RESUMEN
During infection and inflammation, a reduced oxygen level clearly affects cellular functions. Oxygen levels during CNS infections are unknown. Here we established and evaluated an in vivo measurement system to characterize the oxygen level in parallel with bacterial numbers (CFU/mL), the cell number and pH level inside the CSF of healthy compared to Streptococcus suis-infected pigs. The animals were anesthetized over a seven-hour period with isoflurane in air/oxygen at physiologic arterial partial pressure of oxygen. Oxygen levels in CSF of anesthetized pigs were compared to euthanized pigs. The detected partial pressure of oxygen in the CSF remained constant in a range of 47-63 mmHg, independent of the infection status (bacterial or cell number). In contrast, the pH value showed a slight drop during infection, which correlated with cell and bacterial number in CSF. We present physiologic oxygen and pH values in CSF during the onset of bacterial meningitis.
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Infecciones Bacterianas del Sistema Nervioso Central/líquido cefalorraquídeo , Infecciones Bacterianas del Sistema Nervioso Central/fisiopatología , Oxígeno/líquido cefalorraquídeo , Infecciones Estreptocócicas/líquido cefalorraquídeo , Infecciones Estreptocócicas/fisiopatología , Streptococcus suis/aislamiento & purificación , Animales , Femenino , Masculino , PorcinosRESUMEN
Bacteremia is a hallmark of invasive Streptococcus suis infections of pigs, often leading to septicemia, meningitis, or arthritis. An important defense mechanism of neutrophils is the generation of reactive oxygen species (ROS). In this study, we report high levels of ROS production by blood granulocytes after intravenous infection of a pig with high levels of S. suis-specific antibodies and comparatively low levels of bacteremia. This prompted us to investigate the working hypothesis that the immunoglobulin-mediated oxidative burst contributes to the killing of S. suis in porcine blood. Several S. suis strains representing serotypes 2, 7, and 9 proved to be highly susceptible to the oxidative burst intermediate hydrogen peroxide, already at concentrations of 0.001%. The induction of ROS in granulocytes in ex vivo-infected reconstituted blood showed an association with pathogen-specific antibody levels. Importantly, inhibition of ROS production by the NADPH oxidase inhibitor apocynin led to significantly increased bacterial survival in the presence of high specific antibody levels. The oxidative burst rate of granulocytes partially depended on complement activation, as shown by specific inhibition. Furthermore, treatment of IgG-depleted serum with a specific IgM protease or heat to inactivate complement resulted in >3-fold decreased oxidative burst activity and increased bacterial survival in reconstituted porcine blood in accordance with an IgM-complement-oxidative burst axis. In conclusion, this study highlights an important control mechanism of S. suis bacteremia in the natural host: the induction of ROS in blood granulocytes via specific immunoglobulins such as IgM.
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Granulocitos/fisiología , Estallido Respiratorio/fisiología , Streptococcus suis/inmunología , Enfermedades de los Porcinos/microbiología , Acetofenonas/farmacología , Animales , Inhibidores Enzimáticos/farmacología , Granulocitos/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Streptococcus suis/efectos de los fármacos , Porcinos , Enfermedades de los Porcinos/inmunologíaRESUMEN
BACKGROUND: Inhibition of the mitogen-activated protein kinase (MAPK) pathway as well as programmed death 1 receptor (PD-1) blockade was shown to prolong overall survival (OS) in patients with advanced B-Raf proto-oncogene (BRAF)-mutant melanoma. However, due to the lack of head-to-head trials, it remains unclear if one of these therapeutic approaches should be preferred in first-line therapy. Here, we present a retrospective analysis comparing anti-PD-1 monotherapy with BRAF/MAPK/ERK kinase (MEK) combined inhibition used as first-line agents in a real-world clinical setting. PATIENTS AND METHODS: Clinical data, routine blood counts and lactate dehydrogenase (LDH) levels of 301 patients with unresectable or metastatic melanoma harboring an activating mutation in BRAF (V600E/K) were included. Of these, 106 received anti-PD-1 antibodies, while 195 patients were treated with a selective BRAF inhibitor combined with an MEK inhibitor as palliative first-line therapy. Patients were sub-grouped according to previously described predictive and prognostic markers. RESULTS: OS was significantly longer in patients receiving anti-PD-1 monotherapy compared to patients receiving combined MAPK inhibitors. Subsequent therapies were comparable among these groups. The difference in OS was less pronounced in patients with high LDH levels and visceral metastatic spread. CONCLUSION: First-line treatment with a PD-1 blocking antibody might be associated with longer OS than first-line inhibition of the MAPK pathway in patients with advanced melanoma harboring mutant BRAF. These hypothesis-generating data need to be confirmed or rejected in prospective, randomized trials.
