RESUMO
Mounting evidence suggests that autoreactivity and inflammatory processes are involved in the pathogenesis of chronic lymphocytic leukaemia (CLL). Cytoskeletal proteins, including non-muscle myosin heavy chain IIA (MYHIIA), vimentin (VIM) and cofilin-1 (CFL1), exposed on the surface of apoptotic cells have been identified as autoantigens that are recognized by the specific B-cell receptors of the CLL cells. In 212 CLL patients analysed with quantitative reverse transcriptase-polymerase chain reaction we found CFL1 overexpression and low expression of MYH9 in comparison with healthy volunteers. We detected specific cytotoxic immune responses for peptides derived from MYHIIA in 66·7%, VIM in 87·5% and CFL1 in 62·5% CLL patients in an Enzyme-Linked ImmunoSpot assay. Low frequencies of autoreactive peptide-specific T cells were detected against MYHIIA, VIM and CFL1 in CLL patients ex vivo; most of the detected cells had an effector-memory phenotype. Our findings support the existence of cytotoxic immune responses against three autoantigens that have been identified as targets of CLL clonotypic B-cell receptors. The presence of autoreactive CD8(+) T cells against MYHIIA, VIM and CFL1 in CLL patients indicates the involvement of antigen-specific autoreactive T cells in the pathogenesis of CLL.
Assuntos
Autoantígenos/imunologia , Leucemia Linfocítica Crônica de Células B/imunologia , Linfócitos T Citotóxicos/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Cofilina 1/imunologia , Feminino , Humanos , Leucemia Linfocítica Crônica de Células B/patologia , Masculino , Pessoa de Meia-Idade , Cadeias Pesadas de Miosina/imunologia , Vimentina/imunologiaRESUMO
The appropriate function of the immune system depends on the effective regulation of the immune response on multiple levels. The key element of an effective immune response to antigenic stimulation is maintaining a homeostasis between activation and inhibitory function of immunocompetent cells and molecules. In pathological conditions such as chronic infections, autoimmune diseases or cancer there are significant alterations, and prevalence of signals of one type over another. Main markers of these dysfunctions are altered expressions of molecules, such as programmed death-1 (PD-1), Human Leukocyte Antigen G (HLA-G), or changed percentages of T regulatory cells (Treg). These indicators of immune system dysfunction may contribute to disease progression, but also could represent good targets for treatment. Interestingly, in recent years there are many new, interesting reports which showed that the role of PD-1, HLA-G or Treg is ambiguous and not always their higher expression or frequency lead to the progression of disease. Recent studies have shown that Treg can suppress bacteria-driven inflammation which promotes carcinogenesis and thus protect the host from cancer development. Moreover, proliferation of hematological tumor cells expressing ILT-2 receptor can be inhibited by HLA-G, in contrast to solid tumors where HLA-G favors tumor escape. In this paper we present characteristics of expressions of PD-1 and its ligands, HLA-G, and frequency of Treg cells in a variety of physiological and pathological conditions associated with chronic infections, autoimmune diseases and cancer. The understanding of the complex interactions between the functional elements of immune system is essential for a detailed characteristics of the mechanisms leading to the development of diseases and identification of more effective targeted therapies.
Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Terapia de Imunossupressão , Neoplasias/imunologia , Receptor de Morte Celular Programada 1/imunologia , Linfócitos T Reguladores/imunologia , Animais , Doenças Autoimunes/imunologia , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Transdução de Sinais , Evasão TumoralAssuntos
Imunidade Celular , Leucemia Linfocítica Crônica de Células B/imunologia , Linfócitos T Reguladores/imunologia , Idoso , Idoso de 80 Anos ou mais , Vacinas Anticâncer , Estudos de Casos e Controles , Feminino , Humanos , Interleucina-2/farmacologia , Leucemia Linfocítica Crônica de Células B/terapia , Masculino , Pessoa de Meia-Idade , Linfócitos T Citotóxicos/imunologia , Talidomida/farmacologia , Vacinação , Vacinas de Subunidades AntigênicasRESUMO
Functional toll-like receptors (TLRs) could modulate anti-tumor effects by activating inflammatory cytokines and the cytotoxic T-cells response. However, excessive TLR expression could promote tumor progression, since TLR-induced inflammation might stimulate cancer cells expansion into the microenvironment. Myd88 is involved in activation NF-κB through TLRs downstream signaling, hence in the current study we provided, for the first time, a complex characterization of expression of TLR2, TLR4, TLR7, TLR9, and MYD88 as well as their splicing forms in two distinct compartments of the microenvironment of chronic lymphocytic leukemia (CLL): peripheral blood and bone marrow. We found correlations between MYD88 and TLRs expressions in both compartments, indicating their relevant cooperation in CLL. The MYD88 expression was higher in CLL patients compared to healthy volunteers (HVs) (0.1780 vs. 0.128, p < 0.0001). The TLRs expression was aberrant in CLL compared to HVs. Analysis of survival curves revealed a shorter time to first treatment in the group of patients with low level of TLR4(3) expression compared to high level of TLR4(3) expression in bone marrow (13 months vs. 48 months, p = 0.0207). We suggest that TLRs expression is differentially regulated in CLL but is similarly shared between two distinct compartments of the microenvironment.
RESUMO
Psoriasis (Ps), an autoimmune disease, and multiple myeloma (MM), a blood neoplasm, are characterized by immune dysregulation resulting from the imbalance between the effector and regulatory cells, including B regulatory (Breg) lymphocytes. Peripheral blood samples from 80 Ps patients, 17 relapsed/refractory MM patients before and after daratumumab (anti-CD38 monoclonal antibody) treatment, 23 healthy volunteers (HVs), and bone marrow samples from 59 MM patients were used in the study. Bregs were determined by flow cytometry using CD19, CD24, and CD38. Intracellular production of interleukin-10 (IL-10) was assessed by flow cytometry after CD40L, LPS, and CpG stimulation. IL-10 serum or plasma concentrations were tested using ELISA method. The percentage of CD19+CD24hiCD38hi Bregs was not different whereas the production of IL-10 in Bregs was significantly higher in Ps patients in comparison with HVs. The percentage of CD19+CD24hiCD38hi Bregs in MM patients was significantly higher than in HVs (p < 0.0001). The percentage of CD19+CD24hiCD38hi Bregs was significantly higher in MM patients with the ISS stage I (p = 0.0233) while IL-10 production in Bregs was significantly higher in ISS stage III (p = 0.0165). IL-10 serum or plasma concentration was significantly higher in Ps and MM patients when compared to HVs (p < 0.0001). Following the treatment with daratumumab the percentages of CD19+CD24hiCD38hi Bregs significantly decreased (p < 0.0003). Here, in the two opposite immune conditions, despite the differences in percentages of Bregs in Ps and MM we have identified some similarities in the IL-10 producing Bregs. Effective treatment of daratumumab besides the anti-myeloma effect was accompanied by the eradication of Bregs.
Assuntos
ADP-Ribosil Ciclase 1/metabolismo , Antígenos CD19/metabolismo , Linfócitos B Reguladores/imunologia , Antígeno CD24/metabolismo , Mieloma Múltiplo/imunologia , Psoríase/imunologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Linfócitos B Reguladores/efeitos dos fármacos , Feminino , Humanos , Interleucina-10/sangue , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/sangue , Mieloma Múltiplo/tratamento farmacológico , Esclerose Múltipla Recidivante-Remitente/sangue , Esclerose Múltipla Recidivante-Remitente/tratamento farmacológico , Esclerose Múltipla Recidivante-Remitente/imunologia , Psoríase/sangue , Psoríase/tratamento farmacológico , Adulto JovemRESUMO
The status of the immune system of patients with B-cell chronic lymphocytic leukemia (B-CLL) is not yet sufficiently characterized. Clinically, B-CLL patients present immunodeficiency increasing along with disease progression and signs of autoimmunity. In the current study, we evaluated the expression of FOXP3 in CD4+CD25hi T regulatory lymphocytes (Treg) and their influence on immune response against tumor and viral antigens in the complex system of peripheral blood mononuclear cells. In 80 B-CLL patients, the frequency of Treg (CD4+CD25hi FOXP3+) cells was significantly higher in B-CLL patients when compared to healthy volunteers (HV) and increased with the progression of the disease (median: 8.24% in stage A, 11.24% in stage B and 12.57% in stage C according to the Binet classification). The frequency of Treg showed no correlation with prognostic markers such as ZAP-70, CD38 and HLA-G. Notably, Treg frequency correlated with serum levels of TNF (r(2)=0.45, p=0.001). T-cell immune responses against epitopes derived from the tumor-associated antigens survivin, fibromodulin and RHAMM as well as from the influenza matrix protein were evaluated. Functionally, higher frequencies of Treg correlated with decreased T-cell responses against viral and tumor antigens. In conclusion, we detected higher frequencies of Treg in B-CLL patients than in HV. Furthermore, Treg constitute the crucial mechanism of immunosuppression in B-CLL patients.
