Background
Methods
Cell lines
AML patient samples
Ethical approval
P-gp, MRP1, MRP3 and BCRP expression analysis
P-gp, MRP1 and BCRP activity analysis
Cell viability study (MTT assay)
Statistical analysis
Results
Protein expression of P-gp, MRP1, MRP3 and BCRP in K562, HL60 and variant cell lines
P-gp, MRP, and BCRP activity in K562, HL60 and variant cell lines
Modulation of drug resistance in K562 and HL60 variant cell lines
DNR | Idarubicin | Mitox | HHT | Mylotarg | ||||||
---|---|---|---|---|---|---|---|---|---|---|
IC50 (μM) | RMF | IC50 (μM) | RMF | IC50 (μM) | RMF | IC50 (ng/ml) | RMF | IC50 (ng/ml) | RMF | |
K562
| ||||||||||
None | 0.28 ± 0.09 | 0.071 ± 0.026 | 0.60 ± 0.21 | 57.3 ± 6.5 | ||||||
Zosuquidar | 0.26 ± 0.09 | 1.1 | 0.071 ± 0.042 | 1.0 | 0.095 ± 0.029 | 6.3 | 58.4 ± 10.1 | 1.0 | ||
CsA | 0.23 ± 0.12 | 1.2 | 0.068 ± 0.039 | 1.0 | 0.14 ± 0.05 | 4.3 | 56.6 ± 4.2 | 1.0 | ||
K562/HHT40
| ||||||||||
None | 2.3 ± 0.2 | 0.27 ± 0.06 | 1.7 ± 0.2 | 197.6 ± 44.4 | ||||||
Zosuquidar | 0.66 ± 0.08 | 3.5 | 0.20 ± 0.07 | 1.4 | 0.33 ± 0.06 | 5.2 | 85.4 ± 2.9 | 2.3 | ||
CsA | 0.51 ± 0.08 | 4.5 | 0.24 ± 0.11 | 1.1 | 0.45 ± 0.06 | 3.8 | 91.3 ± 9.3 | 2.2 | ||
K562/HHT90
| ||||||||||
None | 3.6 ± 1.5 | 0.17 ± 0.11 | 1.9 ± 0.9 | 409.6 ± 70.2 | ||||||
Zosuquidar | 0.37 ± 0.14 | 9.7 | 0.12 ± 0.09 | 1.4 | 0.13 ± 0.05 | 14.6 | 80.4 ± 8.3 | 5.8 | ||
CsA | 0.32 ± 0.19 | 11.3 | 0.10 ± 0.06 | 1.7 | 0.25 ± 0.18 | 7.6 | 81.8 ± 8.3 | 5.0 | ||
K562/DOX
| ||||||||||
None | >50b
| 1.82 ± 0.17 | >50 | >720 | ||||||
Zosuquidar | 1.1 ± 0.4 | >45.5 | 0.21 ± 0.03 | 8.7 | 0.53 ± 0.10 | >94.3 | 88.8 ± 1.9 | >8.1 | ||
CsA | 10.5 ± 1.6 | >4.8 | 0.30 ± 0.10 | 6.1 | 4.29 ± 0.22 | >11.7 | 639.1 ± 70.1 | >1.1 | ||
HL60
| ||||||||||
None | 0.16 ± 0.03 | 0.053 ± 0.004 | 0.48 ± 0.22 | 40.5 ± 5.2 | 131.9 ± 72.3 | |||||
Zosuquidar | 0.13 ± 0.04 | 1.2 | 0.057 ± 0.008 | 0.9 | 0.09 ± 0.03 | 5.3 | 41.7 ± 3.1 | 1.0 | 83.8 ± 9.5 | 1.6 |
CsA | 0.10 ± 0.02 | 1.6 | 0.033 ± 0.008 | 1.6 | 0.13 ± 0.07 | 3.7 | 33.6 ± 2.6 | 1.2 | 35.5 ± 1.0 | 3.7 |
HL60/ADR
| ||||||||||
None | 2.1 ± 0.4 | 0.51 ± 0.03 | 1.5 ± 0.2 | 60.7 ± 12.4 | >2000 | |||||
Zosuquidar | 2.4 ± 0.5 | 0.9 | 0.50 ± 0.03 | 1.0 | 1.4 ± 0.4 | 1.1 | 58.6 ± 4.6 | 1.0 | >2000 | ND |
CsA | 0.49 ± 0.04 | 4.3 | 0.30 ± 0.09 | 1.7 | 0.51 ± 0.03 | 2.9 | 48.7 ± 5.3 | 1.2 | 67.6 ± 7.6 | >29.9 |
HL60/DNR
| ||||||||||
None | >40 | 0.51 ± 0.05 | >20 | 489.9 ± 24.5 | >2000 | |||||