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Anticuerpos Monoclonales Humanizados/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Melanoma/tratamiento farmacológico , Nivolumab/uso terapéutico , Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Proto-Oncogénicas B-raf/genética , Neoplasias Cutáneas/tratamiento farmacológico , Estudios de Cohortes , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Humanos , Melanoma/mortalidad , Metástasis de la Neoplasia , Receptor de Muerte Celular Programada 1/inmunología , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Estudios Retrospectivos , Transducción de Señal , Neoplasias Cutáneas/mortalidad , Análisis de SupervivenciaRESUMEN
The identification of blood-borne biomarkers correlating with melanoma patient survival remains elusive. Novel techniques such as mass cytometry could help to identify melanoma biomarkers, allowing simultaneous detection of up to 100 parameters. However, the evaluation of multiparametric data generated via time-of-flight mass cytometry requires novel analytical techniques because the application of conventional gating strategies currently used in polychromatic flow cytometry is not feasible. In this study, we have employed 38-channel time-of-flight mass cytometry analysis to generate comprehensive immune cell signatures using matrix boolean analysis in a cohort of 28 stage IV melanoma patients and 17 controls. Clusters of parameters were constructed from the abundance of cellular phenotypes significantly different between patients and controls. This approach identified patient-specific combinatorial immune signatures consisting of high-resolution subsets of the T cell, NK cell, B cell, and myeloid compartments. An association with superior survival was characterized by a balanced distribution of myeloid-derived suppressor cell-like and APC-like myeloid phenotypes and differentiated NK cells. The results of this study in a discovery cohort of melanoma patients suggest that multifactorial immune signatures have the potential to allow more accurate prediction of individual patient outcome. Further investigation of the identified immune signatures in a validation cohort is now warranted.
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Biomarcadores de Tumor/sangre , Citometría de Flujo/métodos , Melanoma/inmunología , Melanoma/patología , Adulto , Anciano , Biomarcadores de Tumor/inmunología , Análisis por Conglomerados , Femenino , Humanos , Estimación de Kaplan-Meier , Masculino , Melanoma/mortalidad , Persona de Mediana EdadRESUMEN
Telomerase is best known for its function in maintaining telomeres but has also multiple additional, non-canonical functions. One of these functions is the decrease of oxidative stress and DNA damage due to localisation of the telomerase protein TERT into mitochondria under oxidative stress. However, the exact molecular mechanisms behind these protective effects are still not well understood. We had shown previously that overexpression of human telomerase reverse transcriptase (hTERT) in human fibroblasts results in a decrease of mitochondrial DNA (mtDNA) damage after oxidative stress. MtDNA damage caused by oxidative stress is removed via the base excision repair (BER) pathway. Therefore we aimed to analyse whether telomerase is able to improve this pathway. We applied different types of DNA damaging agents such as irradiation, arsenite treatment (NaAsO2) and treatment with hydrogen peroxide (H2O2). Using a PCR-based assay to evaluate mtDNA damage, we demonstrate that overexpression of hTERT in MRC-5 fibroblasts protects mtDNA from H2O2 and NaAsO2 induced damage, compared with their isogenic telomerase-negative counterparts. However, overexpression of hTERT did not seem to increase repair of mtDNA after oxidative stress, but promoted increased levels of manganese superoxide dismutase (MnSOD) and forkhead-box-protein O3 (FoxO3a) proteins during incubation in serum free medium as well as under oxidative stress, while no differences were found in protein levels of catalase. Together, our results suggest that rather than interfering with mitochondrial DNA repair mechanisms, such as BER, telomerase seems to increase antioxidant defence mechanisms to prevent mtDNA damage and to increase cellular resistance to oxidative stress. However, the result has to be reproduced in additional cellular systems in order to generalise our findings.
