-lymphoblastic leukemia is described in which fluorescent in situ hybridization (FISH) helped to resolve initially conflicting conventional cytogenetic results. Identification and assessment of the proportion of cells exhibiting trisomy of chromosome 4 in the patient's bone marrow were made using a probe which hybridizes to the centromeric region of chromosome 4. These FISH results were consistent with our retrospective GTG-banded analysis but differed from another study conducted elsewhere. Factors such as culture conditions and duration of culture which may have influenced the proportion of leukemic to nonleukemic cells are discussed. Fluorescent in situ hybridization is a powerful adjunct to conventional cytogenetic analysis and may prove to be a useful tool for monitoring minimal residual disease in this patient.

Anticonvulsant efficacy/safety properties of 2-amino-N-(1,2-diphenylethyl)acetamide hydrochloride.

2-Amino-N-(1,2-diphenylethyl)-acetamide-hydrochloride (FPL 13950) was profiled preclinically in rodents for efficacy against convulsions, as well as for acute safety/behavioral observations. FPL 13950 exhibited good oral efficacy and duration of action with respect to prevention of seizures elicited by maximal electroshock-shock in both rats and mice. Tolerance to protection against maximal electroshock and hexobarbital-induced sleep-time was not evident after subchronic drug administration. FPL 13950 also prevented convulsions/mortality in mice after i.v. dosing with N-methyl-D, L-aspartate, however, it was ineffective against other types of chemically induced convulsions, as well as bicorneal kindling. High oral doses produced neural impairment in both mice and rats and hyperactivity in rats. Sequential administration of yet higher doses elicited tonic/clonic convulsions culminating in death. During i.v. infusion of metrazol in mice, high i.p. doses of FPL 13950 shortened the latency to first twitch and clonus. No increase in the startle response or phencyclidine-like behavior was evident after oral dosing in rats.

In vitro muscarinic activity of spiromuscarones and related analogs.

The cholinergic hypothesis of Alzheimer's disease suggests that cholinergic agonists may have therapeutic potential for treating the attendant memory deficits of the disease. As part of a program aimed at preparing metabolically stable, nonquaternary analogs of muscarone, 1-oxa-2,8-dimethyl-8-azaspiro[4.5]decan-3-one, 2a, and related analogs have been synthesized and their in vitro muscarinic activity evaluated. The synthetic strategy in the formation of the 1-spiro[4.5]decan-3-one ring system of 2a involved cyclization of the diol 4 in the presence of Nafion-Hg. The spiromuscarone 2a was found to displace [3H]Oxo-M binding with a Ki value of 7 nM. Affinities of the oxime and hydrazone analogs of 2a were lower than 2a. The compounds in these series were partial muscarinic agonists as demonstrated by stimulation of phosphatidyl inositol hydrolysis assay, with 2a showing the highest intrinsic intrinsic activity (60% as compared with carbachol). The results from this study indicate that an exo double bond at the C-3 position is essential for the receptor binding.

Biological profile of the metabolites and potential metabolites of the anticonvulsant remacemide.