Zosuquidar | 0.49 ± 0.16 | >81.6 | 0.14 ± 0.01 | 3.6 | 0.22 ± 0.06 | >90.1 | 46.5 ± 9.7 | 10.5 | 620.5 ± 38.9 | 3.2 |
CsA | 2.5 ± 0.8 | >16 | 0.15 ± 0.00 | 3.4 | 1.4 ± 0.6 | >14.3 | 60.3 ± 5.0 | 8.1 | >2000 | ND |
Lack of effect of zosuquidar on wild type BCRP-expressing cells
FTC (10 μM) | Zosuquida | |
---|---|---|
K562/Vec | 0 | 0 |
K562/BCRP | 0.59 ± 0.11 | 0.04 ± 0.07 |
Modulation of drug resistance by zosuquidar in AML patient cells
Age Mean ± SD (range) | 60 ± 17 y (22–81) |
Leukocyte (109/L) | 46 ± 44 (3.2–184) |
FAB subtypes | |
1 | 23% (7/31) |
2 | 23% (7/31) |
4 | 16% (5/31) |
5 | 13% (4/31) |
Myelodysplastic syndrome | 26% (8/31) |
Karyotype | |
Favorable | 6% (2/31) |
Intermediate | 71% (22/31) |
Poor | 19% (6/31) |
Not done | 3% (1/31) |
CD34 | |
Positive | 55% (17/31) |
Negative | 42% (13/31) |
Not done | 3% (1/31) |
P-gp activity | |
Mean D ± SD (range) | 0.1 ± 0.1 (0–0.7) |
Positive D > 0.3 | 23% (7/31) |
Negative D < 0.3 | 77% (24/31) |
RMF | Uptake | ||||
---|---|---|---|---|---|
Resistance Modifiding | DNR/Ida | MITOX | Mylotarg | DIOC2 +Zosu | |
P01 | No | 1.1 | 0.6 | 1.0 | 0.06 |
P02 | Yes/DNR, Ida, Mylotarg | 2.3/3.2 | 20% | 35%e
| 0.53 |
P03 | No | 0.7 | ND | 0.8 | 0.05 |
P04 | No | 0.8 | ND | 1.3 | 0.1 |
P05 | No | 1.1 | 0.92 | 1.1 | 0.05 |
P06 | Yes/Mylotarg | ND | ND | 4.1 | 0.7 |
P07 | Yes/Mylotarg | 1.6 | ND | 29% | 0.46 |
P08 | No | 1.0 | ND | 1.2 | 0.13 |
P09 | No | 0.7/1.2 | ND | 1.0 | 0.24 |
P10 | No | 1.0/1.0 | ND | 1.0 | 0.33 |
P11 | No | 0.9 | ND | 13% | 0.4 |
P12 | Yes/MITOX | ND | 2.1 | ND | 0.6 |
P13 | No | 0.9 | 1.1 | 0.9 | 0.13 |
P14 | No | 1.1 | ND | ND | 0.23 |
P15 | No | 1.1 | 1.3 | 1.0 | 0.13 |
P16 | Yes/DNR, Mitox | 6.6 | 2.1 | ND | 0.04 |
P17 | No | 1.0 | 1.0 | 0.9 | 0.05 |
P18 | No | 1.1 | 0.9 | 1.0 | 0.06 |
P19 | No | 1.0 | 1.0 | 1.0 | 0 |
P20 | Yes/DNR, Mylotarg | 1.4 | ND | 3 | 0.38 |
P21 | No | ND | ND | 0.5 | 0.12 |
P22 | No | 1.2 | 1 | 1 | 0.21 |
P23 | No | 1.1 | 1.0 | 1.0 | 0.21 |
P24 | No | ND | ND | 1.1 | 0.06 |
P25 | Yes/DNR, Mitox, Mylo | 2.5 | 2.3 | 30% | 0.05 |
P26 | No | 1 | ND | ND | 0.04 |
P27 | No | 0.87 | 0.82 | ND | 0.24 |
P28 | No | 0.7 | 1.2 | ND | 0 |
P29 | Yes/Mylotarg | ND | ND | 2.0 | 0.04 |
P30 | No | 1.0 | 1.1 | 1.0 | 0.08 |
P31 | No | 1.2 | ND | 9% | 0.26 |
Modulation* | No modulation | |
---|---|---|
Active P-gp (D > 0.3) | 71% (5/7) | 29% (2/7) |
No active P-gp (D < 0.3) | 13% (3/24) | 87% (21/24) |