Antitumor effect of KT6124, a novel derivative of protein kinase inhibitor K-252a and its mechanism of action

Summary Novel derivatives of K-252a, (8R*,9S*,11S*)-(–)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8, 11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo [a,g]-cycloocta[cde]trinden-1-one, an inhibitor of protein kinases and calmodulin-dependent phosphodiesterase, were synthesized and evaluated for their antitumor activity in vitro and in vivo. Of ten derivatives tested, four were active against the P388 murine leukemia i. p.-i. p. system, although K-252a was inactive. Among these derivatives, KT6124 was selected for further biological evaluation studies because its efficacy was the highest. KT6124 was also active against sarcoma 180 and B16 melanoma. It exerted a relatively broad spectrum of antiproliferative activity against 20 human tumor cell lines in vitro. To determine the mechanism(s) of action underlying the antitumor activity of KT6124, we tested the drug for inhibition of protein kinases, including Ca²⁺-and phospholipid-dependent protein kinase (PKC), in intact A431 human epidermoid carcinoma cells in comparison with the PKC-inhibitory activity of K-252a. KT6124 did not antagonize the action of phorbol 12-myristate 13-acetate (PMA) in A431 cells, whereas K-252a did, suggesting that KT6124 may not act on protein kinases in the cells. The interaction of KT6124 with DNA in living cells was examined by the alkaline elution method. KT6124 apparantly exhibited DNA scission both dose-and time-dependently in the target cells. The DNA breakage was dependent on proteinase K treatment, suggesting its possible interaction with DNA-related enzyme(s). These results indicate that KT6124 exerts antitumor activity by acting on DNA or on DNA-related enzyme(s) in tumor cells rather than via the inhibition of protein kinases.     

Surgical experience of subacute pulmonary thromboembolism with severe pulmonary hypertension.

Surgical treatment for subacute pulmonary arterial thromboembolism has previously been considered to be inappropriate. We undertook a pulmonary arterial thrombectomy and removal of a floating right heart thrombus in a patient who had been symptomatic for over a month. The pulmonary arterial pressure, which had been equal to the systemic pressure preoperatively, decreased gradually and almost normalized one month postoperatively. Pulmonary perfusion scintigraphy revealed a dramatic improvement and the patient returned to normal life activities.     

Use of Glycopeptidolipid Core Antigen for Serodiagnosis of Mycobacterium avium Complex Pulmonary Disease in Immunocompetent Patients

We report the development of a serodiagnostic method for Mycobacterium avium complex (MAC) disease with an enzyme immunoassay (EIA) with the MAC-specific glycopeptidolipid (GPL) core as the antigen. In this study, we confirmed by EIA that the GPL core antibody was in the sera of immunocompetent patients with MAC disease. The EIA for quantifying the GPL core antibody was evaluated as a clinical tool for serodiagnosis of pulmonary MAC disease. A significant increase in GPL core antibodies (immunoglobulins G, A, and M) was detected in sera of patients with MAC pulmonary diseases when they were compared to patients who were colonized with MAC, patients with Mycobacterium kansasii disease or tuberculosis, and healthy subjects. The sensitivities and specificities of the GPL core-based EIA for diagnosis of MAC pulmonary disease were 72.6% and 92.2%, respectively, for IgG, 92.5% and 95.1%, respectively, for IgA, and 78.3% and 91.0%, respectively, for IgM. The best sensitivity and specificity were obtained by measuring immunoglobulin A antibodies against GPL core antigen. The level of GPL core antibodies reflected disease activity, since it decreased in cured MAC patients who had responded to chemotherapy. Measurement of serum antibodies against GPL core is useful for both diagnosis and assessment of disease activity in MAC disease of the lung.     

Clinical study of estramustine phosphate disodium (Estracyt) on prostatic cancer--results of long-term therapy for 38 patients with prostatic cancer

Estramustine phosphate disodium (Estracyt) was used in the treatment of 38 patients with prostatic carcinoma for at least 1 year. Of these patients 37 patients were treated with Estracyt as primary treatment and 1 patient had been treated with another antiandrogenic therapy before the Estracyt treatment. Estracyt was given orally in a dose of 560 mg/day in divided oral doses. The clinical evaluation was done for the change of PAP, the relapse rate, the survival rate and the side effect. Among 22 cases which had shown abnormally high PAP values before the treatment started, the values decreased or normalized in 21 cases (95.5%) in the first year of administration of Estracyt. In 6 cases, however, the values increased again in the second year or later. Relapse was observed in 10 (26.3%) out of 38 cases. Relapse rate was 2.6%, 51.7%, and 51.7%, at the first, third, and fifth year, respectively. Survival rate was 97.4% at the first year, 88.5% at the third year, and 68.8% at the fifth year for the follow-up study. Side effects were observed in 14 (36.8%) out of 38 cases. The main side effect was gynecomastia. Gastro-intestinal disturbance and edema were also observed. However, there were only 2 cases (5.2%) in which administration of Estracyt had to be discontinued.     

