Establishment of a Transformation Coupled in vitro End Joining Assay to Estimate Radiosensitivity in Tumor Cells

Establishment of a Transformation Coupled in vitro End Joining Assay to Estimate Radiosensitivity in Tumor Cells

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Here, we present a modified in vitro end-joining (EJ) assay to quantify EJ capacity, accuracy as well as pathway switch to alternative end-joining (Alt-EJ) or single strand annealing (SSA).A novel transformation assay was established to specifically measure circular repair products, which correlate with classical EJ efficiency.The EJ assay was validated using EJ-deficient mammalian cell lines (Ku80, DNA-PKcs, LigIV, or XRCC4 mutants).

A pathway switch to Alt-EJ and Compression Garments - Stockings and Socks SSA was seen exclusively in Ku-deficient cells.Circular EJ product formation correlated with cell survival and DSB repair capacity after X-irradiation.Investigation of 14 HNSCC cell lines revealed differences in the total EJ capacity Medical and Ambulatory supplies / Ostomy but a broader variation in the amount of circular repair products.

Sequencing of repair junctions in HNSCC cells demonstrated a predominance of high-fidelity EJ and an avoidance of both Alt-EJ and SSA.A significant correlation was observed between the amount of circular repair products, repair of IR-induced DSB and radiosensitivity.Collectively, these data indicate that the presented in vitro-EJ-assay can not only estimate the repair capacity of cancer cells to enable the stratification into radiosensitive or radioresistant, but can also identify repair pathway deregulation such as a switch to Alt-EJ or SSA, which enables tumor targeting.