In vivo anti-tumor activity of a novel indolocarbazole compound, J-107088, on murine and human tumors transplanted into mice.

J-107088 (6-N-(1-hydroxymethyl-2-hydroxy)ethylamino-12,13-dihydro-2,10-dihydroxy- 13-(beta-D-glucopyranosyl)-5H-indolo[2,3-a]-pyrrolo [3,4-c]carbazole-5,7(6H)-dione) is a derivative of NB-506, an indolocarbazole compound previously reported as an anti-tumor agent targeting topoisomerase I. The optimal administration schedule of J-107088 was found to be intermittent injections. The GID75 (75% growth inhibiting total dose) values of J-107088 against LX-1 lung cancer and PC-3 prostate cancer when given by intermittent injection (twice a week for 2 consecutive weeks) were 200 and 15 mg/m2, respectively, whereas the 10% lethal dose (LD10) values of J-107088 against LX-1- and PC-3-bearing mice were 578 and 1200 mg/m2. The ratio of LD10/GID75 indicates the therapeutic window of an anti-tumor agent. Although the ratios of doxorubicin, paclitaxel and cisplatin against PC-3 were <0.3, <0.5 and <0.2, J-107088 showed the widest therapeutic window among the anti-tumor drugs tested. J-107088 was also effective on cells that had acquired resistance related to P-glycoprotein. Furthermore, J-107088 was found to be highly effective in inhibiting proliferation of micro-metastases of tumors to the liver in mice. Therefore, J-107088 is considered to be a promising candidate as an anti-tumor drug for treatment of solid tumors in humans.

Reduction cranioplasty for macrocephaly. Two case reports.

Multi-stage reduction cranioplasty was performed on two children with severe macrocephaly secondary to hydrocephalus. One patient underwent a four-stage operation, and the other underwent a two-stage operation. The postoperative course of both patients was uneventful. Reduction cranioplasty improved quality of life for both patients, and good cosmetic results were achieved. Reduction cranioplasty is effective for the treatment of macrocephaly, and multi-stage surgery can reduce the associated risks.

A new mechanism of acquisition of drug resistance by partial duplication of topoisomerase I.

Topoisomerase (topo)-I targeting antitumor agents are very effective in vivo against various human cancers. The indolocarbazole compound 6-N-formylamino-12,13-dihydro-1,11-dihydroxy-13-(beta-D-glucopyranosyl)- 5H-indolo[2,3-alpha]pyrrolo-[3,4-c]carbazole-5,7(6H)-dione (NB-506) is a potent inhibitor of the religation step of topo I reaction, like camptothecin (CPT). We established a NB-506-resistant cell line from murine leukemia cell line P388. This resistant cell line, P388/F11, exhibited 73-fold higher resistance to NB-506 and 3.5-fold higher cross-resistance to CPT than the parental cell line. No induction of cleavable complex formations induced by NB-506 and CPT were detected by K-SDS precipitation assays in P388/F11 cells. Analysis of nuclear extracts from P388/F11 cells revealed that the relaxation activity of topo I was one-quarter of that of the parental cells, and that the activity was resistant to induction of DNA cleavage by these drugs. Furthermore, Western blot and Northern blot analyses showed the expression of an abnormal-sized 170-kDa topo I protein and its 6.0-kb transcript and the absence of the normal topo I protein and transcript in P388/F11 cells. Analyses of the structure of the abnormal topo I transcript by reverse transcription-PCR and direct sequencing methods revealed that a large portion of the gene from codon 21 to codon 609 was duplicated in its coding region. This internal duplication resulted in in-frame fusion and, thus, production of a partially duplicated protein of 1357 amino acids. Finally, we expressed and purified the recombinant P388/F11 topo I in a baculovirus system. P388/F11 topo I showed similar catalytic activity to wild-type topo I, but reduced sensitivities to NB-506 and CPT. These results show that the altered sensitivity of duplicated topo I is involved in the NB-506 resistance of P388/F11 cells and indicate a novel resistant mechanism which involves duplication of the topo I gene.

Sequence-selective DNA cleavage by a topoisomerase I poison, NB-506.

