Extended Data Fig. 5: Testing of ccD-CMD framework using cell cycle perturbation in vitro and cell cycle modeling in silico, and comparison of temporal inference methods.
From: Temporal and spatial topography of cell proliferation in cancer
a. Left, histograms of fluorescence single-cell log2 normalized signal by plate-based CyCIF (p-CyCIF) from untreated (blue) and 24 h palbociclib 1 µM treated (orange) MCF10A cells grown in culture (n = 10,000 cells per condition). Right, ccD-CMD plot. b. Heat map and time plot of single-cell signals ordered by ccD-CMD (normalized log2 FAU, n = 10,000 cells moving mean over 200 cells). c. Traces from generalized model of mammalian cell cycle80. Top panels, no noise added. Middle panels, Gaussian measurement noise added (additive and multiplicative). Gray areas are the time frame used for plots in panels d. d. ccD-CMD plot of mathematical model results in shaded areas of panel c. Bottom panel, simulation switch to G1 arrest parameters (CDK/CyclinD complex set to 0) after one cell cycle. Colour is the time variable in the mathematical model (n = 10,000 points). e-h. Comparison of time inference methods ccD-CMD, SCORPIUS, Palantir and Cyclum applied to e. synthetic data generated by the mathematical model in panel e. (n = 10,000 observations), f. HER2 + breast cancer tissue data from Fig. 3a-f., and g. MCF10A untreated cells from Fig. 3g. Left, two dimensional visualization output. Right, pseudotime ordering output (n = 5,000 cells, same cells used for all algorithms, normalized log2 FAU, moving mean over 200 cells). h. Comparison between two dimensional reduced space visualization from three time inference algorithms with data from HER2 + breast patient samples #1, 2, and 3 from Fig. 4b-d. i. Coefficient of variation of coherence metrics IOV and CFD in 5 tissues using increasing number of cells (n > 20,000 cells per tissue, CV calculated over 40 independent sub-samplings). j. Comparison of IOV and CFD when one or two markers are removed from ccD-CMD algorithm. Data from untreated MCF10A cells used in panel g. The green dot is the original representation. Orange, one marker removed. Blue, two markers removed. k. Comparison of IOV and CFD when MPI 0 and MPI -1 cells are added at increasing ratios. l. Examples of pseudotime ordering by ccD-CMD algorithm showing inferred CDT1 and Geminin across six tissues areas.