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Medio de Cultivo Libre de Suero/química , Mitocondrias/genética , Superóxido Dismutasa/genética , Telomerasa/genética , Arsenitos/efectos adversos , Células Cultivadas , Reparación del ADN , ADN Mitocondrial/genética , Proteína Forkhead Box O3/metabolismo , Humanos , Peróxido de Hidrógeno/efectos adversos , Mitocondrias/efectos de los fármacos , Superóxido Dismutasa/metabolismo , Telomerasa/metabolismo , Rayos Ultravioleta/efectos adversos , Regulación hacia ArribaRESUMEN
Treatment of advanced melanoma patients with ipilimumab results in improved survival. However, only about 20% of treated patients experience long-term benefit. Combining treatment of ipilimumab with other drugs may improve immune activation and potentially enhance clinical efficacy. The aims of the phase II clinical trial reported here were to investigate tolerability and efficacy of a combined immunotherapeutic strategy comprising standard systemic ipilimumab at 3 mg/kg four times at 3-week intervals and intratumorally injected IL-2 at 9 MIU daily twice weekly for four weeks in pretreated melanoma patients with distant metastasis. The primary endpoint was the disease control rate according to immune-related response criteria at week 12; tolerability according to Common Terminology Criteria for Adverse Events criteria was secondary endpoint. No objective responses were observed in the 15 enrolled patients. Three patients had stable disease 12 weeks after starting treatment, yielding a disease control rate of 20%. Tolerability of this combination treatment was acceptable. Observed adverse events were those expected from the respective monotherapies. Autoimmune colitis was observed in two patients. Grade III/IV adverse events were observed in 40% of patients, and no treatment-related deaths occurred. Thus, this combined immunotherapy is associated with adverse events similar to those associated with the respective monotherapies. However, this study does not provide any evidence of improved efficacy of the combination over ipilimumab alone.
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Anticuerpos Monoclonales/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Inmunoterapia/métodos , Interleucina-2/uso terapéutico , Melanoma/terapia , Neoplasias Cutáneas/terapia , Colitis/etiología , Femenino , Humanos , Inmunoterapia/efectos adversos , Ipilimumab , Masculino , Melanoma/inmunología , Melanoma/patología , Persona de Mediana Edad , Metástasis de la Neoplasia , Estadificación de Neoplasias , Estudios Prospectivos , Neoplasias Cutáneas/inmunología , Neoplasias Cutáneas/patología , Resultado del TratamientoRESUMEN
Considering the large number of studies focused on myeloid-derived suppressor cells (MDSCs) to date, only a handful of well-defined relationships in human cancer have been established. The difficulty of assessing the impact of MDSCs in human cancer is partly due to the relatively small number of studies performed in humans. This is compounded in the literature by a common lack of clear indication of which species is being referred to for each characteristic described. These aspects may result in inappropriate extrapolation of animal studies to those in the human setting. This is especially the case for studies focused on investigating therapies which can be used to target MDSCs or those aimed at understanding their mechanism. Here, we attempt to rectify this by reviewing only studies on MDSC performed in humans. We survey studies which explore (1) whether MDSC levels are altered in cancer patients and if this is correlated with patient survival, (2) the so far identified mechanisms employed by MDSC to exert immune suppression, and (3) whether therapeutic agents can be used to target MDSCs by either altering their level, influencing their differentiation or inhibiting their suppressive function. Despite the fact that these studies clearly show that MDSCs are important in human cancer, the clinical employment of agents intended to target them has not yet been accomplished. We identify factors which have contributed to this and propose steps which may facilitate the translation of these therapies to the clinic in future.
