Effect of fludarabine and arabinosylcytosine on multidrug resistant cells.

BACKGROUND AND OBJECTIVE: Anthracyclines are first-line drugs in the treatment of acute leukemia, but the sensitivity of leukemic cells to anthracyclines can be downmodulated by multidrug resistance (MDR) transport proteins like Pgp. Pgp overexpression is negatively related to treatment response. Alternative drugs may be required to overcome the MDR problem. METHODS: Arabinosylcytosine (ara-C) and 9-beta-D-arabinofuranosyl-2-fluoro-adenine monophosphate (fludarabine, F-ara) were tested alone and in combination in four pairs of leukemia and tumor non-MDR and MDR cell lines. Toxicity was assayed by growth inhibition with the microcultured MTT assay. RESULTS: MDR cells were more sensitive than or as sensitive as non-MDR cells to ara-C and to F-ara alone. The resistance index to ara-C was decreased upon pre-exposure of the MDR cells to low-dose F-ara (10 ng/mL), showing that the combination of ara-C and F-ara was more active on MDR cells than on non-MDR parental ones. INTERPRETATION AND CONCLUSIONS: Neither sensitivity to ara-C nor sensitivity to F-ara was influenced by Pgp overexpression. These data provide a rationale for more extensive and more intensive testing of combinations of ara-C and F-ara in Pgp-mediated MDR acute leukemia. In relapsed/resistant and in secondary acute leukemias, increasing the dose of ara-C and combining ara-C with F-ara might be more rewarding than administering anthracyclines or other Pgp-processable compounds.

Overcoming PGP-related multidrug resistance. The cyclosporine derivative SDZ PSC 833 can abolish the resistance to methoxy-morpholynil-doxorubicin.

BACKGROUND: The results obtained so far in studies designed to neutralize P glycoprotein (PGP)-related multidrug resistance (MDR) by using MDR reversal agents, have not yet fulfilled the promise of the experiments which were performed in vitro. In order to improve PGP-related MDR neutralization, we tested in vitro the activity of the cyclosporine derivative SDZ PSC 833 (PSC) together with doxorubicin (DOX) and with two new DOX derivatives named 4' iodo 4' deoxy-doxorubicin (IODODOX) and methoxy-morpholynil-doxorubicin (MMDOX, FCE 23762) using four different human cell lines and their multi-drug resistant variants. METHODS: Anthracycline toxicity was evaluated by using the MTT method after a 7-day culture with continuous exposure to the antitumor drugs with or without the addition of PSC. RESULTS: PSC significantly downmodulated the toxicity of all three anthracyclines in all the four cell systems. However, despite the great increase caused by PSC in the toxicity of DOX and a more modest effect on the toxicity of the two DOX derivatives, this MDR reversal agent could only completely block the PGP mediated MMDOX resistance whereas DOX refractoriness was only decreased. CONCLUSIONS: The combination of MMDOX or IODODOX with PSC 1.6 microM is more efficient than the combination of DOX plus PSC for the full reversion of PGP-mediated drug resistance. Careful clinical studies are required to evaluate if these associations can also effectively and safely neutralize MDR in vivo.

Plasma cell P170 expression and response to treatment in multiple myeloma.

BACKGROUND: Vincristine and anthracyclines are first-line agents for the treatment of multiple myeloma (MM). P170-related multidrug resistance (MDR) may influence the response to these drugs. MATERIALS AND METHODS: P170 expression of bone marrow plasma cells was assayed by immunocytochemistry (alkaline phosphatase anti-alkaline phosphatase technique) with the MRK-16 monoclonal antibody. A case was considered positive if one per cent or more of plasma cells stained as strongly as positive controls. RESULTS: Six of 17 (35%) cases in relapse and 18/72 (25%) at diagnosis were MDR positive. MDR positivity was not found in micromolecular MM and was significantly associated with the serum beta 2-microglobulin level. Response to treatments including dexamethasone, vincristine and doxorubicin, or idarubicin, or mitoxantrone was independent of MDR positivity (50% in positive cases vs. 56% in negative ones). CONCLUSIONS: The detection of P170 in bone marrow plasma cells with the currently available methodology is not likely to predict response to treatments that include vincristine, anthracyclines or mitoxantrone. Further studies are required to evaluate the relevance of P170-related MDR to the development of MM therapy.

