To give optimal treatment to the patients with early lung cancer, we are trying to establish two new techniques to select the patients of poor prognosis and sensitive to the chemotherapy. To pick up the patients of poor prognosis nevertheless the early stage of their lung cancer, we are developing a new method to detect cancer cells floating in the peripheral blood flow with RT-PCR using cancer specific mRNA. To choose optimal chemotherapy regimens, we have established a new chemosensitivity testing (collagen gel droplet embedded culture drug sensitivity test: CDDST) for non-small cell lung cancer (NSCLC). Median survival time (MST) of the patients (n = 11) with unresectable NSCLC, who were given optimal chemotherapy according to CDDST, was 15.8 months and MST of those (n = 16) who did not have any sensitive agents according to CDDST was 5.6 months. There was significant difference between these two groups (p = 0.0048; logrank).

Worldwide research on the human genome is having a major impact on medical science and advanced medicine, although the detailed function and interaction of most genes remain unclear. The elucidation of human genome data makes it possible to take global views of biological processes and characteristics of cancer and individual drug response. This brings a number of new challenges such as genome based drug development and tailored cancer chemotherapy based on the individual genomic make-up. When a disease gene is identified, truly effective drugs targeting the genes can be developed. When biological characteristics such as polymorphism and gene expression profiles are closely related to drug response, an individually optimized drug therapy can be realized. cDNA microarray technology and new high-throughput single nucleotide polymorphism (SNP) screening offer great hope for such a global view by providing a systematic way to survey DNA and RNA variation. Evolutionary engineering techniques continue to be developed, which significantly promotes studies to understand molecular mechanisms causing diseases, novel disease genes, and genes related to drug response. Such progress in genome research and the functional analysis may revolutionize the anticancer chemotherapy world. The possible contribution of genome research to anticancer chemotherapy and problems of the day are reviewed herein.

Cisplatin and its analogues have been most frequently used for treatment of human cancer including ovarian cancer. Most advanced ovarian cancer which was fatal before introduction of cisplatin have become to be treated for cure by combination chemotherapy containing cisplatin and its analogues. Thus, combination chemotherapy containing cisplatin and carboplatin have become a standard chemotherapy for treatment of ovarian cancer. Initially, platinum-based combination chemotherapy is associated with a 60-70% clinical response rate. However, the overall 5-year survival rate for advanced ovarian cancer patients is still around 20-30%. This low survival rate is due to the fact that some primary tumors and most recurrent tumors develop drug resistance that leads to treatment failure. Thus, overcoming drug resistance is the key to successful treatment of ovarian cancer. The mechanism of cisplatin-resistance in ovarian cancer is multifactorial, and accumulation of multiple genetic changes may lead to the drug-resistant phenotype. In this review, we report several genetic factors conferring cisplatin-resistance which have been elucidated in our laboratory.

A 77-year-old woman was referred to our hospital because of leukocytosis and leukoblastosis in September 1999. She was healthy except for hypertension, and no abnormal findings in the peripheral blood had been observed up to December 1998. Physical examination revealed neither hepatosplenomegaly nor superficial lymphadenopathy. A bone marrow film showed massive proliferation of blast cells (87.8%), some of which contained coarse basophilic granules (38.6%). The cells were negative for peroxidase and esterase (alpha-naphtyl butyrate and ASD-chloroacetate) staining, but the granules showed metachromasia upon toluidine blue staining. As immunophenotypic analysis of the cells showed double positive for CD13/CD19 but negativity for CD33, this case did not meet the diagnostic criteria for biphenotypic acute leukemia. Chromosome and gene analysis showed positivity for the Ph1 chromosome with minor bcr/abl chimeric mRNA. A homogenate of the peripheral mononuclear cells demonstrated a high concentration of histamine. Electron microscopy analysis confirmed that some of the blast cells contained dense granules, which closely resembled "immature basophil granules" morphologically. These results suggested that the blast cells showed basophilic differentiation. As the clinical course and peripheral blood findings were different from blastic crisis of chronic myelogenous leukemia (CML) and CML with minor bcr/abl chimeric mRNA, the present case was diagnosed as "multiphenotypic acute leukemia", a type of acute basophilic leukemia classified by Duchayne.

We describe a complete cytogenetic response to interferon-alpha in a patient with chronic myelogenous leukemia undergoing chronic hemodialysis. Although IFN-alpha therapy has been applied to patients with chronic hepatitis C receiving hemodialysis, the pharmacokinetics of IFN-alpha in patients with poor renal function still remain unclear. In the present patient, the serum IFN-alpha concentration remained high even 48 hours after injection (42.9 IU/ml), and IFN-alpha was almost completely removed by hemodialysis (< 6 UI/ml). The patient was treated with IFN-alpha (3 x 10(6) IU, three times a week), and cytogenetic disappearance (0%) of the Ph-positive clone was confirmed 31 months after the start of therapy. Recombinant human erythropoietin (Epo) was used to treat anemia due to renal failure and IFN-alpha therapy. The anemia was controllable with Epo, and no adverse effect was observed.

