Supplementary Methods and Supplementary Figures S1-S13 from Aurora A is Critical for Survival in HPV-Transformed Cervical Cancer

Contains siRNA primary screen analysis and subsequent supplemental analyses; Figure S1 Analysis was performed using customized R scripts and proceeded via a three step process; Figure S2 Summary of Z scores of the top 54 selected target hits from the primary screen were rescreened using OnTarget Plus siRNAs; Figure S3 Mitosis is not a general target in HPV-transformed cancer cell lines; Figure S4A. Dose response for all the cell lines with the Aurora B inhibitor ZM447439; Figure S4B. Flow cytometry of the DNA content of cells were treated with 5 μM ZM447439 and collected at 24, 48, and 72 h, using DMSO as a control; Figure S5: Either wild type or K14E7 transgenic skin grafted onto wild type mice, either treated with Alisertib in vivo or untreated, was stained for mast cells using touluidine blue; Figure S6: Alisertib treatment of HPV-transformed cancer cells promotes apoptosis; Figure S7A. HPV- and non-HPV-transformed cancer cells were treated with 5 μM Alisertib and sampled at intervals over 72 h and analysed by flow cytometry for DNA content; S7B. Immunofluorescence of representative HPV-transformed (HeLa) and non-HPV C33A cells treated with DMSO (Control) or 5μM of Alisertib for 48 h stained for DNA and microtubules (α-tubulin). S7C. Quantitation of bi-nucleated and multinucleated cells in control and Alisertibtreated cells for 1 and 2 days; Figure S8 Representative time-lapse series of HeLa and C33A with and without 5 μM Alisertib treatment transiting mitosis and undergoing apoptosis; Figure S9: The time lines for 50 HeLa and 50 CaSki cells treated with 5 μM Alisertib; Figure S10: The dose response of parental HeLa cells and HeLa cells over expressing either Bcl-2 or Mcl- 1 with etoposide and Taxol, respectively, are shown; Figure S11: HPV-transformed ME180 and non-HPV C33A cells were treated with 5 μM Alisertib for up to 3 days; Figure S12 Dose response curves for HPV16 E7 expressing C33A and SCC25 cell lines determined (one of four replicate experiments); Figure S13: FACS of the GFP levels in the HPV18 E7-IRES-GFP transduced SCC25 cell lines used in Figure 7B, C.