# Development of BRAF Dimer Inhibitors to Treat Drug Resistant Melanoma

> **NIH NIH R01** · UNIVERSITY OF PENNSYLVANIA · 2021 · $643,599

## Abstract

The overall goal of this proposal is to use the RAS-RAF-MEK-ERK (MAPK/ERK) signaling pathway as a
model to develop small molecule inhibitors that specifically target kinase dimers as lead compounds for
therapeutic development. The MAPK/ERK signaling pathway is frequently activated in human cancer and the
activating BRAF V600E mutant (BRAFV600E) in particular accounts for about 7% of all cancers and about 50%
of malignant melanoma tumors, thus highlighting BRAF and MAPK/ERK signaling as important targets for
therapy. Although many melanoma patients treated with highly selective BRAFV600E mutant inhibitors (BRAFi),
downstream MEK inhibitors (MEKi) or BRAFi/MEKi combinations, initially respond, nearly all develop drug
resistance and progress at a median of about 1 year survival for combination therapy. Intrinsic or acquired
resistance to BRAF inhibitors often arise through potent activation or reactivation of the MAPK pathway,
typically through mutation of upstream RAS or downstream MEK. Biochemically, MAPK activation is driven by
RAF dimerization, whereby drug-bound BRAF or BRAFV600E mediates drug-induced allosteric activation of a
wild-type RAF subunit (BRAF or CRAF) of the kinase dimer, a process called ‘transactivation’ or ‘paradoxical
activation.’ To test if transactivation could be overcome by selectively targeting RAF dimers, we used
BRAFV600E and vemurafenib as a model system to develop bivalent kinase inhibitors to lock RAF dimers in an
inactive conformation that cannot undergo transactivation. This structure-based design effort resulted in the
development of Vem-BisAmide-2, a compound containing two vemurafenib molecules connected by a bis
amide linker. We showed that Vem-BisAmide-2 has comparable inhibitory potency as vemurafenib to
BRAFV600E both in vitro and in cells, but promotes an inactive dimeric BRAFV600E conformation that is unable to
undergo transactivation. In related studies, we also chemically linked vemurafenib to the MEK inhibitor G894,
and demonstrated that this compound inhibited growth of BRAFi-resistant cells more effectively than either
vemurafenib or G894 alone or a vemurafenib/G894 combination. We have also carried out a high-throughput
assay to measure inhibition of the BRAF/MEK interaction with the goal of developing small molecule BRAF-
MEK dimerizatioin inhibitors. With this preliminary data on hand, we will now (1) Develop inhibitors that target
BRAF homo- and hetero-dimers with BRAFV600E, BRAF or CRAF, and (2) Develop inhibitors that target
RAF/MEK complexes. These studies will lead to the development of small molecule inhibitors to target BRAF
homo- and hetero-dimers with BRAF, CRAF, BRAFV600E and MEK that we anticipate will be superior to
inhibitors that target kinase monomers for inhibiting MAPK/ERK signaling in melanoma and melanoma
resistant to therapy. These lead inhibitors will provide a new paradigm for the development of a new family of
melanoma drugs.

## Key facts

- **NIH application ID:** 10058819
- **Project number:** 5R01CA226888-03
- **Recipient organization:** UNIVERSITY OF PENNSYLVANIA
- **Principal Investigator:** Ronen Marmorstein
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $643,599
- **Award type:** 5
- **Project period:** 2018-12-03 → 2023-11-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10058819

## Citation

> US National Institutes of Health, RePORTER application 10058819, Development of BRAF Dimer Inhibitors to Treat Drug Resistant Melanoma (5R01CA226888-03). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10058819. Licensed CC0.

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