Extended Data Fig. 1: Proliferation of primary and metastatic pancreatic cancer cells in culture and cell competition experiments in mice.

(a-c) Proliferation rate (doublings/day) of cells in culture that were isolated from primary tumors, or from liver or lung metastases, as indicated. Data shown were obtained from three independently derived paired cell lines; mean +/− stdev, n=4 technical replicates (a and b), or 3 technical replicates (c) per cell line; representative data from independent biological replicates of at least n=3 per cell line. (d) Representative western blot analysis assessing mutant Kras expression in protein lysates obtained from the indicated pancreatic cell lines and established human cell line controls and pancreatic stellate cells (PSCs). Mutant Kras G12D expression was only validated once per cell line at the time of generating the cell lines, (e) Schematic of experimental procedure where primary or liver metastatic pancreatic cancer cells (liver met) labeled with mCherry or GFP were injected into the pancreas, liver, or subcutaneous flank; either implanted individually or as a 50:50 mixture of the indicated cells. Representation of each cell population in the final tumor was analyzed by flow cytometry or by immunohistochemistry (IHC). Created with BioRender.com. (f) Flow cytometry of individually labeled cell populations (top) and a mixed population (bottom left). Approximately equal representation of each labeled cancer cell population in a mixed population was confirmed prior to injection (representative data shown bottom right). (g) Tumor weight post injection of individually labeled cancer cells (n=3) or a mixed cell population containing equal numbers of both primary (prim) and liver metastasis (met)-derived pancreatic cancer cells (n=8). Mean +/− stdev. (h) Representation of GFP-labeled primary or mCherry-labeled liver metastatic cells in a tumor derived from injection of a mixed population containing equal numbers of labeled primary and liver metastasis-derived pancreatic cancer cells into the pancreas as determined by flow cytometry. Each data point represents one mouse; the numbers associated with each mouse indicate animal ID with each animal being injected with the same mixed population of cells. (i) Representation of mCherry-labeled primary or GFP-labeled liver metastatic cells in a tumor derived from injection of a mixed population containing equal numbers of labeled primary and liver metastasis-derived pancreatic cancer cells into the pancreas as determined by flow cytometry. Each data point represents one mouse; the numbers associated with each mouse indicate animal ID with each animal being injected with the same mixed population of cells. (j) Immunohistochemistry to assess GFP and mCherry expression in a tumor derived from injection of a mixed population containing equal numbers of primary and liver metastasis-derived pancreatic cancer cells into the pancreas as in (h) Scale bar, 250 μm. (k) Representation of GFP-labeled primary or mCherry-labeled liver metastatic cells in a tumor derived from injection of a mixed population containing equal numbers of primary and metastasis-derived pancreatic cancer cells into the liver as determined by flow cytometry. Each data point represents one mouse; the numbers associated with each mouse indicate animal ID with each animal being injected with the same mixed population of cells. (l) Immunohistochemistry to assess GFP and mCherry expression in three different tumors derived from injection of a mixed population containing equal numbers of a primary and liver metastasis-derived pancreatic cancer cells into the liver as in (k), although the mixed population of cells were from an independent experiment where the primary tumor cells were labeled with mCherry, and the liver metastasis cells were labeled with GFP. Scale bar, ~2500 μm. (m) Immunohistochemistry to assess GFP, mCherry, or Cytokeratin-19 (Ck19) expression in whole mount liver tissue sections from a mouse from an independent experiment where a mixed population containing equal numbers of primary, and liver metastasis-derived pancreatic cancer cells were implanted into the liver as in (m). (n) Representation of GFP-labeled primary or mCherry-labeled liver metastatic cells in a tumor derived from subcutaneous injection of mixed population containing equal numbers of labeled primary and liver metastasis-derived pancreatic cancer cells into the flank. Each data point represents one mouse; the numbers associated with each mouse indicate animal ID with each animal being injected with the same mixed population of cells. (o) Representation of mCherry-labeled primary or GFP-labeled liver metastatic cells in a tumor derived from subcutaneous injection of mixed population containing equal numbers of labeled primary and liver metastasis-derived pancreatic cancer cells into the flank. Each data point represents one mouse; the numbers associated with each mouse indicate animal ID with each animal being injected with the same mixed population of cells.