Re-evaluation of refractory anemia with excess blasts in transformation.

The category 'refractory anemia with excess blasts in transformation (RAEBt)' consists of two sub-sets; one group is categorized based on the percentage of blasts in the marrow (> or =20%) and other is based on the percentage of blasts in the peripheral blood (> or =5%). We separated RAEBt patients based on these two criteria and compared hematologic and clinical relevance to assess the reasonable basis for the new classification. All RAEBt patients showing peripheral blood (PB) blasts of > or =5% were re-classified as RAEB by the WHO classification. This subset of RAEBt patients had lower percentages of bone marrow (BM) blasts, and notably they showed frequent complex cytogenetic abnormalities, including -5/5q- and/or -7/7q-. Moreover, the RAEBt patients of this group had shorter survivals compared to RAEBt patients with BM blasts between 20 and 30%. We next assessed hematologic and clinical relevance between refractory anemia with excess blasts (RAEB) and RAEBt patients with PB blasts of > or =5%. Except for the percentage of blasts in the PB (P=0.0037) and BM (P=0.0073), there was no significant difference in hematologic or clinical features between RAEB patients with BM blasts of > or =11% and RAEBt patients with PB blasts of > or =5%. When MDS patients with PB blasts of > or =5% (RAEBt by the FAB classification) were included as RAEB-II based on the "MDS 2000 classification', there was a high frequency of patients with complex chromosome changes, involving 5q and 7q, with significant poorer outcome compared to those with RAEB-I. Although it is still controversial whether MDS patients with BM blasts 20% or more should be considered as acute leukemia, the utilization of the 'MDS 2000 classification' might be useful to designate MDS patients diagnosed based on the percentage of blasts in the peripheral blood.

Combination of granulocyte colony-stimulating factor and low-dose cytosine arabinoside further enhances myeloid differentiation in leukemia cells in vitro.

We examined the differentiation-inducing effect on freshly isolated myeloid leukemia cells in liquid suspension culture by combined treatment with granulocyte colony-stimulating factor (G-CSF) plus low-dose cytosine arabinoside (Ara-C; 5-10 ng/ml) in 25 patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) in leukemic transformation. Culture with G-CSF alone showed leukemic cell growth stimulation in 15 out of the 25 cases (60%) and induction of cell differentiation in 19 out of the 25 cases (76%), respectively. In 23 cases (92%), either growth stimulation and/or differentiation induction of leukemia cells was observed in response to G-CSF. This suggests that most myeloid leukemia cells are able to respond to G-CSF stimulation. In addition, treatment of cells with low-dose Ara-C alone resulted in the enhancement of myeloid specific antigens expression in 16 cases (64%). Treatment of leukemia cells with higher concentrations of Ara-C (over 50 ng/ml) alone resulted in cytocidal effects but not in the induction of differentiation. Furthermore, 15 cases (60%) showed pronounced myeloid differentiation of leukemia cells after combined exposure to G-CSF plus low-dose Ara-C as compared with cells treated with either G-CSF or Ara-C alone. The enhanced effect of differentiation induction by combining G-CSF plus low-dose Ara-C was also observed in a murine myeloid leukemia cell line WEHI-3B in vitro. These data suggest that treatment with G-CSF plus low-dose Ara-C is capable of inducing differentiation of leukemic cells in vitro, and also appears to be useful for the differentiation-based therapy of patients with AML and MDS.

[Rapidly progressive fibrosis and increased CD68-positive cells in the bone marrow at the terminal stage of adult T-cell leukemia accompanied by polycythemia vera].

We report a case of adult T-cell leukemia (ATL) accompanied by polycythemia vera (PV) in which rapid development of myelofibrosis and clinical features of hemophagocytic syndrome (HPS) were observed at the terminal stage. The patient, a 53-year-old man who was born in Oita Prefecture, Japan, was diagnosed as having PV in 1996. He had undergone venesection but had not received any chemotherapy. In June 1997, he showed systemic lymphadenopathy with positivity for serum HTLV-1 antibody (x 10,240). Pathological findings and Southern blotting analysis for detection of monoclonal integration of HTLV-1 provirus DNA in a lymph node biopsy sample revealed that he also had acute-type ATL. Although several courses of chemotherapy were transiently effective, high fever, pancytopenia, increased serum LDH, hypoproteinemia and hyperferritinemia appeared, all of which were compatible with the clinical features of HPS. In addition, cytomegalovirus infection became evident. He died of multiple organ failure with rapid progression of myelofibrosis in May 1998. Detection of both increased CD68-positive histiocytes by immunohistochemistry and iron-stained phagocytic cells in marrow biopsy specimens appeared to be helpful for diagnosis of HPS in this patient, whose marrow showed myelofibrosis with hypocellularity.