Remacemide hydrochloride ((+/-)-2-amino-N-(1-methyl-1,2-diphenylethyl)- acetamide hydrochloride or FPL 1292AA) is a novel compound undergoing clinical trials for patients with generalized tonic/clonic and complex partial epilepsy. Remacemide exhibits efficacy against maximal electroconvulsive shock (MES) in rodents and seizures elicited by N-methyl-D,L-aspartate (NMDLA) in mice. Using rat synaptic membrane fractions, remacemide was shown to possess relatively weak noncompetitive binding to the ionic channel site of the NMDA (N-methyl-D-aspartic acid) receptor complex. With the hypothesis that activity against NMDLA-elicited seizures might be reflected by transformation to a more active metabolic species, the aim of the present study was to evaluate potential pharmacological effects of the 9 identified metabolites of remacemide which were all found in human and dog urine. Moreover, specific entities were recognized in plasma (including the rat's), as well as dog and rat cerebrospinal fluid. Five putative metabolites were also examined. A major route of metabolic transformation of remacemide in rats yields the formation of a pharmacologically active more potent desglycine derivative, namely FPL 12495 (+/-). Potency over the parent compound is revealed in the MES test in mice and rats, the NMDA-induced convulsions/mortality test in mice, and especially involving in vitro displacement of MK801 binding to the channel subsite of the NMDA receptor. The S isomer (FPL 12859) of this desglycinate is even more potent, while the R isomer is less potent than the corresponding racemate. Unlike the non-competitive NMDA antagonist, MK801, these desglycinates did not prevent kindled seizures. Three other identified metabolites show efficacy in the mouse and rat in vivo tests, namely the N-hydroxy-desglycinate (FPL 15053) and the p-hydroxy-desglycinates (FPL 14331 and FPL 14465). FPL 15053 exhibited modest activity in all tests. The only in vivo activity exhibited by the 2 p-hydroxy-desglycinates was evidenced in the MES test following i.p. and i.v. dosing. However, FPL 14331 was active in the MK801 binding assay. An oxoacetate metabolite, PFL 15455, failed to demonstrate any biological activity. Of potential metabolites tested 2 beta-hydroxy-desglycinates (FPL 14991 and FPL 14981) displayed modest activity in the MES test, however, only FPL 14981 prevented NMDLA-induced convulsions/mortality in mice and was 2-fold more active regarding MK801 binding. The hydroxy-methyl derivative of remacemide (FPL 13592) and its desglycinate (FPL 15112) prevented MES-induced convulsions only after i.v. administration; only the desglycine derivative displaced MK801 binding.(ABSTRACT TRUNCATED AT 400 WORDS)

Preclinical profile of the anticonvulsant remacemide and its enantiomers in the rat.

Studies conducted by Fisons Pharmaceuticals and the Antiepileptic Drug Development Program (ADD Program) of the Epilepsy Branch (NINDS, NIH) revealed that 'remacemide' (FPL 12924, formerly PR 934-423) was effective orally in the prevention of maximal electroshock seizures (MES) in rats. In this context (-)stereoisomer (FPL 14145) was of equal potency to the racemate (remacemide), while the (+)stereoisomer (FPL 14144) was 54% less potent. With respect to neurotoxicity, remacemide and its enantiomers possessed more favorable therapeutic indices than phenobarbital and valproate and less favorable indices than phenytoin and carbamazepine. The duration of protection of rats in the MES test at the ED50 or 3 x ED50 of remacemide and the (+)isomer was better or on par with the best reference compounds, phenytoin and phenobarbital. After subchronic administration of either the ED50 or the ED97 of remacemide, no tolerance developed in the hexobarbital sleep test, however, the activities of 3 hepatic microsomal enzymes were elevated. In naive rats high doses of remacemide or its (-)isomer and low doses of phenobarbital caused an increase in spontaneous motor activity. Alternatively, motor activity was depressed subsequent to high doses of phenobarbital and phenytoin. Remacemide was inactive against pentylenetetrazol and 'kindling' seizures. It was without effect in 5 electrophysiological tests (evoked responses, recurrent inhibition, long-term potentiation, penicillin-induced discharge rate and veratridine-induced depolarization) employing the in vitro hippocampal slice technique. Moreover, remacemide failed to demonstrate potent binding in vitro to neuronal L-glutamate, gamma-amino-butyrate A, adenosine A1, benzodiazepine, N-methyl-D-aspartate (strychnine-insensitive glycine and ion channel subsites) or muscarinic receptors. In conclusion, remacemide specifically prevents seizures elicited by MES, an action predicting utility in patients with generalized tonic/clonic convulsions.

Preclinical profile of stereoisomers of the anticonvulsant remacemide in mice.

Stereoisomers of remacemide (racemate form) were compared for anticonvulsant efficacy and safety in mice. In the maximal electroshock seizure (MES) test for oral efficacy, the (-) stereoisomer, FPL 14145, was more potent than the racemate or the (+) stereoisomer, FPL 14144. Respective ED50 values (expressed as mg/kg) were: remacemide, 58; FPL 14145, 45; FPL 14144, 79. In 2 of 3 tests for neural impairment, FPL 14145 yielded significantly better therapeutic indices (toxic dose 50/ED50) than the racemate. The margin of safety (estimated median lethal dose ED50) was more favorable for FPL 14144: remacemide, 15.1; FPL 14144, 18.9; FPL 14145, 15.7. The duration of protection against MES indicated the stereoisomers were longer acting than the racemate. After intravenous administration the order of potency against MES was similar: FPL 14145 greater than remacemide greater than FPL 14144. Following daily administration of the oral ED98 for 4 days, with a dose response curve run on day 5, the MES ED50 values for all compounds were increased. The test indicates tolerance. In the pentylenetetrazol infusion test the racemate and FPL 14144 demonstrated more proconvulsant properties than FPL 14145. Intraperitoneal administration of 50 mg/kg or more produced changes in behavior with all compounds. At higher doses the racemate and FPL 14145 elicited more severe symptoms with death at 200 mg/kg.

