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Biomedical subjects

R A Kramer

Publications and source records attributed to R A Kramer.

At least 55 records · Page 3Linked to original sources
PubMed1989

Protection against cisplatin nephrotoxicity by prochlorperazine.

Prochlorperazine (Compazine; PCPZ) is often used to limit cisplatin (CDDP)-induced emesis. However, recent studies in mice have shown that PCPZ protects against renal injury produced by treatment with various nephrotoxicants (e.g., MethylCCNU, mercuric chloride). Because renal toxicity remains a serious limitation to the effective use of CDDP, we conducted the present study to determine whether PCPZ could also protect against CDDP-induced renal injury. PCPZ treatment was shown to ameliorate CDDP-induced renal lesions in both rats and mice at doses and treatment schedules that were comparable with those used for alleviating chemotherapy-induced emesis. A PCPZ dose of 10 mg/kg x 2 offered complete protection against CDDP-induced increases in blood urea nitrogen (BUN) levels in mice, with significant protection occurring at a PCPZ dose as low as 5 mg/kg. Similarly, PCPZ ameliorated CDDP-induced increases in BUN, glucosouria, and enzymuria in F344 rats. PCPZ treatment did not affect the urinary excretion or renal tissue levels of total platinum or the plasma pharmacokinetics of free platinum. However, it did cause a marked reduction in the concentration of total plasma platinum (free platinum + protein-bound platinum). PCPZ was not found to affect the in vivo antitumor activity of CDDP against P388 leukemia. The present study suggests that PCPZ may be of therapeutic benefit when used with CDDP and provides a rational basis for the selection of antiemetic therapy.

Animals↗
PubMed1989

Nephrotoxicity of 5-(N-phenylcarboxamido)-2-thiobarbituric acid in the Fischer 344 rat.

In the present investigation, administration of a single i.p. dose of the anticancer drug merbarone [5-(N-phenylcarboxamido)-2-thiobarbituric acid] produced an acute and reversible decrease in renal function in female but not male Fischer 344 rats. The renal lesion in female rats was biochemically characterized as a decrease in p-aminohippuric acid accumulation by renal slices along with polyuria, glucosuria, proteinuria, and enzymuria. These functional changes were accompanied by histopathologic changes of focal tubular necrosis that was confined to the deep cortex and outer stripe of the outer medulla. The changes in these parameters were dose-dependent and were observed at doses as low as 0.2 x MELD(10) (12 mg/kg). This low merbarone dose increased urinary glucose and protein excretion by 26- and 9-fold, respectively, in the initial 16-h urine collection in female rats. This increase was accompanied by a 2- to 15-fold increase in the excretion of N-acetyl-beta-D-glucosaminidase (NAG), gamma-glutamyl transpeptidase (gamma-GTP), and lactate dehydrogenase (LDH) activities. No significant changes in renal function were observed in male rats apart from mild enzymuria after a high dose of merbarone (36 mg/kg). The drug did not increase urea nitrogen levels in male or female rats, reflecting the focal nature of this tubular lesion. Merbarone produced small elevations in serum transaminase activities [i.e., glutamic-oxalacetic transaminase (GOT), glutamic-pyruvic transaminase (GPT)] at doses that produced marked alterations in renal function in female rats, suggesting only mild hepatotoxicity. The present study establishes the kidney as a possible dose-limiting target organ for merbarone toxicity.

Alanine Transaminase↗
PubMed1989

The effect of glutathione (GSH) depletion in vivo by buthionine sulfoximine (BSO) on the radiosensitization of SR 2508.

