ABSTRACT
PURPOSE: Our aims are to determine levels of circulating cellular and protein biomarkers in hepatocellular carcinoma (HCC) patients and to analyse any relationships with clinical parameters. METHODS: Fifty-four consenting patients were recruited. Circulating tumour cells (CTCs) were enumerated (by CellSearch) and characterised via filtration [by isolation by size of epithelial tumour cells (ISET)] with downstream immunohistochemistry (IHC). Glypican-3 (GPC3) expression in tumour biopsies and CTCs (by IHC) was compared, and levels of circulating caspase-cleaved and full-length cytokeratin 18 (CK18, measured using M30 and M65 ELISAs) were examined as a putative prognostic factor and marker of tumour burden. RESULTS: CTCs were identified in 14 out of 50 (28%) patients by CellSearch and in 19 out of 19 (100%) patients by ISET. The presence of GPC3-positive CTCs by ISET was 100% concordant with the presence of GPC3-positive cells in the original tumour (n = 5). No statistically significant correlations were observed between CTC number and clinical characteristics, although trends were noted between CTC subtypes, Child-Pugh score and tumour node metastasis stage. Serum M30 and M65 levels (as continuous variables) significantly correlated with overall survival (OS) in a univariate analysis (p = 0.003 and p < 0.001, respectively); M65 levels remained statistically significant in a multivariate analysis (p = 0.029). CONCLUSIONS: This is the first study to detect GPC3-positive CTCs in HCC, important for drug development with this target. The significant association of circulating CK18 with OS in HCC further exemplifies the utility of circulating biomarkers in cancer.
Subject(s)
Biomarkers, Tumor/blood , Carcinoma, Hepatocellular/diagnosis , Liver Neoplasms/diagnosis , Neoplastic Cells, Circulating/pathology , Adult , Aged , Aged, 80 and over , Carcinoma, Hepatocellular/blood , Cell Separation , Female , Follow-Up Studies , Glypicans/blood , Humans , Immunoenzyme Techniques , Keratin-18/blood , Liver Neoplasms/blood , Male , Middle Aged , Neoplasm Staging , PrognosisSubject(s)
CD4 Antigens/biosynthesis , CD4-Positive T-Lymphocytes/immunology , Clonal Anergy , Receptors, Interleukin-2/biosynthesis , T-Lymphocytes, Regulatory/immunology , CD4-Positive T-Lymphocytes/metabolism , Cell Proliferation , Centrifugation, Density Gradient/methods , Flow Cytometry/methods , Humans , T-Lymphocytes, Regulatory/metabolismABSTRACT
PURPOSE: The purpose is to study the immunogenicity of heterologous prime-boost human papillomavirus (HPV) oncogene vaccination in patients with anogenital intraepithelial neoplasia (AGIN). EXPERIMENTAL DESIGN: Twenty-nine women with high-grade AGIN received three i.m. doses of TA-CIN (HPV-16 L2/E6/E7 protein) at four weekly intervals followed by a single dermal scarification of vaccinia HPV-16/18 E6/E7 and were followed up for 12 weeks. Immunity to HPV-16 was assessed by lymphoproliferation, IFN-gamma enzyme-linked immunospot (ELISPOT), and ELISA. RESULTS: The patient group significantly responded to TA-CIN and not to the control antigen HPV-6 L2/E7 at all postvaccination time points when compared with baseline responses (P < or = 0.05). Ten of the patients showed at least a 3-fold increase in TA-CIN-specific proliferation at one or more time points after vaccination. Comparison of stimulation with HPV-16 E6- or E7-GST fusion proteins showed that proliferative responses were biased to HPV-16 E6. This bias was also seen by IFN-gamma ELISPOT using overlapping peptides, with HPV-16 E6- or E7-specific T cells being detected in 9 and 2 patients, respectively. In addition, vaccination resulted in the induction of antibodies against the HPV-16 oncoproteins. Of the 6 clinical responders, 2 patients showed both a proliferative TA-CIN-specific response and an E6-specific IFN-gamma response, whereas 3 other patients displayed E6-specific reactivity only. Stable disease was recorded in 19 patients, 8 of whom showed a concomitant TA-CIN-specific proliferative and/or E6-specific T-cell response. Of the 4 progressors, 2 failed to make a T-cell response and 2 responded by either proliferation or E6 ELISPOT alone. CONCLUSIONS: The prime-boost regimen is immunogenic in AGIN patients (humoral and cellular immunity), but there is no simple relationship between induction of systemic HPV-16-specific immunity and clinical outcome. Other factors that may play a role in the eradication of long-term established AGIN lesions need to be determined to identify the patient group that would benefit from immunotherapy with the vaccines used in this study.