Vitamin K2 induces apoptosis of a novel cell line established from a patient with myelodysplastic syndrome in blastic transformation.

We have previously reported that vitamin K2 (VK2) has a potent apoptosis inducing activity toward various types of primary cultured leukemia cells including acute myelogenous leukemia arising from myelodysplastic syndromes (MDS). We established a novel cell line, designated MDS-KZ, from a patient with MDS in blastic transformation, and further investigated the effects of VK2 using this novel cell line. MDS-KZ shows complex chromosomal anomaly including -4, 5q-, -7, 13q+, 20q-, consistent with that seen in the original patient. Culture of MDS-KZ cells in RPMI1640 medium containing 10% FBS lead to steady but very slow proliferation with a doubling time of 14 days. However, the cellular growth rate was significantly accelerated in the presence of various growth factors such as granulocyte colony-stimulating factor, stem cell factor, granulocyte-macrophage colony-stimulating factor, interleukin-3, and thrombopoietin. Most of the cultured cells show the morphological features of myeloblasts. They are positive for CD7, CD33, CD34, CD45, CD117, and HLA-DR. However, about 10% of the cells are more mature metamyelocytes and neutrophils with various dysplastic characteristics such as pseudo-Pelger nuclear anomaly and hypersegmentation, suggesting a potential for differentiation in this cell line. As previously reported for cultured primary leukemia cells, exposure to VK2, but not to VK1, resulted in induction of apoptosis of MDS-KZ cells in a dose-dependent manner (IC50: 5 microM). In addition, VK2 treatment induced down-regulation of BCL-2 and up-regulation of BAX protein expression with concomitant activation of caspase-3 (CPP32). A tetrapeptide functioning as antagonist of caspase-3, Ac-DEVD-H, suppressed the VK2-induced inhibition of cell growth, suggesting that caspase-3 is, at least in part, involved in VK2-induced apoptosis. These observations suggest that the MDS-KZ cell line can serve as a model for the study of the molecular mechanisms of VK2-induced apoptosis.

[A trial for peripheral blood stem cell harvest by combination of G-CSF with ABVD regimen in the management of Hodgkin's disease].

We studied the possibility of performing peripheral blood stem cell (PBSC) harvests during the course of ABVD therapy by adding G-CSF to the treatment regimen. Six patients with high-risk Hodgkin's disease (HD) (5 untreated cases with bulky mass and 1 relapsed case) received G-CSF (5 micrograms/kg) subcutaneously from day 8 to day 13 of their first course of ABVD treatment; the numbers of CD34+ cells and CFU-GM were monitored. PBSC harvests were performed on day 12 and day 13 of subsequent ABVD plus G-CSF treatment courses. For all patients tested, we were able to harvest CFU-GM (3.78 +/- 1.19 x 10(5) colonies/kg) for peripheral blood stem cell transplants (PBSCT) by performing 2 to 4 cycles. of apheresis, without any modification to the original ABVD protocol. These findings suggest that ABVD plus G-CSF therapy is a strong candidate for the treatment of patients with high-risk HD who may undergo autologous PBSCT.

Vitamin K2 selectively induces apoptosis of blastic cells in myelodysplastic syndrome: flow cytometric detection of apoptotic cells using APO2.7 monoclonal antibody.

We have previously reported that vitamin K2 (VK2) but not VK1 has a potent apoptosis-inducing effect on freshly isolated leukemia cells from patients with various types of leukemia. By multi-color flow cytometric analysis using monoclonal antibody (mAb), APO2.7, which detects mitochondrial 7A6 antigen specifically expressed by cells undergoing apoptosis, we further investigated the apoptosis-inducing effect of VK2 on minor populations of leukemic blast cells in bone marrow from patients with myelodysplastic syndrome (MDS) and overt myeloid leukemia (post-MDS AML). Limiting dilution of CD95 (anti-Fas) mAb-treated apoptotic Jurkat cells with nonapoptotic CTB-1 cells revealed that APO2.7-positive Jurkat cells were consistently detectable by flow cytometry when present at levels of at least 5% in the CTB-1 suspension. In patient samples the gating area for leukemic clone was determined using cell surface antigen-specific mAbs conjugated with either fluorescein isothionate (FITC) or phycoerythrin (PE) and subsequently the cells stained with phycoerythrin cyanine (PE-Cy5)-conjugated APO2.7 mAb were assessed within the gating area of the leukemic clone for monitoring apoptosis. Treatment of the bone marrow mononuclear cells with 3-10 microM of VK2 (menaquinone-3, -4 and -5) in vitro potently induced apoptosis of the leukemic blast cells as compared with the untreated control cells in all 15 MDS patients tested. This effect was more prominent on blastic cells than that on mature myeloid cells such as CD34-/CD33+ gated cells. In addition, VK2 performed much less effectively on CD3-positive lymphoid cells. In contrast to VK2, VK1 did not show apoptosis-inducing activity. These data suggest that VK2 may be used for treatment of patients with MDS in blastic transformation.

