Efficacy of PD-1 blockade in cervical cancer is related to a CD8 + FoxP3 + CD25 + T-cell subset with operational effector functions despite high immune checkpoint levels

• Published in: Journal for immunotherapy of cancer Vol. 7; no. 1; p. 43
• Main Authors: Heeren, A M, Rotman, J, Stam, A G M, Pocorni, N, Gassama, A A, Samuels, S, Bleeker, M C G, Mom, C H, Zijlmans, H J M A A, Kenter, G G, Jordanova, E S, de Gruijl, T D
• Format: Journal Article
• Language: English
• Published: England BioMed Central Ltd 12-02-2019; BMJ Publishing Group LTD; BioMed Central; BMJ Publishing Group
• Subjects: Adult; Antigens; Antineoplastic Agents, Immunological - pharmacology; Biopsy; Cancer; Cancer metastasis; CD4-Positive T-Lymphocytes - drug effects; CD4-Positive T-Lymphocytes - immunology; CD8-Positive T-Lymphocytes - drug effects; CD8-Positive T-Lymphocytes - immunology; Cervical cancer; Criminal investigation; Cytokines; Development and progression; Female; Flow cytometry; Fluorescent antibody technique; FoxP3; Health aspects; Histochemistry; HPV16 E6; Human papillomavirus; Humans; Immunotherapy; Lymph nodes; Lymph Nodes - immunology; Lymphatic system; Lymphocytes; Medical prognosis; Metastasis; Middle Aged; Nivolumab - pharmacology; Oncogene Proteins, Viral - immunology; Patients; PD-1 blockade; Peptides; Programmed Cell Death 1 Receptor - antagonists & inhibitors; Programmed Cell Death 1 Receptor - immunology; Repressor Proteins - immunology; T cells; T-Lymphocyte Subsets - drug effects; T-Lymphocyte Subsets - immunology; Tumors; Uterine Cervical Neoplasms - immunology; PD-1 blockade; FoxP3; T cells; HPV16 E6; Cervical cancer; Lymph nodes
• ISSN: 2051-1426; 2051-1426
• DOI: 10.1186/s40425-019-0526-z

Cervical cancer (CxCa) is mainly a locally invading disease that metastasizes to loco-regional lymph node basins before involving distant organs in more advanced stages. Local immune potentiation of tumor-draining lymph nodes (TDLN) may thus protect against tumor progression.
To identify therapeutic targets for local immune modulation, multi-parameter flow cytometric T-cell profiling of primary cervical tumors (PT) and TDLN (n = 37) was performed. The in-vitro effect of PD-1 blockade on T-cell reactivity to HPV16 E6 oncoproteins was determined in cultures of TDLN and PT single cell suspensions (n = 19). Also, intracellular cytokine staining (ICS) upon anti-CD3 stimulation was performed in metastatic TDLN (LN+) and PT (n = 7), as well as multiplexed immunofluorescence histochemistry staining (n = 8).
Our data revealed elevated rates of activated regulatory T cells (aTregs) and of central or effector memory CD8
T cells in metastatic TDLN (LN+) as compared to tumor-free TDLN (LN-), and equally high or even higher rates of these subsets in PT. Both memory subsets co-expressed multiple immune checkpoints. PD-1 blockade significantly enhanced detectable E6-specific T-cell responses in 4/5 HPV16+ LN+ and in 1/5 HPV16+ PT. Whereas aTreg rates were higher in anti-PD-1 non-responders, in responders elevated levels of CD8
FoxP3
CD25
T cells were observed, which correlated with the efficacy of PD-1 blockade (P = 0.018). This subset was characterized by an early effector memory phenotype with particularly high levels of co-expressed PD-1, CTLA-4, TIM-3 and LAG-3 checkpoints, but, rather than exhausted, was shown upon polyclonal activation to produce higher levels of Granzyme-B and effector cytokines as compared to its CD8
FoxP3
counterparts.
These observations support local PD-(L)1 blockade to interrupt loco-regional immune suppression in CxCa and control metastatic spread to TDLN. Furthermore, our data identify CD8
FoxP3
CD25
T cells as therapeutic targets, which may also serve as predictive biomarker for PD-(L)1 checkpoint blockade.