Associations between oral complications and days to death in palliative care patients.

PURPOSE: Adverse oral symptoms gradually appear in advanced cancer patients as the disease progresses. We retrospectively investigated the associations between the incidence of oral problems and the days to death (DTD) in patients receiving palliative care. METHODS: The dental assessment sheets and medical charts of 105 patients who had been admitted into the palliative care unit at our hospital were examined. Case data included evaluations of organic and functional oral conditions at the time of admission for all patients. The cohort was divided into two groups according to the DTD as the short group (<28 days from the time of dental assessment until death) and the long group (≥28 days). We compared the incidences of organic and functional oral problems between these groups. RESULTS: Dry mouth, tongue inflammation, and bleeding spots were significantly more frequent in the short group than in the long group (78 vs. 54% for dry mouth, 67 vs. 46% for tongue inflammation, 35 vs. 14% for bleeding spots, respectively; p < 0.05). Tongue coating and candidiasis were comparable between the two groups. Dysphagia was significantly more common in the short group (43%) than in the long group (20%) (p = 0.01), as was assistance with oral health care (76 vs. 50%) (p = 0.01). CONCLUSIONS: Our findings suggest that, during palliative care, oral complications appear more frequently when the DTD period is shorter. These symptoms may be useful indicators when deciding on the proper timing of intensive oral care intervention to decrease oral problems and pain in terminally ill patients.

CD26 expression on T cell lines increases SDF-1-alpha-mediated invasion.

BACKGROUND: CD26 is a multifunctional membrane-bound glycoprotein that regulates tumour growth in addition to its other activities. Because disease aggressiveness is correlated with CD26 expression in several T-cell malignancies, we decided to investigate the invasiveness of cells expressing different levels of CD26. METHODS: To assess CD26 involvement in cell invasion, we performed in vitro invasion assays with human T cell lines expressing different levels of CD26. These included the parental CD26-positive T-lymphoblast cell line HSB-2 and clones infected with a retrovirus expressing siRNA vectors that either targeted CD26 or encoded a missense siRNA, and the parental CD26-negative T-leukaemia cell line Jurkat and clones expressing CD26. CD26 expression in these cell lines was evaluated by flow cytometry and western immunoblotting. CXCR4 expression, phosphorylation of signalling kinases, and MMP-9 secretion were also evaluated by western immunoblotting, whereas MMP-9 activity and the effect of kinase and CD45 inhibitors on activity were measured by zymography of conditioned media. RESULTS: The presence of CD26 enhanced stromal-cell-derived factor-1-alpha (SDF-1-alpha)-mediated invasion of T cell lines. This process was regulated in part by the PI-3K and MEK1 pathways, as indicated by increased phosphorylation of p44/42 MAP kinase and Akt in the presence of SDF-1-alpha and the effect of their respective inhibitors on MMP-9 secretion and in vitro invasion. In addition, CD26-associated enhancement of SDF-1-alpha-induced invasion was decreased when CD45 was inhibited. CONCLUSIONS: Our results indicate that the expression of CD26 in T cell lines leads to increased SDF-1-alpha-mediated invasion in an in vitro system and that this is controlled in part by the PI-3K and MEK1 pathways. The data also suggest that CD26 enhancement of invasion may be mediated by CD45, however, more studies are required to confirm this involvement.

Crk-associated substrate lymphocyte type regulates transforming growth factor-beta signaling by inhibiting Smad6 and Smad7.

