HPLC sample pretreatment and preparation.
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Chromatography, High Pressure Liquid↗
Biomedical subjects
L Liebes
Publications and source records attributed to L Liebes.
Phase IB study of low-dose intraperitoneal recombinant interleukin-2 in patients with refractory advanced ovarian cancer: rationale and preliminary report.
The biological activity of recombinant Interleukin-2 (rIL-2) administered intraperitoneally (ip) has not been determined and may differ significantly from the maximum tolerated dose (MTD). In this trial, the pharmacokinetics, toxicity, and biologic activity of a single ip dose were studied initially followed a week later by a 5-day ip rIL-2 given for 2 weeks every 28 days. Planned dose escalation was from 2 x 10(3) to 2 x 10(7) U given in 2 liters of D5W. Drug was obtained from the NCI and was administered through an ip port. Four patients received 1 U/ml and four patients received 10 U/ml. Preliminary data demonstrate an increase in the peritoneal fluid mononuclear cell count. Mononuclear cell phenotyping tested in the first eight patients showed a modest increase in Leu 2a+, Leu 15- cells, corresponding to CTL. A similar increase in Leu 19+ cells was also demonstrated (NK cells). Soluble IL-2 receptor was elevated in peritoneal fluid. Cytotoxicity against K562 and Daudi cell lines was not observed at the first two dose levels. Toxicity of treatment was minimal and related to abdominal distention. No objective responses were seen but in one patient we documented a reduction in serum CA-125 levels. The observed biologic response and lack of toxicity is promising and justifies further exploration of this immune-modulating approach.
Antigens, Differentiation↗
A specific high-performance liquid chromatography assay for dehydroascorbic acid shows an increased content in CLL lymphocytes.
A method for the assay of dehydroascorbic acid using high-performance liquid chromatography with uv detection is described. The dehydroascorbic acid is separated from ascorbic acid and reduced with dithiothreitol, and is then quantitated as ascorbic acid following rechromatography. Since as little as 22 pmol can be detected, sensitivity is at least 40-fold greater than that of other currently available procedures. This method was used to measure the level of dehydroascorbic acid in normal and chronic lymphocytic leukemia lymphocytes. A significantly higher concentration of dehydroascorbic acid was found in leukemic (21.80 +/- 3.55 nmol/10(8) cells, mean +/- SE) than in normal lymphocytes (9.32 +/- 1.15 nmol/10(8) cells) (P less than 0.03). Analysis of extracts from normal B cell lymphocytes revealed comparable dehydroascorbic acid levels to unfractionated lymphocytes, indicating that the elevated level in chronic lymphocytic leukemia was not simply a reflection of the increased percentage of B lymphocytes in this disorder. These studies illustrate that the technique can be used to measure the dehydroascorbic acid content from sources where only scanty material is available or low levels are found.
Ascorbic Acid↗
Increased ascorbic acid content in chronic lymphocytic leukemia B lymphocytes.
Human lymphocyte extracts analyzed by high-performance liquid chromatography reveal a major UV-absorbing peak that was shown to be ascorbic acid by spectral, chemical, and enzymatic criteria. Because this peak appeared very prominent in the elution profile of chronic lymphocytic leukemia (CLL) lymphocyte extracts, we measured the ascorbic acid content in lymphocytes from the blood of normal subjects and untreated patients with chronic lymphocytic leukemia. A significantly higher concentration of 111 +/- 15.3 nmol per 10(8) cells (mean +/- SEM) was found in CLL lymphocytes than in normal blood lymphocytes, which contained 42.2 +/- 3.3 nmol per 10(8) cells. Selective enrichment with B and T cells showed that this difference was limited to the chronic lymphocytic leukemia B cell, which had a 5- to 15-fold higher content of ascorbic acid than normal B cells had. In contrast, the ascorbic acid level was similar in normal and CLL T cells. The very high ascorbic acid content provides the chronic lymphocytic leukemia B cell with a reducing substance that could react with oxidants or free radicals.
Ascorbic Acid↗
The anomalous capping behavior of chronic lymphocytic leukemia lymphocytes: studies with an antilymphocyte antiserum.
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Antilymphocyte Serum↗
Electrophoretic mobilities of RNA tumor viruses. Studies by Doppler-shifted light scattering spectroscopy.
We have used laser beat frequency light scattering spectroscopy to measure, at several pH values, the electrophoretic mobilities of purified avian myeloblastosis (AMV), murine leukemia (MuLV), murine mammary tumor (MuMTV), and feline leukemia (FeLV) viruses. The mobilities of these viruses are similar at pH greater than or equal to7 (-2.7 to -3.2 X 10(-4) (cm/sec)/(V/cm). The isoelectric points of MuLV and AMV are apparently less than pH 3, whereas for FeLV the data could be interpreted to indicate an isoelectric point between 3 and 5. Using a Debye-Hückel model to describe the interaction between electrolytes and virus, we show that our values for the mobility of MuMTV, obtained in ionic strength 0.005, are consistent with the values of Sarkar et al. ((1973), Cancer Res. 33, 2283), obtained in ionic strength of 0.10. This model is then used to calculate surface charge densities. In terms of the density of charged groups, the RNA tumor virus envelope is not very different from the erythrocyte membrane.
Avian Leukosis Virus↗
Hydrodynamic diameters of RNA tumor viruses. Studies by laser beat frequency light scattering spectroscopy of avian myeloblastosis and Rauscher murine leukemia viruses.
