Malignant progenitors from patients with acute myelogenous leukemia are sensitive to a diphtheria toxin-granulocyte-macrophage colony-stimulating factor fusion protein

We have previously demonstrated that human granulocyte-macrophage colony-stimulating factor (GM-CSF) fused to a truncated diphtheria toxin (DT388-GMCSF) kills acute myelogenous leukemia (AML) cell lines bearing the GM-CSF receptor. We now report that exposure of malignant cells from 50 different patients with AML for 48 hours in culture to DT388-GMCSF reduces by a median of 1.6 logs (range, 0 to 3.7 logs) the number of leukemic cells capable of forming colonies in semisolid media (leukemic colony-forming cells [CFU-L]) with a median IC50 of 3 x 10(-12) mol/L (range, 5 to >4,000 x 10(-12) mol/L). Furthermore, the cell kill is dependent on the presence of high-affinity GM-CSF receptors on leukemic blasts, because CFU-L from 27 of 28 AML samples expressing > or = 35 GM-CSF receptors per cell were inhibited by the toxin, whereas the colony growth from all 4 leukemic samples (2 AML, 1 acute lymphoblastic leukemia [ALL], and 1 prolymphocytic leukemia [PLL]) that had less than 35 receptors per cell was unaffected by the drug. Sensitivity of CFU-L to DT388-GMCSF was seen regardless of the clinical responsiveness of the patient's leukemia to standard chemotherapy agents. In contrast, clonogenic cells from normal bone marrow formed colonies at near control numbers after exposure to much higher toxin concentrations (4 x 10(-9) mol/L) than those required to kill CFU-L from most patients. Thus, leukemic progenitors isolated directly from the peripheral blood of most AML patients show the same sensitivity to DT388-GMCSF as previously demonstrated for AML cell lines. Under the same conditions of exposure, normal hematopoietic progenitors are relatively unaffected by DT388-GMCSF, suggesting its potential as a therapeutic agent in AML.     

Synthesis and accumulation of a receptor regulatory protein associated with lipid droplet accumulation in 3T3-L1 cells

Synthesis and accumulation of the recently identified prostaglandin F2alpha receptor regulatory protein (FPRP) was found to correlate closely with lipid droplet accumulation by 3T3-L1 preadipose cells. FPRP, a transmembrane glycoprotein, has been shown to regulate the binding of ligand to certain seven-transmembrane receptors. Anti-FPRP immunohistochemistry, Western blotting, and metabolic labeling/immunoprecipitation experiments demonstrated that FPRP was not detectable in undifferentiated 3T3-L1 cells. Interestingly, low levels of FPRP mRNA were detected in the undifferentiated 3T3-L1 cells. After induction of adipose differentiation, FPRP mRNA increased approximately 3 fold whereas FPRP synthesis increased approximately 50 fold. Differentiation induction with either dexamethasone/insulin/isobutylmethylxanthine or the thiazolidinedione derivative ADD 4743 were both effective at inducing FPRP accumulation and accumulation of lipid droplets. By co-immunohistochemical and lipid staining, greater than 99% of the cells accumulating lipid droplets possessed FPRP. FPRP mRNA and protein are also found in rat adipose tissue. Treatment of 3T3-L1 cells with an FPRP anti-sense oligonucleotide during differentiation decreased FPRP accumulation and resulted in a decrease in lipid droplets without altering the level of induction of a late marker of adipocyte differentiation, glycerol-3-phosphate dehydrogenase activity. Transient expression of an FPRP cDNA in undifferentiated 3T3-L1 cells was insufficient to induce lipid droplet accumulation.     

