Target therapy of multiple myeloma by PTX-NPs and ABCG2 antibody in a mouse xenograft model

Abstract

Multiple myeloma (MM) remains to be an incurable disease. The purpose of this study was to evaluate the effect of ABCG2 monoclonal antibody (McAb) combined with paclitaxel (PTX) conjugated with Fe3O4 nanoparticles (NPs) on MM progressed from cancer stem cells (CSCs) in non-obese-diabetic/severe-combined-immunodeficiency (NOD/SCID) mouse model. Mice were injected with MM CSCs as marked by CD138-CD34- phenotypes through tail veins. The developed MM mice were examined by micro-computer tomography scanning, ultrasonography and enzyme-linked immunosorbent analysis. These mice were then intravenously treated with different combinations of NPs, PTX, McAb, PTX-NPs and melphalan/prednisone once a week for four weeks. The injected mice developed characteristic MM-associated syndromes, including lytic bone lesions, renal damages and proteinuria. All the treated mice showed decrease in bone lesions, renal damages and anemia but increase in apoptosis compared with the mice treated with NPs only. In particular, the treatment with ABCG2 McAb plus PTX-NPs induced the strongest therapeutic response and had an efficacy even better than that of melphalan/prednisone, a conventional regimen for MM patients. These data suggest that PTX-NPs with ABCG2 McAb can be developed into potential treatment regimens for patients with relapsed/refractory MM.

Keywords: ATP-binding cassette sub-family G member 2; cancer stem cells; multiple myeloma; nanoparticles; paclitaxel.

Figure 1. Establishment of a MM murine model

A. Images showed Micro-CT scanning of a normal and a model mouse 18 days after injection of MM CD138⁻CD34⁻cells. BMD values at femur and humerus were indicated by arrows. B. Quantification of BMD as measured by Micro-CT (n = 6). C. A Micro-CT image of kidneys of a model mouse. Arrows indicate renal damaged areas. D. Ultrasound images showing peak systolic BFV of the renal artery of a normal and a model mouse. E. Quantification of peak systolic BFV. F. Urine protein was measured by ELISA. All the data represent mean ± S.D (n = 3). *p < 0.05, **p < 0.01 and ***p < 0.001 were calculated by t test, referring to the statistically significant difference as compared to the normal group.

Figure 2. Characterization of PTX-NPs

A. Schematic illustration of PTX-NPs preparation. PTX, oleic acid-coated iron oxide nanoparticles (Fe3O4@OA NPs) and polyoxyethylene/polyoxypropylene copolymer (Pluronic F68) were dissolved in tetrahydrofuran (THF). The mixture was added into water under sonication, and free THF in the resulting suspension was evaporated under a continuous stirring. B. A TEM image showing prepared nanoparticles conjugated with PTX. The average size of PTX conjugated nanoparticles is about 7.63 nm as estimated by an image analysis program based on more than 300 particles. C. The hydrodynamic diameter distribution of PTX-NPs was measured by dynamic light scattering (DLS).

Figure 3. Significant improvement of BMD by McAb+PTX-NPs in MM mice

A. Micro-CT images showing BMDs in MM mice 4 weeks after treatments with different agents as indicated. B. Quantification of BMD in the mice treated with different agents. The data represent mean ± SD (n = 6) *p < 0.05, **p < 0.01 and ***p < 0.001 referred to the differences as indicated. Bonferroni correction was applied if multiple comparisons were involved.

Figure 4. Bone lesion reduction by McAb+PTX-NPs in MM mice

A. HE staining of femur lesions in MM mice 4 weeks after treatment with different agents as indicated (magnification, 400 ×). Arrows indicate aggregated and infiltrated inflammatory cells. B. Quantification of bone lesions. *p < 0.05, **p < 0.01 and ***p < 0.001, referring to the differences as indicated.

Figure 5. Ultrasound imaging of the BFV of renal arteries in MM mice

A. Images showing peak systolic BFV of renal arteries in MM mice 4 weeks after treatment with different agents as indicated. B. Statistical analysis of peak systolic BFV of MM mice. *p < 0.05, **p < 0.01 and ***p < 0.001.

Figure 6. Histological analysis of kidneys of MM mice

A. Tissue sections derived from MM mice 4 weeks after treatments with different agents were stained with either masson (M) or PAS (P) Arrows indicate representative damaged areas as described in the text. B. Quantitative analysis of renal damages in treated mice. *p < 0.05, **p < 0.01 and ***p < 0.001.

Figure 7. Strong apoptosis induced by McAb+PTX-NPs in BMMCs isolated from MM mice

A. Apoptosis of BMMCs derived from treated MM mice was analyzed as described in the Method. B. Quantification of apoptotic BMMCs. *p < 0.05, **p < 0.01 and ***p < 0.001.