In Ph(+) CML patients who achieved complete cytogenetic response (CCR) with interferon-alpha (IFN) treatment, how long the treatment should be continued has not well been investigated. We report here 2 CML cases who stopped the treatment after CCR had been sustained with IFN for 2-3 years. A 49-year-old male (case 1) achieved CCR 6 months after the initiation of IFN treatment. CCR had been maintained for 3 years, and then the treatment was ceased. CCR has been sustained without any therapy for 4 years. In this case, RT-PCR became negative half a year after achievement of CCR, and since then negative RT-PCR has been maintained. In case 2, a 50-year-old male, CCR was achieved after 8 years of IFN treatment, and maintained for 2 years. One month after cessation of the treatment, CML relapsed cytogenetically. In case 2, negative RT-PCR results were not maintained during the period of CCR. In case 1, the levels of T-cells for PR 1 were undetectable in the peripheral blood.

We reported two cases with germ cell tumor in which new preliminary treatment trials were performed by chemotherapy using anti-cancer drug selected on the basis of multidrug resistance gene mRNA expression, such as MDR1, MRP1, MRP2, MXR1, MGMT, GST pi and TopoII alpha, from RT-PCR assay. A 28-year-old male had gradually developed DI. MR imaging revealed enhanced tumors in the medulla oblongata, the pineal region and the suprasella region. Biopsy of tumor in the medulla oblongata demonstrated germinoma histologically. RT-PCR assay of this tissue revealed overexpression of MRP1, MGMT and GST pi mRNA, but neither MDR1, MRP2 nor MXR1 was observed. The patient was successfully given carboplatin, mitoxanthrone and ifosphamide after irradiation. A 15-year-old male was admitted to our hospital with high intracranial pressure syndrome. MR imaging revealed enhanced tumor in the pineal region. The tumor was diagnosed as a malignant germ cell tumor, histopathologically. RT-PCR assay of this tissue revealed overexpression of MRP1, MRP2, MXR1, MGMT and GST pi mRNA. Only MDR1 was not expressed. The patient was treated by irradiation including radiosurgery combined with chemotherapy, given cisplatin, etoposide and ifosphamide (ICE regimen), but he died because of progressive disease such as CSF dissemination. It seems that preliminary individual adjuvant chemotherapy based on mRNA expression of drug-resistance gene is available for the treatment of germ cell tumors.

Recent data have shown the existence of specific changes in mRNAs in thyroid carcinomas. It has not been clarified, however, why these changes clearly distinguish benign tissues from carcinomas, while genomic alternation such as mutations in the RAS or P53 genes do not. Further, the widely believed hypothesis, multi-step carcinogenesis, does not explain some clinical and experimental evidence of thyroid carcinomas. Considering these facts, we propose a new idea for thyroid carcinogenesis called "germ-cell carcinogenesis", in which cancer cells are derived from the remnant of fetal thyroid germ cells(thyroblasts) instead of normal thyroid follicular cells. Utilizing such mRNAs, we have established a new method for preoperative molecular-based diagnosis of thyroid carcinomas, Aspiration Biopsy Nucleic Acid Diagnosis(ABND). ABND allows us to perform preoperative nucleic acid analyses of the tumors by extracting RNAs or DNAs from tumor cells obtained by fine needle aspiration biopsies(FNABs). Pathological diagnosis of thyroid follicular carcinoma is quite difficult, and the establishment of preoperative molecular-based diagnosis of follicular carcinoma has been long expected. We found that quantification of the trefoil factor 3(TFF3)/galectin-3 mRNA ratio in thyroid tumor cells is a useful tool for distinction between follicular adenomas and carcinomas. Because ABND can be performed without any severe invasion to the patients, in the near future, when more reliable systems of quantitative RNA analysis have been developed, ABND will probably become one of the standard tests for preoperative diagnosis of thyroid carcinoma.

Based on reviews of the concept of diagnostics and in general and in specific tumour areas it was clear that development of diagnostic procedures involving genomics will allow for much better targeted and tailored treatments in the future. This will result in better efficacy and better tolerability of cancer treatments, but will also allow for progress in prediction, diagnosis and dose selection. Large collaborative projects studying the efficacy and safety of drugs on the genome level is promising to bring important benefits to both patients and the national economy by reducing useless drug therapy. In colorectal cancer there are several genetic defects identified that can act as the target for directed therapy in the future. Expressions of tumour specific antigens open the way for immunological targeted therapies. Developments in the understanding of the genomic basis for resistance to anti-tumour therapy is promising to help targeting patients likely to respond and not develop resistance. A Japanese model is being developed to determine the relative risk of breast cancer of Japanese women. Based on this prevention therapies can be instigated. The last four years have seen the introduction of four novel targeted therapies. If this model should become a standard in the future, much stronger collaboration between academic research and pharmaceutical industry need to develop.