The effect of glutathione depletion (GSH) on the efficacy of SR 2508 was evaluated in two murine tumor models with single large doses of radiation or with low doses administered in an accelerated fractionated schedule. To deplete tumor GSH, buthionine sulfoximine (BSO) was administered in the animals drinking water (10 mM) following two i.p. injections of 450 mg/kg. This treatment decreased RIF and MCA tumor GSH concentrations by 95% and 80%, respectively. Mice (C3H/Sed) received BSO for 48-72 hr before the first dose of radiation, and were maintained on BSO drinking water for the duration of the fractionated course of therapy. SR 2508 (200-1000 mg/kg) was injected 45 min prior to each fraction of radiation. Radiation was administered as a single dose of 15 Gy or 20 Gy, for RIF and MCA tumors, respectively. Alternatively, animals received a fractionated course of radiotherapy which consisted of 2.5 Gy/fraction for the RIF, and 3 Gy/fraction for the MCA tumors, b.i.d. for five days (total of 10 fractions). Tumor response with and without BSO, and with and without SR 2508, was determined by regrowth delay. BSO pretreatment increased the efficacy of SR 2508 with single dose radiation in the MCA but not RIF tumor. SR 2508 administered with fractionated radiation produced lower enhancement ratios (SER) than with a single radiation dose. However, BSO significantly enhanced the efficacy of SR 2508 with fractionated radiation. BSO increased the maximum SER for SR 2508 (3 mM/fraction) from 1.2 to 1.4 in the RIF tumor, and from 1.4 to 1.8 in the MCA tumor. BSO also increased the toxicity of SR 2508 by a factor of 2. However, the ability of BSO to increase the efficacy of low doses of sensitizer at clinically relevant doses of radiation suggests that this combined modifier treatment may be of clinical benefit.

Animals↗
PubMed1989

Initial results of a phase I trial of continuous infusion SR 2508 (etanidazole): a radiation therapy oncology group study.

To exploit both the oxygen-mimetic and "pre-incubation" or continuous exposure effects of the 2-nitroimidazole radiosensitizers, we are conducting a Phase I trial of continuous infusion SR 2508 for patients receiving brachytherapy. Following the administration of a loading dose of 2 g/m2, SR 2508 is administered by continuous infusion for 48 hr. Twenty-one patients have completed treatment. The initial total dose was 8 g/m2 with patients currently receiving 15 g/m2. No toxicity has been observed. At the higher doses the steady-state plasma concentrations have been between 50 and 70 micrograms/ml. It is not yet known whether or not hypoxic sensitizers will be of benefit clinically, and if so, when during a course of treatment is the optimal time to use them. Given the lack of toxicity and plasma concentrations achievable with continuous infusion, future studies will be conducted using SR 2508 during both the external beam and brachytherapy aspects of treatment.

Brachytherapy↗
PubMed1988

Role of the glutathione redox cycle in acquired and de novo multidrug resistance.

Drug resistance represents a major obstacle to successful cancer chemotherapy. However, the specific biochemical mechanisms responsible for clinical drug resistance are unknown. In these studies resistance to the antitumor agent adriamycin was found to involve two mechanisms, one that decreased drug accumulation by the P170 mechanism and another that altered the glutathione redox cycle, an important pathway in the detoxification of reactive oxygen. This dual mechanism of drug resistance was demonstrated in cell lines that had acquired the multidrug-resistant phenotype and in human colorectal cancer cells with de novo resistance. These studies support a model of acquired and de novo multidrug resistance that includes alterations in both drug accumulation and the glutathione redox cycle.

Animals↗
PubMed1988

An 11-kDa form of human immunodeficiency virus protease expressed in Escherichia coli is sufficient for enzymatic activity.

In order to define the protease domain of human immunodeficiency virus 1, various regions of the pol open reading frame were cloned and expressed in Escherichia coli. Antiserum directed against the conserved retroviral protease active site was used to identify pol precursor and processed species containing the presumed protease domain. The smallest product that accumulates is about 11 kDa as measured by NaDodSO4/PAGE. This size agrees with that predicted from the presence in this region of two Phe-Pro sequences, which is one of the cleavage sites recognized by HIV protease. DNA encoding only the predicted 11-kDa protein was cloned, bypassing the need for autoprocessing, and the protein was expressed to a high level in E. coli. This form is active as demonstrated by its ability to specifically cleave protease-deficient pol protein in vivo in E. coli. Extracts of E. coli containing the 11-kDa protease also process human immunodeficiency virus gag substrates in vitro. These results demonstrate that the 11-kDa protease is sufficient for enzymatic activity and are consistent with a major role for this form in virus maturation.

Cloning, Molecular↗
PubMed1988

Chemical modifiers of cancer treatment.

