N-Myristoylated PCI peptide, myr-PCI, originally developed based on the PCI sequence of PLC- gamma 2, inhibited activity of purified PLC isoforms in vitro, and external stimuli-dependent IP3 formation in Swiss 3T3 cells. When myr-PCI was added to KMS-8 cells, derived from familial adenomatous polyposis patients, it also suppressed the serum-dependent IP3 formation, DNA synthesis, and cell growth. In order to develop PCI peptides with strong anti-PLC activity, we synthesized the peptides whose N- and/or C-termini were acylated by a variety of compounds, and examined their anti-PLC and anti-proliferative activities. Results would be quite helpful to determine the minimal structure required for PLC inhibition in vivo.

RAS controls at least two signaling pathways, one regulating extracellular signal-regulated kinase (ERK) activation and the other controlling membrane ruffling formation. Activating RAS mutations are commonly found in human tumors, making RAS and its downstream signaling pathways important targets for tumor therapeutics. We have developed a reporter-gene based assay system, utilizing transformation sensitive alpha-actin promoter, to identify compounds that inhibit the transforming activity of RAS either directly or indirectly. SCH51344 is a pyrazolo-quinoline derivative, identified based on its ability to depreprses alpha-actin promoter in RAS-transformed cells and shown to be a potent inhibitor of RAS-transformation. However, this compound had very little effect on the activities of the proteins in the ERK pathway, suggesting that it inhibits RAS-transformation by a novel mechanism and acts on a signaling pathway distinct from ERK pathway. Recently, in collaboration with Dr. Dafna Bar-Sagi's group, we have shown that SCH51344 inhibits membrane ruffling induced by activated forms of H-RAS, K-RAS, N-RAS and RAC. Treatment of fibroblast cells with this compound had very little effect on RAS-mediated activation of ERK and Jun kinase activities. Our results indicate that SCH51344 inhibits a critical component of the membrane ruffling pathway downstream from RAC and suggest that targeting the membrane ruffling pathway may be an effective approach to inhibit transformation by RAS.

DAN gene has been isolated by means of differential screening method between rat fibroblast 3Y1 and Rous sarcoma virus-transformed 3Y1 (SR-Y1) cells. DAN expression was suppressed in some of the transformed cells and its re-expression reverted the various transformed phenotypes such as colony formation in soft agar and tumor formation in mice. When DAN was overexpressed in 3Y1 cells, these cell showed the retardation of the entry into the S phase. DAN cDNA encodes a 27-kD protein which consists of 178 amino acids. The deduced amino acid sequence has no homology with known proteins. The amino terminal of DAN is highly hydrophobic and DAN is indeed secreted into the culture medium. This secreted DAN, when added to the culture of SR-3Y1 cells, suppressed DNA synthesis. Human DAN is mapped to 1p36.11-p36.13, a region known to have highly significant linkage with the genesis and/or progression of human neuroblastoma. Aberrant bands on Southern blot were detected in some of the neuroblastoma cases. Rat and human genomic DANs were isolated. CAT and the electrophoretic mobility shift assays revealed the presence of possible positive and negative regulatory elements at -57 approximately +118 and -1,232 approximately -987 of rat genome, respectively.

TGF-beta is a member of a superfamily associated with development and regulation of cell growth. Recently, its two receptors and intracellular components of signal transduction, a Smad family, have been identified. The type II receptor gene has been found to be mutated specifically in hereditary colon cancers, HNPCC. Alterations of the Smad 4 gene were also detected in pancreatic and colorectal cancers.

