Non-invasive Detection of Immunotherapy-Induced Adverse Events

Abstract

Purpose: Cancer immunotherapy has markedly improved the prognosis of patients with a broad variety of malignancies. However, benefits are weighed against unique toxicities, with immune-related adverse events (irAE) that are frequent and potentially life-threatening. The diagnosis and management of these events are challenging due to heterogeneity of timing onset, multiplicity of affected organs, and lack of non-invasive monitoring techniques. We demonstrate the use of a granzyme B–targeted PET imaging agent (GZP) for irAE identification in a murine model. Experimental Design: We generated a model of immunotherapy-induced adverse events in Foxp3–DTR–GFP mice bearing MC38 tumors. GZP PET imaging was performed to evaluate organs non-invasively. We validated imaging with ex vivo analysis, correlating the establishment of these events with the presence of immune infiltrates and granzyme B upregulation in tissue. To demonstrate the clinical relevance of our findings, the presence of granzyme B was identified through immunofluorescence staining in tissue samples of patients with confirmed checkpoint inhibitor–associated adverse events. Results: GZP PET imaging revealed differential uptake in organs affected by irAEs, such as colon, spleen, and kidney, which significantly diminished after administration of the immunosuppressor dexamethasone. The presence of granzyme B and immune infiltrates were confirmed histologically and correlated with significantly higher uptake in PET imaging. The presence of granzyme B was also confirmed in samples from patients that presented with clinical irAEs. Conclusions: We demonstrate an interconnection between the establishment of irAEs and granzyme B presence and, for the first time, the visualization of those events through PET imaging.

Noninvasive interrogation of CD8+ T cell effector function for monitoring early tumor responses to immunotherapy

“…Third, 68 Ga-grazytracer PET could also be expanded to monitor tumor responses to other immunotherapies related to granzyme B secretion, such as chimeric antigen receptor–T (CAR-T) and CAR-NK cell therapies. Furthermore, considering that the pathology of many other conditions — including transplant rejection ( 47 , 48 ), immune-mediated myocarditis ( 49 ), and other irAEs induced by immunotherapies ( 50 ) — involve granzyme B secretion, 68 Ga-grazytracer could also be expanded for PET imaging of other disorders.…”

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

“…Antigen retrieval was carried out using standard heat-based antigen retrieval techniques [ 18 ]. Immunofluorescence staining was carried out according to previously published procedures [ 15 ]. After blocking with 5% goat serum, rabbit anti-mouse granzyme B primary antibody (ab255598; Abcam, Cambridge, UK) was incubated overnight at 4 °C.…”

“…Antigen retrieval was carried out using standard heat-based antigen retrieval techniques [18]. Immunofluorescence staining was carried out according to previously published procedures [15]. After blocking with 5% goat serum, rabbit anti-mouse granzyme B primary antibody (ab255598; Abcam, Cambridge, UK) was incubated overnight at 4 • C. On the following day, washing was carried out followed by incubation with AlexaFluor 647 conjugated goat anti-rabbit IgG secondary antibody (A32733; ThermoFisher, Waltham, MA, USA) at room temperature in a moist dark chamber for 1 h. DAPI (4 , 6-diamidino-2-phenylindole) was used for staining cell nuclei.…”

“…Our group has previously developed a peptide-based radiotracer ( 68 Ga-GZP) that can selectively bind to granzyme B released by activated CD8 T cells in response to ICI [ 13 , 14 ]. We have shown that granzyme B PET imaging with 68 Ga-GZP can be used as a biomarker for predicting tumoral immunotherapy response [ 14 ] and can non-invasively identify immune-related adverse events [ 15 ]. Since GZP increasingly accumulates in the tumor tissue after immunotherapy, we investigated the use of GZP as a cancer theranostic agent, radiolabeled with 68 Ga ( 68 Ga-GZP) as a PET imaging agent and radiolabeled with 90 Y ( 90 Y-GZP) as a TRT agent for combinational therapy with ICI in murine models of colon cancer.…”

Emerging Biomaterials Imaging Antitumor Immune Response

“…[321] In future, irAEs can be further analyzed using molecular imaging combined with single-cell sequencing to determine the immune regimen most appropriate for patients. [322,323]…”