Assuntos
Linfócitos T CD4-Positivos/patologia , Leucemia Linfocítica Crônica de Células B/imunologia , Leucemia Linfocítica Crônica de Células B/patologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos T CD4-Positivos/imunologia , Feminino , Fatores de Transcrição Forkhead/metabolismo , Humanos , Interferon gama/metabolismo , Leucemia Linfocítica Crônica de Células B/metabolismo , Masculino , Pessoa de Meia-Idade , Fragmentos de Peptídeos/metabolismo , Prognóstico , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Receptores Tipo II do Fator de Necrose Tumoral/metabolismo , Linfócitos T Reguladores/metabolismo , Proteína-Tirosina Quinase ZAP-70/metabolismoRESUMO
INTRODUCTION Currently available prognostic factors determining the course of chronic lymphocytic leukemia (CLL) are not fully efficient, especially for newly diagnosed patients. Investigation of molecular changes may help clarify the reasons for the heterogeneity of the disease. Apart from already confirmed TP53 mutations, the novel candidates: NOTCH1, SF3B1, and MYD88 might represent clinically relevant biomarkers. OBJECTIVES The aim of this study was to evaluate the mutational status of NOTCH1, MYD88, and SF3B1 and to compare the results with confirmed prognostic factors: ZAP70, CD38, and immunoglobulin heavychain variable region (IGHV) mutation in CLL. The study assessed also prognostic significance in terms of the time to first treatment (TTFT) and subset analysis. PATIENTS AND METHODS The study was conducted on samples of 370 newly diagnosed patients with CLL. The analysis was performed using highresolution melting, Sanger sequencing, and polymerase chain reaction methods. RESULTS Patients harboring the NOTCH1 mutation were significantly more often found among patients with an unmutated IGHV gene status and high expression of CD38 and ZAP70. The MYD88 mutation was equally distributed in patients with mutated and unmutated IGHV status (5 vs 7 patients). For MYD88 and SF3B1, there were no significant differences in the levels of CD38 and ZAP70 expression. The tendency for lower median TTFT was revealed in patients with mutated SF3B1 (P = 0.08). The analysis showed the presence of 14 different types of the subsets of IGHV in 50 of 345 patients (14.5%). The most frequent were subsets #1 and #2. CONCLUSIONS The NOTCH1 and SF3B1 mutations accompany biological markers of unfavorable prognosis in patients with CLL. The mutations may contribute to the identification of patients with highrisk CLL.