Crk-associated substrate lymphocyte type (Cas-L) is a 105 kDa docking protein with diverse functional properties, including regulation of cell division, proliferation, migration and adhesion. Cas-L is also involved in beta1 integrin- or antigen receptor-mediated signaling in B and T cells. In the present study, we demonstrate that Cas-L potentiates transforming growth factor-beta (TGF-beta) signaling pathway by interacting with Smad6 and Smad7. Immunoprecipitation experiments reveal that single domain deletion of full-length Cas-L completely abolishes its docking function with Smad6 and Smad7, suggesting that the natural structure of Cas-L is necessary for its association with Smad6 and Smad7. On the other hand, both N-terminal and C-terminal deletion mutants of Smad6 and Smad7 still retain their docking ability to Cas-L, suggesting that Smad6 and Smad7 possess several binding motifs to Cas-L. Moreover, Cas-L interaction with Mad-homology (MH)2 domain, but not with MH1 domain of Smad6 or Smad7, ameliorates TGF-beta-induced signaling pathway. Finally, depletion of Cas-L by small-interfering RNA oligo attenuates TGF-beta-induced growth inhibition of Huh-7 cells, with a concomitant reduction in phosphorylation of Smad2 and Smad3. These results strongly suggest that Cas-L is a potential regulator of TGF-beta signaling pathway.

Close correlation of 1-beta-D-arabinofuranosylcytosine 5'-triphosphate, an intracellular active metabolite, to the therapeutic efficacy of N(4)-behenoyl-1-beta-D-arabinofuranosylcytosine therapy for acute myelogenous leukemia.

N(4)-Behenoyl-1-beta-D-arabinofuranosylcytosine (BHAC), a prodrug of 1-beta-D-arabinofuranosylcytosine, is used effectively for the treatment of leukemia in Japan. BHAC therapy may be more effective if it is delivered in conjunction with monitoring of 1-beta-D-arabinofuranosylcytosine 5'-triphosphate (ara-CTP), the intracellular active metabolite of ara-C derived from BHAC. However, previous monitoring methods for ara-CTP were insufficiently sensitive. Here, using our new sensitive method, we evaluated the ara-CTP pharmacokinetics in relation to the therapeutic response in 11 acute myelogenous leukemia patients who received a 2-h infusion of BHAC (70 mg / m(2)) in combination remission induction therapy. ara-CTP could be monitored at levels under 1 mM. BHAC maintained effective levels of plasma ara-C and intracellular ara-CTP for a longer time, even compared with historical values of high-dose ara-C. The area under the concentration-time curve of ara-CTP was significantly greater in the patients with complete remission than in the patients without response. This greater amount of ara-CTP was attributed to the higher ara-CTP concentrations achieved in the responding patients. There was no apparent difference of plasma ara-C pharmacokinetics between the two groups. Thus, for the first time, the ara-CTP pharmacokinetics was evaluated in relation to the therapeutic effect of BHAC, and the importance of ara-CTP was proven. Administration of optimal BHAC therapy may require monitoring of the ara-CTP pharmacokinetics in each individual patient.

Antiproliferative activity of ecteinascidin 743 is dependent upon transcription-coupled nucleotide-excision repair.

While investigating the novel anticancer drug ecteinascidin 743 (Et743), a natural marine product isolated from the Caribbean sea squirt, we discovered a new cell-killing mechanism mediated by DNA nucleotide excision repair (NER). A cancer cell line selected for resistance to Et743 had chromosome alterations in a region that included the gene implicated in the hereditary disease xeroderma pigmentosum (XPG, also known as Ercc5). Complementation with wild-type XPG restored the drug sensitivity. Xeroderma pigmentosum cells deficient in the NER genes XPG, XPA, XPD or XPF were resistant to Et743, and sensitivity was restored by complementation with wild-type genes. Moreover, studies of cells deficient in XPC or in the genes implicated in Cockayne syndrome (CSA and CSB) indicated that the drug sensitivity is specifically dependent on the transcription-coupled pathway of NER. We found that Et743 interacts with the transcription-coupled NER machinery to induce lethal DNA strand breaks.

Monitoring of intracellular 1-beta-D-arabinofuranosylcytosine 5'-triphosphate in 1-beta-D-arabinofuranosylcytosine therapy at low and conventional doses.

