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. 2016 Oct;65(4):849-855.
doi: 10.1016/j.jhep.2016.06.027. Epub 2016 Jul 7.

Personalized Peptide Vaccine-Induced Immune Response Associated With Long-Term Survival of a Metastatic Cholangiocarcinoma Patient

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Free PMC article

Personalized Peptide Vaccine-Induced Immune Response Associated With Long-Term Survival of a Metastatic Cholangiocarcinoma Patient

Markus W Löffler et al. J Hepatol. .
Free PMC article

Erratum in

Abstract

Background & aims: We report a novel experimental immunotherapeutic approach in a patient with metastatic intrahepatic cholangiocarcinoma. In the 5year course of the disease, the initial tumor mass, two local recurrences and a lung metastasis were surgically removed. Lacking alternative treatment options, aiming at the induction of anti-tumor T cells responses, we initiated a personalized multi-peptide vaccination, based on in-depth analysis of tumor antigens (immunopeptidome) and sequencing.

Methods: Tumors were characterized by immunohistochemistry, next-generation sequencing and mass spectrometry of HLA ligands.

Results: Although several tumor-specific neo-epitopes were predicted in silico, none could be validated by mass spectrometry. Instead, a personalized multi-peptide vaccine containing non-mutated tumor-associated epitopes was designed and applied. Immunomonitoring showed vaccine-induced T cell responses to three out of seven peptides administered. The pulmonary metastasis resected after start of vaccination showed strong immune cell infiltration and perforin positivity, in contrast to the previous lesions. The patient remains clinically healthy, without any radiologically detectable tumors since March 2013 and the vaccination is continued.

Conclusions: This remarkable clinical course encourages formal clinical studies on adjuvant personalized peptide vaccination in cholangiocarcinoma.

Lay summary: Metastatic cholangiocarcinomas, cancers that originate from the liver bile ducts, have very limited treatment options and a fatal prognosis. We describe a novel therapeutic approach in such a patient using a personalized multi-peptide vaccine. This vaccine, developed based on the characterization of the patient's tumor, evoked detectable anti-tumor immune responses, associating with long-term tumor-free survival.

Keywords: Anti-tumor T cell response; Cholangiocarcinoma; HLA; Immunopeptidome; Immunotherapy; Peptides; Primary liver cancer.

Figures

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Fig. 1
Fig. 1
Clinical course and therapy. The course of the disease is subdivided in quarters of the year (Q) and the interventions performed (surgery/vaccination) are shown. Clinical imaging results for relevant events are depicted exemplarily in chronological order. Further exemplary imaging results depicting the course of disease can be accessed in Supplementary Figs. 1–15. The multi-peptide vaccination schedule is also indicated chronologically annotated for days (after initiation) and count of vaccinations. Seg, segment; R-classification, R0: no residual tumor; R1: microscopic residual tumor; R2: macroscopic residual tumor; lhs/rhs, left/right hand side; classification of malignant tumours (TNM Classification) (UICC): T = tumor extent (0–4); N = positive lymphnodes (0–1); M = Metastasis (0–1); L = lymphinvasion (0–1); V = venous invasion (0–1); G = grading (1–3); p = histopathological staging; c = clinical staging; Ø diameter; s.c., subcutaneous; i.d., intradermal; nV = nth vaccination.
Fig. 2
Fig. 2
Tumor characterization. Characterization of the various surgically resected tumors is shown. (A) Tumor location and putative ancestry based on shared driver mutations are indicated with a color code. (B) Tumors were assessed by immunohistochemistry and qualitative staining patterns (− negative; + slightly positive; ++ moderately positive; +++ strongly positive) are given for different immune and tumor markers. Staining for Hep Par 1 and CK7 supported the initial diagnosis of cholangiocarcinoma (L06/10) and staining of P03/13 for thyroid transcription factor-1 (TTF1) and Napsin A ruled out a primary lung adenocarcinoma. Further, counts for immune cell infiltrates in the epithelial compartment are given for each tumor and microscopic pictures (×100) of perforin staining are shown. More detailed data is provided in Supplementary Table 2.
Fig. 3
Fig. 3
Immunomonitoring of patient PBMCs for vaccine induced T cell responses. PBMCs were pre-stimulated using either peptide pool I (RGS-5, ADFP-2, ADFP-3, MMP7-(1) and HIV-A03) or peptide pool II (CCND1, IGFBP3, MMP7-(2) and Filamin-A), and expanded for 12 days using IL-2. (A) 300,000 cells (Class I binding peptides) or 200,000 (for class II binding peptides) were re-stimulated in triplicates (duplicates for pre-vaccination ‘scr’ time point) in an IFN-γ ELISPOT assay using individual peptides. The top panel shows the normalized spot counts for individual peptide stimulations and the bottom panel shows examples of scanned ELISPOT wells. At least 700,000 cells were re-stimulated with the respective peptides, in the presence of Brefeldin A and GolgiStop and 12 h later, intracellular IFN-γ and TNF were stained and analysed for B and C. (B) Exemplary dot plots of IFN-γ and TNF production by CD4(+) cells in response to RGS5 and CCND1 peptides, collected at the 25th vaccination (25V) are shown in the left panel. TNF production by CD8(+) cells in response to MMP7-(1) peptide are shown in the right panel (top). MMP7-(1) multimer staining of the corresponding vaccination time point (13V) is shown in the right panel (bottom). (C) Frequencies of the RGS5 peptide induced cytokine(+) live CD4(+) lymphocytes are shown (left y axis: IFN-γ right y axis: TNF). (D) At least 600,000 cells were analyzed by HLA-peptide multimers (class I). MMP7-(1) multimer(+) cells within live CD4(−) lymphocytes are shown and the % of multimer(+) CD8(+) lymphocytes are indicated.

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