Metabolites of (+)-dehydroabietic acid in rabbits

The seven metabolites of (+)-dehydroabietic acid (DHA) were newly isolated from rabbit urine by liquid chromatography. On the basis of chemical and spectral data their structures were established to be (15S)-8,11.13-abietatrien-16,18-dioic acid, 2 alpha-hydroxy-8,11,13,15-abietatetraen-18-oic acid, (15R)-15,16-dihydroxy-8,11,13-abietatrien-18-oic acid, 2 beta,15-dihydroxy-8,11,13-abietatrien-18-oic acid, (15S)-2 beta,16-dihydroxy-8,11,13-abietatrien-18-oic acid, 2 alpha,15-dihydroxy-8,11,13-abietatrien-18-oic acid, and (15S)-2 alpha,16-dihydroxy-8,11,13-abietatrien-18-oic acid. The possible hydroxylation routes of DHA in rabbits and the difference between the metabolism of DHA in microorganisms and that in rabbits are discussed.     

Myocardial protective effect of lidocaine during experimental off-pump coronary artery bypass grafting.

Off-pump coronary artery bypass grafting (OPCABG) has recently gained popularity. During OPCABG, patients remain vulnerable to ischemic-reperfusion injury due to a temporary coronary occlusion without any active cardioprotection. Some strategies such as ischemic preconditioning (IP) and an intracoronary shunt have been applied with a view to minimizing the effects of ischemia, but the effects of these strategies remain controversial. This study was carried out to investigate the protective effect of lidocaine against myocardial ischemic-reperfusion injury. Twenty-one pigs were assigned to three groups, each consisting of seven pigs. In the control group, using a left internal thoracic artery (LITA) bypass circuit, the left anterior descending coronary artery (LAD) was occluded for 45 min followed by two hours of reperfusion. In the IP group, five min of occlusion followed by 15 min of reperfusion was performed. In the lidocaine group, 2 mg/kg of lidocaine was administered directly into the LAD just before the LAD occlusion. Infarct size expressed as a percentage of the area at risk was significantly smaller in the lidocaine group (2.7+/-4.2%) than in the control group (79.9+/-6.0%, p<0.001) or the IP group (57.0+/-25.9%, p<0.001). Lidocaine exhibited a potent myocardial protective effect in the present OPCABG model.     

Ketamine Suppresses Platelet Aggregation Possibly by Suppressed Inositol Triphosphate Formation and Subsequent Suppression of Cytosolic Calcium Increase

Background: Ketamine has been shown to suppress platelet aggregation, but its mechanisms of action have not been defined. The purpose of the current study is to clarify the effects of ketamine on human platelet aggregation and to elucidate the underlying mechanisms of its action.

Methods: Platelet aggregation was measured using an eight-channel aggregometer, and cytosolic free calcium concentration was measured in Fura-2/AM-loaded platelets using a fluorometer. Inositol 1,4,5-triphosphate (IP3) was measured with use of a commercially available IP3 assay kit. To estimate thromboxane A2 (TXA2) receptor binding affinity and expression, Scatchard analysis was performed using [3H]S145, a specific TXA2 receptor antagonist. TXA2 agonist binding assay was also performed. The membrane-bound guanosine 5'-triphosphatase activity was determined using [[gamma]-32P]guanosine triphosphate by liquid scintillation analyzer.

Results: Ketamine (500 [mu]m) suppressed aggregation induced by adenosine diphosphate (0.5 [mu]m), epinephrine (1 [mu]m), (+)-9,11-epithia-11,12-methano-TXA2 (STA2) (0.5 [mu]m), and thrombin (0.02 U/ml) to 39.1 +/- 30.9, 46.3 +/- 4.3, -2.0 +/- 16.8, and 86.6 +/- 1.4% of zero-control, respectively. Ketamine (250 [mu]m-1 mm) also suppressed thrombin- and STA2-induced cytosolic free calcium concentration increase dose dependently. Although ketamine (2 mm) had no effect on TXA2 receptor expression and its binding affinity, it (1 mm) suppressed intracellular peak IP3 concentrations induced by thrombin and STA2 from 6.60 +/- 1.82 and 4.39 +/- 2.41 to 2.41 +/- 0.98 and 1.90 +/- 0.86 pmol/109 platelets, respectively, and it suppressed guanosine triphosphate hydrolysis induced by thrombin (0.02 units/ml) and STA2 (0.5 [mu]m) to 50.3 +/- 3.2 and 67.5 +/- 5.5%versus zero-control, respectively.     

