The long-term goals of this work are the development of therapeutic strategies to improve the survival of patients with disseminated melanoma. There is already good evidence that mutated BRAF is a bona fide therapeutic target in over 50% of melanomas, and that impressive but short-lived clinical responses can be achieved with small molecule BRAF inhibitors (such as PLX4032). Clinically, BRAF inhibitor resistance follows a course where tumor regression is followed by quiescence and eventual relapse. The overall hypothesis is that BRAF inhibition remodels both the melanoma and host microenvironments to provide a protective "sanctuary" for the minor populations of melanoma cells that escape therapy. Conceptually, it is believed that there are at least two forms of environment-mediated therapeutic escape;the first, defined as "tumor intrinsic", was observed in melanoma cell lines that lack PTEN function (PTEN-) and involved the establishment of autocrine integrin/extracellular matrix (ECM) signaling loops that bypass BRAF signaling and downregulate apoptosis. The second was host-mediated, where BRAF inhibition in primary human fibroblasts paradoxically activated AKT and MEK signaling leading to the expression of PDGF-D, VEGF and the Notch ligand Jagged as well as increasing the deposition of fibronectin, collagens and laminin.
The first aim will investigate how BRAF inhibition leads to the acquisition of autocrine ECM-driven signaling loops and will determine how these altered adhesion signals "re-wire" the signaling of the escaping population and leads to the adoption of a phenotype that is slow-proliferating, apoptosis resistant and pro-invasive. We will then test whether therapeutic targeting of the melanoma cell/ECM interactions ameliorates intrinsic resistance by preventing the microenvironment- mediated reorganization of the melanoma signaling network.
In aim 2, we will test the hypothesis that BRAF inhibition activates host fibroblasts leading to the creation of a "refuge" vascular microenvironment that allows PTEN+ melanoma cells to escape from therapy. We will address how BRAF inhibition in normal host fibroblasts leads to the establishment of autocrine growth factor signaling loops that drives their activation. We will then use novel 3D melanoma/fibroblast/endothelial cell co-culture and in vivo xenograft models to investigate the mechanisms by which inhibition of BRAF in fibroblasts drives the angiogenic response and will elucidate the role of the vascular niche as a protective "sanctuary" for the escaping melanoma cells. It is expected that knowledge gained from this work will provide novel therapeutic strategies for overcoming BRAF inhibitor resistance in the clinic. 1

Public Health Relevance

Although small molecule BRAF inhibitors are showing great promise as novel melanoma therapies, their effectiveness is severely limited by both acquired and intrinsic drug resistance. The aim of the proposal is to elucidate the mechanisms by which the host and tumor microenvironments contribute towards both of these resistance mechanisms. We expect this research to define novel microenvironment-directed strategies for delaying and abrogating drug resistance in melanoma that can be evaluated in future clinical trials.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA161107-03
Application #
8479132
Study Section
Basic Mechanisms of Cancer Therapeutics Study Section (BMCT)
Program Officer
Forry, Suzanne L
Project Start
2011-08-01
Project End
2016-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
3
Fiscal Year
2013
Total Cost
$325,734
Indirect Cost
$130,684
Name
H. Lee Moffitt Cancer Center & Research Institute
Department
Type
DUNS #
139301956
City
Tampa
State
FL
Country
United States
Zip Code
33612
Smyth, Tomoko; Paraiso, Kim H T; Hearn, Keisha et al. (2014) Inhibition of HSP90 by AT13387 delays the emergence of resistance to BRAF inhibitors and overcomes resistance to dual BRAF and MEK inhibition in melanoma models. Mol Cancer Ther 13:2793-804
Haarberg, H Eirik; Smalley, Keiran S M (2014) Resistance to Raf inhibition in cancer. Drug Discov Today Technol 11:27-32
Rebecca, Vito W; Smalley, Keiran S M (2014) Change or die: targeting adaptive signaling to kinase inhibition in cancer cells. Biochem Pharmacol 91:417-25
Johnson, Douglas B; Smalley, Keiran S M; Sosman, Jeffrey A (2014) Molecular pathways: targeting NRAS in melanoma and acute myelogenous leukemia. Clin Cancer Res 20:4186-92
Rebecca, Vito W; Alicea, Gretchen M; Paraiso, Kim H T et al. (2014) Vertical inhibition of the MAPK pathway enhances therapeutic responses in NRAS-mutant melanoma. Pigment Cell Melanoma Res 27:1154-8
Rebecca, Vito W; Massaro, Renato R; Fedorenko, Inna V et al. (2014) Inhibition of autophagy enhances the effects of the AKT inhibitor MK-2206 when combined with paclitaxel and carboplatin in BRAF wild-type melanoma. Pigment Cell Melanoma Res 27:465-78
Fedorenko, Inna V; Fang, Bin; Koomen, John M et al. (2014) Amuvatinib has cytotoxic effects against NRAS-mutant melanoma but not BRAF-mutant melanoma. Melanoma Res 24:448-53
Rebecca, Vito W; Wood, Elizabeth; Fedorenko, Inna V et al. (2014) Evaluating melanoma drug response and therapeutic escape with quantitative proteomics. Mol Cell Proteomics 13:1844-54
Beck, Daniela; Niessner, Heike; Smalley, Keiran S M et al. (2013) Vemurafenib potently induces endoplasmic reticulum stress-mediated apoptosis in BRAFV600E melanoma cells. Sci Signal 6:ra7
Gibney, Geoffrey T; Smalley, Keiran S M (2013) An unholy alliance: cooperation between BRAF and NF1 in melanoma development and BRAF inhibitor resistance. Cancer Discov 3:260-3

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