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Células Supresoras de Origen Mieloide/patología , Neoplasias/metabolismo , Neoplasias/patología , Humanos , Sistema Inmunológico/patología , Terapia de Inmunosupresión , Modelos Biológicos , Terapia Molecular DirigidaRESUMEN
BACKGROUND: The need for reliable clinical biomarkers to predict which patients with melanoma will benefit from immune checkpoint blockade (ICB) remains unmet. Several different parameters have been considered in the past, including routine differential blood counts, T cell subset distribution patterns and quantification of peripheral myeloid-derived suppressor cells (MDSC), but none has yet achieved sufficient accuracy for clinical utility. METHODS: Here, we investigated potential cellular biomarkers from clinical routine blood counts as well as several myeloid and T cell subsets, using flow cytometry, in two independent cohorts of a total of 141 patients with stage IV M1c melanoma before and during ICB. RESULTS: Elevated baseline frequencies of monocytic MDSCs (M-MDSC) in the blood were confirmed to predict shorter overall survival (OS) (HR 2.086, p=0.030) and progression-free survival (HR 2.425, p=0.001) in the whole patient cohort. However, we identified a subgroup of patients with highly elevated baseline M-MDSC frequencies that fell below a defined cut-off during therapy and found that these patients had a longer OS that was similar to that of patients with low baseline M-MDSC frequencies. Importantly, patients with high M-MDSC frequencies exhibited a skewed baseline distribution of certain other immune cells but these did not influence patient survival, illustrating the paramount utility of MDSC assessment. CONCLUSION: We confirmed that in general, highly elevated frequencies of peripheral M-MDSC are associated with poorer outcomes of ICB in metastatic melanoma. However, one reason for an imperfect correlation between high baseline MDSCs and outcome for individual patients may be the subgroup of patients identified here, with rapidly decreasing M-MDSCs on therapy, in whom the negative effect of high M-MDSC frequencies was lost. These findings might contribute to developing more reliable predictors of late-stage melanoma response to ICB at the individual patient level. A multifactorial model seeking such markers yielded only MDSC behavior and serum lactate dehydrogenase as predictors of treatment outcome.
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Melanoma , Células Supresoras de Origen Mieloide , Humanos , Melanoma/patología , Biomarcadores , Resultado del Tratamiento , Citometría de FlujoRESUMEN
The host-pathogen interaction during meningitis can be investigated with blood-cerebrospinal-fluid-barrier (BCSFB) cell culture models. They are commonly handled under atmospheric oxygen conditions (19-21% O2), although the physiological oxygen conditions are significantly lower in cerebrospinal fluid (CSF) (7-8% O2). We aimed to characterize oxygen levels in a Streptococcus (S.) suis-infected BCSFB model with transmigrating neutrophils. A BCSFB model with human choroid plexus epithelial cells growing on transwell-filters was used. The upper "blood"-compartment was infected and blood-derived neutrophils were added. S. suis and neutrophils transmigrated through the BCSFB into the "CSF"-compartment. Here, oxygen and pH values were determined with the non-invasive SensorDish® reader. Slight orbital shaking improved the luminescence-based measurement technique for detecting free oxygen. In the non-infected BCSFB model, an oxygen value of 7% O2 was determined. However, with S. suis and transmigrating neutrophils, the oxygen value significantly decreased to 2% O2. The pH level decreased slightly in all groups. In conclusion, we characterized oxygen levels in the BCSFB model and demonstrated the oxygen consumption by cells and bacteria. Oxygen values in the non-infected BCSFB model are comparable to in vivo values determined in pigs in the CSF. Infection and transmigrating neutrophils decrease the oxygen value to lower values.