Chemical modification is a concept in cancer therapy in which the state of tumor cells or normal tissues is modified such that a therapeutic gain can be achieved using conventional therapeutic modalities. Hypoxic zones targeted as cells within them may be radiation resistant, poorly perfused by chemotherapeutic agents, and possibly drug resistant due to hypoxia-related gene amplification. Nitroimidazoles have gained particular attention as chemical modifiers because they can increase the radiation sensitivity of hypoxic cells, are cytotoxic to hypoxic cells, can increase sensitivity to chemotherapeutic agents, and are useful for imaging hypoxic cells. While both radiosensitization and chemosensitization require hypoxia, the mechanism of the enhancement of each of the modalities is different. The 2-nitroimidazole hypoxic sensitizers SR 2508 and Ro-03-8799, which are less toxic than the prototype misonidazole (Miso), are in clinical trials, and dual function molecules that include a hypoxic sensitizer and alkylating function are being developed. The presence of both acutely and chronically hypoxic cells in animal tumors has been demonstrated by new imaging techniques. Oxygen delivery to tumors is being altered by the use of perfluorocarbons, and agents that alter hemoglobin affinity for oxygen. Compounds that are selectively toxic to hypoxic cells are being developed. Nonhypoxic modifiers are also being investigated. Thiol modification, particularly the alteration of glutathione concentration, has complex effects on the cell's biochemistry, in addition to affecting the competition between oxygen and thiol groups for the restoration and fixation of radiation-induced radicals. WR-2721 is being studied as a means of reducing the normal tissue toxicity of radiation and chemotherapy. Increased thiol concentration may be a mechanism of cross-resistance between certain chemotherapeutic agents and radiation.

Antineoplastic Agents↗
PubMed1987

Chemosensitization of L-phenylalanine mustard by the thiol-modulating agent buthionine sulfoximine.

Glutathione (GSH) plays a crucial role in the protection of normal and tumor tissue against the toxic effects of numerous chemotherapeutic drugs. Therefore, the possible therapeutic benefit of thiol depletion in cancer treatment is dependent upon the relative degree to which tumor or normal tissue is sensitized to the toxic effects of subsequent chemotherapy. To address this issue, the following studies on the chemosensitization of melphalan (L-PAM) by the thiol-depleting agent buthionine sulfoximine (BSO) were conducted in vivo in BDF mice inoculated with L-PAM-resistant murine L1210 leukemia. Different dosing regimens of BSO were found to potentiate L-PAM toxicity in a manner that depended upon the degree of GSH depletion. Multiple i.p. injections of BSO (450 mg/kg every 6 h X 5) were found to reduce GSH concentrations in most tissues by 70-80%, and to decrease the LD50 for L-PAM from 22 to 14 mg/kg. No two organs were found to behave entirely the same with respect to the rate of depletion or recovery of GSH, or to the maximum depletion that could be obtained by BSO. In this regard, the bone marrow was found to be the most resistant tissue to thiol depletion by BSO and was found to tolerate the combination of BSO and therapeutic doses of L-PAM. However, BSO pretreatment markedly inhibited the recovery of the peripheral WBC population at the LD10 dose of L-PAM. Differences also were found in the in vivo metabolism of GSH by L-PAM-sensitive and -resistant murine L1210 leukemia cells. The intracellular concentration of GSH in the resistant cell line was 1.6-fold higher than in the sensitive tumor. Moreover, GSH levels were depleted more rapidly in the resistant tumor relative to the sensitive cell line. A single injection of BSO decreased GSH concentrations in both tumors to equivalent levels (20 nmol/10(7) cells) within 24 h. However, multiple i.p. injections of BSO failed to produce a significant increase in the life-span of L-PAM-treated animals despite a 90% reduction in tumor GSH concentrations (5.5 nmol/10(7) cells). In contrast to the median day survival data, BSO was found to enhance the antitumor activity of L-PAM as determined by an in vivo/in vitro clonogenic assay or by in vivo thymidine incorporation. Using decreased thymidine incorporation as an index of antitumor activity, BSO was found to increase the therapeutic index (LD10/ED50) of L-PAM from 3.6 to 6.5.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

Signal peptide specificity in posttranslational processing of the plant protein phaseolin in Saccharomyces cerevisiae.