An indolocarbazole compound, NB-506, inhibits the activity of topoisomerase I by stabilizing the DNA-topoisomerase I complex (cleavable complex). NB-506 inhibited the religation step of topoisomerase I activity more potently than camptothecin or its derivative, topotecan. A cleavage assay using an end-labeled fragment of DNA revealed that the pattern of cleavage induced by NB-506 was different from that induced by camptothecin. The preferred cleavage sites of NB-506 were found to be not only T but also A or C at the 3'-terminus of the cleaved DNA (position -1), while the DNA cleavage sites of camptothecin always had T at position -1. At the 5'-terminus of the cleaved DNA (position +1), NB-506 showed a preference for G, which is a feature shared in common with camptothecin. Therefore, the difference in cleavage patterns was most likely due mainly to the preferred base at position -1. Moreover, the re-ligation rate was significantly slower at NB-506-selective sites, which had C at position-1, than at camptothecin-selective sites or at sites cleaved by both NB-506 and camptothecin. Our data suggest that NB-506 is an unique topoisomerase I poison and that its potent inhibition of topoisomerase I is partly dependent on retardation of re-ligation at sites selectively induced by NB-506.

Antimetastatic effect of a novel indolocarbazole (NB-506) on IMC-HM murine tumor cells metastasized to the liver.

IMC-HM cells were isolated from spontaneously induced ascitic IMC carcinoma cells that had been maintained intraperitoneally in CDF1 mice. Metastasis to the liver of subcutaneously implanted IMC-HM cells was detected 10 days after implantation into the flanks of mice (day 10), but metastasis to other organs was limited. Thereafter, however, tumor cells spread rapidly to lymph nodes, lung, spleen, ovary and other organs, and the mice died on day 13 to 18. We report here, together with the properties of IMC-HM cells, the effects of adriamycin, cisplatin, etoposide and a new indolocarbazole antitumor compound (NB-506) on this model of metastasis. Although these anticancer agents all inhibited the growth of the subcutaneous tumors, their effects on the life span of the tumor-bearing mice varied. Treatment with NB-506, started on day 1, more than doubled the survival period at doses 30 mg/m2 to 900 mg/m2. Further, treatment with NB-506, started on day 4 after resection of the primary tumor, inhibited growth of the metastasized tumor in the liver and other organs. Etoposide also increased the life span at a limited range of doses. However, the life-prolonging effects of adriamycin and cisplatin were marginal. These results demonstrate that IMC-HM carcinoma is a good model for spontaneous metastasis to the liver followed by lethal spread to many organs. Moreover, NB-506 was found to be highly effective against the growth not only of subcutaneous tumors, but also of tumors metastasized to the liver.

Potent antitumor activity of quinolone compounds with an unsaturated aminoazabicyclo group at the C-7 position of the quinolone ring.

Relationships between the substituents on the quinolone nucleus of 2 and related compounds and their biological activities were studied. 2, 3 and 1 carrying a (1R, 2R, 6R)-2-amino-8-azabicyclo[4.3.0.]non-3-en-8-yl group at the C-7 position increased the rate of formation of DNA-protein complexes in cells, and inhibited the growth of tumor cells more strongly than the compounds with other substituents. The introduction of a fluorine atom or a methoxy group at the 8-position and an amino group at the 5-position increased the activity still further. The three compounds listed were all effective against P388 leukemia in mice. Subcutaneous injection of 2 at 2 mg/kg strongly suppressed the growth of human MX-1 breast cancer cells in nude mice. 1 has various functional groups that increase the cytotoxic potential of quinolone derivatives: a (1R, 2R, 6R)-2-amino-8-azabicyclo[4.3.0.]non-3-en-8-yl moiety at C-7, a cyclopropyl group at the 1-position, fluorine atoms at the 6- and 8-positions, and an amino group at the 5-position of the quinoline carboxylic acid. These data suggest that this series of compounds provide good models for the further design of potent antitumor quinolones.

New surgical treatment of middle fossa arachnoid cyst.

BACKGROUND: A new surgical treatment of middle fossa arachnoid cyst is reported. The benefit of our surgical procedure is to maintain the physiologic pathway of the cerebrospinal fluid (CSF) without a shunting procedure. METHODS: Following a small frontotemporal craniotomy, a linear incision in the outer wall of the cyst is made. The basal subarachnoid cisterns and distal sylvian cisterns are widely opened microsurgically. The outer wall of the cyst is then closed to prevent CSF leakage (arachnoidplasty). RESULTS: We have experienced six patients who underwent this new surgical treatment. All their cysts showed nor or delayed influx of the contrast medium on preoperative computed tomography cisternogram. A favorable outcome was attained in the long-term follow-up without any complications. CONCLUSIONS: The present technique was designed to secure physiologic CSF circulation and to avoid the use of a shunting system. A total of six patients treated with our new technique have been followed-up 6 years, and a good clinical outcome has been observed in all of them without complications.