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Barrera Hematoencefálica/química , Técnicas de Cultivo de Célula/métodos , Líquido Cefalorraquídeo/química , Oxígeno/química , Infecciones Estreptocócicas/microbiología , HumanosRESUMEN
The detection of antigen-reactive T cells has shown great utility for patient clinical monitoring. However, many of the methods commonly used to detect these cells face major limitations, like the predetermination of the given HLA type. The herein described protocol provides a means of overcoming many of the obstacles associated with the use of multimers and other common approaches in this field. In order to be able to detect rare cells which are below the detection limit of direct ex vivo measurement, in the present protocol, antigen-reactive T cells are first expanded in vitro using libraries of overlapping peptides which span the entire protein of interest and consist of 15 amino acid-long peptides that share a 12-amino-acid overlap. This theoretically allows the detection of T cells to any epitope within a protein of interest and consequently does not require the patient's HLA type to be characterized. Furthermore, this method simultaneously detects CD4+ and CD8+ T cells that produce cytokines upon encounter with antigen and thereby provides a functional insight into the behavior of the responding T cells. In our case, we have investigated the pro- or anti-inflammatory cytokines IL-2, IL-5, IL-10, IL-17, TNF-α, and IFN-γ.
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Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Citometría de Flujo/métodos , Neoplasias/inmunología , Antígenos de Neoplasias/inmunología , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/metabolismo , Técnicas de Cultivo de Célula/instrumentación , Técnicas de Cultivo de Célula/métodos , Células Cultivadas , Citocinas/inmunología , Citocinas/metabolismo , Citometría de Flujo/instrumentación , Humanos , Activación de Linfocitos/inmunología , Neoplasias/sangreRESUMEN
The role of the assessment of peripheral T-cell phenotypes in predicting overall survival (OS) after ipilimumab treatment is unclear. Here, we analysed mononuclear cells in the blood before and at different time points during treatment with ipilimumab in 137 late-stage melanoma patients. The proportions of baseline naïve and memory T-cells were measured by flow cytometry and correlated with OS, with an emphasis on PD-1 expression. High frequencies (>13%) of CD8 effector-memory type 1 (EM1) T-cells at baseline correlated with longer OS (p = 0.029) and higher clinical response rates (p = 0.01). The frequency of these EM1 cells and the M category had independent impacts on OS (hazard ratio = 1.5, p = 0.033; and hazard ratio = 1.9, p = 0.007). In contrast, high baseline frequencies of late stage-differentiated effector memory CD8 cells (>23.8%) were negatively associated with OS (p = 0.034) but did not correlate with clinical response. Following treatment, a decrease of CD8 cells from baseline to the time of the second drug dose and at later time points was strongly and consistently correlated with a high clinical response rate. Our observations thus suggest an important predictive role of baseline CD8 EM1 cells and changes in CD8 cells for clinical response of ipilimumab. Further validation of these biomarker candidates is warranted.
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Anticuerpos Monoclonales/uso terapéutico , Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/metabolismo , Linfocitos T CD8-positivos/metabolismo , Memoria Inmunológica , Melanoma/tratamiento farmacológico , Neoplasias Cutáneas/tratamiento farmacológico , Adulto , Anciano , Antígenos de Neoplasias/metabolismo , Linfocitos T CD4-Positivos/metabolismo , Femenino , Citometría de Flujo , Humanos , Ipilimumab , Estimación de Kaplan-Meier , Masculino , Melanoma/inmunología , Melanoma/metabolismo , Melanoma/patología , Persona de Mediana Edad , Receptor de Muerte Celular Programada 1/metabolismo , Neoplasias Cutáneas/inmunología , Neoplasias Cutáneas/metabolismo , Neoplasias Cutáneas/patologíaRESUMEN
BACKGROUND: Mucosal melanoma (MM) is a rare but diverse cancer entity. Prognostic factors are not well established for Caucasians with MM. PATIENTS AND METHODS: We analysed the disease course of 444 patients from 15 German skin cancer centres. Disease progression was determined with the cumulative incidence function. Survival times were estimated with the Kaplan-Meier method. Prognostic parameters were identified with multivariate Cox regression analysis. RESULTS: Common anatomic sites of primary tumours were head and neck (MMHN, 37.2%), female genital tract (MMFG, 30.4%) and anorectal region (MMAN, 21.8%). MMAN patients showed the highest vertical tumour thickness (p = 0.001), had a more advanced nodal status (p = 0.014) and a higher percentage of metastatic disease (p = 0.001) at diagnosis. Mutations of NRAS (13.8%), KIT (8.6%) and BRAF (6.4%) were evenly distributed across all tumour site groups. Local relapses were observed in 32.4% and most commonly occurred in the MMHN group (p = 0.016). Male gender (p = 0.047), advanced tumour stage (p = 0.001), nodal disease (p = 0.001) and incomplete resection status (p = 0.001) were independent risk factors for disease progression. Overall survival (OS) was highest in the MMFG group (p = 0.030) and in patients without ulceration (p = 0.004). Multivariate risk factors for OS were M stage at diagnosis (p = 0.002) and incomplete resection of the primary tumour (p = 0.001). CONCLUSION: In this large series of MM patients in a European population, anorectal MM was associated with the poorest prognosis.