[Chronic myelogenous leukemia received unrelated bone marrow transplantation following surgical resection of cerebral arteriovenous malformation--first case report].

We report a 15-year-old boy with chronic myelogenous leukemia who received unrelated bone marrow transplantation (uBMT) after surgical resection of cerebral arteriovenous malformation (AVM). The incidence of cerebral hemorrhage caused by rupture of cerebral ABM in cases of BMT is uncertain. However, since the risk of rupture of AVM was supposed to increase due to both severe thrombocytopenia after intensive chemotherapy and increased intracranical pressure because of total body irradiation (TBI) as preconditioning therapy for BMT, we have first carried out surgical resection of the cerebral AVM, and subsequently performed uBMT. This resulted in a favorable clinical course without serious complications.

Cytological detection of telomerase activity using an in situ telomeric repeat amplification protocol assay.

A previously reported highly sensitive assay for measuring telomerase activity on cell and tissue extracts indicates that most human tumor tissues, but not cells adjacent to tumors, have detectable telomerase activity. Although this assay has provided a significant amount of information about the presence or absence of telomerase activity, it does not indicate whether all cells within a tumor have telomerase activity or whether only a subset does. The present report demonstrates the ability to advance this technology to an in situ assay. Using fluorescent telomerase primers and in situ PCR, we show that telomerase activity can be detected at the cellular level. This study demonstrates that telomerase activity is not detected in normal cells but is detected in tumor cells of clinical specimens and in tumor-derived cell lines.

Vitamin K2 and its derivatives induce apoptosis in leukemia cells and enhance the effect of all-trans retinoic acid.

Geranylgeraniol, a polyprenylalcohol composing the side chain of vitamin K2 (VK2), was previously reported to be a potent inducer of apoptosis in tumor cell lines (Ohzumi H et al, J Biochem 1995; 117: 11-13). We examined the apoptosis-inducing ability of VK2 (menaquinone 3 (MK3), MK4 and MK5) and its derivatives such as phytonadione (VK1), as well as polyprenylalcohols with side chains of various lengths including farnesol (C15-OH; FO), geranylgeraniol (C20-OH; GGO), and geranylfarnesol (C25-OH; GFO) toward leukemia cells in vitro. MK3, MK4, MK5 and GFO (at 10 microM) showed a potent apoptosis-inducing activity for all freshly isolated leukemia cells tested and for leukemia cell lines such as NB4, an acute promyelocytic leukemia (APL)-derived cell line and MDS92, a cell line derived from a patient with myelodysplastic syndrome, although there were some differences depending on the cells tested. In contrast, VK1 showed no effect on any of the leukemia cells. The combination of MK5 plus all-trans retinoic acid (ATRA) resulted in enhanced induction of apoptosis in both freshly isolated APL cells and NB4 cells as compared to each reagent alone. These data suggest the possibility of using VK2 and its derivatives for the treatment of myelogenous leukemias, including APL.

Inhibitory effect of a nucleoside analog, acyclovir, on leukemia cells.

Acyclovir (ACV), a nucleoside analog, has been demonstrated previously to suppress selectively the proliferation of NIH3T3 fibroblastic cells transformed by either v-abl or bcr-abl gene transfection. From a viewpoint of clinical application of ACV, we investigated whether ACV inhibited the growth of leukemia cells expressing either p210 BCR-ABL or p185BCR-ABL. Acyclovir exerted an inhibitory effect on OM9;22 cells, p185BCR-ABL expressing cells, in a dose-dependent manner. Despite no down-modulation of a BCR-ABL tyrosine kinase activity or its expression was observed after treatment with ACV, cell cycle analysis demonstrated synchronization of OM9;22 cells at the G0/G1 phase. This suggests that, although ACV does not directly act on BCR-ABL tyrosine kinase, ACV may exert its inhibitory effect on some leukemia cell lines via alterations of the cell cycle. Although selective inhibition of Philadelphia chromosome-positive leukemia cell growth was not apparent, our data provides a therapeutic possibility for ACV in the treatment for leukemia.