We examined the expression of gelsolin in a murine ras tumor and a number of human stomach, colon, bladder, and lung cancer cell lines and tissues. In most of the cell lines and tumor tissues, gelsolin expression was undetectable or extremely low in comparison with its expression in normal epithelial cells. Upon the introduction of the exogenous human wild-type gelsolin cDNA into human cancer cell lines, the gelsolin transfectants had greatly reduced colony-forming ability and tumorigenicity in vivo. After UVC irradiation, the gelsolin-overexpressing bladder cancer cells demonstrated increased accumulation and/or protracted delay in G2 phase as compared to neotransfected cells. UVC-induced production of diacylglycerol was reduced in gelsolin-overexpressed UMUC-2 cells as compared to neo-transfected UMUC-2 cells. Levels of cyclin B in the synchronized and gelsolin-overexpressing UMUC-2 cells remained low during the G2 delay. To investigate the in vivo efficacy of gene therapy with the gelsolin tumor suppressor, we treated human urinary bladder cancers (UMUC-2 and DAB-1), inoculated in nude mice, with recombinant retrovirus packaging cells containing the human gelsolin cDNA. This gene therapy resulted in remarkable tumor growth inhibition, and prolonged survival time in the majority of animals. These observations suggest that gelsolin plays a key role as a tumor suppressor by regulating a G2 checkpoint function of cancer cells through phosphoinositol lipid metabolism, and demonstrate the potential of using the gelsolin tumor suppressor in human urinary bladder carcinoma.

The APC gene is mutated in the majority of colorectal tumors. The product of this gene is a 300 kDa protein associated with beta-catenin and the human homolog of the Drosophila tumor suppressor DLG. The study of the function of APC provides important insights into the mechanisms of the development of colorectal tumor.

Neurofibromatosis type 2 (NF2) is an autosomal dominantly inherited disorder, strongly associated with development of benign intracranial tumors, including bilateral vestibular schwannomas and meningiomas. The NF2 gene located on the human chromosome 22q12 was recently cloned, and the protein it encodes (termed merlin/schwannomin) was found to be strikingly similar to the moesin-ezrin-radixin (MER) family of cytoskeleton-associated proteins. Mutations of NF2 gene have been found not only in the NF2 patient but also in NF2-related sporadic tumors such as acoustic schwannomas and meningiomas, suggesting that the NF2 gene functionally acts as a tumor suppressor gene. To elucidate the biological function of merlin, we have detected five merlin binding cellular proteins. The N-terminal region of merlin, the entire merlin-ezrin-radixin-moesin (MERM) homology domain, was found to be essential for binding to all five proteins. Since most reported NF2 mutations in the region were determined to be necessary for binding, the mutations probably impair binding. The majority of NF2 mutations are nonsense mutations or frameshifts that resulted in premature termination of merlin. The lack of these interactions caused by such mutations of NF2 gene may affect the cellular signals, resulting in benign intracranial tumors in NF2 patients.

A DNA mismatch repair system exists that repairs mispaired bases formed during DNA replication and genetic recombination. Genetic defects in this mismatch repair system are known to increase the rate of spontaneous mutation in Escherichia coli. Some cases of inherited cancer are associated with inherited defects of mismatch repair genes, showing the importance of the mismatch repair system in maintenance of genetic stability and avoidance of cancer susceptibility. This review focused on what is known about the mechanisms of mismatch repair in human cells and the relationship between defects in mismatch repair and carcinogenesis.

Hereditary cancer is rare, but it has served as a useful model in the understanding of carcinogenesis. A number of cancer genes have been identified by the study of hereditary human cancers and implicated in sporadic forms of the some tumors. In this article, I reviewed von Hippel-Lindau disease (VHL) gene and tuberous sclerosis 2 (TSC 2) gene.

A 66-year-old woman complained of chest discomfort in January 1995. In March the accelerated phase of chronic myeloid leukemia (CML) was diagnosed. Chromosomal analysis demonstrated negative Ph and positive t(9;16) (q34;p11) with positive major BCR/ABL chimeric mRNA. Administration of hydroxycarbamide was initiated, but in May she developed high fever and severe left hypochondralgia. Her WBC was 62,100/microliter (blast 64%), and LDH was 3,590 IU/l. Bone marrow examination showed 78.6% blasts, with a nucleated cell count of 74 x 10(3)/microliter. Blasts were negative for esterase stain and partially positive for both peroxidase stain and PAS reaction. Surface marker analysis revealed that blasts were positive for CD13, CD19, CD33, CD34, and HLA-DR. A diagnosis of blast crisis was made and she was treated with the VDS-CP regimen with heparin for DIC. After temporary improvement her disease recurred rapidly with severe DIC. Treatment with low molecular weight heparin and fresh frozen plasma failed to control DIC and she died of subarachnoid hemorrhage on the 48th hospital day. This is the first veprted of case Ph-negative, M-BCR/ABL-positive CML with t(9;16) accompanied by severe DIC.