Platinum compounds as radiosensitizers in strontium-89 metabolic radiotherapy

PURPOSE: Strontium-89 is currently used for the treatment of painful bone metastases. This study reports on two preliminary experiences with low-dose platinum compounds, carboplatin and cisplatin, as radiosensitizers in 89Sr therapy. PATIENTS AND METHODS: 30 patients entered the carboplatin study: 15 patients (Group A) were treated with 148 MBq 89Sr followed by carboplatin (100 mg/m2 at 7 and 21 days) and 15 patients (Group B) were treated with 89Sr alone. 12 patients entered the cisplatin study: six patients (Arm 1) received 148 Mq 89Sr plus cisplatin (50 mg/m2) in two administrations (immediately before and 10 days after 89Sr injection) and six patients (Arm 2) received 89Sr plus two placebo administrations. Pain response was assessed 8 weeks after the therapy on the Wisconsin score modifications. RESULTS: No clinically significant adverse effects or myelosuppression by platinum compounds were observed. In carboplatin study a pain response was observed in 20 of 27 (74%) evaluable patients, 13/15 in group A and 7/12 in group B. The pain response in the patients treated with 89Sr and carboplatin was clearly superior to that seen in the patients treated with 89Sr alone (P = 0.025), whereas survival was only marginally better in the combined treatment group (8.1 vs 5.7 months, P = 0.19). In cisplatin study a pain response was observed in 10 of 12 (83%) evaluable patients, 5/6 in Arm 1 and 4/6 in Arm 2. CONCLUSIONS: Low-dose platinum compounds seem to enhance the effects of 89Sr radioisotope therapy on pain from bone metastases without relevant hematological toxicity.     

Video-assisted minimally invasive mitral valve surgery

OBJECTIVE: This study was done to determine the potential benefits of minimally invasive mitral surgery performed with intraoperative video assistance. METHODS: From May 1996 until March 1997, a minithoracotomy and video assistance were used in 31 consecutive patients undergoing mitral repair (n = 20) and replacement (n = 11). Their ages ranged from 18 to 77 years (59 +/- 2.6 years; mean +/- standard error of the mean). Ejection fractions were 35% to 62% (55% +/- 1.5%). Operations were done with either antegrade/retrograde (n = 10) or antegrade (n = 19) cold blood cardioplegia and a new transthoracic crossclamp or with ventricular fibrillation (n = 2). Peripheral arterial cannulation (n = 28) and pump-assisted right atrial drainage (n = 26) were used most often. RESULTS: No hospital deaths occurred, but the 30-day mortality was 3.2%. Complications included deep venous thrombosis and a phrenic nerve palsy in one patient each. No patient had a stroke or required reoperation for bleeding. Postoperative echocardiography showed excellent valve function in all but one patient. Cardiopulmonary bypass and arrest times averaged 183 +/- 7.2 and 136 +/- 5.5 minutes, respectively. Compared with 100 patients having conventional mitral valve operations, these patients had significantly shorter hospitalization times (8.6 +/- 0.5 vs 5.1 +/- 0.9 days, p = 0.05). Moreover, 81% of the later cohort were discharged between day 3 and 5 (3.6 +/- 0.2 days). Hospital charges (decreases 27%, p = 0.05) and costs (decreases 34%, p < 0.05) were less than in conventional operations. Patient follow-up suggested minimal perioperative pain and rapid recovery. CONCLUSIONS: Early results suggest that video-assisted minimally invasive mitral operations can be done safely. These methods may benefit patients through less morbidity, earlier discharge, and lower cost.     

Percutaneous absorption and disposition of [14C]chlordecone in young and adult female rats

The objective of this study was to evaluate the effect of age and dosage on percutaneous absorption and disposition of [14C]chlordecone (Kepone) and to describe results using a physiological based pharmacokinetic (PBPK) model. Female Fischer 344 rats 33 and 82 days old were used as the young and adult animal models, respectively, and were studied over a 10-fold dose range. [14C]Chlordecone (0.286 micromol/cm2) was applied to dorsal skin (2. 3% BSA) and radioactivity was quantified in selected tissues and excreta up to 120 h. Absorption and disposition were also determined at three dose levels up to 2.68 micromol/cm2; fraction absorbed decreased as dose increased. In vitro percutaneous absorption was measured by static and flow-through methods; these yielded similar penetration rates, which were lower than those obtained in vivo. In vivo percutaneous absorption over 120 h was 14.4+/-0.99 and 14.2+/-1. 5% dose in young and adults, respectively. Organ and tissue content increased over time (carcass>liver>kidney), indicating prolonged absorption. Statistical differences between young and old were found for liver, skin, and urine, but not for absorption. Excretion occurred primarily in feces, but also in urine. A biophysically based percutaneous model was fitted to both young and adult in vivo absorption data. This was embedded in a whole body PBPK model which, upon optimization with SAAM II, estimated apparent tissue partition coefficients, urinary and fecal excretion rates, and parameters characterizing hepatic nonlinear uptake of bound chlordecone. The model reasonably predicted tissue chlordecone content at higher doses, when decreased absorption was accounted for.     