Immobilized anti-KIT monoclonal antibody induces ligand-independent dimerization and activation of Steel factor receptor: biologic similarity with membrane-bound form of Steel factor rather than its soluble form.

Interaction of a tyrosine kinase type receptor and its ligand induces receptor-dimerization or -oligomerization followed by transphosphorylation and activation of its intrinsic kinase, which leads to a series of intracellular signals. We have previously reported that the membrane-bound form of Steel factor (SLF) induces more persistent tyrosine kinase activation and longer life span of c-kit encoded protein (KIT) than its soluble form (Miyazawa et al, Blood 85:641, 1995). In this study, we used YB5.B8 monoclonal antibody (MoAb) that recognizes the extracellular domain of KIT to investigate whether immobilized anti-KIT MoAb can substitute for SLF as a potent activator of KIT by cross-linking receptors and further compared its effect with each SLF isoform in a factor-dependent cell line M07e. YB5.B8 MoAb in a soluble state suppressed SLF-induced M07e cell proliferation in a dose-dependent manner. By contrast, once this antibody was immobilized on the goat-antimouse MoAb (GAM)-coated culture plates, it supported the growth of M07e cells in the absence of any growth factors, whereas culture the cells in GAM alone or YB5.B8 without GAM-coated plates resulted in rapid cell-death within 24 hours. As with the natural ligand SLF, immobilized YB5.B8 MoAb synergized with granulocyte-macrophage colony-stimulating factor (GM-CSF) in inducing cell proliferation compared with either YB5.B8 MoAb or GM-CSF alone. Immunoblotting with antiphosphotyrosine MoAb showed that interaction of M07e cells with immobilized YB5.B8 induced tyrosine phosphorylation of a series of intracellular proteins including KIT (145 kD). In addition, cross-linking studies using a water-soluble cross linking reagent bis-sulfosuccinimidyl-suberate showed that immobilized YB5.B8 MoAb induced dimerization and activation of KIT. However, as with stimulation by the membrane-bound form of SLF, the kinetics of KIT activation with YB5.B8 MoAb was more prolonged compared with the cells treated with recombinant soluble SLF. Flow cytometry showed that, unlike the cells treated with soluble SLF, no downmodulation of cell-surface KIT expression was observed in M07e cells cultured with immobilzed YB5.B8 MoAb. These data suggest that immobilized antibodies against hematopoietic receptors may replace their ligand-stimulators; however, their activities may resemble the membrane-bound form rather than the soluble form of natural ligands.

[Aplastic anemia associated with ovarian bleeding].

A 22-year-old female was admitted on June 19, 1993 with pancytopenia. Aplastic anemia was diagnosed and she was treated with anti-thymocyte globulin. After one month, she started to feel abdominal pain. Her platelet count was 1.3 x 10(4)/microliters at that time. Computed tomography revealed vast accumulation of blood in the abdomen and pelvis. A total of 1,000 ml blood was obtained from the right ovarium and she underwent right partial oophorectomy. Although ovarian bleeding is rarely reported in aplastic anemia, we should always keep in mind this possibility when abdominal pain occurs in a female patient.

Membrane-bound Steel factor induces more persistent tyrosine kinase activation and longer life span of c-kit gene-encoded protein than its soluble form.

Alternative splicing of exon 6 results in the production of two isoforms of Steel factor (SLF): the membrane-bound and soluble forms. To investigate differences in the kinetics of c-kit tyrosine kinase activated by these two isoforms, we used a stromal cell line (SI/SI4) established from SI/SI homozygous murine embryo fetal liver and its stable transfectants containing either hSCF248 cDNA (including exon 6; secreted form) or hSCF220 cDNA (lacking exon 6; membrane-bound form) as the source of each isoform. Interaction of factor dependent myeloid cell line MO7e with stromal cells producing either isoform resulted in activated c-kit tyrosine kinase and induction of the same series of tyrosine phosphorylated cellular proteins in MO7e cells. However, SI4-h220 (membrane-bound form) induced more persistent activation of c-kit kinase than SI4-h248 (soluble form) did. Flow cytometric analysis and pulse-chase studies using [35S]methionine showed that SI4-h248 induced rapid downmodulation of cell-surface c-kit expression and its protein degradation in MO7e cells, whereas SI4-h220 induced more prolonged life span of c-kit protein. Addition of soluble recombinant human SLF to SI4-h220 cultures enhanced reduction of cell-surface c-kit expression and its protein degradation. Because the kinetics of c-kit inactivation strikingly fits with the protein degradation rates of c-kit under the conditions described above, rapid proteolysis of c-kit protein induced by soluble SLF stimulation may function as a "turn-off switch" for activated c-kit kinase.

