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.     

Prognostic value of blood glucose in patients with cardiogenic shock.

BACKGROUND: Although an elevated blood glucose has prognostic value in cardiovascular disease, few data are available regarding its prognostic value for patients across the spectrum of cardiogenic shock. METHOD AND RESULTS: A total of 81 patients with cardiogenic shock whose blood glucose and adrenaline were measured on arrival at the emergency room (ER) were enrolled in this prospective study. The primary endpoint was death from any cause in hospital. The rate of death was 12.3% (10/81), and the glucose level was lower among patients who were discharged alive than among those who died (8.7+/-3.7 mmol/L vs 13.8+/-6.7 mmol/L, p<0.001). The unadjusted rate of death increased in a stepwise fashion among patients in increasing quartiles of glucose level (p<0.05). The blood glucose level of 9.2 mmol/L had the highest combined sensitivity and specificity for the identification of death. In the multiple logistic-regression analysis for the primary outcome, the adjusted odd ratio for a glucose level of 9.2 mmol/L or more was 5.8 (95% confidence interval, 1.0-32.8, p=0.047). There was a significant positive correlation between the glucose and adrenaline levels (R=0.726, p<0.0001). CONCLUSION: The measurement of blood glucose level on ER arrival provides predictive information for use in risk stratification across the spectrum of cardiac emergencies complicated by cardiogenic shock.     

Z‐axis video for cytology database is a useful tool for the case presentation prior to the cytology training workshop

We have built a database on the Internet managing z‐axis video for cytology (Zavic), and report on a new style of case discussion supported by the Zavic database. Z‐axis video for cytology is a movie file derived from the video recording of a microscopic field with changes in the focus. We used it for the case presentation of EUS‐FNA of pancreatic lesions on the Internet prior to a training workshop. The attendees were asked to observe the Zavic and to make diagnoses of 20 cases before the workshop. Fourteen attendees also observed lesions under a microscope on that day, and the results were compared with those of Zavic observation. The evaluation of the Zavic database (DB) was surveyed by a questionnaire. The average number of accurate diagnoses by 46 Zavic observers was 10.8. These accuracies for those who observed both the Zavic and glass slides were 11.57 and 11.43, respectively, for the videos and slides. Compared with Zavic observation alone, the diagnoses with glass slide observation were shifted to a correct diagnosis in two cases, but were shifted to an incorrect diagnosis in two cases. Approximately 60% of Zavic observers replied in the questionnaire that the movies on Zavic DB started to play within 3–4 seconds after clicking the play button. We successfully carried out the new style of case discussion supported by the Zavic DB. It was evaluated favorably by many attendees, who were psychologically still dependent on the glass slide observation. Diagn. Cytopathol. 2013. © 2011 Wiley Periodicals, Inc.     

An autopsied case of Creutzfeldt‐Jakob disease with mutation in the prion protein gene codon 232 and type 1+2 prion protein

A 68‐year‐old Japanese man gradually showed abnormal behavior and gait disturbance with bradykinesia. Slowly progressive dementia, including memory disturbance and disorientation, was also observed. Cerebral cortical hyperintensity on diffusion‐weighted MRI was observed 6 months after onset. The patient progressed to an akinetic mutism state with mild myoclonus, and atypical periodic sharp‐wave complexes were observed by electroencephalogram 13 months after onset. He was clinically suspected of having atypical CJD and died after 19 months total disease duration. The brain weighed 1160 g and showed mild atrophy of the cerebrum and cerebellum with ventricular dilatation. Spongiform changes with varying vacuole size and gliosis was extensive in the cerebral cortex and basal ganglia. Neuron loss in the cerebral cortex, basal ganglia and thalamus was relatively mild. The cerebellum showed mild spongiform changes of the molecular layer and mild neuron loss in the Purkinje cell layer. PrP immunostaining showed mainly coarse‐type combined with diffuse synaptic‐type PrP deposition in the cerebral gray matter. Some perivacuolar‐type PrP deposition was also present. Numerous plaque‐type PrP depositions were observed in the molecular layer of the cerebellum. Analysis of the PrP gene revealed a methionine‐to‐arginine (Met‐to‐Arg) substitution at codon 232 (M232R) with Met homozygosity at codon 129. Western blot analysis of protease‐resistant PrP indicated type 2 dominant PrP combined with type 1. Genetic CJD with M232R substitution in the PrP gene has only been reported in Japan. Although two clinical phenotypes (rapid‐type and slow‐type) were suggested in the M232R CJD cases (despite the presence of the same PrP genotype), the pathological and molecular backgrounds have not been well understood because there have only been a few autopsied case reports. This is the first case report of M232R CJD presenting with 1 + 2 PrP.     

