Immune Checkpoint Inhibitor-Mediated Cancer Theranostics with Radiolabeled Anti-Granzyme B Peptide

Abstract

Although immune checkpoint inhibitors (ICI) have revolutionized cancer management, patient response can be heterogeneous, and the development of ICI resistance is increasingly reported. Novel treatment strategies are necessary not only to expand the use of ICI to previously unresponsive tumor types but also to overcome resistance. Targeted radionuclide therapy may synergize well with ICIs since it can promote a pro-inflammatory tumor microenvironment. We investigated the use of a granzyme B targeted peptide (GZP) as a cancer theranostic agent, radiolabeled with ⁶⁸Ga (⁶⁸Ga-GZP) as a PET imaging agent and radiolabeled with ⁹⁰Y (⁹⁰Y-GZP) as a targeted radionuclide therapy agent for combinational therapy with ICI in murine models of colon cancer. Our results demonstrate that GZP increasingly accumulates in tumor tissue after ICI and that the combination of ICI with ⁹⁰Y-GZP promotes a dose-dependent response, achieving curative response in some settings and increased overall survival.

Keywords: immune checkpoint inhibitors; targeted radionuclide therapy; theranostics.

Figure 1

(A) Representative PET/MR MIP images of CT26 and MC38 tumor-bearing mice with and without ICI treatment (1 h post injection of 68 Ga-NOTA-GZP). (B) Uptake values of blood pool and major organs, showing significantly higher tumor uptake with ICI, represented as %ID/g. * p < 0.05.

Figure 2

Tumor growth curves of (A) CT26 and (B) MC38 tumor-bearing mice after administration of the different therapeutic groups. Checkpoint inhibitors were given days 3, 6 and 9 after tumor implantation and ⁹⁰Y-GZP was given on day 12 (indicated by arrows). High dose of ⁹⁰Y-GZP (22.2 MBq) after ICI achieved complete tumor response in both tumor models. * Statistically significant compared to all other groups (p < 0.05).

Figure 3

(A) Overall survival plots of MC38 and CT26 tumor-bearing mice. Significantly increased overall survival was achieved for ICI followed by 22.2 MBq of ⁹⁰Y-GZP (high dose) in both tumor models as well as ICI followed by 2.2 MBq of ⁹⁰Y-GZP (low dose) in animals bearing MC38 tumors. (B) Body weight measurements carried out throughout the study reveal no signs of overall toxicity in any of the treatment groups. * Statistically significant when compared to all other groups at the end of the study (p < 0.05).

Figure 4

Histological evaluation of major organs reveals no systemic toxicity after both doses of ⁹⁰Y-GZP at either high or low dose. Scale bar = 200 µm.

Figure 5

Immunofluorescence staining against granzyme B (red) of tumor samples after ICI therapy only and ICI + ⁹⁰Y-GZP (high dose) demonstrating high levels of granzyme B presence within the tissue. Nuclei are stained with DAPI (blue). Scale bar = 200 µM.