Drug resistance, which often occurs during chemotherapy, is still a great obstacle to the success of human malignancy treatment. Among many possible mechanisms of drug resistance (biological, biochemical, kinetic or pharmacological), both typical and atypical multidrug-resistance (MDR) have been extensively studied. We picked up MDR-1, MXR, MRP1, MRP2, TopoII alpha, MGMT, and GST-pi as drug-resistant gene, based on experimental data and previous reports. Expression of these genes were measured in 14 malignant glioma specimens by reverse transcription polymerase chain reaction assay. We chose anticancer drugs for each patient, based on results of drug resistant gene expression to acquire good response to drugs. Though our follow-up periods are not long enough to analyze the results of our chemotherapy, 78% (7/9) of our glioma patients who were treated with our chemotherapy are free from tumor progression. The assays, which measure the expression of drug resistant genes, are necessary to allow rapid detection of the drug-sensitivity to chemotherapy in malignant glioma patients.

A 3-year-old boy with poorly prognostic acute megakaryoblastic leukemia (AML M7) showing t(16;21)(p11;q22) karyotype underwent unrelated bone marrow transplantation (U-BMT) during his first hematological remission. The conditioning regimen consisted of BU, VP-16 and L-PAM. Engraftment was smooth, but the patient developed grade I acute GVHD. During hematological remission before U-BMT, the TLS/FUS-ERG chimeric transcript of t(16;21)(p11;q22) was consistently detectable as minimal residual disease (MRD) by RT-PCR. However, after U-BMT it soon became undetectable. There was no detectable MRD until 7 months after U-BMT, but bone marrow relapse occurred 10 months after U-BMT. We consider that U-BMT is a promising treatment for t(16;21)(p11;q22) AML. However, an intensified conditioning regimen or modification of GVHD prophylaxis is needed.

Serial monitoring of chimerism after allogeneic hematopoietic stem cell transplantation (HSCT) can be performed easily and rapidly using PCR-based assays analyzing informative tandem repeat genetic markers. Sequential analysis of individual chimerism status was performed in 34 patients who underwent myeloablative allo-HSCT using a commercial multiplex short tandem repeat (STR) kit. Mixed chimerism (MC) was found in 14 of the patients for more than one month. The incidence of MC seemed to be dependent on the type of disease or pretransplantation regimen. There was no significant difference in relapse rates between MC and complete donor chimerism (CC) in all patients. However, the relapse rate was significantly higher in MC than in CC among patients with acute leukemia. The severity of acute graft-versus-host disease (aGVHD) was significantly reduced in the patients with MC. Most of the MC patients with hematologic malignancies had transient mixed T-lymphoid chimerism, and CC was achieved within 6 months after HSCT in such cases. Patients with MC beyond 6 months after HSCT and patients with reappearance of autologous signals (MC after CC) may have an enhanced risk of relapse.

The concept of Molecular Science of the Living Organism was described, where the living state is explained as the purposive flows of the quantum mechanically controlled chemical reaction systems which support the homeostasis of the living organism. In the 21st century, the post genomic sequence era, the concept may be a self-evident truth. Molecular Science of the Living Organism was presented in the case of G-proteins: i.e., the atomically controlled mechanism of 1. the carcinogenesis which originates from the point mutation of ras p21, 2. the activation of a receptor protein at the cell membrane, especially in the case of bacteriorhodopsin, 3. the activation of an inactive G-protein by the activated receptor protein.

Telomerase is an enzyme that replaces repetitive (TTAGGG)n sequences on the ends of chromosomes that would otherwise be lost during successive cell divisions. Telomerase activity is closely linked to attainment of cellular immortality, a step in carcinogenesis, while lack of such activity contributes to cellular senescence. Telomerase is activated in more than 85% of malignant tumors. However, with the exception of some self-renewing tissues with high regenerative potential, telomerase activity is usually repressed in normal somatic tissues. Based on these reports, we investigated telomerase activity in gastric mucosal tissues. Telomerase activity is highest in cancer, followed by intestinal metaplasia, chronic gastritis, and normal mucosa. In patients with intestinal-type gastric cancer, telomerase activity was higher in those with intestinal metaplasia and H. pylori infection than in patients without infection. Our results suggest that H. pylori infection may influence telomerase activity in cancer and noncancerous tissue. Genes encoding three major components of human telomerase have been recently cloned. They included those for human telomerase RNA component (hTR), human telomerase reverse transcriptase (hTERT), and telomerase-associated protein 1 (TEP1). More recently, two human telomeric repeat binding factors (TRFs) have also been cloned: TRF1, considered to inhibit the action of telomerase at the telomeric region, and TRF2, believed to prevent fusion of chromosome ends and, in vitro, to remodel linear telomeric DNA into large duplex loops. However, the details of mechanisms regulating telomerase activity are still poorly understood, and specific components or binding proteins that might represent suitable targets for cancer gene therapy have not yet been identified. Therefore, we established quantitative assays using a TaqMan RT-PCR for mRNAs encoding the telomerase components hTR, hTERT, and TEP1, as well as for those encoding TRF1 and TRF2. By using our quantitative assays, we found the following results: 1) Expression of TRF1 and TRF2 mRNA was greater in the normal cells than in human malignant hematopoietic cell lines or in patients with acute leukemia, 2) hTERT mRNA expression showed changes paralleling telomerase activity and became undetectable with HL60 cell differentiation, 3) initially low expression of TRF1 and TRF2 mRNA increased during differentiation. Our results suggest that not only hTERT but also TRF1 and 2 are important regulators of telomerase activity.