Dana-Farber Cancer Institute: Discovery of Resistance Mechanisms

Resistance to targeted therapy is emerging as a bottleneck to achieving durable drug responses in cancer. The goal of the CTD2 Center at Dana Farber Cancer Institute is to identify mechanisms of resistance for both existing therapeutics as well as for emerging targets even prior to the identification of lead compounds. They aim to use this information to inform combinatorial treatments. In representative examples they have found that YAP1 leads to resistance after KRAS targeting and that PRKACA mediates resistance to HER2 therapy.

A. Kinase ORF Library Screening for resistance to HER2 targeted therapy

A systematic interrogation of mechanisms of resistance to suppression of HER2 was used to identify the major mechanisms of resistance to HER2-directed therapy.

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Experimental Approaches

Two kinome open reading frame (ORF) screens were conducted in parallel to identify genes that confer resistance to the lapatinib-like dual EGFR/HER2 inhibitor AEE788 and to suppression of HER2 with a short hairpin RNA (shRNA). Then a V5 epitope-tagged kinase ORF collection was used to identify genes that mediate resistance to these manipulations.

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B. Determining Functional Substitutes for KRAS in KRAS-Dependent Cancer Cell Lines

We performed a genome-scale genetic rescue screen to identify genes that support the survival of KRAS-dependent cancer cells upon suppression of KRAS.

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Experimental Approaches

A genome-scale genetic rescue screen was performed to identify genes that support the survival of KRAS-dependent cancer cells upon suppression of KRAS. A cell line for screening was generated by stably introducing a doxycycline-inducible shRNA targeting the KRAS 3′ untranslated region (UTR) into the HCT116 KRAS mutant colon cancer cell line. Then 15,294 ORFs were introduced into these cells in an arrayed format in triplicate under optimized conditions. Suppression of KRAS was induced and then cell proliferation/survival was assessed.

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If you cannot access the manuscript, or if you have additional questions, please email Joshua Dempster.

Data

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Last updated: July 01, 2017