We linked the cDNA coding region for the bean storage protein phaseolin to the promoter and regulatory region of the Saccharomyces cerevisiae repressible acid phosphatase gene (PHO5) in multicopy expression plasmids. Yeast transformants containing these plasmids expressed phaseolin at levels up to 3% of the total soluble cellular protein. Phaseolin polypeptides in S. cerevisiae were glycosylated, and their molecular weights suggested that the signal peptide had been processed. We also constructed a series of plasmids in which the phaseolin signal-peptide-coding region was either removed or replaced with increasing amounts of the amino-terminal coding region for acid phosphatase. Phaseolin polypeptides with no signal peptide were not posttranslationally modified in S. cerevisiae. Partial or complete substitution of the phaseolin signal peptide with that from acid phosphatase dramatically inhibited both signal peptide processing and glycosylation, suggesting that some specific feature of the phaseolin signal amino acid sequence was required for these modifications to occur. Larger hybrid proteins that included approximately one-half of the acid phosphatase sequence linked to the amino terminus of the mature phaseolin polypeptide did undergo proteolytic processing and glycosylation. However, these polypeptides were cleaved at several sites that are not normally used in the unaltered acid phosphatase protein.

Acid Phosphatase↗
PubMed1987

Nephrotoxicity of S-(2-chloroethyl)glutathione in the Fischer rat: evidence for gamma-glutamyltranspeptidase-independent uptake by the kidney.

S-(2-chloroethyl)glutathione (CEG; 270 mumol/kg) produced renal lesions that were confined to the proximal tubules of the outer stripe of the outer medulla and were similar to those lesions produced by the cysteine analog S-(2-chloroethyl)cysteine or by the nephrotoxic glutathione (GSH) adduct of 2-bromohydroquinone. These histopathologic changes in the kidney were correlated with alterations in renal function as reflected by dose- and time-dependent elevations in blood urea nitrogen levels as well as by the increased urinary excretion of protein, glucose and lactate dehydrogenase activity. The role of renal GSH metabolism as a mediating factor in the nephrotoxicity of these GSH conjugates was investigated by administering the gamma-glutamyltranspeptidase inhibitor AT-125 [L-(alpha-S,5S)-alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid]. Treatment with AT-125 led to a dose-dependent decrease in renal gamma-glutamyltranspeptidase activity that correlated inversely with increased GSH concentrations in the urine and kidney. Pretreatment with AT-125 ameliorated 2-bromohydroguinone-induced renal toxicity but did not protect against the CEG-induced renal lesion. In fact, pretreatment with AT-125 produced a dose-dependent potentiation of CEG renal toxicity. The CEG-induced renal lesion was dependent on a probenecid-sensitive transport system that was not involved in the toxicity of 2-bromohydroguinone. These studies demonstrate that CEG need not be metabolized by gamma-glutamyltranspeptidase to the corresponding cysteine adduct [S-(2-chloroethyl)cysteine] in order to enter renal tubule cells and ultimately exert its nephrotoxic action.

Animals↗
PubMed1986

In vivo studies on the relationship between hepatic metabolism and the renal toxicity of 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea (MeCCNU).

Administration of a single sc dose of the nephrotoxic anticancer agent 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea (MeCCNU) to rats led to a time-dependent decrease in renal function (i.e., renal slice anion accumulation, renal concentrating ability, and urinary output) which was correlated with the accumulation of carbamylating and alkylating intermediates of 14C-labeled MeCCNU that bound irreversibly to kidney protein. MeCCNU also produced a dose-dependent decrease in glutathione (GSH) preferentially in liver, but not in kidney. Pretreatment with piperonyl butoxide (PIP) decreased the renal toxicity and covalent binding of MeCCNU, and ameliorated the MeCCNU-dependent decrease in liver and kidney GSH. Radioactivity detected in the urine from the PIP-pretreated group was markedly lower than that in the MeCCNU-only group. In contrast, PIP pretreatment increased the accumulation of parent MeCCNU into fatty tissue. Pretreatment with phenobarbital (PB) increased the renal toxicity of MeCCNU. Moreover, PB pretreatment resulted in the increased alkylation of both liver and kidney macromolecules and to an increase in the urinary clearance of ethylene-labeled MeCCNU. In all experiments, modifiers of hepatic biotransformation produced changes in the level of covalent binding by ethylene-labeled MeCCNU (alkylation) which correlated with the degree of toxicity of MeCCNU in the kidney. Additional evidence supporting a role for hepatic biotransformation in the toxicity of MeCCNU was provided by an in vivo/in vitro colony-forming assay which demonstrated the presence of a cytotoxic metabolite in the bile of a MeCCNU-administered rat. These studies suggest that hepatic metabolism contributes significantly to the alkylating activity of MeCCNU in the liver and the kidney, and indicate that a liver-derived metabolite may be responsible for the renal toxicity of MeCCNU.

