Details

Autor(en) / Beteiligte

• Whiteaker, Jeffrey R.
• Sharma, Kanika
• Hoffman, Melissa A.
• Kuhn, Eric
• Zhao, Lei
• Cocco, Alexandra R.
• Schoenherr, Regine M.
• Kennedy, Jacob J.
• Voytovich, Ulianna
• Lin, Chenwei
• Fang, Bin
• Bowers, Kiah
• Whiteley, Gordon
• Colantonio, Simona
• Bocik, William
• Roberts, Rhonda
• Hiltke, Tara
• Boja, Emily
• Rodriguez, Henry
• McCormick, Frank
• Holderfield, Matthew
• Carr, Steven A.
• Koomen, John M.
• Paulovich, Amanda G.

Titel

Targeted mass-spectrometry-based assays enable multiplex quantification of receptor tyrosine kinase, MAP kinase, and AKT signaling

Ist Teil von

• Cell reports methods, 2021-07, Vol.1 (3), p.100015, Article 100015

Ort / Verlag

United States: Elsevier Inc

Erscheinungsjahr

2021

Link zum Volltext

Quelle

Alma/SFX Local Collection

Beschreibungen/Notizen

• A primary goal of the US National Cancer Institute's Ras initiative at the Frederick National Laboratory for Cancer Research is to develop methods to quantify RAS signaling to facilitate development of novel cancer therapeutics. We use targeted proteomics technologies to develop a community resource consisting of 256 validated multiple reaction monitoring (MRM)-based, multiplexed assays for quantifying protein expression and phosphorylation through the receptor tyrosine kinase, MAPK, and AKT signaling networks. As proof of concept, we quantify the response of melanoma (A375 and SK-MEL-2) and colorectal cancer (HCT-116 and HT-29) cell lines to BRAF inhibition by PLX4720. These assays replace over 60 western blots with quantitative mass-spectrometry-based assays of high molecular specificity and quantitative precision, showing the value of these methods for pharmacodynamic measurements and mechanism-of-action studies. Methods, fit-for-purpose validation, and results are publicly available as a resource for the community at assays.cancer.gov. [Display omitted] •Quantitative protein assays are required to understand cancer signaling networks•We develop a suite of multiplexed mass-spectrometry-based assays•The assays offer specific and precise quantification of key networks and PTMs•The assays provide a resource for mechanism-of-action and pharmacodynamic measurements A lack of quantitative, multiplexable assays for phosphosignaling limits comprehensive investigation of aberrant signaling in cancer and evaluation of novel treatments. To alleviate this limitation, we sought to develop assays by using targeted mass spectrometry for quantifying protein expression and phosphorylation through the receptor tyrosine kinase, MAPK, and AKT signaling networks. The resulting assays provide a resource for replacing over 60 western blots in examining cancer signaling and tumor biology with high molecular specificity and quantitative rigor. Whiteaker et al. describe a suite of mass-spectrometry-based assays for quantification of protein expression and phosphorylation in receptor tyrosine kinase, AKT, and MAP-kinase networks. The assays provide a resource for replacing over 60 commonly used cancer signaling and tumor biology western blots with high molecular specificity and quantitative rigor.