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Three ERBB4-Dependent Human Melanoma Cell Lines Are Also Dependent on ERBB2

Madison Zelan¹, Vipasha Dwivedi¹, Lauren M. Lucas¹, Ella Wilson¹, and David J. Riese II^1,2

¹Department of Drug Discovery and Development, Harrison College of Pharmacy, Auburn University, Auburn, AL 36849 USA

²Cancer Biology and Immunology Program, O’Neal Comprehensive Cancer Center, The University of Alabama at Birmingham, AL 35294 USA

Background/Purpose:

Our overall goal is the discovery of targeted therapies for metastatic melanomas that possess wild-type (WT) BRAF alleles. The proliferation of four BRAF-WT human melanoma cell lines is dependent on ERBB4, which encodes a protein that is closely related to the epidermal growth factor receptor (EGFR), ERBB2 (HER2), and ERBB3 (HER3). Yet, a synthetic, constitutively active and homodimerized ERBB4 mutant (Q646C) inhibits the proliferation of a BRAF-WT human melanoma cell line.

Hypothesis:

Thus, here we begin to test the hypothesis that heterodimers of ERBB4 with either EGFR or ERBB2 drive the proliferation of BRAF-WT human melanoma cell lines.

Methods:

We have developed recombinant retroviral expression vectors that express a neomycin gene along with the wild-type EGFR, the dominant-negative EGFR (K721A) mutant, wild-type ERBB2, or the dominant-negative ERBB2 mutant (references 1 and 2).

Methods (continued):

We have developed a process for packaging these recombinant retroviral expression vectors into amphotropic retrovirus particles (references 3 and 4).

We have developed an assay to determine the effects of EGFR or ERBB2 alleles on the clonogenic proliferation of four ERBB4-dependent, BRAF-WT human melanoma cell lines (references 4 and 5).

References:

DJ Riese II. Recombinant retroviral expression vectors based on pLXSN that encode EGFR, ERBB2, ERBB3, and ERBB4. Protocols.io (2023). https://doi.org/10.17504/protocols.io.261gedd97v47/v1
E Wilson, M Scott, V Dwivedi, DJ Riese II, and M. Zelan. Recombinant retroviral expression vectors that encode dominant-negative mutant alleles of EGFR and ERBB2. Protocols.io (2024). https://doi.org/10.17504/protocols.io.bp2l6x5w1lqe/v1
V Dwivedi, J Davis, E Wilson, M Scott, M Zelan, N DeFeo, V Huffman, Rees Cooke, K O’Daniel, and DJ Riese II. Stable transfection of plasmid DNA into adherent rodent cell lines using calcium phosphate. Protocols.io (2024). https://doi.org/10.17504/protocols.io.rm7vzxz5rgx1/v1
LM Lucas, RL Cullum, JN Woggerman, V Dwivedi, JA Markham, CM Kelley, EL Knerr, LJ Cook, HC Lucas II, DS Waits, TM Ghosh, KM Halanych, RB Gupta, and DJ Riese II. ERBB4 drives the proliferation of BRAF WT melanoma cell lines. medRxiv, 2022 (updated 2023). https://doi.org/10.1101/2022.06.20.22276663
V Dwivedi, LM Lucas, R Cooke, J Davis, E Wilson, M Scott, K O'Daniel, C Dion, J Kerley, M Zelan, N DeFeo, V Huffman, MN Ingrao, and DJ Riese II. Human melanoma cell lines that possess wild-type BRAF alleles but are dependent on ERBB4 and ERBB2. bioRxiv (2024). https://doi.org/10.1101/2024.08.22.609260

Disclosures:

Dr. Riese has been a consultant for the United States National Institutes of Health, the Israel Science Foundation, Bristol-Myers Squibb, Eli Lilly and Company, ImClone Systems, the European Union, the Austrian Science Fund (FWF Der Wissenschaftsfonds), the Children’s Tumor Foundation, and General Dynamics Information Technology.

Contact Information:

Madison Zelan: mmz0008@auburn.edu

David J. Riese II: djr0009@auburn.edu

Results:

The dominant-negative ERBB2 mutant inhibits the proliferation of the IPC-298, MEL-JUSO, and MeWo BRAF-WT human melanoma cell lines. The dominant-negative EGFR mutant inhibits the proliferation of the MeWo BRAF-WT human melanoma cell line. Finally, the proliferation of the SK-MEL-2 BRAF-WT human melanoma cell line is not inhibited by the dominant-negative ERBB2 mutant nor the dominant-negative EGFR mutant (reference 5).

Conclusions and Future Directions:

These data strongly suggest that ERBB4-ERBB2 heterodimers drive the proliferation of the female IPC-298 and MEL-JUSO BRAF-WT melanoma cell lines. Likewise, both ERBB4-ERBB2 and ERBB4-EGFR heterodimers appear to drive the proliferation of the male MeWo BRAF-WT melanoma cell line. Finally, ERBB4 appears to drive the proliferation of the male SK-MEL-2 BRAF-WT melanoma cell line via a mechanism other then ERBB4 heterodimerization. We are pursuing orthogonal approaches to test these hypotheses and we will also investigate the role of endogenous ERBB receptor ligands in driving the proliferation of ERBB4-dependent BRAF-WT melanomas.