Animals↗
PubMed1986

HTLV-III gag protein is processed in yeast cells by the virus pol-protease.

The gag-pol gene of HTLV-III (human T-lymphotropic virus), the virus linked to AIDS (acquired immune deficiency syndrome), was expressed in yeast, and processing of the gag precursor into proteins of the same size as those in the virion was observed. Processing of the gag gene in yeast cells mimics the process that naturally occurs in mammalian cells during maturation of virions. Therefore it was possible to perform mutational analysis of the virus genome to localize the gene that codes for the protease function to the amino terminal coding region of the pol gene. Since this region overlaps the gag gene, it is likely that ribosomal frameshifting occurs from gag to pol. Antibodies in all of the AIDS patients' sera tested recognized the yeast synthesized gag proteins, although the sera showed differences in relative reactivity to the individual gag proteins and the precursor. This yeast system should be valuable not only for production of viral proteins for diagnostic or vaccine purposes but also for analysis of the genetics and biochemistry of viral gene functions--parameters that are difficult to study otherwise with this virus.

Acquired Immunodeficiency Syndrome↗
PubMed1986

Comparative nephrotoxicity of 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea (MeCCNU) and chlorozotocin: functional-structural correlations in the Fischer 344 rat.

1-(2-Chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea (MeCCNU) and chlorozotocin (CZ; 2-[3-(2-chloroethyl)-3-nitrosoureido]-D-glucopyranose) are structurally related anticancer agents which differ by virtue of the increased water solubility, and comparatively low carbamylating activity, of CZ relative to MeCCNU. In the present study, a single sc injection of either of these chloroethylnitrosoureas was nephrotoxic to male Fischer 344 rats. However, at equimolar doses, CZ was shown to be a much more potent nephrotoxicant. A lethal 40-mg/kg dose of CZ (127 microM) initially resulted in acute tubular necrosis of the proximal tubules of the cortex, followed later by a necrosis of papillary collecting ducts. In contrast, lethal doses of MeCCNU (100-180 mg/kg; 400-730 microM) produced only minimal proximal tubule injury. A 250-mg/kg (1 mM) dose of MeCCNU resulted in massive papillary necrosis within 7 days, with only limited necrosis to the proximal tubules. Sublethal doses of either drug, resulted in a similar, chronic, progressive nephropathy which was delayed in onset and was characterized by polyuria, enzymuria, a decrease in urine concentrating ability, and in renal slice organic ion accumulation. Alterations in less sensitive indicators of renal toxicity (i.e., proteinuria, glucosuria, and elevated blood urea nitrogen) were observed no earlier than 3 to 7 days after administration of only the highest tested doses of CZ (40 mg/kg) or MeCCNU (250 mg/kg). At sublethal doses, administration of either drug resulted in karyomegaly to the collecting ducts in the renal medulla within 2 to 4 weeks. These studies demonstrate that carbamylation-mediated reactions may not be necessary for nephrotoxicity to develop following administration of this class of antitumor agent.

Animals↗
PubMed1986

Reciprocal regulation of the tandemly duplicated PHO5/PHO3 gene cluster within the acid phosphatase multigene family of Saccharomyces cerevisiae.

