BRAF mutations occur in a significant proportion of human tumors, and represent a mechanism of constitutive activation of the MAPK pathway. We have demonstrated that activating mutations of BRAF confer sensitivity to small molecule inhibitors of the pathway. In contrast, we showed that HER2-overexpressing breast carcinomas were resistant to MEK inhibition, despite effective pharmacologic inhibition of MAPK activity. Our preliminary data suggest that tumors with HER kinase activation (and WT BRAF) and those with oncogenic BRAF have similar levels of phosphorylated ERK;however, BRAF mutant tumors have higher levels of phosphorylated MEK and selected MEK-ERK dependent transcripts. Further, MEK inhibitor-induced feedback upregulation of the pathway is seen only in the receptor-activated tumors, but not in tumors with activating BRAF mutation. We hypothesize that increased output of the MAPK pathway in B- RAF mutant tumors compared to HER kinase-activated tumors is due to the impairment of feedback inhibition of the pathway, upstream of, and/or at the level of, RAF. We hypothesize that disabling of upstream feedback in BRAF mutant tumors causes an increase in pathway throughput, resulting in increased expression of ERK effectors, and targets responsible for pathway feedback (DUSP, SPRY proteins). This increase in DUSPs (MAP kinase phosphatases) may be critical for the downregulation of ERK to physiologically tolerated levels. The increase in both feedback and effector proteins may together be responsible for aspects of the transformed phenotype. In this proposal, we describe further preliminary data which support these assertions, and describe a research plan to determine the mechanism of feedback response to MEK inhibition. We will determine whether phosphorylated ERK represents an accurate reflection of pathway activation. We will use small molecule inhibitors of the pathway, as well as RNA interference to determine the role of critical proteins in the feedback program. Finally, we will develop isogenic systems transformed by oncogenic BRAF and activated receptor tyrosine kinases to determine if specific feedback and effector protein expression patterns can be generated. The translational goals of these studies are to identify feedback pathways modulating the response to RAF and MEK inhibitors which will impact the effectiveness of these compounds in clinical trials.

Public Health Relevance

The RAS-RAF-MEK-ERK signaling pathway is an important regulator of tumor cell growth and proliferation. The goals of this research proposal are to understand feedback mechanisms which regulate the activation of this pathway. Understanding this regulatory mechanism will help investigators use drugs which target this pathway to select patients whose tumors are most likely to respond to them. PHS 398/2590 (Rev. 11/07) Page Continuation Format Page

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08CA127350-04
Application #
8321029
Study Section
Subcommittee G - Education (NCI)
Program Officer
Perkins, Susan N
Project Start
2009-09-21
Project End
2014-08-31
Budget Start
2012-09-01
Budget End
2013-08-31
Support Year
4
Fiscal Year
2012
Total Cost
$173,232
Indirect Cost
$12,832
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
Xing, F; Persaud, Y; Pratilas, C A et al. (2012) Concurrent loss of the PTEN and RB1 tumor suppressors attenuates RAF dependence in melanomas harboring (V600E)BRAF. Oncogene 31:446-57
Ambrosini, Grazia; Pratilas, Christine A; Qin, Li-Xuan et al. (2012) Identification of unique MEK-dependent genes in GNAQ mutant uveal melanoma involved in cell growth, tumor cell invasion, and MEK resistance. Clin Cancer Res 18:3552-61
Pratilas, Christine A; Solit, David B (2010) Targeting the mitogen-activated protein kinase pathway: physiological feedback and drug response. Clin Cancer Res 16:3329-34