Figure 1.
Engraftment and growth of U266luciferase cells as monitored by BLI, serum paraprotein and MRI. A.
(i) Dorsal and (ii) ventral BLI acquired from IVIS over weeks 3–7 post-inoculation. B. Quantitative measurement of radiance from BLI. Radiance reflects the intensity of luciferase luminescence and therefore number of luciferase-tagged cells present. Results show that radiance increases in a time-dependent manner over the course of the experiment and that a significant increase in radiance occurs over weeks 5–7 (p<0.05, 1-way ANOVA with Bonferroni post-test). C. Paraprotein levels in the serum increases in a time dependent manner and correlates with the increase seen in BLI. MRI-derived tumour volumes determined at approximately weeks 5 and 10 confirmed tumour progression seen with BLI.
Figure 2.
CD138 expression and histology validate BLI, paraprotein and MRI results. A.
Flow cytometry histograms show the presence of CD138+ cells in the tibia, femur and spine of myeloma mice. Cytometric evaluation of the organs confirmed that the cells were confined to the bone marrow. B. Histological analysis of the tibia and femur confirmed the presence of a high number of CD138+ plasma cells (green arrows), resulting in the loss of classical bone marrow architecture.
Figure 3.
BLI and serum paraprotein changes in response to therapy.
A. (i) Pre-treatment and (ii) post-treatment BLI of mice at weeks 5 and 9. B. Quantitative measurement of radiance from BLI. (i) No significant difference in radiance between treatment groups was seen at the start of the treatment schedule. (ii) Post-treatment radiance levels revealed a significant attenuation of tumour spread by both BZB and tosedostat (p<0.05, 1-way ANOVA with Bonferroni post-test). C. Paraprotein levels during treatment schedule. Positive control mice showed an exponential increase in serum levels of Igλ over 9 weeks. In comparison, both treatment groups did not exhibit the same increase, with significantly lower levels by the end of treatment (p<0.05).
Figure 4.
CD138 expression changes in response to therapy.
A. Percentage of CD138+ human myeloma cells measured by flow cytometry in bone aspirates of mice (n = 3), showing a significantly lower percentage of positive cells in both tibias and spine of mice in the two treatment groups than in untreated mice (p<0.05, 2-way ANOVA with Bonferroni post-test). No CD138+ cells were observed in the organs of any of the mice. B. Histological analysis of sections from the tibias of mice from each group showed distinct differences. (i) Sections from healthy mice displayed classical architecture, with no CD138+ cells. (ii) In comparison, sections from untreated myeloma mice showed a high infiltration of CD138+ cells with loss of normal architecture. (iii) Treatment of mice with BZB resulted in the return of normal architecture and loss of CD138+ cells. (iv) A similar result was observed in mice treated with tosedostat, but with occasional scattered CD138+ cells.
Figure 5.
MRI changes in response to therapy.
MRI-derived tumour volumes. A. Tumour was identified as a hyperintense signal enclosed within the cortical bone on T2-weighted images. MRI images showed a reduction in signal intensity in both treatment groups compared to positive control in both the tibia (T) and femur (F). B. Tumour volume was quantified from regions of interest drawn on the periphery of the hyperintense signal. Data are mean ± SEM, n≥6. Both BZB and tosedostat (TDT) treatment resulted in a significantly lower tumour volume compared to control (p<0.05, 1-way ANOVA with Bonferroni post-test). In addition, there was no significant difference in tumour volume between the BZB treated and negative control group.
Table 1.
Summary of current mouse models for studying the development and treatment of myeloma.