PB101, a VEGF- and PlGF-targeting decoy protein, enhances antitumor immunity and suppresses tumor progression and metastasis

Eun-Jin Go, Hannah Yang, Seung Joon Lee, Hyun Gul Yang, Jin A Shin, Won Suk Lee, Hye Seong Lim, Hong Jae Chon, Chan Kim, Eun-Jin Go, Hannah Yang, Seung Joon Lee, Hyun Gul Yang, Jin A Shin, Won Suk Lee, Hye Seong Lim, Hong Jae Chon, Chan Kim

Abstract

Antiangiogenic therapy is a recognized method for countering the immunosuppressive tumor microenvironment (TME) and improving anti-tumor immunity. PB101 is a glycosylated decoy receptor that binds to VEGF-A and PlGF with high affinity, based on the VEGFR1 backbone. Here, we elucidated PB101-induced remodeling of tumor angiogenesis and immunity, which enhances anti-PD-L1 immune checkpoint blockade. PB101 inhibited tumor growth by suppressing angiogenesis and enhancing CD8+ T cell infiltration into the tumors. PB101 induced robust reprogramming of antitumor immunity and activates intratumoral CD8+ T cells. Anti-tumor efficacy of PB101 is mostly dependent on CD8+ T cells and IFN-γ. PB101 reprograms tumor immunity in a manner distinct from that of the conventional VEGF decoy receptor, VEGF-trap. With its potent immune-modulating capability, PB101 synergizes with an anti-PD-L1, triggering strengthened antitumor immunity. Combining PB101 and anti-PD-L1 could establish durable protective immunity against tumor recurrence and metastasis. The findings of this study offer scientific rationales for further clinical development of PB101, particularly when used in combination with immune checkpoint inhibitors, as a potential treatment for advanced cancers.

Keywords: Angiogenesis; PB101; PlGF; VEGF-A; combination immunotherapy; immune checkpoint inhibitor; tumor immunity.

Conflict of interest statement

CK and HJC has received research grants from Panolos Bioscience. HGY, JAS, and HSL are full-time employees of Panolos Bioscience. The other authors have no potential conflicts of interest to disclose.

© 2023 The Author(s). Published with license by Taylor & Francis Group, LLC.

Figures

Figure 1.
Figure 1.
PB101 suppresses tumor angiogenesis and enhances intratumoral CD8+ T cell infiltration. a. Diagram depicting treatment schedule in MC38 colon cancer. Red and black arrows indicate treatment and sacrifice, respectively. b. Comparison of tumor growth in mice treated with phosphate-buffered saline (PBS) or PB101. Pooled data from three independent experiments with N = 10 per group. c. Representative images and quantifications of CD31+ blood vessels and CD8+ T cells within tumors. d. Representative images and quantifications of caspase 3+ apoptosis cells within tumors. N = 6 per group. Scale bars, 100 µm. Values are shown as the mean ± SD. *P < .05 vs control.
Figure 2.
Figure 2.
PB101 reprograms antitumor immunity. a – c. Representative flow cytometric plot and comparisons of CD11c+ dendritic cells (a), CD11b+F4/80+ tumor-associated macrophages (TAMs), and of NOS2+ M1-like and Arginase 1+ M2-like TAMs (c) within tumors. d. Representative images and quantification of NOS2+ M1-like macrophages and CD206+ M2-like macrophages within the tumors. Scale bars, 20 µm. E and F. Representative flow cytometric plot and comparison of tumor-infiltrating CD3+ T cells (e) and CD8+ T cells (f). G. Representative images and quantification of granzyme B (GzB)-expressing CD8+ T cells. Scale bars, 100 µm. N = 6–7 per group. Values are shown as the mean ± SD. *P < .05 vs control.
Figure 3.
Figure 3.
PB101 depends on CD8+ T cells and interferon (IFN)-γ to suppress tumor growth. a. Heatmap of NanoString immune-related gene expression in PB101-treated or PBS-treated control tumors. N = 4 per group. b. Volcano plot showing the changes of gene expression in PB101-treated tumors compared to control tumors. c. Dot plot showing enrichment of gene ontology (GO) biological processes for immune-related genes in tumors treated with PB101. The size of each dot represents the number of genes significantly associated with a GO term, and the color of the dots represent the P-adjusted value. d. Comparisons of gene expression related to the tumor microenvironment (TME), lymphocyte (LC)-endothelial cell (EC) interaction, dendritic cell, macrophage, IFN, Th1/Th2 responses, and T cell activation. *P < .05 vs. control. e. Diagram depicting the treatment schedule of the PB101, anti-CD4, anti-CD8 and anti-IFN-γ. f. Comparison of individual tumor growth over time. N = 7 per group. *P < .05 vs PB101.
Figure 4.
Figure 4.
PB101 shows distinct immunologic mechanisms of action compared to VEGF-trap. a. Diagram depicting treatment schedule. b. Comparison of MC38 tumor growth. C – G. Comparisons of dendritic cell (c), TAMs, M1/M2-like macrophages, M1/M2 ratio (d), activated CD8+ T cell (e), CD4+ T cell, regulatory T cell (Treg) (f), and CD8+ T cell (CD8+) and Treg ratio within the tumor (g). H – K. Comparisons of gene expressions related to innate immunity (mavs, Tmem173, and Myd88) (h), cytotoxic lymphocytes (Gzma, Gzmb, Prf1) (i), angiogenesis (Vegfa, Plgf) (j), and transcription factors (Hif1α and Creb1) (k). Values are shown as the mean ± SD. *P < .05.
Figure 5.
Figure 5.
Combination immunotherapy of PB101 and anti-PD-L1 further suppressed tumor progression and prolonged overall survival. a. Diagram depicting treatment schedule in MC38 colon cancer. b. Comparison of MC38 colon cancer growth. Mean and individual tumor growth curves over time. N = 7 per group. c. Diagram depicting treatment schedule in Hepa-V liver cancer. d. Comparison of liver cancer growth. Mean and individual tumor growth curves over time e. Kaplan-Meier survival curves showing overall survival of liver cancer-bearing mice. Table shows P-values determined by log-rank test. Values are shown as the mean ± SD. *P < .05 vs. control, *P < .05 vs. PB101. N = 8–11 per group.
Figure 6.
Figure 6.
Combination immunotherapy of PB101 and anti-PD-L1 provide long-term immune protection against recurrence and metastasis. a. Diagram depicting re-challenge schedule in Hepa-V liver cancer. Red and blue arrows indicate treatments and black arrow indicates subcutaneous and intravenous re-challenge of Hepa-V liver cancer cells. b. Comparison of Hepa-V tumor growth. Mean and individual tumor growth curves over time. N = 4–10 per group. c. Representative gross images and hematoxylin and eosin (H&E) staining images of lung. Arrow heads indicate pulmonary metastatic lesions. d. Comparison of the number of metastatic colonies per lung section. Values are shown as mean ± SD. *P < .05 vs naïve. N = 4–6 per group.

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