Fig. 1: Identification of AURKA as a synthetic lethal partner of RB1 in lung cancer cells.

a, b Western blot analyses to verify RB1 knockout in RB1^−/− lung cancer cells. MDA-MB468, a RB1-null breast cancer cell line, was used as a positive control for RB1 knockout. GAPDH was used as a loading control. c Flow chart of the synthetic lethality screenings in epigenetics small molecule and RNAi libraries. d Scatter plot of the averaged Z-scores for epigenetic siRNA screening. The Z-score of −3 was used as the cut-off (dotted line) to identify synthetic lethality hits. The three top hits, AURKA, HDAC1, and MYC are shown with red, green, and black dots, respectively. The screening was done in duplicate. e Total 11 candidates were found as potential synthetic lethality partners with the averaged Z-scores less than −2. The cut-off of Z-score −3 is marked with a dotted line. f Scatter plot of averaged log-IC₅₀ values of each small molecule for A549 RB1^+/+ and RB1^−/− from small molecule screening. The diagonal line represents small molecules that do not exhibit any selectivity toward A549 RB1^+/+ or RB1^−/− cell lines. Colored dots are candidate small molecules that selectively inhibited the viability of A549 RB1^−/− cells. The average IC₅₀ values from the duplicate screenings are shown. g Selectivity indices (SI) of the synthetic lethal candidates for RB1. SI = IC₅₀ ^RB1+/+/ IC₅₀ ^RB1−/−. Top 11 candidates with SI >4 (dotted line) are shown in the graph. Validation of the synthetic lethality between RB1 and AURKA using AURKA siRNA (h) and small molecule AURKA inhibitors, including ENMD-2076 (i) and alisertib (j). Data are presented as mean ± SD (n = 3 independent experiments). Synthetic lethal effects of AURKA siRNA (k), ENMD-2076 (l) and alisertib (m) in a panel of lung cancer cell lines with different RB1 status. The cell viability was measured with AlamarBlue assay. Data are presented as mean ± SD (n = 3 independent experiments).