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Melanoma , Membrana Mucosa/patología , Neoplasias , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos/uso terapéutico , Terapia Combinada , Femenino , Humanos , Masculino , Melanoma/mortalidad , Melanoma/patología , Melanoma/terapia , Persona de Mediana Edad , Neoplasias/mortalidad , Neoplasias/patología , Neoplasias/terapia , Pronóstico , Análisis de Regresión , Estudios Retrospectivos , Factores de Riesgo , Análisis de Supervivencia , Adulto JovenRESUMEN
Biomarkers are strongly needed for diagnostic surveillance of patients with metastatic melanoma. On the basis of its known association with tumor metastasis and its ability to induce cancer cachexia, we investigated serum levels of growth and differentiation factor 15 (sGDF-15) as a marker for overall survival (OS). sGDF-15 was retrospectively measured by ELISA in 761 samples obtained at distinct time points during routine clinical care of patients with stage III/IV melanoma. In the entire cohort, sGDF-15 ≥ 1.5 ng/ml was strongly associated with reduced OS after assessment. Subsequent analyses were performed separately for tumor-free stage III, tumor-free stage IV, and unresectable stage IV patients. For patients with unresectable distant metastasis (n = 206), sGDF-15 was independently associated with OS when considered together with the M-category and superior to serum level of lactate dehydrogenase. Analysis in tumor-free stage III patients during routine surveillance (n = 468) revealed sGDF-15 to be associated with OS and an independent factor when considered together with S100B and the pattern of locoregional metastasis. Only in tumor-free stage IV patients (n = 87) sGDF-15 was not associated with OS. sGDF-15 should thus be further validated as a marker for early detection of recurrence in stage III patients and as a prognostic or predictive marker particularly in the context newly available treatments in unresectable stage IV patients.
