Visualizing Engrafted Human Cancer and Therapy Responses in Immunodeficient Zebrafish.

Xenograft cell transplantation into immunodeficient mice has become the gold standard for assessing pre-clinical efficacy of cancer drugs, yet direct visualization of single-cell phenotypes is difficult. Here, we report an optically-clear prkdc-/-, il2rga-/- zebrafish that lacks adaptive and natural killer immune cells, can engraft a wide array of human cancers at 37°C, and permits the dynamic visualization of single engrafted cells. For example, photoconversion cell-lineage tracing identified migratory and proliferative cell states in human rhabdomyosarcoma, a pediatric cancer of muscle. Additional experiments identified the preclinical efficacy of combination olaparib PARP inhibitor and temozolomide DNA-damaging agent as an effective therapy for rhabdomyosarcoma and visualized therapeutic responses using a four-color FUCCI cell-cycle fluorescent reporter. These experiments identified that combination treatment arrested rhabdomyosarcoma cells in the G2 cell cycle prior to induction of apoptosis. Finally, patient-derived xenografts could be engrafted into our model, opening new avenues for developing personalized therapeutic approaches in the future.

Stromal Microenvironment Shapes the Intratumoral Architecture of Pancreatic Cancer.

Single-cell technologies have described heterogeneity across tissues, but the spatial distribution and forces that drive single-cell phenotypes have not been well defined. Combining single-cell RNA and protein analytics in studying the role of stromal cancer-associated fibroblasts (CAFs) in modulating heterogeneity in pancreatic cancer (pancreatic ductal adenocarcinoma [PDAC]) model systems, we have identified significant single-cell population shifts toward invasive epithelial-to-mesenchymal transition (EMT) and proliferative (PRO) phenotypes linked with mitogen-activated protein kinase (MAPK) and signal transducer and activator of transcription 3 (STAT3) signaling. Using high-content digital imaging of RNA in situ hybridization in 195 PDAC tumors, we quantified these EMT and PRO subpopulations in 319,626 individual cancer cells that can be classified within the context of distinct tumor gland "units." Tumor gland typing provided an additional layer of intratumoral heterogeneity that was associated with differences in stromal abundance and clinical outcomes. This demonstrates the impact of the stroma in shaping tumor architecture by altering inherent patterns of tumor glands in human PDAC.

Monocyte Anisocytosis Changes in Patients after Major Burn Injuries.

The recovery of patients after severe burns is a long and complex process. Recently, genomic analysis of white blood cells from burn and trauma patients revealed excessive and prolonged innate immune activation in patients with complicated outcomes. However, translating this knowledge into practical biomarkers has not been possible yet. Although several biomarkers for monitoring burn patients have been proposed, their ability to accurately distinguish between inflammation stemming from initial tissue destruction, infections, and organ failure complications is limited. Here, we focused on monocytes, critical innate immune cells in the response to burn injured tissues. We measured the monocyte anisocytosis (quantified as monocyte distribution width, MDW, a recently emerged marker of sepsis) throughout the recovery of patients from the time of burn injury until the end of the hospital stay. We observed that MDW increases in patients during the first week after major burns. Among the patients with major burns who survive, MDW starts decreasing in the second week and normalizes by the end of the hospital stay. The duration of hospital stay appears to be proportional to how fast MDW decreases during the second week after the injury. We also found that MDW decreases significantly in most patients after excision and debridement surgeries but not after allo- and auto-graft surgeries. Moreover, high MDW values correlated with a higher rate of positive microbiology blood culture samples and respiratory infections. These findings underscore the importance of monitoring MDW as a potential biomarker for the risk of complications during burn patient recovery.