title: Monitoring metabolic responses to chemotherapy in single cells and tumors using nanostructure-initiator mass spectrometry (NIMS) imaging.
abstract: BACKGROUND
  Tissue imaging of treatment-induced metabolic changes is useful for optimizing cancer therapies, but commonly used methods require trade-offs between assay sensitivity and spatial resolution. Nanostructure-Initiator Mass Spectrometry imaging (NIMS) permits quantitative co-localization of drugs and treatment response biomarkers in cells and tissues with relatively high resolution. The present feasibility studies use NIMS to monitor phosphorylation of 3'-deoxy-3'-fluorothymidine (FLT) to FLT-MP in lymphoma cells and solid tumors as an indicator of drug exposure and pharmacodynamic responses.
  
  
  METHODS
  NIMS analytical sensitivity and spatial resolution were examined in cultured Burkitt's lymphoma cells treated briefly with Rapamycin or FLT. Sample aliquots were dispersed on NIMS surfaces for single cell imaging and metabolic profiling, or extracted in parallel for LC-MS/MS analysis. Docetaxel-induced changes in FLT metabolism were also monitored in tissues and tissue extracts from mice bearing drug-sensitive tumor xenografts. To correct for variations in FLT disposition, the ratio of FLT-MP to FLT was used as a measure of TK1 thymidine kinase activity in NIMS images. TK1 and tumor-specific luciferase were measured in adjacent tissue sections using immuno-fluorescence microscopy.
  
  
  RESULTS
  NIMS and LC-MS/MS yielded consistent results. FLT, FLT-MP, and Rapamycin were readily detected at the single cell level using NIMS. Rapid changes in endogenous metabolism were detected in drug-treated cells, and rapid accumulation of FLT-MP was seen in most, but not all imaged cells. FLT-MP accumulation in xenograft tumors was shown to be sensitive to Docetaxel treatment, and TK1 immunoreactivity co-localized with tumor-specific antigens in xenograft tumors, supporting a role for xenograft-derived TK1 activity in tumor FLT metabolism.
  
  
  CONCLUSIONS
  NIMS is suitable for monitoring drug exposure and metabolite biotransformation with essentially single cell resolution, and provides new spatial and functional dimensions to studies of cancer metabolism without the need for radiotracers or tissue extraction. These findings should prove useful for in vitro and pre-clinical studies of cancer metabolism, and aid the optimization of metabolism-based cancer therapies and diagnostics.
authors.affiliation: Pfizer Worldwide Research and Development, La Jolla Laboratories, La Jolla, CA, USA. peter.obrien2@pfizer.com.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: O'Brien
authors.forename: Michelle
authors.initials: M
authors.lastname: Lee
authors.forename: Mary E
authors.initials: ME
authors.lastname: Spilker
authors.forename: Cathy C
authors.initials: CC
authors.lastname: Zhang
authors.forename: Zhengming
authors.initials: Z
authors.lastname: Yan
authors.forename: Timothy C
authors.initials: TC
authors.lastname: Nichols
authors.forename: Wenlin
authors.initials: W
authors.lastname: Li
authors.forename: Caroline H
authors.initials: CH
authors.lastname: Johnson
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Patti
authors.forename: Gary
authors.initials: G
authors.lastname: Siuzdak