1-beta-D-Arabinofuranosylcytosine (ara-C) is used empirically at a low, conventional, or high dose. Ara-C therapy may be optimal if it is directed by the clinical pharmacokinetics of the intracellular active metabolite of ara-C, 1-beta-D-arabinofuranosylcytosine 5'-triphosphate (ara-CTP). However, ara-CTP has seldom been monitored during low- and conventional-dose ara-C therapies because detection methods were insufficiently sensitive. Here, with the use of our newly established method (Cancer Res., 56, 1800 -- 1804 (1996)), ara-CTP was monitored in leukemic cells from acute myelogenous leukemia patients receiving low- or conventional-dose ara-C [subcutaneous ara-C administration (10 mg / m(2) ) (3 patients), continuous ara-C infusion (20 or 70 mg / m(2) / 24 h) (7 patients), 2-h ara-C infusion (70 mg / m(2) ) (4 patients), and 2-h infusion of N(4)-behenoyl-1-beta-D-arabinofuranosylcytosine, a deaminase-resistant ara-C derivative (70 mg / m(2) ) (6 patients)]. Ara-CTP could be determined at levels under 1 microM. There was a close correlation between the elimination half-life values of the plasma ara-C and the intracellular ara-CTP. The presence of ara-C in the plasma was important to maintain ara-CTP. The continuous ara-C and the 2-h N(4)-behenoyl-1-beta-D-arabinofuranosylcytosine infusions maintained ara-CTP and the plasma ara-C longer than the subcutaneous ara-C or the 2-h ara-C infusion. They also afforded relatively higher ara-CTP concentrations, and consequently produced ara-CTP more efficiently than the 2-h ara-C infusion. Different administration methods produced different quantities of ara-CTP even at the same dose.

Characterization of a novel topoisomerase I mutation from a camptothecin-resistant human prostate cancer cell line.

In this study, we characterized the structure and function of topoisomerase I (top1) protein in the camptothecin (CPT)-resistant prostate cancer cell lines, DU-145/RC0.1 and DU-145/RC1 (RC0.1 and RC1, respectively). Both of the cell lines were previously selected by continuous exposure to 9-nitro-CPT. The RC0.1 and RC1 cells have high cross-resistance to CPT derivatives including SN-38 and topotecan, but are not cross-resistant to the non-top1 inhibitors etoposide, doxorubicin, and vincristine. Although the top1 protein levels were not decreased in the resistant cells compared with the parental cells, CPT-induced DNA cleavage was markedly reduced in the RC0.1 and RC1 nuclear extracts. The resistant-cell-line nuclear extracts also demonstrated top1 catalytic activity and resistance to CPT, in in vitro assays. Reverse transcription-PCR products from the resistant cell lines were sequenced, and revealed a point mutation resulting in a R364H mutation in the top1 of both RC0.1 and RC1. No wild-type top1 RNA or genomic DNA was detected in the resistant cell lines. Using a purified recombinant R364H top1, we found that the R364H mutant top1 was CPT resistant and fully active. In the published top1 crystal structure, the R364H mutation is close to the catalytic tyrosine and other well-known mutations leading to CPT resistance.

Use of camptothecin-resistant mammalian cell lines to evaluate the role of topoisomerase I in the antiproliferative activity of the indolocarbazole, NB-506, and its topoisomerase I binding site.

NB-506 is a topoisomerase I (top1) inhibitor in clinical trials. In this study, we used a series of camptothecin (CPT)-resistant cell lines with known top1 alterations. We show that three mutations in different domains of the top1 enzyme that confer CPT resistance also confer cross-resistance to NB-506. The CPT-resistant cell lines and corresponding mutations were: human prostate carcinoma cells DU-145/RC1 (mutation R364H), Chinese hamster fibroblasts DC3F/C10 (mutation G503S), and human leukemia CEM/C2 cells (N722S). This result suggests that NB-506 and CPT share a common binding site in the top1-DNA complex. We next used these three cell lines and their parental cells to study the relationship between top1 poisoning by NB-506 and antiproliferative activity. We found that the CPT-resistant cells were only 2-10-fold resistant to NB-506, which suggests that NB-506 targets other cellular processes/pathways besides top1. This conclusion was further supported by the limited cross-resistance of top1-deficient murine leukemia P388/CPT45 cells (2-fold). Cross-resistance was also limited for J-109,382, an isomer of NB-506 that does not intercalate into DNA, indicating that the non-top1-mediated antiproliferative activity of NB-506 is not attributable to DNA intercalation. Together, these data indicate that NB-506 and indolocarbazoles are promising agents to overcome CPT resistance.

