Evaluation of prognostic significance of granulocyte-related factors in cancer patients undergoing personalized peptide vaccination

Since cancer vaccines do not always elicit beneficial effects in treated patients, identification of biomarkers for predicting clinical outcomes would be highly desirable. We previously reported that abnormal granulocytes present in peripheral blood mononuclear cells (PBMC) may contribute to poor prognosis in advanced prostate cancer patients receiving personalized peptide vaccination (PPV). In the current study, we examined whether soluble factors derived from granulocytes, such as matrix metalloproteinase 9 (MMP-9), myeloperoxidase (MPO), and arginase 1 (ARG1), and inhibitory cytokine TGFβ in pre-vaccination plasma were useful for predicting prognosis after PPV in advanced cancer patients. In biliary tract cancer (n=25), multivariate Cox regression analysis demonstrated that patients with higher plasma MMP-9 levels had a significantly worse overall survival (OS) [hazard ratio (HR) = 4.637, 95% confidence interval (CI) = 1.670 - 12.877, P = 0.003], whereas MPO, ARG1, or TGFβ levels were not correlated with OS. Similarly, patients with higher MMP-9 levels showed worse prognosis than those with lower MMP-9 levels in other types of advanced cancers, including non-small cell lung cancer (n=32, P = 0.037 by log-rank test), and pancreatic cancer (n=41, P = 0.042 by log-rank test). Taken together, plasma MMP-9 levels before vaccination might be potentially useful as a biomarker for selecting advanced cancer patients who would benefit from PPV.This study was supported by a research program of the Project for Development of Innovative Research on Cancer Therapeutics (P-Direct), Ministry of Education, Culture, Sports, Science and Technology of Japan; a research program of the Regional Innovation Cluster Program of the Ministry of Education, Culture, Sports, Science and Technology of Japan; and Kurozumi Medical Foundation

Immunological evaluation of peptide vaccination for cancer patients with the HLA-A26 allele

To develop a peptide vaccine for cancer patients with the HLA-A26 allele, which is a minor population worldwide, we investigated the immunological responses of HLA-A26+ ⁄ A26+ cancer patients to four different CTL epitope peptides under personalized peptide vaccine regimens. In personalized peptide vaccine regimens, two to four peptides showing positive peptide-specific IgG responses in pre-vaccination plasma were selected from the four peptide candidates applicable for HLA-A26+ ⁄ A26+ cancer patients and administered s.c. Peptide-specific CTL and IgG responses along with cytokine levels were measured before and after vaccination. Cell surface markers in PBMCs and plasma cytokine levels were also measured. In this study, 21 advanced cancer patients, including seven lung, three breast, two pancreas, and two colon cancer patients, were enrolled. Their HLAA26 genotypes were HLA-A26:01 (n = 24), HLA-A26:03 (n = 10), and HLA-A26:02 (n = 8). One, 14, and 6 patients received two, three, and four peptides, respectively. Grade 1 or 2 skin reactions at the injection sites were observed in the majority of patients, but no severe adverse events related to the vaccination were observed. Peptide-specific CTL responses were augmented in 39% or 22% of patients after one or two cycles of vaccination, respectively. Notably, peptide-specific IgG were augmented in 63% or 100% of patients after one or two cycles of vaccination, respectively. Personalized peptide vaccines with these four CTL epitope peptides could be feasible for HLA-A26+ advanced cancer patients because of their safety and higher rates of immunological responses.This study was supported in part by the Japan Agency for Medical Research and development, AMED, a research program of the Regional Innovation Cluster Program of the Ministry of Education, Culture, Sports, Science and Technology of Japan, and a grant from the Sendai Kousei Hospital

Clinical Study Randomized Phase II Study of Docetaxel plus Personalized Peptide Vaccination versus Docetaxel plus Placebo for Patients with Previously Treated Advanced Wild Type EGFR Non-Small-Cell Lung Cancer

Objectives. To evaluate the efficacy and safety of personalized peptide vaccination (PPV) combined with chemotherapy for patients with previously treated advanced non-small-cell lung cancer (NSCLC). Patients and Methods. Previously treated PS0-1 patients with IIIB/IV EGFR (epidermal growth factor receptor) wild genotype NSCLC were randomly assigned to docetaxel (60 mg/m 2 on Day 1) plus PPV based on preexisting host immunity or docetaxel plus placebo. Docetaxel administration was repeated every 3 weeks until disease progression. Personalized peptides or placebo was injected subcutaneously weekly in the first 8 weeks and biweekly in subsequent 16 weeks. The primary efficacy endpoint was progression-free survival (PFS). Results. PPV related toxicity was grade 2 or less skin reaction. The median PFS for placebo arm and PPV arm was 52 days and 59 days, respectively. There was no significant difference between two arms by log-rank test ( = 0.42). Interestingly, PFS and overall survival (OS) in humoral immunological responder were significantly longer than those in nonresponder. Conclusion. PPV did not improve the survival in combination with docetaxel for previously treated advanced NSCLC. However, PPV may be efficacious for the humoral immunological responders and a further clinical investigation is needed

