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ALK / ROS1 Pathway

Anaplastic lymphoma kinase (ALK) and c-ros oncogene 1 (ROS1) are related receptor tyrosine kinases (RTKs). ALK and ROS1 have been identified as mutant C-terminal (kinase domain) fusion proteins in research studies of a wide range of human cancers (1-10). Table 1 lists some of the ALK and ROS1 fusion partners that have been identified and the associated cancer types.

RTKs Inhibitors Fusion Partners Associated Cancers
ALK Crizotinib5 EML42,11, TFG1, KIF5B6, NPM7 NSCLC1,2, ALCL7, CRC11
ROS1 Crizotinib5 FIG8, CD749, SLC34A29,11 Glioblastoma8, Cholangiocarcinoma10, Ovarian cancer3, NSCLC1,4, CRC11
Table 1. Summary of ALK and ROS1 fusion partners and associated cancers. Abbreviations:  ALCL=Anaplastic large cell lymphoma  EML4=Echinoderm microtubule-associated protein-like 4  NPM=Nucleophosmin  NSCLC=Non-small cell lung cancer  TFG=TRK-fused gene
  1. Rikova, K. et al. (2007) Cell 131, 1190–1203.
  2. Takeuchi, K. et al. (2008) Clin Cancer Res 14, 6618–6624.
  3. Birch, A.H. et al. (2011) PLoS One 6, e28250.
  4. Rimkunas, V.M. et al. (2012) Clin Cancer Res 18, 4449–4457.
  5. D’Arcangelo, M. et al. (2013) Curr Opin Oncol 25, 121–129.
  6. Takeuchi, K. et al. (2009) Clin Cancer Res 15, 3143–3149.
  7. Morris, S.W. et al. (1994) Science 263, 1281–1284.
  8. Charest, A. et al. (2003) Genes Chromosomes Cancer 37, 58–71.
  9. Stumpfova, M. and Jänne, P.A. (2012) Clin Cancer Res 18, 4222–4224.
  10. Gu, T.L. et al. (2011) PLoS One 6, e15640.
  11. Aisner, D.L. et al (2013) Mol Cancer Res (Epub ahead of print)

ALK and ROS1 Antibodies from CST

Cell Signaling Technology recommends two highly sensitive antibodies for detection of ALK and ROS1 full-length proteins and C-terminal fusion proteins.

Features and Benefits:

  • C-terminal Epitopes – enable detection of full-length protein and C-terminal fusion oncoproteins.
  • Highly Sensitive Antibodies – allow detection of endogenous levels of ROS1 or ALK.
  • No Cross-reactivity with Other Family Members – ensures specific detection of ROS1 or ALK protein.
  • Rabbit Monoclonal Antibodies – CST’s proprietary XMT® technology allows careful selection of antibodies for relevant applications.

ROS1 (D4D6) Rabbit mAb #3287 IHC-P (paraffin)

3287 IHC

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using ROS1 (D4D6) Rabbit mAb #3287. Note: Staining is of FIG-ROS1 fusion (4).

3287 WB

Western blot analysis of extracts from HCC78 (SLC34A2-ROS1), U-118 MG (FIG-ROS1), and HeLa (ROS1 negative) cells using ROS1 (D4D6) Rabbit mAb #3287 (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower). Note: HCC78 cells express the 85, 70, and 59 kDa forms of the SLC34A2-ROS1 fusion protein (1).

ALK (D5F3) XP® Rabbit mAb #3633 IHC-P (paraffin)

3633 IHC

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using ALK (D5F3) XP® Rabbit mAb #3633.

3633 WB

Western blot analysis of extracts from NCI-H2228 and NCI-H3122 cells using ALK (D5F3) XP® Rabbit mAb #3633. Variants denoting fusions of different EML4 exons (v1 or v3) are indicated.

Visit our product pages for more information and additional validation data:

CST's Research to Identify ALK and ROS1 Fusion Proteins in NSCLC

Step 1

CST performed an unbiased, large-scale survey of tyrosine kinase activity in lung cancer using PTMScan® Technology. This proprietary technology, developed at Cell Signaling Technology(11), uses a CST™ Motif Antibody for immunoaffinity purification of peptides from digested cell extracts combined with LC tandem mass spectrometry to identify and quantify changes in post-translational modifications such as phosphorylation, acetylation, or ubiquitination. For this study, we used a phospho-tyrosine motif antibody to analyze changes in phosphorylation across the proteome in NSCLC cell lines and tissues.

PTMScan Flow Chart
 

Step 2

Using PTMScan® Technology, we surveyed the phosphotyrosine status of receptor tyrosine kinases (RTK) and non-receptor tyrosine kinases in 41 NSCLC cell lines and over 150 NSCLC tumors. Over 50 tyrosine kinases and more than 2,500 downstream substrates that play roles in NSCLC growth and progression were identified. Two very exciting findings from this study were the identification of novel anaplastic lymphoma kinase (ALK) and c-ros oncogene 1 (ROS1) C-terminal fusion proteins in some NSCLC cell lines and tumors.

PTMScan Flow Chart
 

Step 3

Rabbit monoclonal antibodies specific to ALK and ROS1 [ALK (D5F3) XP® Rabbit mAb #3633; ROS1 (D4D6) Rabbit mAb #3287] were developed to detect both full-length and C-terminal fusion proteins. These antibodies have been validated for IHC and can detect ALK and ROS1 fusion protein expression in NSCLC sample(12, 4).

ALK IHC

IHC analysis of paraffin-embedded human lung carcinoma using ROS1 (D4D6) Rabbit mAb #3287. Note: Staining is of FIG-ROS1 fusion protein(4).

 

Step 4

A partnership with Pfizer, Inc., creator of the ALK inhibitor crizotinib, and Ventana Medical Systems, Inc., a leader in companion diagnostic testing, was formed to develop the use of ALK (D5F3) XP® Rabbit mAb in an automated diagnostic IHC screening assay to detect ALK fusion proteins in NSCLC patient samples.

Patient samples that stain positive for ALK expression may be candidates for crizotinib, which was approved for use in the U.S. in August 2011.

ALK IHC

IHC analysis of paraffin-embedded human lung carcinoma with high (upper) and low levels (lower) of ALK expression using ALK (D5F3) XP® Rabbit mAb #3633.

 

ALK and ROS1-related Publications from CST’s Research Group

CST scientists are highlighted in bold.

2012

2011

2010

2009

2008

  • Guo A, Villén J, Kornhauser J, Lee KA, Stokes MP, Rikova K, Possemato A, Nardone J, Innocenti G,Wetzel R,Wang Y, MacNeill J, Mitchell J, Gygi SP, Rush J, Polakiewicz RD, Comb MJ (2008) Signaling networks assembled by oncogenic EGFR and c-Met. Proc. Natl. Acad. Sci. U.S.A. 105(2), 692-7.

2007

2005

2004

2002