Ecteinascidin 743 induces protein-linked DNA breaks in human colon carcinoma HCT116 cells and is cytotoxic independently of topoisomerase I expression.

Ecteinascidin 743 (Et743; NSC 648766) is a potent antitumor agent presently in clinical trials. Et743 selectively alkylates guanine N2 from the minor groove of duplex DNA and bends the DNA toward the major groove. This differentiates Et743 from other DNA-alkylating agents presently in the clinic. To date, the cellular effects of Et743 have not been elucidated. Recently, Et743 DNA adducts have been found to suppress gene expression selectively and to induce topoisomerase I (top1) cleavage complexes in vitro and top1-DNA complexes in cell culture. In the present study, we characterized the DNA damage and the cell cycle response induced by Et743 in human colon carcinoma HCT116 cells. Alkaline elution experiments demonstrated that micromolar concentrations of Et743 produced comparable frequencies of DNA-protein cross-links and DNA single-strand breaks. The single-strand breaks were protein-cross-linked and were not associated with detectable DNA double-strand breaks. By contrast with camptothecin, these lesions persisted for several hours after drug removal and were not formed at 4 degrees C. Et743 treatment induced transient p53 elevation, dose-dependent cell cycle accumulation in G2-M and in G1- and S-phase, and inhibition of DNA synthesis. The sensitivity of camptothecin-resistant mouse leukemia P388/ CPT45 cells, which fail to express detectable top1, was similar to the sensitivity of wild-type P388 cells, suggesting that top1 is not a critical target for the antiproliferative activity of Et743.

Simultaneous treatment with 1-beta-D-arabinofuranosylcytosine and daunorubicin induces cross-resistance to both drugs due to a combination-specific mechanism in HL60 cells.

We have established a human myelogenous leukemia cell line (HL60/AD) that is 10-fold cross-resistant to both 1-beta-D-arabinofuranosylcytosine (ara-C) and daunorubicin; the cell line was isolated from HL60 by simultaneous treatment with these two agents at low drug concentrations attainable in clinical trials. HL60/AD was found to have multiple resistance mechanisms. With regard to ara-C, HL60/AD cells showed decreased deoxycytidine kinase activity but did not show elevation of cytidine deaminase activity or a decrease in ara-C influx. With regard to daunorubicin, a decrease in topoisomerase II activity was found. A decrease in intracellular accumulation of daunorubicin was also found. P-glycoprotein was not detected, but the multidrug resistance-associated protein was expressed. Furthermore, an increase of total cellular glutathione (GSH) content was found. Interestingly, the resistance of HL60/AD cells not only to daunorubicin but also to ara-C was markedly reversed by treatment with L-buthionine-(S,R)-sulfoximine (BSO), a potent inhibitor of GSH synthesis. After exposure of HL60/AD to ara-C, mitochondrial membrane potential and reactive oxygen intermediates showed no significant change, but a considerable loss of mitochondrial membrane potential and an increase in reactive oxygen intermediate generation were caused by pre-incubation with BSO. Neither elevation of GSH nor reversal of resistance by BSO was found in ara-C-resistant HL60 cells that were selected only with ara-C. These findings suggest that in addition to the summation of the mechanisms of resistance to each agent reported previously, an increased level of GSH plays an important role in the cross-resistance induced in HL60/AD cells by simultaneous exposure to both drugs.

Topoisomerase I-mediated cytotoxicity of N-methyl-N'-nitro-N-nitrosoguanidine: trapping of topoisomerase I by the O6-methylguanine.