Novel indolocarbazole compound 6-N-formylamino-12,13-dihydro-1,11-dihydroxy- 13-(beta-D-glucopyranosyl)-5H-indolo[2,3-a]pyrrolo-[3,4-c]carbazole- 5,7(6H)-dione (NB-506): its potent antitumor activities in mice.

NB-506 [6-N-formylamino-12,13-dihydro-1,11-dihydroxy-13-(beta-D- glucopyranosyl)-5H-indolo[2,3-a]pyrrolo[3,4-c]carbazole-5,7(6H)-di one d is a new antitumor indolocarbazole compound. The growths of murine M5076 and Ehrlich solid tumors were inhibited 76 and 96%, respectively, by i.v. injection at doses of 300 mg/m2. Furthermore, NB-506 caused regression of nodules of human PC-13 lung cancer and MKN-45 stomach cancer cells at i.v. doses of 90 mg/m2. Human HCT 116 and LS 180 colon cancers also regressed with injections of NB-506. Repeated injections of NB-506 had a stronger antitumor effect than intermittent injections in mice with MKN-45. The cumulative toxicity of NB-506 was low in terms of lethality in mice, i.e., the LD50s on single and 10 repeated i.v. injections into CDF1 mice were 990 and 810 mg/m2/injection, respectively. In conclusion, NB-506 is considered to be an interesting possible candidate as an anticancer drug for treatment of solid tumors in humans.

Novel antitumor indolocarbazole compound 6-N-formylamino-12,13-dihydro-1,11- dihydroxy-13-(beta-D-glucopyranosyl)-5H-indolo[2,3-a]pyrrolo[3,4- c]carbazole-5,7(6H)-dione (NB-506): induction of topoisomerase I-mediated DNA cleavage and mechanisms of cell line-selective cytotoxicity.

A new indolocarbazole antitumor agent, NB-506 [6-N-formylamino-12,13-dihydro-1,11-dihydroxy-13-(beta-D-glucopyranosyl) -5H- indolo[2,3-a]pyrrolo[3,4-c]carbazole-5,7(6H)-dione], enhanced the DNA cleavage catalyzed by HeLa S3 topoisomerase I at 0.01 microM but not the cleavage by topoisomerase II at 300 microM. It also caused single-strand DNA breakage in intact cells at 0.08 microM and more. Unlike the known topoisomerase I inhibitor camptothecin, NB-506 intercalated with DNA. However, the binding affinity to DNA and the inhibition against DNA polymerase alpha and RNA polymerase II were marginal compared with those of Adriamycin or actinomycin D. NB-506 inhibited the growth of various tumor cell lines at two micromoles or less, and its cytotoxicity was found to be cell line selective. This selective cytotoxicity of NB-506 was not fully explained by the differences in topoisomerase I activity in these cell lines, but there was some relationship between the amount of NB-506 accumulated in these cell lines and its cytotoxicity toward them. In conclusion, NB-506 is a potent topoisomerase I poison, acting selectively on tumor cell lines accumulating NB-506.

Inhibition of topoisomerase II by a novel antitumor cyclic depsipeptide, BE-22179.

BE-22179, a novel cyclic depsipeptide antibiotic having two 3-hydroxyquinoline moieties, inhibited the DNA-relaxing activity of L1210 topoisomerase II completely at 0.08 microM. This effect was far stronger than that of VP-16. However, it did not show any marked effect on topoisomerase II-mediated DNA cleavage. BE-22179 was ineffective in inhibiting the DNA relaxation by topoisomerase I at concentrations up to 10 microM, but showed DNA-intercalating ability (DNA unwinding) at 30 microM. The structure of BE-22179 is quite novel for a topoisomerase II inhibitor. Echinomycin, a quinoxaline antibiotic structurally related to BE-22179, interfered with DNA relaxation by topoisomerase II, though the effect was not due to inhibition of the catalytic activity of topoisomerase II but to conformational change of DNA based on its intercalation into DNA. Therefore, the potent inhibitory activity on topoisomerase II might not be a common activity of quinoxaline antibiotics, but might rather be specific to BE-22179. BE-22179 prevented DNA synthesis as well as RNA synthesis in L1210 cells and inhibited the growth of the cells. However, it remains unclear to what extent the topoisomerase II inhibition was responsible for the cytotoxicity of BE-22179.

Hemiatrophy of the tongue due to hypoglossal schwannoma shown by MRI.