MDR-related P170-glycoprotein modulates cytotoxic activity of homoharringtonine.

Homoharringtonine (HHT) is a new drug with antileukemic activity which is currently tested for treatment of acute and chronic leukemias, either alone or in combination with other agents. Since HHT showed a low efficacy in refractory and relapsed acute leukemia and in the blastic phase of chronic myeloid leukemia (CML) which are frequently characterized by an overexpresion of the multidrug resistance (MDR)-related P170-glycoprotein, we postulated a relationship between the poor antileukemic effect of HHT in these leukemias and the expression of P170-glycoprotein. For this reason, sensitive (LOVO109 and CEM) and MDR (LOVO DX and CEM VLB) cell lines were exposed to HHT with or without some MDR modifiers, namely, Cyclosporine A (CyA), the Cyclosporine derivative SDZ PSC 833 (PSC), and the D-isomer of Verapamil (DVRP). It was found that MDR cells were about 15 times more resistant to HHT than non-MDR cells, and that resistance to HHT was significantly decreased by all the MDR modifiers that were tested. This in vitro study showed that HHT belongs to the category of MDR-related drugs, like anthracyclines, vinca alkaloids, epipodophylline derivatives, and taxol.

Evaluation of the clinical relevance of the anionic glutathione-s-transferase (GST pi) and multidrug resistance (mdr-1) gene coexpression in leukemias and lymphomas.

By using RNA slot-blot technique, the frequency and the degree of GST pi and mdr-1 gene coexpression were investigated in 23 AML patients, 9 ALL, 9 CLL and 11 cases of NHL in an attempt to study their clinical and prognostic relevance. GST pi and mdr-1 levels were expressed as arbitrary units (U) with respect to the negative controls (U = 0), MCF7 and HL60 sensitive cell lines, and the positive controls (U = 10), MCF7/DOXO and HL60/DNR resistant cell lines. The concomitant GST pi/mdr-1 gene overexpression showed a negative prognostic value in the set of newly diagnosed AML pts (10 cases), furthermore higher GST pi and mdr-1 mRNA levels were averagely detected in the relapsed/resistant ALL pts (4 cases), and in CLL (7 cases) and NHL (8 cases) heavily pretreated patients who were unresponsive to chemotherapy and with a disease progression. These preliminary data show that two different mechanisms of drug resistance can be coexpressed at the same time in those leukemias and lymphomas with a clinically unfavourable course.

Overexpression of MDR-related p170 glycoprotein in chronic myeloid leukemia.

BACKGROUND AND METHODS: Philadelphia (Ph) positive chronic myeloid leukemia (CML) cannot be induced into a true remission with conventional chemotherapy. Blast cells and precursors obtained from 51 Ph+ CML cases were assayed for expression of the multidrug resistance (MDR)-associated glycoprotein (p170) by immunocytochemistry (APAAP) with the MRK-16 monoclonal antibody. RESULTS AND CONCLUSIONS: Positive cells were found in 11/17 cases in chronic phase (65%), in 4/8 cases in accelerated phase, and in 23/26 cases in blastic phase (89%). The proportion of positive cells, which ranged between less than 1% and 95%, was higher in blastic phase (mean 32 +/- 29.9) than in chronic phase (mean 3 +/- 5.3) (p = 0.006). These findings show that p170 overexpression is common in Ph+ CML, especially after progression to blastic phase, and suggest that p170-related MDR may contribute significantly to treatment failure.

p170-dependent multidrug resistance. Restoring full sensitivity to idarubicin with verapamil and cyclosporin A derivatives.