We report on the leukemic cell karyotype of 180 children with acute myeloblastic leukemia (AML). They were treated by the protocols of chemotherapy for the Children's Cancer and Leukemia Study Group (CCLSG) in the last decade. Of 132 cases with adequate banding analysis, 24.2% had normal karyotype, 21.2% had miscellaneous clonal abnormalities and 54.6% were classified into known cytogenetic subgroups: t(8;21) (n = 35), t(15;17) (n = 23), inv (16) (n = 6), t(11q23;V) (n = 6), -7/7q-(n = 2). Each karyotype was closely correlated with a particular FAB subtype such as t(8;21) in M2, t(15;17) in M3, inv (16) in M4, t(11q23;V) in M5. In the M1+M2 group, although patients with t(8;21) had favorable clinical features such as low WBC counts and less frequent lymphadenopathy, their treatment outcome was not significantly better than those of patients with a normal karyotype (3-year EFS: 58 +/- 11% vs. 47 +/- 12%). Patients with miscellaneous chromosomal abnormalities had a significantly shorter EFS (22% +/- 10%) (p < 0.05) than those with t(8;21) or normal karyotype. In M4+M5 group, 2-year EFS of patients with inv (16) (40 + 30%) was longer than that of patients with normal karyotype (25 +/- 19%), and t(11q23;V) or miscellaneous chromosomal abnormalities (0 +/- 25%). These results suggest that cytogenetic data may be useful for risk-based treatment assignments for children with AML.

From a series of 176 patients with acute myeloid leukemia (AML), we have identified 11 patients with HLA-DR-negative AML excluding acute promyelocytic leukemia. All patients showed not (15; 17) or consistent chromosomal abnormalities. According to the French-British-American criteria, seven, three, and one patients were classified into M1, M2, and M4, respectively. Blasts from these patients were CD33+, HLA-DR-, and lymphoid-antigen-. All patients entered complete remission after induction therapy. Of these, blasts from four patients had strikingly invaginated nuclear membrane and finely granular peroxidase activity. The phenotype of the blasts was CD33+, CD34-, CD2-, and HLA-DR-. All four patients showed hyperleukocytosis on initial presentation. One patient received all-trans retinoic acid at relapse, however, the drug did not induce the differentiation of the blasts. These four patients were suspected to have acute myeloid/natural killer (NK) leukemia because of the prominent morphological feature of the blasts and the initial hyperleukocytosis, although NK cell activity of the blasts was not examined. Since HLA-DR-negative AML is heterogeneous, it is necessary for identifying acute myeloid/ NK leukemia within the disorder.

The DNA ploidy patterns of fresh specimens from 145 patients with squamous cell carcinoma of the oral cavity and the pharynx were analyzed. The specimens were embedded in paraffin and the DNA ploidy patterns were analyzed mainly by flow cytometry. Aneuploid patterns were found in 70%, and they were found more frequently in hypopharyngeal carcinomas than in oral cavity carcinomas. Correlation between DNA ploidy and tumor size was statistically significant but metastasis to lymph nodes and clinical stage were not correlated. In stage I and II oral cavity carcinomas and stage III and IV epi and oropharyngeal carcinomas, survival was much poorer in the aneuploid than in the diploid group. Analysis of 30 patients showed better response to preoperative chemotherapy in the aneuploid than in the diploid group; the mean efficacy rates were as high as 68% in the aneuploid and 48% in the diploid group. Aneuploidy with a low DNA index did not respond well to chemotherapy compared with those with a high DNA index. DNA heterogeneity was found in 22% (4/18) of the oral cavity carcinomas and in 50% (6/ 12) of the hypopharyngeal carcinomas.