High expression of thymidylate synthase is associated with the drug resistance of gastric carcinoma to high dose 5-fluorouracil-based systemic chemotherapy

BACKGROUND: In the past 4 years, the weekly 24-hour infusion of high dose 5-fluorouracil (5-FU) and leucovorin in the treatment of patients with advanced gastric carcinoma has been prospectively studied at the authors' institution. This has enabled them to explore the possibility that the level of expression of thymidylate synthase (TS), the target enzyme of 5-FU, is related to the drug sensitivity of gastric carcinoma to 5-FU-based chemotherapy. METHODS: To be eligible for this study, patients were required to have received high dose 5-FU and leucovorin chemotherapy (weekly 24-hour infusions of 5-FU, 2,600 mg/m2, and leucovorin, 300 mg/m2) and to have had adequate prechemotherapy gastric carcinoma tissues for immunohistochemical study. TS106 monoclonal antibody was used to detect the expression of TS. A visual scoring system, which ranged from 0 to 3+, was adopted by 2 independent pathologists to semiquantitate the intensity of TS expression. RESULTS: Between 1993 and 1996, a total of 30 patients, 18 men and 12 women, with a median age of 61.5 years, were enrolled. Of these patients, 16 (53.3%) and 14 (46.7%) had high and low expression of TS, respectively. Two of the 16 patients (12.5%) with high expression of TS and 13 of the 14 patients (92.9%) with low expression of TS responded to chemotherapy (P < 0.001, chi-square test). The median overall survival was 10 months for patients with low TS expression and 4 months for patients with high TS expression (P < 0.01, log rank test). CONCLUSIONS: The data from this study suggest that the expression of TS, as determined by immunohistochemistry, is a relatively reliable indicator of whether 5-FU should be used in the treatment of patients with gastric carcinoma.     

Infection with human immunodeficiency virus type 1 upregulates DNA methyltransferase, resulting in de novo methylation of the gamma interferon (IFN-gamma) promoter and subsequent downregulation of IFN-gamma production

The immune response to pathogens is regulated by a delicate balance of cytokines. The dysregulation of cytokine gene expression, including interleukin-12, tumor necrosis factor alpha, and gamma interferon (IFN-gamma), following human retrovirus infection is well documented. One process by which such gene expression may be modulated is altered DNA methylation. In subsets of T-helper cells, the expression of IFN-gamma, a cytokine important to the immune response to viral infection, is regulated in part by DNA methylation such that mRNA expression inversely correlates with the methylation status of the promoter. Of the many possible genes whose methylation status could be affected by viral infection, we examined the IFN-gamma gene as a candidate. We show here that acute infection of cells with human immunodeficiency virus type 1 (HIV-1) results in (i) increased DNA methyltransferase expression and activity, (ii) an overall increase in methylation of DNA in infected cells, and (iii) the de novo methylation of a CpG dinucleotide in the IFN-gamma gene promoter, resulting in the subsequent downregulation of expression of this cytokine. The introduction of an antisense methyltransferase construct into lymphoid cells resulted in markedly decreased methyltransferase expression, hypomethylation throughout the IFN-gamma gene, and increased IFN-gamma production, demonstrating a direct link between methyltransferase and IFN-gamma gene expression. The ability of increased DNA methyltransferase activity to downregulate the expression of genes like the IFN-gamma gene may be one of the mechanisms for dysfunction of T cells in HIV-1-infected individuals.