Schwannomas account for 8.5% of all intracranial tumours; more than 90% arise from the 8th cranial nerve. Only 42 cases of schwannoma of the hypoglossal nerve have been reported. A 59 year-old woman developed right hemiatrophy of the tongue, clearly demonstrated on MRI, as was a small hypoglossal schwannoma. High signal was seen in the atrophic side of the tongue on both T1- and T2-weighted images, as described in the literature.

[A case of convexity arachnoid cyst associated with chronic subdural hematoma and intracystic hemorrhage].

Arachnoid cysts account for about 1% of all intracranial tumors, in which about 50% arise from the middle cranial fossa, and rarely occur at the cerebral convexity. They sometimes are associated with chronic subdural hematoma (CSDH) but the exact mechanism of their development is still unclear. A 15-year-old boy was admitted to our hospital with an arachnoid cyst at the right frontal convexity. When he experienced recurrent generalized seizure, CT and MRI revealed CSDH localized at the surface of the arachnoid cyst. Radical operation for the cyst with CSDH was successfully performed. As far as we know, it is rare that CSDH and hematoma are colocalized over an arachnoid cyst at the cerebral convexity. In this report, the operative findings of this patient and possible mechanism of CSDH formation were reported.

BE-23372M, a novel and specific inhibitor for epidermal growth factor receptor kinase.

The fungal metabolite BE-23372M is a structurally novel protein kinase inhibitor. Its IC50 for epidermal growth factor (EGF) receptor kinase was 0.03 microM. IC50 values of BE-23372M for other protein tyrosine kinases, erbB-2, p43v-abl, insulin receptor kinase, and p60c-src were 0.42, 1.0, 3.3, and 4.5 microM, respectively, and the IC50 for protein kinase C, a serine/threonine kinase, was 4.1 microM. Cdc2 kinase, casein kinases I and II and cAMP-dependent protein kinase were not inhibited by 20 microM BE-23372M. A kinetic study showed that BE-23372M was competitive with respect to the substrate peptide and to ATP. Autophosphorylation of solubilized EGF receptor kinase was clearly inhibited by 0.1 microM BE-23372M. Autophosphorylation of EGF receptor in A431 cells was also inhibited. These results show that BE-23372M is a potent and specific EGF receptor kinase inhibitor. It should be a valuable tool for EGF receptor kinase research.

BE-23372M, a novel protein tyrosine kinase inhibitor. I. Producing organism, fermentation, isolation and biological activities.

BE-23372M, a novel protein tyrosine kinase inhibitor, was isolated from the culture broth of a fungus. The producing strain, F23372, was identified as Rhizoctonia solani, based on the cultural and morphological characteristics. The active principle was extracted from the mycelium with acetone and purified by solvent extraction, silica gel column chromatography and Sephadex LH-20 column chromatography. BE-23372M showed strong inhibitory activity against EGF receptor kinase with IC50 values of 0.02 and 0.03 microM on two different substrates, whereas IC50 values against protein kinase C and cAMP-dependent protein kinase were 4.5 and > 20 microM, respectively. The compound inhibited the growth of A431 human epidermoid carcinoma and MKN-7 human stomach cancer cell lines with IC50 values of 8 and 24 microM, respectively.

A new topoisomerase II inhibitor, BE-22179, produced by a streptomycete. I. Producing strain, fermentation, isolation and biological activity.

A new topoisomerase II inhibitor, designated BE-22179, was isolated from the culture broth of Streptomyces sp. A22179, which resembles "Streptomyces gangtokensis". The inhibitor was extracted from the mycelial cake of the culture broth with organic solvent and successively purified by silica gel chromatography. BE-22179 inhibited topoisomerase II strongly but not topoisomerase I and showed potent antitumor activity against various tumor cell lines both in vitro and in vivo.

Benign intracerebellar cyst without epithelial lining.

A case of benign intracerebellar cyst is reported in a 61-year-old woman who presented progressive neurological deficits. MRI and operative findings could not reveal any evidence of neoplastic growth. The biopsied cyst wall revealed normal cerebellar tissue. The simple cyst of the cerebellum is rare, thus we discuss the clinical features and etiology of this rare entity.

A new antitumor substance, BE-18591, produced by a streptomycete. I. Fermentation, isolation, physico-chemical and biological properties.

New antitumor substance, designated BE-18591, was isolated from the culture broth of a streptomycete, strain BA18591. The active principle was extracted from mycelium by methanol and purified by silica gel chromatography. BE-18591 inhibited the growth of MKN-45 human stomach cancer cell line as well as P388 cell line. In in vivo experiments, BE-18591 inhibited the growth of Ehrlich ascites tumor.BE-18591 showed antimicrobial activity against Gram-positive and some Gram-negative bacteria.