RIPA Lysis Buffer br The in vivo therapeutic e ect of combined ABT
3.5. The in vivo therapeutic eﬀect of combined ABT199 and IRIN
To further confirm the in vivo cytotoxic eﬀect of ABT263 + AXIT combination, KRAS mutant A549 flank tumor nude mice were treated with ABT199 alone, IRIN, and ABT199 + IRIN combination for
Fig. 2. The combined ABT199 and IRIN inhibit cell growth. (A–B) Clonogenic assay was performed in KRAS-Wide type (A) and mutant (B) H661 cells. (C–D) Clonogenic assay was performed in A549 and H441 RIPA Lysis Buffer t treated with single or combination drugs. (E–F) Clonogenic assay was performed in KRAS-Wide type H838 and H522 cells treated with single or combination drugs.
Fig. 3. The combined ABT199 and IRIN leads to enhanced apoptosis. (A–B) KRAS-Wide type (A) and mutant (B) H661 cells treated with single or combination drugs were collected and analyzed for annexin-V and PI labelling. (C–D) A549 cells and H441 cells treated with single or combination drugs were collected and analyzed for annexin-V and PI labelling. The percentage of cell that was labeled with annexin-V (primary apoptosis) and with annexin-V plus PI (secondary apoptosis) was measured. (E–F) H838 and H522 cells treated by single or combination drugs were subject to the Annexin-V/propidium iodide apoptosis assay by flow cytometry.
4 weeks. ABT199 + IRIN combination resulted in smaller tumors (Fig. 5A), reduced tumor weight (Fig. 5B) and reduced tumor growth (Fig. 5C), demonstrating the potent in vivo eﬃcacy of the combination drugs (Fig. 5A, B). Measurement of the tumor weight at the end point of the treatment study confirmed the finding as well (p < 0.001, Fig. 5C). These results suggested that ABT199 + IRIN combination has sub-stantial preclinical in vivo eﬃcacy in KRAS-mutant cancer mouse models and provide a rationale for this combination in clinical trials.
Rebecca L. Siegel reported that the most common causes of cancer death are cancers of the lung, prostate, and colorectum in men and the lung, breast, and colorectum in women. Lung cancer has become the most prevalent neoplasm throughout the world with 1,2 million deaths per year (Siegel et al., 2018). Recently, many oncogenic mutations have been identified in lung cancer (Yang et al., 2016). Molecular genetic analyses have suggested that KRAS mutation is much more frequency in
Fig. 5. The in vivo therapeutic eﬀect of combined ABT199 and IRIN. (A) Photographs of the xenograft A549 tumors that had been treated by single or combination drugs. (B) The tumors were recorded for the weight. * p < 0.05, ** p < 0.01, *** p < 0.001. Data are presented as mean ± sem. (C) Tumor volume was measured every other day for diﬀerent drug treatment regiments (i.e. single or combination drugs).
NSCLC (Eberhard et al., 2005). Therefore, synthetic screening inhibitors will be a good avenue for the therapeutic treatment of KRAS-trans-formed tumors.
In the present study, we provide evidence that we used the combi-natorial strategy to target oncogene addiction for RAS-mutant lung cancer cells. We confirm that combined ABT199 and irinotecan leads to RAS-mutant lung cancer cell growth inhibition and enhances apoptosis. Given the importance of combinations of inhibitors, the major me-chanism may be promising strategies to treat KRAS-mutant cancers. Some studies indicate that resistance of EGFR-KRAS tumors is a con-sequence into the PI3K-AKT signaling, which is known to stimulate cell survival through down-regulation of cell death pathways (Grana et al., 2002). Gupta et al. showed that blocking of PI3K with LY294002 radio sensitizes Ras-transformed REFs and two human tumor cell lines con-taining activated ras alleles (Gupta et al., 2001). Here, we also show that the combination of ABT199 and irinotecan can block PI3K/AKT signaling, suggesting it can be therapeutically exploited to inhibit KRAS-driven lung cancer. However, whether other signaling pathways are also changed need further study.