Figure 3 from Antigen-Loaded Extracellular Vesicles Induce Responsiveness to Anti–PD-1 and Anti–PD-L1 Treatment in a Checkpoint Refractory Melanoma Model

https://doi.org/10.1158/2326-6066.30721183

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posted on 2025-11-26, 12:46 authored by Rosanne E. Veerman, Gözde Güclüler Akpinar, Annemarijn Offens, Loïc Steiner, Pia Larssen, Andreas Lundqvist, Mikael C.I. Karlsson, Susanne Gabrielsson

EVs alone or combined with checkpoint blockade treatment induces antigen-specific immune responses in a melanoma model. A, Mice with OVA-expressing B16 melanoma were treated with PBS or EVs intravenously and anti–PD-1/PD-L1 intraperitoneally as indicated. B, After inoculation, tumors were measured every 2 to 3 days, and mice were sacrificed when the tumors reached 1,000 mm3 in size. The results represent the mean size of the tumors in mice in each group (n = 10–14). Survival was plotted using a Kaplan–Meier survival curve. C, At the endpoint, tumors were collected and analyzed for immune cell infiltration using flow cytometry. Immune cells were identified as CD45+ and B16 melanoma cells as CD45−. * represents significance compared with the control group. D, B16 melanoma cells were analyzed for surface MHC class I and PD-L1 expression. E, The proportion of infiltrating total, CD8+ and antigen-specific T cells in tumors was analyzed by flow cytometry. F, PD-1 expression on the infiltrating CD8+ T cells was determined using flow cytometry. The graphs show the results of two independent experiments, with 4 to 5 mice per group. Dots represent a single mouse, and data are presented as the mean ± SD. Data were analyzed using the Kruskal–Wallis test with Dunn test for multiple comparisons. The Mantel–Cox test was applied for the survival curve. *, P < 0.05; **, P < 0.01; and ***, P < 0.001.

Funding

Vetenskapsrådet (VR)

Cancerfonden (Swedish Cancer Society)

Radiumhemmets Forskningsfonder (Cancer Research Foundations of Radiumhemmet)

Stockholm läns landsting (Stockholm County Council)

Hjärt-Lungfonden (Swedish Heart-Lung Foundation)

Karolinska Institutet (KI)

History

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DOI - Is supplement to Antigen-Loaded Extracellular Vesicles Induce Responsiveness to Anti–PD-1 and Anti–PD-L1 Treatment in a Checkpoint Refractory Melanoma Model

ARTICLE ABSTRACT

Extracellular vesicles (EV) are important mediators of intercellular communication and are potential candidates for cancer immunotherapy. Immune checkpoint blockade, specifically targeting the programmed cell death protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) axis, mitigates T-cell exhaustion, but is only effective in a subset of patients with cancer. Reasons for therapy resistance include low primary T-cell activation to cancer antigens, poor antigen presentation, and reduced T-cell infiltration into the tumor. Therefore, combination strategies have been extensively explored. Here, we investigated whether EV therapy could induce susceptibility to anti–PD-1 or anti–PD-L1 therapy in a checkpoint-refractory B16 melanoma model. Injection of dendritic cell–derived EVs, but not checkpoint blockade, induced a potent antigen-specific T-cell response and reduced tumor growth in tumor-bearing mice. Combination therapy of EVs and anti–PD-1 or anti–PD-L1 potentiated immune responses to ovalbumin- and α-galactosylceramide–loaded EVs in the therapeutic model. Moreover, combination therapy resulted in increased survival in a prophylactic tumor model. This demonstrates that EVs can induce potent antitumor immune responses in checkpoint refractory cancer and induce anti–PD-1 or anti–PD-L1 responses in a previously nonresponsive tumor model.