Myelomatous pleural effusion: report of an unusual occurrence.

Pleural effusion complicating the course of multiple myeloma is common. Myelomatous pleural effusion, in contrast, is distinctly rare. Searching the Medline data base, a total of only 35 previously reported cases could be located. A recent case we cared for prompted us to review the literature in this area. The incidence of myelomatous effusion, possible mechanisms of its formation, and implications for therapy and prognosis are discussed.

Preclinical profile of remacemide: a novel anticonvulsant effective against maximal electroshock seizures in mice.

Anticonvulsant tests in mice revealed specific, potent actions of remacemide for protection of mice against maximal electroshock seizures (MES). Comparisons of oral efficacy to reference compounds yielded the following ED50 values (expressed as mg/kg): remacemide = 33, phenytoin = 11, phenobarbital = 20, carbamazepine = 13 and valproate = 631. The duration for protection by remacemide was longer than carbamazepine or valproate, but shorter than phenytoin or phenobarbital. In neural impairment tests (inverted screen or rotorod) to determine the oral toxic dose 50 (TD50) the following therapeutic indices (TD50/ED50) were obtained: (1) inverted screen--remacemide = 17.6, phenytoin = 57.4, phenobarbital = 5.1, carbamazepine = 10.2, and valproate = greater than 3; and (2) rotorod--remacemide = 5.6, phenytoin = 9.6, phenobarbital 4.8, and valproate = 1.9. Remacemide was devoid of sedative actions and possessed a favorable 28.1 margin of safety value (median estimated lethal dose/ED50 for MES). An intermediate potency against either audiogenic- or N-methyl-D-aspartate-induced seizures was exhibited by remacemide. Tolerance to MES was not apparent after 5 days of oral daily dosing of remacemide. Remacemide was inactive in vitro against gamma-aminobutyrate or benzodiazepine receptors and adenosine uptake mechanisms. Therapeutic utility for generalized tonic/clonic seizures is predicted for remacemide.

The treatment of progressive non-Hodgkin's lymphoma with intensive chemoradiotherapy and autologous marrow transplantation.

Intensive chemoradiotherapy, with or without additional local radiotherapy, and unpurged autologous marrow transplantation was given to 68 patients with progressive non-Hodgkin's lymphoma. Responses were attained in 44 patients (65%, 95% confidence intervals [CI], 52% to 76%), including 37 who achieved complete responses. Fifteen patients (22%, 95% C.I. 13% to 34%) remain free of disease (including 11 continuously) at a median of 5.3 (range 3.1 to 9.1) years later. Higher Karnofsky scores (P less than .01, Mann-Whitney U test) and the absence of a history of prior radiotherapy (P = .02, chi 2 test) were associated with achievement of complete plus partial responses. Higher Karnofsky scores (P less than .01, Mann-Whitney U test) and less resistant disease status at transplantation (P = .04, chi 2 test) were significant when calculations were limited to complete responses. Karnofsky scores were also associated with the probability of freedom from progression (P = .02, log-rank) for responding patients. Also, Karnofsky scores and the absence of prior radiotherapy (P less than .01 and P = .01, respectively, log-rank) were associated with improved survival. Progressive lymphoma was the chief cause of failure; progression usually occurred less than 6 months after transplantation, most often at the sites of active disease before the transplant. However, five patients (including four with high-grade non-Hodgkin's lymphoma) suffered hematogenous patterns of relapse; four of these five patients had no prior history of marrow involvement. Other causes of mortality included interstitial pneumonitis, sepsis, hemorrhage and renal failure. Intensive chemoradiotherapy and autologous marrow transplantation produces durable remissions in some patients with progressive non-Hodgkin's lymphoma. Since such therapy is more effective when given to patients with signs of less advanced disease, earlier treatment would be the simplest way to produce improved results. However, improved conditioning regimens will also be needed, and measures to reduce occult lymphoma stem cell contamination with the autograft may also be required to increase the likelihood of cure in some patients.

