TLR2 ligand-synthetic long peptide conjugates effectively stimulate tumor-draining lymph node T cells of cervical cancer patients

Abstract

The potency of human papillomavirus type 16 (HPV16)-encoded synthetic long peptides (SLP), conjugated to an optimized Toll-like receptor 2 ligand (TLR2-L), was assessed in ex vivo activation of HPV16+ cancer patient-derived T cells. Two highly immunogenic SLP sequences derived from the oncogenic E6 protein of HPV16 were selected and conjugated to a Pam3CSK4-based TLR2-L under GMP conditions. Both conjugates were able to mature human DCs in vitro and to activate human skin-derived antigen-presenting cells (APCs) upon intradermal injection in an ex vivo skin model, associated with induction of a favorable chemokine profile to attract and activate T cells. The conjugated SLPs were efficiently processed by APCs, since HPV16-specific CD4+ and CD8+ T-cell clones isolated from HPV16+ cervical tumors proliferated in response to both conjugates. The TLR2-L SLP conjugates significantly enhanced ex vivo T helper type 1 T-cell activation in cell suspensions obtained from tumor-draining lymph nodes (LN) resected during hysterectomy of HPV16+ cervical cancer patients. These results show that TLR2-L SLP conjugates can activate circulating or LN-derived tumor-specific T cells, a promising outcome for studying these two conjugates in a phase I/II clinical safety and immunogenicity trial.

Bioconjugation Approaches to Producing Subunit Vaccines Composed of Protein or Peptide Antigens and Covalently Attached Toll-Like Receptor Ligands

“…Some lipopeptide TLR2 agonists, such as Pam2Cys and Pam3Cys, contain an amino acid scaffold, and hence can be easily conjugated onto peptide antigens during SPPS. 14,[117][118][119][120] In addition, TLR7, 8 and 9 agonists have also been conjugated onto peptides to produce self-adjuvanting vaccines. 118,[121][122][123] Fully synthetic vaccines composed of a model OVA CTL antigen (DEVSGLEQLESIINFEKL; OVA 247-264 ) fused with the TLR2 ligand Pam3CysSK 4 and TLR9 ligand CpG ODN (5'-TCCATGACGTTCCTGACGTT-3') have been designed and produced with high quality, and without contamination by other TLR ligands such as LPS.…”

Identification of a neo-epitope dominating endogenous CD8 T cell responses to MC-38 colorectal cancer

“…Therapeutic vaccination against Rpl18 using synthetic peptides induced delay of tumor outgrowth and extended survival although overall survival was not affected. We hypothesized that a conjugation to an optimized TLR-2 ligand Upam or Amplivant® 25,26,31,35,36 would improve immunogenicity and survival. As expected, vaccination of MC-38 tumor-bearing mice with this construct resulted in a significant improvement of survival when compared to adjuvant alone, thus underscoring the relevance of the Rpl18 CD8 + neo-epitope in the MC-38 colorectal cancer model.…”

“…The method used for the mice treated with anti-PD -L1 antibodies can be found in the publication of Sow, H. S. et al 23 Vaccinated mice were injected once with 20 nmol Adpgk or Rpl18 synthetic peptides intradermally in 50 µL PBS adjuvanted with 20 µg of CpG (ODN 1826 -TLR9 ligand InvivoGen tlrl-1826-1) or 10 nmol Rpl18-conjugated with equimolar Upam-ligand (peptides and Upam where conjugated as described previously in). [24][25][26] In vitro (re-)stimulation T cell cultures were started with splenocytes from several experiments. Therefore, spleens were mashed on single cell strainers with the blunt end of a 5 mL syringe and washed with culture medium.…”

Development of a vaccine based on bacteria‐mimicking tumor cells coated with novel engineered toll‐like receptor 2 ligands

“…Tumor cells alone cannot induce strong immune responses because although tumors express many specific antigens (such as cancer-testis antigens and tumor-specific neoantigens) they do not express TLR ligands. To induce strong immune responses, simple mixtures of TLR ligands and antigens [24][25][26][27][28] or TLR ligands conjugated with antigen peptide 29,30 are widely used as vaccines.…”