The diffusion constants of avian myeloblastosis virus (AMV) and murine leukemia virus (MuLV) (Rauscher) suspensions in buffer and in 30% sucrose were determined by laser beat frequency light scattering spectroscopy at a series of temperatures ranging rom 5 to 25 degrees. By the use of the Stokes-Einstein equation, the following hydrodynamic diameters are calculated at 20 degrees: MuLV, 154 plus or minus 3 nm in sucrose and 145 plus or minus 7 nm in buffer; AMV, 144 plus or minus 3 nm in sucrose and 138 plus or minus 4 nm in buffer. While the diameters measured in buffer were temperature independent, the diameters measured in sucrose decreased by about 20% as the temperature was raised from 5 to 25 degrees. The concentration of virus particles in the suspensions ranged from 10 7 to 10 9 particles/ml. The absolute particle concentrations are estimated within plus or minus 30% by determining the dilution needed to reach a concentration sufficiently low that the particle number fluctuation contribution was comparable to that of the interference scattering. Particle weights of 3.9 x 10 8 daltons for MuLV and 4.0 x 10 8 daltons for AMV were calculated from the diffusion constants and from our own experimentally determined sedimentation coefficients. From these particle weights and the hydrodynamic diameters of the viruses, we calculated the per cent of the hydrodynamic volume of the viruses which could be freely penetrated by water, viz., 57% for AMV and 69% for MuLV.
Avian Leukosis Virus↗
Nucleoside transport by perfused human placenta.
Nucleoside transport and metabolism by human placenta was studied using the dual perfusion technique. With [3H] thymidine added to the maternal perfusate and neither perfusate recirculated (steady-state studies) around 40 per cent of the thymidine in the maternal outflow and 50 per cent of the transferred thymidine was degraded. In similar studies with adenosine, over 95 per cent of the nucleoside was degraded. Even with the bolus technique which sharply limits the duration of contact with the placenta, degradation of adenosine was over 95 per cent. Uptake as calculated by the dual-tracer method ([3H] adenosine/[14C] L-glucose) was equally rapid from the maternal and fetal perfusates, was saturable and inhibited by nitrosobenzylthioinosine, consistent with the facilitated diffusion system for nucleosides. Thymidine was taken up at one-third the rate of adenosine. Thymidine in large excess (500 microM) reduced adenosine uptake suggesting a common transporter. Zidovudine, a thymidine analogue used for the treatment of AIDS in which the ribose is modified at the 2' 3' position, did not compete with adenosine for uptake consistent with previous reports that zidovudine is transferred across the placenta by simple diffusion.
Adenosine↗
Role of complement activation in hypersensitivity reactions to doxil and hynic PEG liposomes: experimental and clinical studies.
Pegylated liposomal doxorubicin (Doxil) and 99mTc-HYNIC PEG liposomes (HPL) were reported earlier to cause hypersensitivity reactions (HSRs) in a substantial percentage of patients treated i.v. with these formulations. Here we report that (1) Doxil, HPL, pegylated phosphatidylethanolamine (PEG-PE)-containing empty liposomes matched with Doxil and HPL in size and lipid composition, and phosphatidylglycerol (PG)-containing negatively charged vesicles were potent C activators in human serum in vitro, whereas small neutral liposomes caused no C activation. (2) Doxil and other size-matched PEG-PE and/or PG-containing liposomes also caused massive cardiopulmonary distress with anaphylactoid shock in pigs via C activation, whereas equivalent neutral liposomes caused no hemodynamic changes. (3) A clinical study showed more frequent and greater C activation in patients displaying HSR than in non-reactive patients. These data suggest that liposome-induced HSRs in susceptible individuals may be due to C activation, which, in turn, is due to the presence of negatively charged PEG-PE in these vesicles.
Animals↗
Analysis of the antimetastatic effects of synthetic muramyl tripeptide (CGP 19835A) encapsulated in liposomes in combination with other immunomodulatory agents and chemotherapeutic drugs.
The synthetic molecule muramyl tripeptide (CGP 19835A) encapsulated in liposomes is effective in increasing the survival of mice with spontaneous experimental lung metastases induced by the RENCA renal adenocarcinoma and B16 melanoma tumor models. The present study was aimed at extending the effects of CGP 19835A to another highly metastatic carcinoma model and at evaluating the efficacy of combination therapy with standard cytotoxic agents and other immunomodulators. C57BL/6 mice received whole tumor implants of PancO2, a spontaneously metastasizing pancreatic adenocarcinoma, subcutaneously in the hind leg. Therapeutic effects were measured by increased survival which is a direct function of the growth of spontaneous lung metastases in this system. No therapeutic efficacy was observed with CGP 19835A alone or in combination with any of a series of cytotoxic or biological agents, including cis-platinurn (cis-Pt), mitomycin C (MMC), adriamycin (ADR), cyclophosphamide (CP), interferon gamma (IFN gamma), and interleukin 2 (IL-2). In accord with previous studies, when the B16-F10 melanoma was used as an experimental metastatic tumor model, CGP 19835A, alone and in combination with CP, significantly reduced the number of pulmonary metastases. Cis-Pt, however, partially negated the effects of CGP 19835A when a combination of the two agents was used. The results indicate that CGP 19835A is an effective therapeutic agent in some models of spontaneous or experimental lung metastases, but not others, and that the effects of CGP 19835A are not enhanced by the accompanying cytotoxic drugs tested here.
Acetylmuramyl-Alanyl-Isoglutamine↗