Immunoblastic lymphoma of T-cell type in a chronically immunosuppressed renal transplant recipient.

A case of non-Hodgkin's lymphoma of T-cell type occurring in a renal transplant recipient is described. This lymphoma was classified as large cell, immunoblastic type and presented as a mediastinal mass, although it was demonstrated to be disseminated at autopsy two weeks later. Lymphoma cells expressed the immunologic profile of a mature, activated cytotoxic/suppressor T-lymphocyte. T-cell receptor gene rearrangement and karyotypic studies confirmed a clonal T-cell genotype. Molecular studies failed to demonstrate any association with Epstein-Barr virus or human T-cell lymphotropic virus type I or type II. Pathogenesis of lymphomas in this clinical setting is discussed.

Treatment of progressive Hodgkin's disease with intensive chemoradiotherapy and autologous bone marrow transplantation.

Twenty-six patients with progressive Hodgkin's disease after conventional chemotherapy received intensive chemoradiotherapy and autologous bone marrow transplantation (ABMT); 19 also received additional involved-field radiotherapy. Twenty-one patients [81%, 95% confidence intervals (CI) 61% to 94%] attained complete (n = 18) or partial responses. Ten patients (38%, 95% CI 20% to 59%) are disease-free a median of 4.5 years later (range 3.5 to 7.0 years), including seven patients with continuous complete responses. The likelihood of overall response was not significantly influenced by any clinical or treatment variable examined. However, there was a trend favoring patients with higher Karnofsky scores, and higher scores were associated with attainment of complete responses (P = .06 and P = .02, respectively, Mann-Whitney U test). Both higher Karnofsky scores and shorter durations of disease before transplantation were associated with improved survival in a stepwise Cox multivariate analysis. The chief cause of failure was progression at sites previously involved with Hodgkin's disease. No patient relapsed in the marrow, and two of three patients with a history of marrow involvement with Hodgkin's disease achieved durable complete responses after transplantation. These data suggest that inadequate pretransplant conditioning, and not the reinoculation of occult tumor cells in the autologous marrow, caused most relapses. Fatal treatment-related toxicity occurred in six patients. Three patients died of idiopathic interstitial pneumonitis; each had previously received local mediastinal irradiation before intensive chemoradiotherapy. Intensive chemoradiotherapy and ABMT produces durable responses in some patients with Hodgkin's disease incurable with conventional therapy. Use of such therapies at the first sign of failure with conventional chemotherapy and development of more effective conditioning regimens should further improve results.

Loss of heterozygosity for the short arm of chromosome 1 in human neuroblastomas: correlation with N-myc amplification.

Partial monosomy of the short arm of chromosome 1 is the most consistent cytogenetic abnormality found in human neuroblastomas, but its overall frequency and significance are unclear. Using a panel of chromosome-1-specific DNA probes that identify restriction fragment length polymorphisms, we demonstrate that 13 of 47 human neuroblastomas (28%) have somatic loss of heterozygosity (LOH) at one or more loci on the distal short arm of chromosome 1. the chromosomal region that shows LOH most consistently is between 1p36.1 and 1p36.3; loss of a gene or genes in this region may be critical for the development or progression of neuroblastomas. The region of LOH in human neuroblastoma may resemble that described for pheochromocytoma, medullary thyroid carcinoma, and melanoma, which are also tumors of neural-crest origin. Although LOH for distal chromosome 1p can occur in early stages of neuroblastoma, the loss usually occurs in tumors of advanced clinical stages. LOH for the short arm of chromosome 1 correlates significantly with N-myc amplification, suggesting that these two genetic events are related. Indeed, these two lesions appear to characterize a genetically distinct subset of particularly aggressive neuroblastomas.

Cation transport in lymphoblastoid cell lines established from bipolar manic-depressive patients.

Lymphoblastoid cell lines established from patients suffering from bipolar manic depression have been used to study the possible involvement of cation transport in the aetiology of this illness. No significant difference was found in the K+ fluxes mediated by the ouabain-sensitive sodium pump, the diuretic-sensitive cotransport system and the passive leak pathway of cell lines established from either control or bipolar subjects. The mean value for the specific binding of 3H-ouabain (sodium pump site number) was significantly higher in the bipolar group (approximately 30%) than in the control group.