Alkylating agents such as N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) are known to covalently link alkyl groups at the position 6 of guanines (O6MG) in DNA. O6-alkylguanine-DNA alkyltransferase (AGT) specifically removes the methyl group of the O6MG. Using purified human topoisomerase I (Top1), we found an 8-10-fold enhancement of Top1 cleavage complexes when O6MG is incorporated in oligonucleotides at the +1 position relative to a unique Top1 cleavage site. Top1 poisoning by O6MG is attributable to a decrease of the Top1-mediated DNA religation as well as an increase in the enzyme cleavage step. Increased cleavage is probably linked to a change in the hydrogen bonding pattern, such as in the case of the 8-oxoguanine, whereas inhibition of religation could be attributed to altered base pairing, such as abasic sites or base mismatches, because incorporation of a 6-thioguanine did not affect Top1 activity. Top1-DNA covalent complexes are also induced in MNNG-treated CHO cells constitutively lacking the AGT enzyme. Conversely, no increase could be detected in CHO cells transfected with the wild-type human AGT. Moreover, we show that yeasts overexpressing the human Top1 are more sensitive to MNNG, whereas knock-out Top1 strain cells display some resistance to the drug. Altogether, these results suggest a role for Top1 poisoning by alkylated bases in the antiproliferative activity of alkylating agents as well as in the DNA lesions resulting from endogenous and carcinogenic DNA modifications.

Long-term follow-up of the clinical efficacy of chemotherapy for acute myeloid leukemia at a single institute.

A retrospective study was performed on 125 patients with de-novo acute myeloid leukemia (AML) who had received first remission induction therapy at Fukui Medical University Hospital in the 16 years between 1983 and 1998. For remission induction therapies, patients in the 1980s received mainly behenoylcytarabine (BHAC), 6-mercaptopurine (6-MP), and prednisolone (PSL), plus aclarubicin (ACR) or daunorubicin (DNR). Patients in the 1990s received mainly BHAC, 6-MP, and etoposide (VP-16) plus DNR or mitoxantrone (MIT) or idarubicin (IDA). Patients with hypoplastic bone marrow received low-dose cytarabine (Ara-C) therapy or cytarabine ocfosfate (SPAC). Since 1992, patients with French-American-British disease classification of M3 have received all-trans retinoic acid (ATRA) (+/-chemotherapy). In the 1990s, more intensified postremission therapy was performed compared with that done in the 1980s. The complete remission (CR) rate of all patients was 58%. Predicted 6-year overall survival (OS) and disease-free survival (DFS) rates in the CR patients were 22% and 28%, respectively. Multivariate analysis showed age and leukocyte counts as significant prognostic factors regarding CR, OS, and DFS rates. The CR and OS rates in the 1990s were improved significantly from those in the 1980s, at 69% versus 48% (P = 0.016), and 32% versus 15% (P = 0.0014), respectively. The early death rate, within 30 days, was decreased from 26% in the 1980s to 9% in the 1990s (P = 0.013). This decrease was thought to be the main cause of the high CR rate in the 1990s. However, DFS was not significantly improved. It is necessary to establish more effective postremission therapies in order to reduce the relapse rate and improve the prognosis.

Activity of a novel camptothecin analogue, homocamptothecin, in camptothecin-resistant cell lines with topoisomerase I alterations.

Homocamptothecin (hCPT), which differs from camptothecin (CPT) by the presence of an additional methylene group in the E-ring, was evaluated in CPT-resistant cell lines. Topoisomerase I (top1)-deficient leukemia P388/CPT45 cells were highly resistant to hCPT, which demonstrates that top1 is the primary target of hCPT. Three CPT-resistant cell lines with top1 point mutations (Chinese hamster lung fibroblast DC3F/C10, human prostate carcinoma DU-145/RC1, and human leukemia CEM/C2) and their top1 enzymes were cross-resistant to hCPT. The antiproliferative activity of hCPT was greater than that of CPT in both parental and CPT-resistant cell lines, particularly in the prostate cell lines. The top1 cleavage complexes formed in the presence of hCPT appear to be more stable than those induced by CPT. Together, these data indicate that hCPT is a specific top1 inhibitor, which shares a common binding site with CPT in the topl-DNA cleavage complexes. Because of its potency, hCPT might overcome resistance to CPT in some cancer cells.