BACKGROUND: Cell sensitivity to anthracyclines and other drugs depends on several factors, including overexpression of a 170Kd transmembrane glycoprotein (P170) that enhances drug efflux from the cells. Since the result of treatment is negatively related to the expression of P170 in leukemia, malignant lymphoma and other tumors, it is important to investigate drugs and methods that can modify multidrug resistance (MDR). MATERIALS AND METHODS: Using an MTT-microcultured tetrazolium colorimetric method, we assayed sensitivity to daunorubicin (DNR) and to its 4-demethoxy derivative idarubicin (IDA) in two MDR cell lines (CEM VLB and LOVO DX) and in their respective non-MDR parental lines (CEM and LOVO 109), with and without three MDR modifiers, namely the D-isomer of verapamil (DVRP), cyclosporin A (CyA) and the new CyA derivative SDZ PSC 833. RESULTS: We showed that down-modulation of resistance with MDR modifiers was greater for DNR than for IDA in MDR cells. However, we also demonstrated that restoration of full sensitivity could only be achieved for IDA, not for DNR. DVRP and CyA in combination were more effective than either compound alone and could abolish P170-related resistance to IDA at concentrations of 1-2 microM and 1.6 microM, respectively. SDZ PSC 833 alone was even more effective and set MDR to zero at a concentration ranging between 0.8 and 1.6 microM. CONCLUSIONS: These data suggest that combinations of IDA and MDR modifiers may improve the results of cancer and leukemia treatment and that they are worth investigating in vivo, with attention to possible effects on drug pharmacokinetics and on normal tissue damage.

The expression of the multidrug resistance related glycoprotein in adult acute lymphoblastic leukemia.

BACKGROUND: More than 50% of adults with acute lymphoblastic leukemia (ALL) actually die with resistant leukemia. The multidrug resistance (MDR) related P170 glycoprotein may be involved in treatment failure. METHODS: P170 content of leukemic cells was assayed by immunocytochemistry with the MRK-16 monoclonal antibody in 41 consecutive cases of adult ALL (27 at onset and 14 at relapse). RESULTS: Positive cells were found in 11/14 cases at relapse and in 18/27 cases at onset. All cases with a high WBC, FAB L3, B-mature phenotype, and t(9; 22) were positive. In the 27 patients who were studied at onset and were treated with the same protocol, the overall failure rate was 44% in negative cases and 83% in positive cases. CONCLUSION: P170 overexpression may contribute to drug resistance and to chemotherapy failure in adult ALL.

D-verapamil downmodulates P170-associated resistance to doxorubicin, daunorubicin and idarubicin.

Verapamil (VRP) is an effective modulator of P170-associated multidrug resistance (MDR), but its clinical application is limited by cardiovascular side-effects. The D-isomer of VRP (D-VRP) is 10 times less active than the racemic mixture on the cardiovascular system, but retains a MDR modulating activity. Daunorubicin (DNR) and doxorubicin (DX) are two anthracyclines whose cytotoxicity is strongly related with the expression of P170, while their respective lipophylic derivatives idarubicin (IDA) and iododoxorubicin (IDX) are less P170-dependent. We studied the effect of D-VRP on intracellular retention and on the cytotoxicity of these four anthracyclines in two MDR cell systems (LOVO and CEM) by flow cytometry and by a microcultured tetrazolium colorimetric assay (MTT). We found that in MDR cells D-VRP increased intracellular anthracycline concentration and increased the cytotoxicity of DNR, IDA and DX but not of IDX. The effect of D-VRP was dose-related, but it was already consistent at D-VRP concentrations that can be readily maintained in vivo (2-3 microM). These data suggest that at a clinically tolerable concentration D-VRP can downmodulate the resistance to DNR and DX and can restore full sensitivity to IDA.