Advancing wide implementation of precision oncology: A liquid nitrogen-free snap freezer preserves molecular profiles of biological samples.

PURPOSE: In precision oncology, tumor molecular profiles guide selection of therapy. Standardized snap freezing of tissue biospecimens is necessary to ensure reproducible, high-quality samples that preserve tumor biology for adequate molecular profiling. Quenching in liquid nitrogen (LN2 ) is the golden standard method, but LN2 has several limitations. We developed a LN2 -independent snap freezer with adjustable cold sink temperature. To benchmark this device against the golden standard, we compared molecular profiles of biospecimens. METHODS: Cancer cell lines and core needle normal tissue biopsies from five patients' liver resection specimens were used to compare mass spectrometry (MS)-based global phosphoproteomic and RNA sequencing profiles and RNA integrity obtained by both freezing methods. RESULTS: Unsupervised cluster analysis of phosphoproteomic and transcriptomic profiles of snap freezer versus LN2 -frozen K562 samples and liver biopsies showed no separation based on freezing method (with Pearson's r 0.96 (range 0.92-0.98) and >0.99 for K562 profiles, respectively), while samples with +2 h bench-time formed a separate cluster. RNA integrity was also similar for both snap freezing methods. Molecular profiles of liver biopsies were clearly identified per individual patient regardless of the applied freezing method. Two to 25 s freezing time variations did not induce profiling differences in HCT116 samples. CONCLUSION: The novel snap freezer preserves high-quality biospecimen and allows identification of individual patients' molecular profiles, while overcoming important limitations of the use of LN2 . This snap freezer may provide a useful tool in clinical cancer research and practice, enabling a wider implementation of (multi-)omics analyses for precision oncology.

Cooling of a vial in a snapfreezing device without using sacrificial cryogens.

A fresh and frozen high-quality patient bio-sample is required in molecular medicine for the identification of disease-associated mechanism at molecular levels. A common cooling procedure is immersing the tissue enclosed in a vial in a coolant such as liquid nitrogen. This procedure is not user friendly and is laborious as reducing the lag time from excision time to freezing depends on the logistic organizational structure within a hospital. Moreover snapfreezing must be done as soon as possible after tissue excision to preserve the tissue quality for molecular tests. Herein, we report an electrically powered snap freezing device as an alternative to quenching the vial in liquid nitrogen and therefore can be used directly at the location where the tissue is acquired. This device also facilitates the study of the effect of freezing conditions on the various molecular processes in the samples. Cooling experiments of a vial in the snap freezing device show that the cooling rates similar to or faster than quenching in liquid nitrogen are feasible. We performed experiments with several set point conditions and compared the results with a mathematical model.