Towards the introduction of the ‘Immunoscore’ in the classification of malignant tumours

The American Joint Committee on Cancer/Union Internationale Contre le Cancer (AJCC/UICC) TNM staging system provides the most reliable guidelines for the routine prognostication and treatment of colorectal carcinoma. This traditional tumour staging summarizes data on tumour burden (T), the presence of cancer cells in draining and regional lymph nodes (N) and evidence for distant metastases (M). However, it is now recognized that the clinical outcome can vary significantly among patients within the same stage. The current classification provides limited prognostic information and does not predict response to therapy. Multiple ways to classify cancer and to distinguish different subtypes of colorectal cancer have been proposed, including morphology, cell origin, molecular pathways, mutation status and gene expression-based stratification. These parameters rely on tumour-cell characteristics. Extensive literature has investigated the host immune response against cancer and demonstrated the prognostic impact of the in situ immune cell infiltrate in tumours. A methodology named ‘Immunoscore’ has been defined to quantify the in situ immune infiltrate. In colorectal cancer, the Immunoscore may add to the significance of the current AJCC/UICC TNM classification, since it has been demonstrated to be a prognostic factor superior to the AJCC/UICC TNM classification. An international consortium has been initiated to validate and promote the Immunoscore in routine clinical settings. The results of this international consortium may result in the implementation of the Immunoscore as a new component for the classification of cancer, designated TNM-I (TNM-Immune). © 2013 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland

EGFR T790M Mutation as a Possible Target for Immunotherapy; Identification of HLA-A*0201-Restricted T Cell Epitopes Derived from the EGFR T790M Mutation

Treatment with epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs), such as gefitinib and erlotinib, has achieved high clinical response rates in patients with non–small cell lung cancers (NSCLCs). However, over time, most tumors develop acquired resistance to EGFR-TKIs, which is associated with the secondary EGFR T790M resistance mutation in about half the cases. Currently there are no effective treatment options for patients with this resistance mutation. Here we identified two novel HLA-A*0201 (A2)-restricted T cell epitopes containing the mutated methionine residue of the EGFR T790M mutation, T790M-5 (MQLMPFGCLL) and T790M-7 (LIMQLMPFGCL), as potential targets for EGFR-TKI-resistant patients. When peripheral blood cells were repeatedly stimulated in vitro with these two peptides and assessed by antigen-specific IFN-γ secretion, T cell lines responsive to T790M-5 and T790M-7 were established in 5 of 6 (83%) and 3 of 6 (50%) healthy donors, respectively. Additionally, the T790M-5- and T790M-7-specific T cell lines displayed an MHC class I-restricted reactivity against NSCLC cell lines expressing both HLA-A2 and the T790M mutation. Interestingly, the NSCLC patients with antigen-specific T cell responses to these epitopes showed a significantly less frequency of EGFR-T790M mutation than those without them [1 of 7 (14%) vs 9 of 15 (60%); chi-squared test, p = 0.0449], indicating the negative correlation between the immune responses to the EGFR-T790M-derived epitopes and the presence of EGFR-T790M mutation in NSCLC patients. This finding could possibly be explained by the hypothesis that immune responses to the mutated neo-antigens derived from T790M might prevent the emergence of tumor cell variants with the T790M resistance mutation in NSCLC patients during EGFR-TKI treatment. Together, our results suggest that the identified T cell epitopes might provide a novel immunotherapeutic approach for prevention and/or treatment of EGFR-TKI resistance with the secondary EGFR T790M resistance mutation in NSCLC patients

Plasma Levels of High-Mobility Group Box 1 during Peptide Vaccination in Patients with Recurrent Ovarian Cancer

High-mobility group box 1 (HMGB1) is a nuclear protein that is known to be secreted into extracellular fluids from injured cells, activated macrophages, and tumor cells. The clinical correlation of circulating HMGB1 levels with various diseases including cancer has been reported. However, there is no information on HMGB1 levels in cancer patients treated with peptide vaccination. In the present study, we investigated the plasma levels of HMGB1 during personalized peptide vaccination in patients with recurrent ovarian cancer. Frozen plasma samples of 39 patients from previously conducted clinical trials were used in this study. HMGB1 levels were decreased after the 1st cycle of vaccination from their prevaccination levels. However, no correlation was observed between HMGB1 and overall survival (OS). The correlation between plasma HMGB1 levels and other biomarker levels was further analyzed by scatter plot, revealing that HMGB1 levels after the 1st cycle of vaccination were significantly correlated with myeloid-derived suppressor cell (MDSC) frequency after the 1st cycle of vaccination (r=0.357, p=0.032). Chi-square test showed that epitope spreading was significantly related with changes of HMGB1 (p=0.030). These results suggest that plasma HMGB1 is a possible biomarker for cancer vaccine therapy, although direct correlation with OS has not been obtained. This trial is registered with Clinical Trial Registry under trial numbers UMIN000003083 and UMIN000001482