Thermostable DNA polymerase chain amplification of t(14;18) chromosome breakpoints and detection of minimal residual disease.

Achieving the capacity to detect minimal numbers of neoplastic cells is a major cancer diagnostic challenge. Chromosomal translocations such as the t(14;18)(q32;q21) found in follicular and some nonfollicular lymphomas provide a tumor-specific molecular marker. The 14;18 breakpoints are focused at one of six immunoglobulin heavy chain joining (JH) regions on chromosome 14 and a small major breakpoint region (MBR) of the BCL2 gene on chromosome 18. We utilized universal oligonucleotide primers of a region 5' to the BCL2 MBR and at the 3' end of JH segments to initiate a DNA polymerase chain reaction that amplified these BCL2-JH junctures. Use of thermostable DNA polymerase enabled annealing and synthesis steps at temperatures approaching the melting point of the primers, providing a sensitive and specific assay capable of detecting 1 lymphoma cell in 10(6) normal cells. This technique identified the subclinical presence of leukemic cells in all seven patients examined, including two in clinical remission. It also assessed the effectiveness of protocols designed to purge malignant cells from marrow. Moreover, this approach enabled the rapid DNA sequencing of chromosomal breakpoints without their molecular cloning. This assay markedly refines the capacity to detect minimal residual disease and should improve the ability to determine the stage of disease, stratify treatment, and evaluate therapy.

A lymphoproliferative disorder caused by human T-lymphotropic virus type I. Demonstration of a continuum between acute and chronic adult T-cell leukemia/lymphoma.

A 35-year-old black man is described who had a human T-lymphotropic virus type I (HTLV-I) infection while living in a non-endemic region. A lymphoproliferative disorder developed that might be considered as a transition stage between acute and chronic adult T-cell leukemia/lymphoma. This suggests that HTLV-I-induced neoplasias represent a continuous disease spectrum.

6-Methyl-6-azabicyclo[3.2.1]octan-3 alpha-ol 2,2-diphenylpropionate (azaprophen), a highly potent antimuscarinic agent.

The synthesis and antimuscarinic properties of 6-methyl-6-azabicyclo[3.2.1]octan-3 alpha-ol 2,2-diphenylpropionate (1, azaprophen) are described. Azaprophen is 50 times more potent than atropine as an antimuscarinic agent as measured by the inhibition of acetylcholine-induced contraction of guinea pig ileum and is more than 1000 times better than atropine in its ability to block alpha-amylase release from pancreatic acini cells induced by carbachol. In addition, azaprophen is 27 times more potent than atropine as an inhibitor of binding of [N-methyl-3H]scopolamine to muscarinic receptors, with human IMR-30 neuroblastoma cells. The potencies of azaprophen and atropine in altering operant behavior were similar. The structural features of 1 are compared to the standard anticholinergic drugs atropine and quinuclidinyl benzilate by using energy calculations and molecular modelling studies. A modification of the pharmacophore model hypothesis for cholinergic agents is suggested.

A morphologic study of childhood lymphoma of the lymphoblastic type. The pediatric Oncology Group experience.

For this study 227 non-Hodgkin's lymphomas, registered through the Pediatric Oncology Group clinical studies between 1976 and 1982, were morphologically subclassified into major histologic types and subtypes, and their histopathologic and clinical features were compared. These lymphomas were distributed primarily into only three of the recognized major histologic types: lymphoblastic (LB), 106 (47%); undifferentiated (DU), 49 (21%); and diffuse histiocytic (DH), 72 (32%). These patient groups were found to differ in several ways: the LB lymphomas contained most of the patients under two years of age; the LB lymphomas tended to present in higher clinical stages; the LB lymphomas tended to involve lymph node groups and the bone marrow more often than did the DU and DH lymphomas; and the DU lymphomas had a greater tendency for gastrointestinal tract and other major organ system involvement. The complete remission rate of 96%, for the LB lymphomas was better than for either the DU or the DH lymphomas. The disease-free survival of the LB lymphomas was significantly better than the DU group, but not the DH group. The LB were histologically divisible into three subtypes: convoluted (C), nonconvoluted (NC), and large cell variant (LCV). The C and NC subtypes preferentially involved the mediastinum and peripheral lymph nodes initially, while the LCV tended to involve the abdomen. However, none of the subtypes differed in clinical stage. The complete remission, and the disease-free survival rates between these subtypes were not statistically different.