Assuntos
Leucemia Linfoide/genética , Mutação , Fosfoproteínas/genética , Fatores de Processamento de RNA/genética , Receptor Notch1/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/metabolismo , Leucemia Linfoide/diagnóstico , Leucemia Linfoide/metabolismo , Leucemia Prolinfocítica de Células T/diagnóstico , Leucemia Prolinfocítica de Células T/genética , Leucemia Prolinfocítica de Células T/metabolismo , Masculino , Pessoa de Meia-Idade , Fator 88 de Diferenciação Mieloide/genética , Polônia , PrognósticoRESUMO
Programmed death-1 (PD-1) is one of the most important inhibitory co-receptors expressed predominantly on activated T and B lymphocytes whose expression could be sustained by permanent antigenic stimulation accompanying chronic or recurrent tonsillitis. The expression of PD-1 and PD-1L was analyzed using flow cytometry on hypertrophied tonsils collected from 57 children. We observed high expression of PD-1 and PD-1L on certain lymphocytes subpopulations of hypertrophied tonsils; among T cells, the expression of PD-1 on protein level was higher on CD4+ cells (70.3 %) than on CD8+ cells (35 %). Interestingly, a limited expression of PD-1 was observed on CD19+ B lymphocytes (6.5 %), while CD5+CD19+ B cells overexpressed PD-1 (52.5 %). Moreover, the expression of PD-1L was also higher on CD5+CD19+ B cells (16.5 %) than on CD19+ B cells (3.5 %) and on CD4+ T cells (20 %) than on CD8+ T cells (10 %). PD-1 and PD-1L expressions correlated only on CD5+CD19+ cells. In conclusion, high expression of PD-1 and PD-1L on T and B cells could represent hallmark of immune system adaptation to chronic antigenic exposition in patients with tonsillitis.
Assuntos
Tonsila Faríngea/imunologia , Linfócitos B/metabolismo , Antígeno B7-H1/metabolismo , Proteína 2 Ligante de Morte Celular Programada 1/metabolismo , Receptor de Morte Celular Programada 1/metabolismo , Tonsilite/cirurgia , Tonsila Faríngea/metabolismo , Tonsila Faríngea/cirurgia , Adolescente , Antígenos CD19/metabolismo , Linfócitos B/imunologia , Antígenos CD5/metabolismo , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Masculino , Linfócitos T/metabolismo , Tonsilite/imunologia , Regulação para CimaRESUMO
Programmed death-1 (PD-1) is a negative receptor expressed on lymphocytes including malignant B cells in chronic lymphocytic leukemia (CLL). In this work, we found that patients with CLL had a higher expression of PD-1 transcript (PDCD1) than healthy volunteers (p < 0.0001). PDCD1 expression was comparable between CLL cells from accumulation (peripheral blood) and proliferation (bone marrow) disease compartments. In blood samples of patients with mutated IGHV genes PDCD1 expression was higher than with unmutated IGHV (p = 0.0299). We demonstrated that phosphorylation of SYK and LYN, key B-cell receptor signaling kinases, was independent of PD-1 expression in patients with CLL, while ZAP-70 phosphorylation in negative tyrosine residue 292 showed strong inverse correlation (r = - 0.8, p = 0.0019). No associations between five single nucleotide polymorphisms of PDCD1, their expressions and susceptibility to CLL were found. In conclusion, PD-1 might be an independent, universal marker of CLL cells and a part of their activated phenotype, and subsequently might modulate the function of ZAP-70.