Effect of PSC 833 on the cytotoxicity and pharmacodynamics of mitoxantrone in multidrug-resistant K562 cells.

We examined the effect of PSC 833, a nonimmunosuppressive cyclosporin analogue, on the cytotoxicity, accumulation and retention of an anthraquinone antileukemia drug mitoxantrone (MIT). This was done in P-glycoprotein (PGP)-overexpressing multidrug-resistant K562/D1-9 cells and compared with the effect of cyclosporin A (CsA). We also compared MIT with the effect of PSC 833 on the cytotoxicity of daunorubicin (DNR) and doxorubicin (DOX). While PSC 833 and CsA had no effect on the cytotoxicity, accumulation and retention of MIT in the parent K562 cells, PSC 833 and CsA restored accumulation and retention of MIT in K562/D1-9 cells dose-dependently. Consequently, there was increased sensitivity of K562/D1-9 cells to MIT. The reversing activity of PSC 833 on the cytotoxicity of MIT was stronger than that of CsA, and was almost the same as the reversing activity of PSC 833 on the cytotoxicity of DNR and DOX. The resistance index of MIT decreased from 43.9-fold to 2.8-fold by 0.4 microM PSC 833, which is a clinically achievable plasma concentration. These results suggest that the combination of PSC 833 with MIT could be a promising treatment in reversing PGP-mediated MDR in leukemia patients.

Induction of topoisomerase I cleavage complexes by 1-beta -D-arabinofuranosylcytosine (ara-C) in vitro and in ara-C-treated cells.

1-beta-d-Arabinofuranosylcytosine (Ara-C) is a nucleoside analog commonly used in the treatment of leukemias. Ara-C inhibits DNA polymerases and can be incorporated into DNA. Its mechanism of cytotoxicity is not fully understood. Using oligonucleotides and purified human topoisomerase I (top1), we found a 4- to 6-fold enhancement of top1 cleavage complexes when ara-C was incorporated at the +1 position (immediately 3') relative to a unique top1 cleavage site. This enhancement was primarily due to a reversible inhibition of top1-mediated DNA religation. Because ara-C incorporation is known to alter base stacking and sugar puckering at the misincorporation site and at the neighboring base pairs, the observed inhibition of religation at the ara-C site suggests the importance of the alignment of the 5'-hydroxyl end for religation with the phosphate group of the top1 phosphotyrosine bond. This study also demonstrates that ara-C treatment and DNA incorporation trap top1 cleavage complexes in human leukemia cells. Finally, we report that camptothecin-resistant mouse P388/CPT45 cells with no detectable top1 are crossresistant to ara-C, which suggests that top1 poisoning is a potential mechanism for ara-C cytotoxicity.

Enlargement of mandibular canal without hypesthesia caused by extranodal non-Hodgkin's lymphoma: a case report.

A rare condition of enlargement of the mandibular canal caused by an extra-nodal non-Hodgkin's lymphoma in a 59-year-old Japanese woman was reported. The patient had a swelling of the hard palate and protrusion of both ocular bulbs, which had been present for 10 years. A panoramic radiograph revealed that the right mandibular canal was widely enlarged, extending from the mandibular foramen to the mental foramen, without bone destruction. The continuous dilation of the mandibular canal to an approximate 15-mm width was associated with peripheral bony sclerosis. Computed tomography and magnetic resonance imaging showed a soft tissue tumor inside the mandibular canal. The lesion demonstrated expansive growth in the orbits, extending to the skull base through the superior orbital fissures and cavernous sinus. The lymphoma was suspected to have grown so slowly that the adjacent mandibular canal and ocular bulbs enlarged without destroying the normal bone and nervous tissue.