Based on recent advancements in biotechnology and molecular oncology, genetic tests for molecular markers have been widely used for diagnosis and therapeutic monitoring in patients with hematological malignancies. However, incorporation of such tests has been limited in the care of patients with non-hematopoietic malignancies. The reasons for this include difficulty in sampling of the tumor tissue, and a lack of evidence for cost-effectiveness in the test utilization. A practical approach is the use of bodily fluids, such as blood, sputum, urine and feces, as specimens for early detection and monitoring of tumor-derived nucleic acid sequences. The prospect has been raised for development of tests for individualized medicine based on genetic information such as prediction of responsiveness to chemotherapeutic agents for selection of treatment, which will be promoted by gene expression profiling using DNA microarray.

Aberrant epigenetic information, represented by aberrant DNA methylation, plays a large role in carcinogenesis. Hypermethylation of CpG islands in promoter regions can lead to silencing of tumor-suppressor genes, and hypomethylation of the genome could lead to its instability. Epigenetic information is inherited from a cell to its daughter cells without any decay in somatic cells, but shows dynamic changes during embryonic development. Based on the possibility that epigenetic information can be theoretically reversed, attempts are being made to use the aberrant epigenetic information as therapeutic targets. A demethylating agent, 5-aza-2'-deoxycytidine, has been reported to be effective for some hematological malignancies, and its derivatives are being developed. Finally, reports are coming out on methods that can induce sequence-specific methylation changes. Rapid progress in this field is expected.

21-year-old male was diagnosed as having acute lymphoblastic leukemia (ALL) in March 2001. Standard G-banding analysis revealed normal karyotype. He achieved complete remission after the first course of induction therapy and underwent allogeneic PBSCT in August 2001. However, leukemic cells appeared in the peripheral blood demonstrating complex chromosomal abnormalities with Philadelphia chromosome (Ph1) and expressing minor BCR/ABL transcripts. We thus retrospectively examined the bone marrow sample at presentation with both interphase FISH and RT-PCR methods. Surprisingly, almost all leukemic blasts at presentation turned out to possess the minor type of BCR/ABL fusion gene. In spite of the second allogeneic PBSCT, the patient died of pneumonia and graft-versus-host disease in the liver in September 2002. This case is rare in that cytogenetic analysis failed to detect Ph1 due to leukemic cells dormancy. The application of both FISH and RT-PCR studies would appear to be essential to avoid overlooking Ph1-positive ALL.

We report an unusual case of the 'reverse' variant of follicular lymphoma in which the nodules had central parts that stained dark and cuffs that stained pale. Because diagnosis was difficult relying only on formalin-fixed histopathology, we examined the cell surface markers and karyotype. The patient, a 65-year-old man, presented with multiple lymphadenopathy, low-grade fever, night sweats, anorexia, dry cough and sense of chest oppression. Cell surface marker analysis showed that pathologic lymphocytes were positive for CD 10, CD 19, CD 20, HLA-DR, IgM/IgD and kappa, and t (14; 18) (q 32; q 21) was detected by karyotypic analysis. The 'reverse' variant of follicular lymphoma, clinical stage IVB was diagnosed the rearrangement band was detected with PCR-based clonality analysis in not only the immunoglobulin heavy chain gene but also the T cell receptor gamma chain gene, thus confirming monoclonal proliferation of both B cells and T cells.

