Data analysis for the manuscript, "Osteosarcoma is dependent on ERK-driven MCL1 production for survival in the early metastatic niche"
Introduction
We sought to identify dependencies of slow growing, chemo-resistant osteosarcoma cells that anchor in the metastatic niche. These dependencies yield insight into what allows early cells to survive in the hostile lung environment, as well as potential therapeutic targets. In our studies of metastatic colonization, we identified two distinct populations of osteosarcoma cells. The first, dubbed “anchor cells”, are prevalent in the early metastatic niche where they are hypo-proliferative and hyper-secretory. The second subpopulation, “growth cells”, proliferate rapidly and predominate as lesions begin to grow rapidly. By targeting “anchor” cells in addition to growth cells, we hope to reduce or eliminate otherwise chemoresistant metastases. We identified and exploited vulnerabilities of these anchor cells.
Methods
We used scRNAseq, IHC-F, live cell reporters and ICC to interrogate and confirm vulnerabilities of anchor cells in the metastatic niche as well as identify sources of pro-survival signaling in the metastatic niche. We tested these vulnerabilities in vitro using tumor-on-lung spheroids and in vivo using a murine model of metastasis.
Results
We found anchor cells have upregulated ERK activity and MCL1 expression compared to growth cells. This activity is associated with production of several growth factors in the metastatic niche that can activate receptors expressed by osteosarcoma cells. Growth-factor induced ERK activity is necessary for increased MCL1 expression in osteosarcoma cells. We found expression of many growth factors produced by niche cells is augmented as osteosarcoma cells colonize the lung. Additionally, host cells within the niche express comparatively more growth factors early in the metastatic process, with macrophages representing a predominant source of growth factors. We found cells expressing higher levels of MCL1 are vulnerable to its inhibition in vitro. Metastatic burden decreased as the number of AZD5991 doses increased in vivo.
Conclusion
Our data suggest growth factors in the lung activate ERK in the early metastatic niche. This ERK signaling has multiple outcomes, one of which is increased MCL1 expression in early metastatic cells. Targeting MCL1 is effective at reducing metastatic burden in mice, and works synergistically with cyclophosphamide to drastically increase survival.
This set of code is intended to document the analyses performed to generate the data presented in the above manuscript, serving to investigate the early metastatic signaling that assists in tumor cell survival in the lung. This is not intended as a plug-and-play pipeline for use by others; however, this code may be useful as a reference for how to perform some of the analyses.