Complementary DNA encoding the human T-cell FK506-binding protein, a peptidylprolyl cis-trans isomerase distinct from cyclophilin.

The recently discovered macrolide FK506 has been demonstrated to have potent immunosuppressive activity at concentrations 100-fold lower than cyclosporin A, a cyclic undecapeptide that is used to prevent rejection after transplantation of bone marrow and organs, such as kidney, heart, and liver. After the recent discovery that the cyclosporin A-binding protein cyclophilin is identical to peptidylprolyl cis-trans isomerase, a cellular binding protein for FK506 was found to be distinct from cyclophilin but to have the same enzymatic activity. In this study, we isolated a cDNA coding for FK506-binding protein (FKBP) from human peripheral blood T cells by using mixed 20-mer oligonucleotide probes synthesized on the basis of the sequence, Glu-Asp-Gly-Lys-Lys-Phe-Asp, reported for bovine FKBP. The DNA isolated contained an open reading frame encoding 108 amino acid residues. The first 40 residues of the deduced amino acid sequence were identical to those of the reported amino-terminal sequence of bovine FKBP, indicating that the DNA sequence isolated represents the gene coding for FKBP. Computer-assisted analysis of the deduced amino acid sequence indicates that FKBP exhibits no internal homology and does not have significant sequence similarity to any other amino acid sequences of known proteins, including cyclophilin. This result suggests that two catalytically similar proteins, cyclophilin and FKBP, evolved independently. In Northern blot analysis, mRNA species of approximately 1.8 kilobases that hybridized with human FKBP cDNA were detected in poly(A)+ RNAs from brain, lung, liver, and placental cells and leukocytes. Induction of Jurkat leukemic T cells with phorbol 12-myristate 13-acetate and ionomycin did not affect the level of FKBP mRNA. Southern blot analysis of human genomic DNA digested with different restriction enzymes suggests the existence of only a few copies of the DNA sequence encoding FKBP. This is in contrast to the result that as many as 20 copies of the cyclophilin gene and possible pseudogenes may be present in the mammalian genome.

Immunogenicity of a recombinant vaccinia virus expressing envelope a glycoprotein of bovine leukaemia virus.

We constructed a recombinant vaccinia virus (RVV) expressing envelope (env) glycoprotein (gp51) of bovine leukaemia virus (BLV): the expression of gp51 was detected by Western blot of the lysates of rabbit kidney cells infected with the RVV. The rabbits inoculated intradermally with the RVV alone failed to induce detectable anti-gp51 antibodies even 10 weeks after immunization. However, when these animals were boosted with inactivated BLV virion in saline, significant levels of anti-gp51 antibodies were induced as shown both in Western blot and immunodiffusion analyses. In these animals, antibodies against gag product (p24) were not detected. On the other hand, the rabbits inoculated with wild-type vaccinia virus and boosted similarly three times with the BLV virion in saline did not induce detectable anti-gp51 antibodies at all. The present experiment revealed that the RVV possessed the capability to endow immunological memory without inducing apparent anti-gp51 antibody responses, meaning that the RVV activated helper T cells far more strongly than B cells. The applicability of the RVV to vaccine development is discussed.

Increased resistance to the anticellular effect of interferon in an ultraviolet light-resistant human cell line, UVr-1.

