title: Quantification of Anaplastic Lymphoma Kinase Protein Expression in Non-Small Cell Lung Cancer Tissues from Patients Treated with Crizotinib.
abstract: BACKGROUND
  Crizotinib has antitumor activity in ALK (anaplastic lymphoma receptor tyrosine kinase)-rearranged non-small cell lung cancer (NSCLC). The current diagnostic test for ALK rearrangement is breakapart fluorescence in situ hybridization (FISH), but FISH has low throughput and is not always reflective of protein concentrations. The emergence of multiple clinically relevant biomarkers in NSCLC necessitates efficient testing of scarce tissue samples. We developed an anaplastic lymphoma kinase (ALK) protein assay that uses multiplexed selected reaction monitoring (SRM) to quantify absolute amounts of ALK in formalin-fixed paraffin-embedded (FFPE) tumor tissue.
  
  
  METHODS
  After validation in formalin-fixed cell lines, the SRM assay was used to quantify concentrations of ALK in 18 FFPE NSCLC samples that had been tested for ALK by FISH and immunohistochemistry. Results were correlated with patient response to crizotinib.
  
  
  RESULTS
  We detected ALK in 11 of 14 NSCLC samples with known ALK rearrangements by FISH. Absolute ALK concentrations correlated with clinical response in 5 of 8 patients treated with crizotinib. The SRM assay did not detect ALK in 3 FISH-positive patients who had not responded to crizotinib. In 1 of these cases, DNA sequencing revealed a point mutation that predicts a nonfunctional ALK fusion protein. The SRM assay did not detect ALK in any tumor tissue with a negative ALK status by FISH or immunohistochemistry.
  
  
  CONCLUSIONS
  ALK concentrations measured by SRM correlate with crizotinib response in NSCLC patients. The ALK SRM proteomic assay, which may be multiplexed with other clinically relevant proteins, allows for rapid identification of patients potentially eligible for targeted therapies.
authors.affiliation: OncoPlex Diagnostics, Rockville, MD; NantOmics, LLC, Rockville, MD;
authors.forename: Todd
authors.initials: T
authors.lastname: Hembrough
authors.affiliation: OncoPlex Diagnostics, Rockville, MD; NantOmics, LLC, Rockville, MD;
authors.forename: Wei-Li
authors.initials: WL
authors.lastname: Liao
authors.affiliation: Department of Pathology, Dartmouth-Hitchcock Medical Center, Lebanon, NH; current affiliation: Department of Pathology and Laboratory Medicine, Medical College of Wisconsin, Milwaukee, WI;
authors.forename: Christopher P
authors.initials: CP
authors.lastname: Hartley
authors.affiliation: Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, Department of Hematology and Medical Oncology, Taussig Cancer Institute, and current affiliation: Mary Babb Randolph Cancer Center, West Virginia University, Morgantown, WV.
authors.forename: Patrick C
authors.initials: PC
authors.lastname: Ma
authors.affiliation: Department of Hematology and Medical Oncology, Taussig Cancer Institute, and.
authors.forename: Vamsidhar
authors.initials: V
authors.lastname: Velcheti
authors.affiliation: Robert J. Tomsich Pathology and Laboratory Medicine Institute, Cleveland Clinic, Cleveland, OH;
authors.forename: Christopher
authors.initials: C
authors.lastname: Lanigan
authors.affiliation: OncoPlex Diagnostics, Rockville, MD; NantOmics, LLC, Rockville, MD;
authors.forename: Sheeno
authors.initials: S
authors.lastname: Thyparambil
authors.affiliation: OncoPlex Diagnostics, Rockville, MD; NantOmics, LLC, Rockville, MD;
authors.forename: Eunkyung
authors.initials: E
authors.lastname: An
authors.affiliation: Mary Babb Randolph Cancer Center, West Virginia University, Morgantown, WV;
authors.forename: Manish
authors.initials: M
authors.lastname: Monga
authors.affiliation: OncoPlex Diagnostics, Rockville, MD; NantOmics, LLC, Rockville, MD;
authors.forename: David
authors.initials: D
authors.lastname: Krizman
authors.affiliation: OncoPlex Diagnostics, Rockville, MD;
authors.forename: Jon
authors.initials: J
authors.lastname: Burrows
authors.affiliation: Department of Pathology, Dartmouth-Hitchcock Medical Center, Lebanon, NH; Geisel School of Medicine at Dartmouth, Hanover, NH; laura.j.tafe@hitchcock.org.
authors.forename: Laura J
authors.initials: LJ
authors.lastname: Tafe
mesh_terms: D000077548:Anaplastic Lymphoma Kinase; D000970:Antineoplastic Agents / Q000494:pharmacology / Q000627:therapeutic use*; D002289:Carcinoma, Non-Small-Cell Lung / Q000175:diagnosis / Q000188:drug therapy* / Q000235:genetics; D045744:Cell Line, Tumor; D000077547:Crizotinib; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D007150:Immunohistochemistry; D017404:In Situ Hybridization, Fluorescence; D008175:Lung Neoplasms / Q000175:diagnosis / Q000188:drug therapy* / Q000235:genetics; D011720:Pyrazoles / Q000494:pharmacology / Q000627:therapeutic use*; D011725:Pyridines / Q000494:pharmacology / Q000627:therapeutic use*; D020794:Receptor Protein-Tyrosine Kinases / Q000032:analysis* / Q000037:antagonists & inhibitors / Q000235:genetics*
chemical_list: D000970:Antineoplastic Agents; D011720:Pyrazoles; D011725:Pyridines; D000077547:Crizotinib; C000626173:ALK protein, human; D000077548:Anaplastic Lymphoma Kinase; D020794:Receptor Protein-Tyrosine Kinases