We characterized the organization and expression of PHO5 and PHO3, the tightly linked repressible and constitutive acid phosphatase genes of Saccharomyces cerevisiae. The "constitutive" gene, PHO3, is expressed only when PHO5 is not. Altering PHO5 expression, either through promoter deletions or through mutations in trans-acting regulatory genes, showed that PHO5 expression is sufficient to block transcription of PHO3. An active genomic copy of PHO5 was able to block expression of PHO3 from a high-copy-number plasmid, showing that some trans-acting product of PHO5 is involved. This is probably a translation product, since the presence of a nontranslatable PHO5 RNA did not inhibit transcription of PHO3.

Acid Phosphatase↗
PubMed1986

Sequential morphologic analysis of the nephrotoxicity produced in rats by single doses of chlorozotocin.

Chlorozotocin is a chloroethylnitrosourea antitumor agent that is in clinical trial for a variety of human tumors. Renal failure has been a reported side effect of treatment with several of the chloroethylnitrosoureas, including chlorozotocin. To better understand the pathogenesis of this target organ toxicity, we have studied the nephrotoxicity of a single high, intermediate, or low dose of chlorozotocin in male F344 rats. We report here the sequence of histopathologic changes seen over a 1-10-day (high dose) or 1-28-day (intermediate or low dose) period. The single high dose (40 mg/kg, s.c.) produced an acute cortical necrosis involving the proximal tubules, followed by later necrotic changes in the collecting ducts in the inner medulla. Karyomegaly was noted at 10 days in occasional cells of the papillary collecting ducts and urinary epithelium lining the papilla. A single intermediate dose (25 mg/kg, s.c.) caused a similar but less severe injury of later onset. Proximal tubule injury was less severe and more limited. Necrosis of papillary collecting ducts was not seen; however, karyomegaly was pronounced in cells of the collecting ducts in the inner stripe of the outer medulla and inner medulla, and in the urinary epithelium covering the papilla. No discernible histopathology was present following the low dose (12.5 mg/kg, s.c.) of chlorozotocin. The histopathology was correlated with biochemical parameters. Our findings have possible implications for monitoring the severity of nephrotoxic side effects in patients, as well as provide preliminary evidence that this antineoplastic agent may itself cause preneoplastic changes, a finding with important long term implications.

Animals↗
PubMed1985

Differential distribution and covalent binding of two labeled forms of methyl-CCNU in the Fischer 344 rat.

The present study compares the organ distribution and covalent binding of MeCCNU labeled either within the carbamylating [( cyclohexyl-1-14C]MeCCNU; Chx-14C-MeCCNU) or alkylating [( 2-chloroethyl-1,2-14C]MeCCNU; Cle-14C-MeCCNU) region of the compound in an animal model shown to be suitable for studying the nephrotoxicity of the nitrosoureas. Extraction of tissue homogenates with organic solvents of increasing polarity, and subsequent analysis of these extracts by HPLC showed fat to accumulate the highest concentration of parent compound. Kidney accumulated the highest levels of the more polar ether- and methanol-extractable metabolites and/or degradation products of either cyclohexyl-derived or chloroethyl-derived 14C-MeCCNU. Striking differences were apparent in the accumulation, degradation and/or metabolism, and tissue distribution of covalently bound radioactivity for the chloroethyl and cyclohexyl moieties. For example, approximately twice as much cyclohexyl-derived 14C was bound covalently to protein of kidney than to protein of liver or lung. In contrast, approximately twice as much chloroethyl-derived 14C was bound to lung protein than to liver of kidney protein. No radioactivity was bound covalently to tissue DNA following Chx-14C-MeCCNU administration. On the other hand, at 4 h, chloroethyl-derived 14C was irreversibly bound to DNA in the relative amounts of kidney (5.0 nmol/mg), liver (2.7 nmol/mg), and lung (1.25 nmol/mg). These results demonstrate that MeCCNU metabolites and/or degradation products are preferentially accumulated in kidney, a primary target organ for MeCCNU toxicity. Moreover, kidney protein and DNA were subject to extensive carbamylation and alkylation reactions as measured by irreversibly bound cyclohexyl-derived and chloroethyl-derived 14C, respectively. These data suggest that the extent of irreversibly bound drug to tissue macromolecules may be a valid predictor of MeCCNU toxicity. However, the relative toxicological significance of either protein carbamylation or DNA alkylation in mediating MeCCNU-induced nephropathy is not yet understood.

Adipose Tissue↗