The crystal structure of an inactive dimer of PDZ-binding kinase.

The overexpression of PDZ-binding kinase/T-LAK cell-originated protein kinase (PBK/TOPK) has been associated with hematologic tumors, breast cancer and various other cancers. However, the three-dimensional structure of PBK has not been solved. In this study, we determined the crystal structure of human PBK, which has two phospho-mimicking mutations T9E and T198E. The structural data indicated that PBK may assemble into an inactive dimer in alkaline conditions. Analytical size-exclusion chromatography and analytical ultracentrifugation confirmed that PBK exists in a conformational transition between dimers and monomers at different pH conditions. Co-IP and kinase assays suggested that the active state of PBK is a monomer and does not form a dimer even under alkaline conditions. These results showed that the conformational transition of PBK is important for its kinase activity regulation. Collectively, our observations may provide a novel starting point for structure-based functional studies.

Neoantigen vaccination induces clinical and immunologic responses in non-small cell lung cancer patients harboring EGFR mutations.

BACKGROUND: Neoantigen (NeoAg) peptides displayed at the tumor cell surface by human leukocyte antigen molecules show exquisite tumor specificity and can elicit T cell mediated tumor rejection. However, few NeoAgs are predicted to be shared between patients, and none to date have demonstrated therapeutic value in the context of vaccination. METHODS: We report here a phase I trial of personalized NeoAg peptide vaccination (PPV) of 24 stage III/IV non-small cell lung cancer (NSCLC) patients who had previously progressed following multiple conventional therapies, including surgery, radiation, chemotherapy, and tyrosine kinase inhibitors (TKIs). Primary endpoints of the trial evaluated feasibility, tolerability, and safety of the personalized vaccination approach, and secondary trial endpoints assessed tumor-specific immune reactivity and clinical responses. Of the 16 patients with epidermal growth factor receptor (EGFR) mutations, nine continued TKI therapy concurrent with PPV and seven patients received PPV alone. RESULTS: Out of 29 patients enrolled in the trial, 24 were immunized with personalized NeoAg peptides. Aside from transient rash, fatigue and/or fever observed in three patients, no other treatment-related adverse events were observed. Median progression-free survival and overall survival of the 24 vaccinated patients were 6.0 and 8.9 months, respectively. Within 3-4 months following initiation of PPV, seven RECIST-based objective clinical responses including one complete response were observed. Notably, all seven clinical responders had EGFR-mutated tumors, including four patients that had continued TKI therapy concurrently with PPV. Immune monitoring showed that five of the seven responding patients demonstrated vaccine-induced T cell responses against EGFR NeoAg peptides. Furthermore, two highly shared EGFR mutations (L858R and T790M) were shown to be immunogenic in four of the responding patients, all of whom demonstrated increases in peripheral blood neoantigen-specific CD8+ T cell frequencies during the course of PPV. CONCLUSIONS: These results show that personalized NeoAg vaccination is feasible and safe for advanced-stage NSCLC patients. The clinical and immune responses observed following PPV suggest that EGFR mutations constitute shared, immunogenic neoantigens with promising immunotherapeutic potential for large subsets of NSCLC patients. Furthermore, PPV with concurrent EGFR inhibitor therapy was well tolerated and may have contributed to the induction of PPV-induced T cell responses.