Interferon (alpha, beta and gamma) susceptibility was tested in a human cell line, UVr-1, a u.v. light-resistant variant of RSa cells; the latter have high sensitivity to both u.v. lethality and the cell proliferation inhibition (anticellular) effect of human interferon (HuIFN) preparations. UVr-1 cells were less sensitive than the parental RSa cells to the inhibitory effects of HuIFN preparations, as measured by cell proliferation and the incorporation of [3H]deoxythymidine and [3H]deoxyadenosine into acid-insoluble cellular material. Nevertheless, UVr-1 cells exposed to HuIFN showed almost the same enhanced levels of antiviral activity and pppA(2'p5'A)n synthetase activity as similarly treated RSa cells. Further, UVr-1 cells had much the same binding capacity for 125I-labelled HuIFN-alpha A. Thus, it seems likely that the variant has an increased resistance to the anticellular effect but not to the antiviral effect of HuIFN preparations. UVr-1 cells showed no significant difference from RSa cells in u.v.-induced DNA repair synthesis. However, when a comparison was made between the susceptibility of normal fibroblasts and fibroblasts from patients with Cockayne's syndrome, characterized by an altered u.v. sensitivity but no alteration of DNA repair replication synthesis, the Cockayne's syndrome fibroblasts, CCK-3 and CCK-4, were more susceptible to HuIFN-beta as judged by cell proliferation and deoxythymidine incorporation tests.

Cross-sensitivity between interferon and uv in human cell strains: IFr, HEC-1, and CRL1200.

Correlation between susceptibility to the anticellular effect of human interferon (HuIFN) and ultraviolet (uv) lethality was examined in a set of isogeneous human cell lines (RSa and IFr cells), a human endometrial cancer cell line (HEC-1 cells), and a Xeroderma pigmentosum-derived fibroblast cell line ( CRL1200 cells). IFr cells, previously established as a HuIFN-alpha-resistant subline by exposing HuIFN-alpha- and uv-sensitive RSa cells to HuIFN-alpha preparations, appeared more refractory to uv than did the parent RSa cells in the cell proliferation and colony-formation studies. The extent of recovery from uv-inhibited total cellular DNA synthesis and uv-induced DNA-repair synthesis was enhanced to a greater extent in IFr cells than in RSa cells. The preexposure of RSa cells to HuIFN-alpha enhanced uv-induced DNA-repair replication activities. HEC-1 cells, which are reportedly totally refractory to HuIFN actions, appeared most resistant to uv, in all the tests. The uv-sensitive CRL1200 cells appeared highly susceptible to HuIFN-beta, in both cell proliferation and DNA-synthesis inhibition tests. These results support and extend our previous finding (N. Suzuki, J. Nishimaki , and T. Kuwata (1982). Mutat . Res. 106, 357-376) that susceptibility to the anticellular effect of HuIFN closely relates with uv lethality.

Characterization of a UV-resistant strain, UVr-10, established from a human clonal cell line, RSb, with high sensitivity to UV, 4NQO, MNNG and interferon.

Characterization was performed of a UV-resistant variant strain. UVr-10, derived from a human clonal cell line, RSb, with high sensitivity not only to the lethal effect of 254-nm far-ultraviolet (UV) irradiation but also to the effects of 4-nitroquinoline 1-oxide (4NQO) and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), and to the cell proliferation inhibition (CPI) effect of human leukocyte interferon (HuIFN-alpha) preparations. Colony-formation assays confirmed the increased resistance of UVr-10 cells to both UV and 4NQO, but no increased resistance to MNNG. The marked recovery from the inhibition of the total cellular DNA synthesis of UVr-10 cells, estimated by [methyl-3H]thymidine ([3H]dThd) uptake into the cellular DNA materials, was seen during 6 h after UV irradiation or 4NQO treatment even under the conditions without the recovery uptake into those of the parent RSb cells, but not during 6 h after MNNG treatment. Comparative studies on the activity of DNA repair synthesis between UVr-10 and RSb cells, by measuring the extent of UV-, 4NQO- or MNNG-induced unscheduled DNA synthesis (UDS) and DNA repair replication, revealed an increased activity of UVr-10 cells to UV and 4NQO but no significant increase of the activity to MNNG. These results suggest that increased DNA repair activities of a UVr-10 cell line may account for its becoming resistant to the lethal effect of UV and 4NQO. Concerning the CPI effect of HuIFN-alpha, UVr-10 cells showed increased resistance. Further, the DNA synthesis activity of UVr-10 cells was not so inhibited by HuIFN-alpha exposure as that of RSb cells. However, HuIFN-alpha-exposed UVr-10 cells showed more enhanced levels of activity of pppA(2'p5'A)n synthetase (2-5A synthetase) than the exposed RSb, thus suggesting that HuIFN-alpha could exert enough intracellular effect even in UVr-10 cells. The implication of the increased resistance of UVr-10 cells to the effects of UV, 4NQO and HuIFN-alpha, but not to those of MNNG, is discussed.