One-hundred and four patients with previously untreated tongue cancer seen in our department between 1986 and 1998 were enrolled in a clinical study. The DNA ploidy patterns observed in fresh frozen specimens obtained from 41 patients were analyzed, and prognostic factors were investigated. According to the TNM classification (UICC 1997), 43 patients had stage I tumors, 29 had stage II tumors, 17 had stage III tumors, and 15 had stage IV tumors. The 5-year cause-specific survival rates for each stage were 94.7%, 64.4%, 50.0% and 45.7%, respectively. The most frequent cause of death associated with the original disease was the recurrence of the disease in cervical lymph nodes (19/27, 70.4%). The occurrence of late cervical metastasis was high among patients with a T2N0 disease. Patients with stage II disease should undergo elective neck dissection or be carefully monitored using ultrasonography. Among the 41 cases in which the DNA ploidy pattern was analyzed, diploid patterns were found in 30 cases and aneuploid patterns were found in 11. The 5-year cause-specific survival rate and the 5-year locoregional control rate were significantly lower for the aneuploid cases (18.2%, 38.9%) than for the diploid cases (66.5%, 69.8%) (p = 0.0003, p = 0.0339). The incidence of distant metastasis was significantly higher among the aneuploid cases (6/11, 54.5%) than among the diploid cases (3/30, 10.0%) (p = 0.0058). The ploidy pattern, as determined by flow cytometric DNA analysis, may reflect the malignancy grade of tongue cancers.

Imatinib Mesylate, a specific inhibitor of BCR/ABL tyrosine kinase, was developed as a molecularly targeted drug for the treatment of patients with chronic myelogenous leukemia. We evaluated effectiveness of the drug on cytogenetic response for monitoring residual disease using fluorescence in situ hybridization(FISH), reverse transcription-nested-polymerase chain reaction(RT-nested-PCR) and competitive PCR strategy. Of 9 patients in chronic phase, 7 achieved major cytogenetic response(CR) and 2 achieved minor CR by FISH. In 3 out of 6 patients with complete CR, no BCR/ABL gene was detected by RT-nested-PCR in peripheral blood or bone marrow specimens. Of 4 patients in accelerated phase, 1 achieved complete CR but 3 developed blast crisis. Despite high efficacy of Imatinib, 5 out of 13 patients showed resistance to the drug. To clarify the mechanism of resistance, we have newly developed a method for investigating a point mutation of T315I in tyrosine kinase domain of BCR/ABL gene using RT-PCR restriction digested analysis. None of them showed T315I mutation. The sensitivity of the method was as low as 10 copies of mutant gene. The method is useful for screening the mutation when there are many clinical samples or low copy number of BCR/ABL gene. BCR/ABL value obtained by FISH was of use to predict cytogenetic response when residual disease was above 2 to 3%. Below this level, the routine use of RT-nested-PCR was suffices to monitor minimal residual disease.

A 71-year-old man with chronic myelogenous leukemia received interferon therapy for 21 months, but did not have complete cytogenetic response. Imatinib was then added to the former therapy. Imatinib was prescribed for only 12 days and discontinued because of severe erythema. Since then, however, marked reduction of bcr-abl fusion chromosome positive cells was observed, and this state has been maintained to the present day even though treatment has consisted only of interferon. we discuss efficacy of the combined therapy, composed of interferon and imatinib, for chronic myelogenous leukemia and other bcr-abl fusion chromosome positive diseases.

Recent progress in molecular biology and cancer biology has revealed that many molecules, which are heavily involved in the un-limited growth, anti-apoptotic effects and invasion of tumors, would be targets of cancer therapy. Part of the drugs which inhibit these molecules have shown clinical response as well as clinical benefits. In this review article, the summary of mechanism based drug screening for cancer as well as recent status of new molecular target drugs for cancer, are described.

Tumor-reactive effector lymphocytes were generated using tumor-derived amplified RNA and cultured dendritic cells (DC). Tumor RNAs were extracted from gastric cancer cell line MKN45 or cancer cells of malignant effusions, and were processed with T7 amplification. DCs were induced from an adherent cell population of peripheral blood mononuclear cells (PBMCs) with GM-CSF and IL-4. Tumor-RNA was introduced into DCs using electroporation. Effector cells were generated by stimulating a non-adherent fraction of PBMCs with tumor RNA-introduced DCs. It was observed that milligram RNA could efficiently be amplified from microgram RNA. The effector cells, designated as tumor-RNA-introduced DC-activated killer (TRiDAK) cells, showed IFN-gamma spots in a tumor-specific manner when examined using ELISPOT analysis, and demonstrated cytotoxic activities against tumor cells from which RNA was extracted. TRiDAK cells produced more tumor-specific IFN-gamma spots when stimulated repeatedly. These results suggest that TRiDAK cells are practical and may be effective lymphocytes for adoptive cancer immunotherapy.