Enhancing Effects of Harman and Norharman on Induction of Preneoplastic and Neoplastic Kidney Lesions in Rats Initiated with N-Ethyl-N-hydroxyethylnitrosamine

The modifying potential of two nephrotoxic agents, harman and norharman, on N-ethyl-N-hydroxyethylnitrosamine (EHEN)-induced renal and hepatic carcinogenesis was investigated in male F344/DuCrj rats. Animals were given 0.1% EHEN in their drinking water for the first 2 weeks as an initiator. Subsequently, starting 3 weeks from the commencement, they were fed diet containing these compounds at concentrations of 1000, S00 or 0 ppm until week 26, and then killed for light microscopic examination. The mean numbers of renal tubular cell hyperplasias/cm² and those of tumors/cm² in rats given harman and norharman at 1000 ppm after initiation, but not at 500 ppm, were significantly increased as compared to the control values. However, neither compound modified liver carcinogenesis. It is concluded that harman and norharman show enhancing effects on rat kidney carcinogenesis, when ingested at dose levels which cause renal tubular damage.     

HISTOPATHOLOGIGAL STUDY OF HYPERTROPHIC CARDIOMYOPATHY WITH PROGRESSION TO LEFT VENTRICULAR DILATATION

The heart of seven cases of fatal congestive heart failure with dilated left ventricle, developing in 5 patients with symptomatic hypertrophic cardiomyopathy (HCM) and 2 patients with histologically widespread disarray of both ventricles, was morphologically investigated. These 7 cases showed myocardial widespread disarray and massive fibrosis, the mean percent area of fibrosis was 40.6% and 59.4% at upper and lower levels of left ventricles, respectively. Fibrosis was most extentsive in the lateral wall, and followed by anterior, posterior and interventricular walls. The severity of cell infiltration in left ventricle was completely matched to that of fibrosis and was most extensive in subepicardial area followed by middle and subendocardial areas of left ventricle. The intima and medial thickness of intramural small arteries in the fibrotic areas was significantly larger (p<0.05) than that of nonfibrotic areas, which suggested that the effect of intramural small artery was not essential for pathogenesis of massive fibrosis. ACTA PATHOL. JPN. 37: 1041 -1052, 1987.     

Inhibitory Effect of Cryptoporic Acid E, a Product from Fungus Cryptoporus volvatus, on Colon Carcinogenesis Induced with N-Methyl-N-Nitrosourea in Rats and with 1,2-Dimethylhydrazine in Mice

The antitumorigenic effect of cryptoporic acid E (CPA-E), a dimeric drimane sesquiterpenoid isolated from the fungus Cryptoporus volvatus , on colon carcinogenesis was investigated. Female F344 rats given an intrarectal instillation of 2 mg of N-methyl-N-nitrosourea 3 times weekly in weeks 1 and 2 were fed diet containing 0.2% CPA-E from week 3. Femal ICR mice given 15 weekly intraperitoneal injections of 10 mg of 1,2-dimethylhydrazine/kg body weight during weeks 1 to 15 were fed diet containing 0.06% CPA-E from week 1. The experiment was terminated at week 35 for rats and at week 25 for mice. The incidence and the number of tumors per animal were reduced in CPA-E-fed animals compared to the controls: 31% vs. 75% (P<0.05) and 0.4±0.2 (SEM) vs. 0.9 ± 0.2 (0.1> P >0.05) in rats, and 31% vs. 63% (0.1>P>0.05) and 0.4±0.2 vs. 2.4 ± 0.8 (P<0.05) in mice (16 animals in each group). Intrarectal deoxycholic acid-induced colonic mucosal ornithine decarboxylase activity was significantly lowered in CPA-E-fed animals compared to controls. This shows an antipromoting activity of CPA-E against colon carcinogenesis. Thus, it was concluded that CPA-E inhibits colon cancer development in both rats and mice treated with 2 different colon carcinogens.