Pancreatic Cancer Organoids in the Field of Precision Medicine: A Review of Literature and Experience on Drug Sensitivity Testing with Multiple Readouts and Synergy Scoring.

Pancreatic ductal adenocarcinoma (PDAC) is a silent killer, often diagnosed late. However, it is also dishearteningly resistant to nearly all forms of treatment. New therapies are urgently needed, and with the advent of organoid culture for pancreatic cancer, an increasing number of innovative approaches are being tested. Organoids can be derived within a short enough time window to allow testing of several anticancer agents, which opens up the possibility for functional precision medicine for pancreatic cancer. At the same time, organoid model systems are being refined to better mimic the cancer, for example, by incorporation of components of the tumor microenvironment. We review some of the latest developments in pancreatic cancer organoid research and in novel treatment design. We also summarize our own current experiences with pancreatic cancer organoid drug sensitivity and resistance testing (DSRT) in 14 organoids from 11 PDAC patients. Our data show that it may be necessary to include a cell death read-out in ex vivo DSRT assays, as metabolic viability quantitation does not capture actual organoid killing. We also successfully adapted the organoid platform for drug combination synergy discovery. Lastly, live organoid culture 3D confocal microscopy can help identify individual surviving tumor cells escaping cell death even during harsh combination treatments. Taken together, the organoid technology allows the development of novel precision medicine approaches for PDAC, which paves the way for clinical trials and much needed new treatment options for pancreatic cancer patients.

Parallel evolution of Pseudomonas aeruginosa phage resistance and virulence loss in response to phage treatment in vivo and in vitro.

With rising antibiotic resistance, there has been increasing interest in treating pathogenic bacteria with bacteriophages (phage therapy). One limitation of phage therapy is the ease at which bacteria can evolve resistance. Negative effects of resistance may be mitigated when resistance results in reduced bacterial growth and virulence, or when phage coevolves to overcome resistance. Resistance evolution and its consequences are contingent on the bacteria-phage combination and their environmental context, making therapeutic outcomes hard to predict. One solution might be to conduct 'in vitro evolutionary simulations' using bacteria-phage combinations from the therapeutic context. Overall, our aim was to investigate parallels between in vitro experiments and in vivo dynamics in a human participant. Evolutionary dynamics were similar, with high levels of resistance evolving quickly with limited evidence of phage evolution. Resistant bacteria-evolved in vitro and in vivo-had lower virulence. In vivo, this was linked to lower growth rates of resistant isolates, whereas in vitro phage resistant isolates evolved greater biofilm production. Population sequencing suggests resistance resulted from selection on de novo mutations rather than sorting of existing variants. These results highlight the speed at which phage resistance can evolve in vivo, and how in vitro experiments may give useful insights for clinical evolutionary outcomes.

Functional Profiling of FSH and Estradiol in Ovarian Granulosa Cell Tumors.

Adult-type granulosa cell tumors (AGCTs) are sex-cord derived neoplasms with a propensity for late relapse. Hormonal modulators have been used empirically in the treatment of recurrent AGCT, albeit with limited success. To provide a more rigorous foundation for hormonal therapy in AGCT, we used a multimodal approach to characterize the expressions of key hormone biomarkers in 175 tumor specimens and 51 serum samples using RNA sequencing, immunohistochemistry, RNA in situ hybridization, quantitative PCR, and circulating biomarker analysis, and correlated these results with clinical data. We show that FSH receptor and estrogen receptor beta (ERß) are highly expressed in the majority of AGCTs, whereas the expressions of estrogen receptor alpha (ERα) and G-protein coupled estrogen receptor 1 are less prominent. ERß protein expression is further increased in recurrent tumors. Aromatase expression levels show high variability between tumors. None of the markers examined served as prognostic biomarkers for progression-free or overall survival. In functional experiments, we assessed the effects of FSH, estradiol (E2), and the aromatase inhibitor letrozole on AGCT cell viability using 2 in vitro models: KGN cells and primary cultures of AGCT cells. FSH increased cell viability in a subset of primary AGCT cells, whereas E2 had no effect on cell viability at physiological concentrations. Letrozole suppressed E2 production in AGCTs; however, it did not impact cell viability. We did not find preclinical evidence to support the clinical use of aromatase inhibitors in AGCT treatment, and thus randomized, prospective clinical studies are needed to clarify the role of hormonal treatments in AGCTs.