3.3. αCT1 successfully sensitizes GBM organoids to temozolomide
Like αCT11, αCT1 is a Cx43 mimetic peptide. However, unlike αCT11, it contains a cell penetration sequence that should allow it to better permeate the cells (Jiang et al., 2019). We hypothesized that because of the presence of this additional sequence, it would significantly increase GBM cell sensitivity to TMZ compared to αCT11. Using an ATP assay to quantify cell viability, we found that when the αCT1 peptide is present at high concentrations (αCT1 = 10 and 100 μM) in A172 and BT169 organoids, viability is considerably decreased compared to the control groups (αCT1 = 0) for several TMZ concentrations (Figure 3A and 3C ). Surprisingly, αCT1 peptide at a concentration of 10 μM is more effective in the A172 organoids than the highest concentration (100 μM). In the U87 MG organoids, however, the αCT1 had little influence on cell growth (Figure 3B ).
The LIVE/DEAD images (Figure 4 ) show comparable results as the ATP assay. The highest concentration of TMZ (1000 µM) combined with highest concentration of αCT1 (100 µM) indicates the most significant cell killing in A172 and BT169 cell lines (Figure 4A and 4C ). This combination of TMZ and αCT1 treatment group, however, did not increase cell killing efficacy in the U87 cell line, which is consistent with the results of the ATP assay (Figure 4B ). LIVE/DEAD images of combinatorial treatment with TMZ and lower concentrations of αCT1 (1 and 10 µM) are shown in Supplemental Figures 4-6 .
This finding indicates that the combinatorial treatment with TMZ and αCT1 may only be effective in certain GBM tumor populations. It’s also worth noting that even when the TMZ concentration is zero, αCT1 shows a significant decrease in viability in the A172 organoids. This could indicate that αCT1 is impacting several important cell pathways, not just those related to TMZ efficacy in GBM cells.