Our previous study using a cDNA microarray demonstrated that positive identification of differently expressed genes among gastric cancer cells involved in peritoneal dissemination could be accomplished. One of these genes with overexpression is inositol 1, 4, 5-trisphosphate receptor type 3 (IP3R3). IP3R3 is an intracellular Ca2+ release channel responsible for mobilizing stored Ca2+. Three different receptor types have been molecularly cloned, and their genes have been classified into a family. But the role of the IP3 signaling pathway in the peritoneal dissemination of gastric cancers is still unclear. In this study, IP3R3 is overexpressed in gastric cancer cell lines established from malignant ascites, but weakly expressed in gastric cancer cell lines established from primary tumor as well as in normal gastric epithelial cells. IP3R1 and 2 are expressed only weakly or not at all in these cells. The antagonist of IP3R, 2-APB, inhibited cell proliferation and induced apoptosis of gastric cancer cells from malignant ascites at concentrations of 100 nM to 100 microM in a dose dependent manner. Conversely, 2-APB showed a weak effect on other gastric cancer cells established from primary tumors (SNU1), lymph node metastases or liver metastases (MKN1 or 74), methothelial cell lines Met5A and myeloid leukemia cell HL60 cells. This suggests that this inhibitory effect depends on the level of IP3R3 expression. As cells that express IP3R3 mRNA (i.e., pancreatic aciner cells) are known to have a secretory function in which IP3/Ca2+ signaling has been shown to be involved, IP3R3 may be a prerequisite for secretion in gastric cancer cells. These results indicate that IP3R3 may be specifically involved in gastric cancer peritoneal dissemination and that IP3R3 may be a molecular target of the peritoneal dissemination of gastric cancer. Its antagonist, 2-APB, may thus be useful for the treatment of gastric cancer, especially for peritoneal dissemination.

Despite major recent advances in the understanding of the molecular biology of adult acute myeloblastic leukemia(AML), the treatment of the disease remains challenging. This review summarizes literature in the field of curative chemotherapy for adult acute myeloblastic leukemia(AML). In particular, detailed analysis of curing leukemia in patients treated with risk-oriented chemotherapy and in patients with APL was presented. In addition, results using our curative chemotherapy approach for adult AML were reported. Lastly, future directions for curing adult AML were discussed.

Using cDNA expression array and tissue microarray, we comprehensively and systematically analyzed the expression profile to investigate the lymphomagenesis. We detected the significant decrease of hematopoietic cell specific protein-tyrosine phosphatase SHP1 gene expression in various malignant lymphomas and leukemias. High-frequent silencing of the SHP1 gene by promoter methylation was detected in various kinds of leukemias and lymphomas as well as in many hematopoietic cell lines. The promoter methylation of the SHP1 gene was significantly correlated with the clinical stage. Loss of heterozygosity with microsatellite markers around the SHP1 gene was detected in 79% of informative ALL cases. These results suggest that loss of functional SHP1 is closely associated with the pathogenesis of leukemias/lymphomas.

A 69-year-old man was found to have leukocytosis and a bleeding tendency, when he underwent surgery for hemorrhoids in November 1992, at the age of 69. The patient was referred to our department for further examination, and was admitted on December 4. On admission, he had hepatomegaly (5 cm) and splenomegaly (12 cm). Laboratory data on admission showed that the leukocyte count was 173,400/microliter, erythrocyte count, 314 x 10(4)/microliter, hemoglobin level, 10.5 g/dl, hematocrit value, 29.7%, and platelet count, 14.4 x 10(4)/microliter, respectively. Peripheral hemogram revealed neutrophilia with a shift to the left to promyelocytes, and the positivity of neutrophil alkaline phosphatase (NAP) was very low. The bone marrow was hyperplastic with a high M/E ratio (5.8). As the chromosome analysis revealed that he had 9:22 translocation in all 20 karyotypes, chronic myelogenous leukemia in the chronic phase, was diagnosed. After the daily intramuscular administration of 9 megaunits interferon alpha-2b was started on December 9, 1992, his leukocyte count stabilized between 5,000 and 8,000/microliter. Thereafter, intramuscular administration of IFN alpha has been continued regularly almost twice a week at the outpatient clinic until now. The leukocyte count ranges from 3,000 to 6,000/ml and he is asymptomatic. In April 1995, complete cytogenetic response was achieved 28 months after the start of interferon alpha therapy. The recent bone marrow chromosomes examination showed Philadelphia-negative metaphases until now, December, 2002, although major bcr-abl still remains positive. This case suggests that treatment with interferon alpha may still be useful in some elderly patients with chronic myelogenous leukemia.