Successful treatment of cytomegalovirus retinitis in a patient with malignant lymphoma: a case report and review of the literature.

A 52-year-old Japanese woman was diagnosed as having angioimmunoblastic T-cell lymphoma (stage IV-B). She received 6 courses of chemotherapy including cyclophosphamide, doxorubicin, vincristine, and prednisolone every two weeks (biweekly CHOP), and was considered to be in partial remission. She complained of loss of visual acuity in her right eye during her last cycle of chemotherapy. Cytomegalovirus (CMV) retinitis was suspected from the characteristic ophthalmoscopic appearance. This diagnosis was further supported by the detection of CMV DNA in blood and antigens in polymorphonuclear leukocytes, a sign of CMV reactivation. Although DNAemia and antigenemia became negative, retinitis remained slightly active despite a 4-week systemic treatment of ganciclovir. Intraocular injection of ganciclovir was started and continued until the retinitis became inactive ophthalmoscopically. The patient received high-dose chemotherapy with peripheral blood stem cell transplantation and achieved complete remission. During the after this therapy no recurrence of CMV infections was observed. This case shows that 1) a quick and accurate diagnosis of CMV retinitis was possible by applying DNAemia and antigenemia and 2) intensive treatment for the CMV infection enabled the accomplishment of cure-oriented chemotherapy of the lymphoma without the recurrence of CMV retinitis.

Effect of PSC 833 on the cytotoxicity of idarubicin and idarubicinol in multidrug-resistant K562 cells.

We examined the effect of PSC 833, a non-immunosuppressive cyclosporin analogue, on the cytotoxicity, accumulation and retention of idarubicin (IDA) and its 13-dihydro metabolite, idarubicinol (IDAol). P-glycoprotein (PGP)-overexpressing multidrug-resistant K562/D1-9 cells were used for these studies. PSC 833 had no effect on the cytotoxicity, intracellular accumulation, or retention of IDA and IDAol in the parent K562 cells. However, intracellular accumulation of IDA and IDAol in K562/D1-9 cells after a 60-min incubation was restored by 0.4 microM PSC 833 to 104% and 116%, respectively, of the level in parent K562 cells. The retention of IDA and IDAol in K562/D1-9 cells was also restored by 0.4 microM PSC 833. Consequently, 0.4 microM PSC 833 increased the sensitivity of K562/D1-9 cells to IDA and IDAol. The resistance index (RI) of IDA decreased from 20-fold to 4.0-fold, and the RI of IDAol decreased from 104-fold to 1.5-fold. These results suggest that the combination of IDA and PSC 833 may be effective in reversing PGP-mediated multidrug resistance in leukemia cells.

Multidrug resistance due to impaired DNA cleavage in a VP-16-resistant human leukemia cell line.

We established a VP-16-resistant line of human leukemia cells, K562/VP-H2, derived from K562 cells. K562/VP-H2 cells were 44-fold more resistant to VP-16 than were K562 cells. K562/VP-H2 cells were also resistant to doxorubicin, daunorubicin and mitoxantrone, but showed little or no resistance to vincristine, aclarubicin, idarubicin, idarubicinol, cytosine arabinoside, cis-platinum or camptothecin. K562/VP-H2 cells did not over-express P-glycoprotein or multidrug resistance protein, and showed intracellular accumulation of VP-16 similar to that in K562 cells. While the expressions of topoisomerase II-alpha gene and topoisomerase II-beta gene, or catalytic activity in nuclear extract of K562/VP-H2 cells were similar to that of K562 cells, the VP-16 induced DNA cleavage was reduced in K562/VP-H2 cells compared to K562 cells, suggesting that the reduction of topoisomerase II-mediated DNA cleavage through qualitative alteration of topoisomerase II may be the main mechanism of acquired multidrug resistance for K562/VP-H2 cells. The K562/VP-H2 cell line is an interesting model for studying resistance to antileukemia drugs targeting topoisomerase II.