Figure 4.

RSPO3-taxane regimen reduces CSC self-renewal properties by reducing β-catenin transduction and potentiating paclitaxel mediated mitotic blockade. (A) Limiting dose assay in OMP-C8 tumor model, from Fig. 3. The combination of anti-Rspo3 (25 mpk) on day 1 and nab-paclitaxel (30 mpk) on day 3 and day 10 in three-two week cycles reduces the cancer stem cell frequency as compared to nab-paclitaxel, by 40-fold. Tumors were serially passaged on day 38, seven days after the last treatment cycle, at a cell dose of 30 or 300 cells into recipient mice, 10 mice per cell dose and 20 mice per treatment group. On day 85 of the LDA, tumor take was determined and the CSC frequency was calculated. (B) Anti-Rspo3 + nab-paclitaxel reduced nuclear β-catenin expression, disrupted mitosis, and promoted the expansion of differentiated mucin producing goblet cells. OMP-C8 tumors were assayed for β-catenin expression (brown) and mitotic activity (red, PHH3) with nuclear counterstain (blue), 20x magnification. Goblet cells were detected by Alcian Blue counterstain (blue) with nuclei in purple, 10x magnification. (C) Nab-paclitaxel expands for mitotic cells while combination with anti-Rspo3 promotes for endoreduplication and the selection of differentiated goblet cells. From OMP-C9 study in Fig. 3. OMP-C9 tumors were assayed for mitotic activity (PHH3, brown) with nuclear counterstain (blue), 20x magnification. Goblet cells were detected by Alcian Blue counterstain (blue) with nuclei in purple, 10x magnification. D. RSPO3-taxane treatment reduces LGR5 expression and promotes suppression of Wnt signaling. Taqman qPCR, n = 2–4 per treatment group, expressed as mean + SEM, *p < 0.05.