Toxicity-specific peripheral blood T and B cell dynamics in anti-PD-1 and combined immune checkpoint inhibition

Abstract

Background Immune checkpoint inhibitors (ICI) have revolutionized the treatment landscape of advanced malignancies, but come with a diverse spectrum of immune-related adverse events (irAEs). Studies into irAE mechanisms are needed to make a transition from expert-opinion to evidence-based irAE treatment strategies. Methods We aimed to longitudinally characterize peripheral blood T and B cell dynamics in ICI-treated patients developing irAEs and remaining irAE-free. PBMCs were immunophenotyped and functionally assessed with multicolor flow cytometry at baseline, after ±3 weeks and ±6 weeks or upon clinically relevant irAEs. Additionally, serum cytokine concentrations for 23 analytes were measured by multiplex immunoassay at the same timepoints. Results We analyzed samples from 44 ICI-treated patients (24 anti-PD-1 monotherapy, 20 combined anti-PD-1 and anti-CTLA-4; cICI), of whom 22 developed clinically relevant irAEs, and 10 healthy donors. IrAEs after cICI were characterized by significantly enhanced proliferation of Th1-associated, mainly effector memory T cells, as well as Th17-associated and possibly antibody-mediated immune responses. The latter response was reflected in cICI toxicity by rising CXCL13 and IL-21 levels, but without changes in CD21lo, memory, class-switched or newly activated B cell subsets. Anti-PD-1 monotherapy-inflicted irAEs were associated with a modestly enhanced Th1-associated response compared to irAE-free patients, only reflected by increasing serum CXCL9 and CXCL10, but without cellular changes in Th1/Tc1 subsets. PD-1+LAG-3+double-positive (DP) CD8+T cells retained their proinflammatory potential in all treatment groups, regardless of irAE development. Especially DP CD8+T cells showed enhanced cytotoxic capacity in patients with irAEs after cICI. In all patients with irAEs, highly cytotoxic CD57+CD8+T cells were abundant. Conclusions Peripheral blood immune responses are substantially different between cICI and anti-PD-1 monotherapy-treated patients. ICI-induced toxicity is clearly dominated by an enhanced Th1-associated response, but in cICI we also found evidence for Th17-associated and possibly antibody-mediated responses. CD4+effector memory T cells were the principal cycling cells in irAEs after cICI. Together, our data add to the growing body of evidence that irAEs may be antibody-mediated and driven by newly activated CD4+helper T cells, specifically after cICI.

CD4 T cells and toxicity from immune checkpoint blockade

“…In contrast, other studies correlated with ≥ grade 2 toxicities 99,104,110 or with any toxicity including grade 1 98,101 . A second contributing factor could be that other studies mostly normalized CD4 T‐cell populations relative to parent lineages (i.e., T cells, CD4 T cells, or memory CD4 T cells) 98,99,101,104,110 . In contrast, we found that over‐normalizing the CD4 T EM population led to a loss of association with severe irAE risk 54 .…”

“…In support of our findings, this study found that higher pretreatment CD4 T EM proliferation was associated with a shorter time to toxicity development in patients receiving combination ICIs. Moreover, in the combination ICI group, T h1 memory cells were highly proliferative and T h17 cells increased significantly in on‐treatment patients that developed irAEs 110 (Figure 1). These cellular findings were corroborated by increases in serum T h1 ‐associated cytokines (CXCL9 and CXCL10), 110 similar to Nuñez et al 98 …”

“…Moreover, in the combination ICI group, T h1 memory cells were highly proliferative and T h17 cells increased significantly in on‐treatment patients that developed irAEs 110 (Figure 1). These cellular findings were corroborated by increases in serum T h1 ‐associated cytokines (CXCL9 and CXCL10), 110 similar to Nuñez et al 98 …”

“…While there is overall consistency between the aforementioned studies implicating CD4 T‐cell states in irAE development, the association between activated CD4 T EM /T M cells was strongest in Lozano et al One potential reason for this is that Lozano et al specifically focused on identifying cellular characteristics associated with severe irAE development, defined by grade 3 or higher toxicity 54 . In contrast, other studies correlated with ≥ grade 2 toxicities 99,104,110 or with any toxicity including grade 1 98,101 . A second contributing factor could be that other studies mostly normalized CD4 T‐cell populations relative to parent lineages (i.e., T cells, CD4 T cells, or memory CD4 T cells) 98,99,101,104,110 .…”

“…Furthermore, a recently submitted preprint by van Eijs and colleagues leveraged multicolor flow cytometry and a multiplex serum immunoassay to longitudinally profile B and T cells in a 44‐patient multi‐cancer cohort treated with combination ICI therapy or anti‐PD‐1 alone and found an early on‐treatment increase in CD4 T EM proliferation 110 (Figure 1). In support of our findings, this study found that higher pretreatment CD4 T EM proliferation was associated with a shorter time to toxicity development in patients receiving combination ICIs.…”