Assuntos
Leucemia Linfocítica Crônica de Células B/metabolismo , Receptor de Morte Celular Programada 1/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Medula Óssea/patologia , Células da Medula Óssea/metabolismo , Células da Medula Óssea/patologia , Aberrações Cromossômicas , Feminino , Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Predisposição Genética para Doença , Humanos , Imunofenotipagem , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Leucemia Linfocítica Crônica de Células B/diagnóstico , Leucemia Linfocítica Crônica de Células B/genética , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/patologia , Desequilíbrio de Ligação , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Fosforilação , Polimorfismo de Nucleotídeo Único , Receptor de Morte Celular Programada 1/genética , Proteínas Tirosina Quinases/metabolismo , Quinase Syk , Proteína-Tirosina Quinase ZAP-70/metabolismo , Quinases da Família src/metabolismoRESUMO
BACKGROUND: Thalidomide may represent a novel therapeutic strategy in the treatment of chronic lymphocytic leukemia (CLL). Since the activation of nuclear factor kappa B (NF-κB) causes not only malignant transformation and tumor progression, but also allows tumor cells to evade immune surveillance, NF-κB signaling components might constitute a potential target for future therapy in CLL. OBJECTIVES: The current study is an attempt to characterize proteins regulated by thalidomide. Thalidomide's influence on NF-κB proteins and on regulatory T cells (Treg) in CLL was investigated. MATERIAL AND METHODS: A total of 15 patients with CLL were treated with a combined thalidomide/fludarabine regimen. Peripheral blood mononuclear cells were separated by Ficoll density gradient centrifugation. To evaluate glucocorticoid induced tumour-necrosis-factor-receptor-related protein (GITR) expression in regulatory T cells, cells incubated with anti-CD3, ani-CD4 and anti-CD25 were permeabilized and then stained with anti-FOXP3 and analyzed using flow cytometry. Human TNF enzyme-linked immunosorbent assay (ELISA) was used to determine the tumor necrosis factor (TNF) levels in the serum. To evaluate NF-κB activity, chemiluminescent oligonucleotide-based ELISA was performed. RESULTS: It was found that thalidomide regulates NF-κB activity differentially, and the activity of certain NF-κB components correlated with TNF levels and T regulatory cell (CD4 + CD25 high GITR + ). CONCLUSIONS: These results might indicate that thalidomide not only regulates TNF but also directly interferes with NF-κB components.
Assuntos
Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , NF-kappa B/fisiologia , Linfócitos T Reguladores/imunologia , Talidomida/farmacologia , ADP-Ribosil Ciclase 1/análise , Idoso , Feminino , Humanos , Leucemia Linfocítica Crônica de Células B/imunologia , Masculino , Glicoproteínas de Membrana/análise , Pessoa de Meia-Idade , Proteína-Tirosina Quinase ZAP-70/análiseRESUMO
Suppressed immune status facilitates immune escape mechanisms that allow chronic lymphocytic leukemia cells to proliferate and expand. The expression of HLA-G could effectively inhibit the immune response. In immune response inhibitory signals follow activation of immune system which might be occur during bacterial or viral infection in CLL patients. In the current study we characterized two components of immune system, inhibitory molecule HLA-G with its receptor - CD85j and Toll-like receptor 9. The material was obtained from 41 CLL patients and 41 HV with similar median age. In CLL patients expression of intracellular and surface HLA-G and soluble HLA-G levels were significantly higher than in HV. We found higher expression of CD85j compared to HV and the positive correlation between expression of HLA-G and CD85j. All the CLL cells expressed TLR-9, and the level of expression positively correlated with expression of HLA-G and CD85j. Patients with higher expression of intracellular expression of TLR-9 have significantly longer treatment-free survival than patients with low expression of TLR-9 (57 months vs. 8 months, respectively). Summarizing in CLL we characterized activatory and inhibitory components of immune system that might be connected functionally. Analysis of TLR-9 expression might have additional prognostic value for CLL patients.
Assuntos
Antígenos HLA-G/metabolismo , Leucemia Linfocítica Crônica de Células B/metabolismo , Receptor Toll-Like 9/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/metabolismo , Membrana Celular/metabolismo , Feminino , Antígenos HLA-G/sangue , Humanos , Imunofenotipagem , Leucemia Linfocítica Crônica de Células B/sangue , Leucemia Linfocítica Crônica de Células B/mortalidade , Receptor B1 de Leucócitos Semelhante a Imunoglobulina , Masculino , Pessoa de Meia-Idade , Prognóstico , Receptores Imunológicos/metabolismo , Adulto JovemAssuntos
Envelhecimento/genética , Ilhas de CpG/genética , Metilação de DNA , Leucemia Linfocítica Crônica de Células B/sangue , Acetiltransferases/genética , Idoso , Estudos de Casos e Controles , Elongases de Ácidos Graxos , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Fatores de Transcrição Kruppel-Like , Proteínas com Homeodomínio LIM/genética , Proteínas de Membrana/genética , Metaloproteínas/genética , Pessoa de Meia-Idade , Proteínas Musculares/genética , Fatores de Transcrição Sp/genética , Fatores de Transcrição/genética , Proteínas com Motivo TripartidoRESUMO
Human lymphocyte antigen-G (HLA-G) is an immunosuppressive molecule that induces functional silencing of immune component cells and can be responsible for immunosuppression in patients with multiple myeloma (MM). Immune dysfunction is an important feature of MM and leads to infections as well as may promote disease progression. Ninety-five patients were included in this study. In MM, the sHLA-G levels were increased when compared to healthy volunteers and the levels of sHLA-G correlated with concentration of creatinine. Interestingly, we detected high levels of sHLA-G in patients with renal insufficiency without any malignant disease but levels were lower than in MM.