Strong cytoplasmic expression of NF-κB/p65 correlates with a good prognosis in patients with triple-negative breast cancer

Purpose

Recent studies have indicated that constitutive NF-κB activity could be involved in the proliferation of triple-negative breast cancer.

Methods

The NF-κB/p65 expression and the effects of a NF-κB inhibitor, (?)-DHMEQ, were examined in triple-negative MDA-MB-231 breast cancer cells. Women with triple-negative breast cancer treated with neoadjuvant chemotherapy between 2002 and 2012 were retrospectively analyzed for their expression of NF-κB/p65, Bcl2 and Ki67 by immunohistochemistry in pre- and post-treatment specimens. The factors predicting the response to neoadjuvant chemotherapy and the prognosis were analyzed.

Results

NF-κB/p65 was predominantly expressed in the cytoplasm of MDA-MB-231 cells. Of 34 triple-negative breast cancer patients, positive staining for NF-κB/p65 expression was detected in the nuclei of a few cells in seven tumors before neoadjuvant chemotherapy, while the expression of NF-κB/p65 in the cytoplasm was detected in almost all tumor cells of 33 tumors. The expression levels of NF-κB/p65 were not associated with the response to neoadjuvant chemotherapy, although the cytoplasmic NF-κB/p65 staining intensity was significantly decreased in the post-treatment tumor samples compared with the pretreatment samples. All patients whose tumors showed strong cytoplasmic NF-κB/p65 expression before neoadjuvant chemotherapy are currently disease free.

Conclusion

Our results suggest that strong cytoplasmic NF-κB/p65 expression could be a prognostic marker for patients with triple-negative breast cancer.

     

GABA-induced intracellular Ca2+ elevation in the embryonic chick brainstem slice

We examined the intracellular Ca2+ ([Ca2+]i) elevation evoked by GABA in an 8-day embryonic chick brainstem slice using a Ca imaging technique with Ca green-1 AM. When small quantities of GABA were pressure-ejected on the surface of the slice, the [Ca2+]i elevation was clearly detected. The GABA-induced [Ca2+]i elevation was eliminated in a Ca2+-free solution, whereas the previously reported GABA-induced light-scattering change was independent of extracellular Ca2+. Although, micro-application of glycine or glutamate also induced [Ca2+]i elevation, these changes were smaller than that by GABA. These results suggest that the GABA-induced [Ca2+]i elevation is due to Ca2+ entry resulting from membrane depolarization and may play an important role in the development of the central nervous system (CNS).     

Safety and efficacy of metallic stent for unresectable distal malignant biliary obstruction in elderly patients

AIM To study the safety of insertion of metallic stents in elderly patients with unresectable distal malignant biliary obstruction.METHODS Of 272 patients with unresectable distal malignant biliary obstruction, 184 patients under the age of 80 were classified into Group A, and 88 subjects aged 80 years or more were classified into Group B. The safety of metallic stent insertion, metal stent patency period, and the obstruction rate were examined in each group.RESULTS In Group B, patients had a significantly worse per-formance status, high blood pressure, heart disease, cerebrovascular disease, and dementia; besides the rate of patients orally administered antiplatelet drugs or anticoagulants tended to be higher(P 0.05). Metallic stents were successfully inserted in all patients. The median patency period was 265.000 ± 26.779(1-965) d; 252.000 ± 35.998(1-618) d in Group A and 269.000 ± 47.885(1-965) d in Group B, with no significant difference between the two groups. Metallic stent obstruction occurred in 82 of the 272(30.15%) patients; in 53/184(28.80%) patients in Group A and in 29/88(32.95%) of those in Group B, showing no significant difference between the two groups. Procedural accidents due to metal stent insertion occurred in 24/272(8.8%) patients; in 17/184(9.2%) of patients in Group A and in 7/88(8.0%) of those in Group B, with no significant difference between the two groups, either.CONCLUSION These results suggested that metallic stents can be safely inserted to treat unresectable distal malignant biliary obstruction even in elderly patients aged 80 years or more.