Abstract
Purpose
Despite unprecedented responses to immune checkpoint inhibitors and targeted therapy in melanoma, a major subset of patients progresses and have few effective salvage options. We have previously demonstrated robust, selective uptake of the peptidomimetic LLP2A labeled with Cu-64 ([64Cu]-LLP2A) for positron emission tomography (PET) imaging in subcutaneous and metastatic models of B16F10 murine melanoma. LLP2A binds with high affinity to very late antigen-4 (VLA-4, integrin α4β1), a transmembrane protein overexpressed in melanoma and other cancers that facilitates tumor growth and metastasis. Yet B16F10 fails to faithfully reflect human melanoma biology, as it lacks certain oncogenic driver mutations, including BRAF mutations found in ≥ 50 % of clinical specimens. Here, we evaluated the PET tracer [64Cu]-CB-TE1A1P-PEG4-LLP2A ([64Cu]-LLP2A) in novel, translational BRAFV600E mutant melanoma models differing in VLA-4 expression—BPR (VLA-4−) and BPRα (VLA-4+).
Procedures
BPR cells were transduced with α4 (CD49d) to overexpress intact cell surface VLA-4 (BPRα). The binding affinity of [64Cu]-LLP2A to BPR and BPRα cells was determined by saturation binding assays. [64Cu]-LLP2A internalization into B16F10, BPR, and BPRα cells was quantified via a plate-based assay. Tracer biodistribution and PET/CT imaging were evaluated in mice bearing subcutaneous BPR and BPRα tumors.
Results
[64Cu]-LLP2A demonstrated high binding affinity to BPRα (Kd = 1.4 nM) but indeterminate binding to BPR cells. VLA-4+ BPRα and B16F10 displayed comparable time-dependent [64Cu]-LLP2A internalization, whereas BPR internalization was undetectable. PET/CT showed increased tracer uptake in BPRα tumors vs. BPR tumors in vivo, which was validated by significantly greater (p < 0.0001) BPRα tumor uptake in biodistribution analyses.
Conclusions
[64Cu]-LLP2A discriminates BPRα (VLA-4+) vs. BPR (VLA-4−) melanomas in vivo, supporting translation of these BRAF-mutated melanoma models via prospective imaging and theranostic studies. These results extend the utility of LLP2A to selectively target clinically relevant and therapy-resistant tumor variants toward its use for therapeutic patient care.
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Acknowledgements
The authors are grateful to Susan Rottinghaus and the Cell and Immunobiology Core at the University of Missouri for technical assistance. Figure 1a was created with BioRender.com.
Funding
This work was funded by K08 CA241319 and R01 CA214018. This work utilized the Hillman Cancer Center In Vivo Imaging Facility, a shared resource at the University of Pittsburgh supported by the CCSG P30 CA047904. RP received support from the Hillman Cancer Center Early Career Fellowship for Innovative Cancer Research.
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MB and LN: Research design, data acquisition and analysis, drafting and revising the paper. JL, K-VH, RF, and JT: Data acquisition and analysis. KD, MP, FG, and SN: Data acquisition. K-VH, RSE, WS, RP, and CJA: Research design and revising the paper.
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Michael C. Bellavia and Lea Nyiranshuti contributed equally to this work.
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Bellavia, M.C., Nyiranshuti, L., Latoche, J.D. et al. PET Imaging of VLA-4 in a New BRAFV600E Mouse Model of Melanoma. Mol Imaging Biol 24, 425–433 (2022). https://doi.org/10.1007/s11307-021-01666-1
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DOI: https://doi.org/10.1007/s11307-021-01666-1