Assuntos
Antígenos HLA-G/sangue , Mieloma Múltiplo/sangue , Insuficiência Renal/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Estudos de Coortes , Creatinina/análise , Creatinina/sangue , Feminino , Antígenos HLA-G/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/metabolismo , Concentração Osmolar , Insuficiência Renal/imunologia , Insuficiência Renal/metabolismo , Solubilidade , Adulto JovemRESUMO
Programmed death-1 (PD-1) is an immunoreceptor predominantly expressed on exhausted T cells, which through an interaction with its ligand (PD-L1), controls peripheral tolerance by limiting effector functions of T lymphocytes. qRT-PCR for PD-1, PD-L1 and their splicing forms as well as flow cytometric assessment of surface expression was performed in a cohort of 58 chronic lymphocytic leukemia (CLL) patients. In functional studies, we assessed the influence of the proliferative response of leukemic B-cells induced by IL-4 and CD40L on PD-1 transcripts and expression on the protein level. The median level of PD-1, but not PD-L1, transcripts in CLL patients was higher in comparison to healthy volunteers (HVs, n = 43, p = 0.0057). We confirmed the presence of PD-1 and PD-L1 on the CLL cell surface, and found the expression of PD-1, but not PD-L1, to be higher among CLL patients in comparison to HVs (47.2% vs. 14.8%, p<0.0001). The Kaplan-Meier curves for the time to progression and overall survival in groups with high and low surface expression of PD-1 and PD-L1 revealed no prognostic value in CLL patients. After stimulation with IL-4 and CD40L, protein expression of PD-1 was significantly increased in samples that responded and up-regulated CD38. PD-1, which is aberrantly expressed both at mRNA and cell surface levels in CLL cells might represent a novel immunotolerant molecule involved in the pathomechanism of the disease, and could provide a novel target for future therapies.
Assuntos
Linfócitos B/metabolismo , Antígeno B7-H1/metabolismo , Leucemia Linfocítica Crônica de Células B/metabolismo , Receptor de Morte Celular Programada 1/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Antígeno B7-H1/genética , Antígenos CD40/farmacologia , Feminino , Humanos , Interleucina-4/farmacologia , Leucemia Linfocítica Crônica de Células B/imunologia , Masculino , Pessoa de Meia-Idade , Receptor de Morte Celular Programada 1/genéticaRESUMO
Although the immune status of chronic lymphocytic leukemia (CLL) patients is mostly characterized by immunosuppression, there is an accumulation of in vivo (graft-versus-leukemia effect) and in vitro (spontaneous remissions after infections) data that indicates that CLL might be effectively targeted by T-cell based immunotherapy. Recently, we characterized receptor for hyaluronic acid mediated motility (RHAMM) as a preferential target for immunotherapy of CLL. We also completed a RHAMM-derived peptide vaccination phase I/II clinical trial in CLL. Here, we present a detailed immunological analysis of six CLL patients vaccinated with HLA-A2 restricted RHAMM-derived epitope R3 (ILSLELMKL). Beside effective induction of R3-specific cytotoxic T-cells, peptide vaccination caused profound changes in different T-cell subsets as well as cytokines. We present longitudinal analyses of Th17, CD8(+)CD103(+), CD8(+)CD137(+) and IL-17 producing CD8(+) T cells (CD8(+)IL- -17(+)) as well as important cytokines involved in regulation of immune response such as TGF-ß, IL-10, IL-2 and TNF throughout the peptide vaccination period.