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Biomarcadores de Tumor/sangre , Factor 15 de Diferenciación de Crecimiento/sangre , Melanoma/sangre , Melanoma/patología , Neoplasias Cutáneas/sangre , Neoplasias Cutáneas/patología , Adulto , Factores de Edad , Anciano , Análisis de Varianza , Enfermedad Crítica/terapia , Supervivencia sin Enfermedad , Femenino , Humanos , Estimación de Kaplan-Meier , Masculino , Melanoma/mortalidad , Melanoma/terapia , Persona de Mediana Edad , Análisis Multivariante , Invasividad Neoplásica/patología , Estadificación de Neoplasias , Pronóstico , Medición de Riesgo , Factores Sexuales , Neoplasias Cutáneas/mortalidad , Neoplasias Cutáneas/terapia , Análisis de Supervivencia , Melanoma Cutáneo MalignoRESUMEN
PURPOSE: To investigate changes of peripheral blood biomarkers and their impact on clinical outcome following treatment with ipilimumab in advanced melanoma patients. EXPERIMENTAL DESIGN: Changes in blood counts and the frequency of circulating immune cell populations analyzed by flow cytometry were investigated in 82 patients to compare baseline values with different time-points after starting ipilimumab. Endpoints were overall survival (OS) and best clinical response. Statistical calculations were done by Wilcoxon-matched pairs tests, Fisher exact test, Kaplan-Meier analysis, and Cox regression analysis. RESULTS: Increases in absolute lymphocyte counts (ALC) 2 to 8 weeks (P = 0.003) and in percentages of CD4+ and CD8+ T cells 8 to 14 weeks (P = 0.001 and P = 0.02) after the first dose of ipilimumab were correlated with improved survival. These associations did not meet significance criteria, when conservatively adjusted for multiple testing, but were additionally correlated with clinical responses (all P < 0.05). However, validation is required. Increases in all three factors were observed in 36% of patients, who had a favorable outcome and survival probabilities of 93.3% and 63.8% at 12 and 24 months, respectively. A partial or complete response was observed in 71% of these patients compared with only 8% in patients with decreases in ≥1 of the 3 factors, respectively. Changes of regulatory T cells or myeloid-derived suppressor cells were not associated with OS. CONCLUSIONS: Increases of ALC observed 2 to 8 weeks after initiation of ipilimumab and delayed increases in CD4+ and CD8+ T cells reflect changes associated with positive outcome. These changes represent surrogate marker candidates and warrant further validation. Clin Cancer Res; 22(19); 4848-58. ©2016 AACR.
Asunto(s)
Antineoplásicos Inmunológicos/uso terapéutico , Ipilimumab/uso terapéutico , Melanoma/tratamiento farmacológico , Melanoma/inmunología , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/inmunología , Adulto , Anciano , Biomarcadores de Tumor/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Femenino , Humanos , Estimación de Kaplan-Meier , Recuento de Linfocitos , Masculino , Melanoma/mortalidad , Persona de Mediana Edad , Modelos de Riesgos Proporcionales , Neoplasias Cutáneas/mortalidadRESUMEN
PURPOSE: Biomarkers for outcome after immune-checkpoint blockade are strongly needed as these may influence individual treatment selection or sequence. We aimed to identify baseline factors associated with overall survival (OS) after pembrolizumab treatment in melanoma patients. EXPERIMENTAL DESIGN: Serum lactate dehydrogenase (LDH), routine blood count parameters, and clinical characteristics were investigated in 616 patients. Endpoints were OS and best overall response following pembrolizumab treatment. Kaplan-Meier analysis and Cox regression were applied for survival analysis. RESULTS: Relative eosinophil count (REC) ≥1.5%, relative lymphocyte count (RLC) ≥17.5%, ≤2.5-fold elevation of LDH, and the absence of metastasis other than soft-tissue/lung were associated with favorable OS in the discovery (n = 177) and the confirmation (n = 182) cohort and had independent positive impact (all P < 0.001). Their independent role was subsequently confirmed in the validation cohort (n = 257; all P < 0.01). The number of favorable factors was strongly associated with prognosis. One-year OS probabilities of 83.9% versus 14.7% and response rates of 58.3% versus 3.3% were observed in patients with four of four compared to those with none of four favorable baseline factors present, respectively. CONCLUSIONS: High REC and RLC, low LDH, and absence of metastasis other than soft-tissue/lung are independent baseline characteristics associated with favorable OS of patients with melanoma treated with pembrolizumab. Presence of four favorable factors in combination identifies a subgroup with excellent prognosis. In contrast, patients with no favorable factors present have a poor prognosis, despite pembrolizumab, and additional treatment advances are still needed. A potential predictive impact needs to be further investigated. Clin Cancer Res; 22(22); 5487-96. ©2016 AACR.
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Anticuerpos Monoclonales Humanizados/uso terapéutico , Anticuerpos Monoclonales/uso terapéutico , Biomarcadores de Tumor/metabolismo , Melanoma/tratamiento farmacológico , Melanoma/metabolismo , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/metabolismo , Anciano , Estudios de Cohortes , Femenino , Humanos , Estimación de Kaplan-Meier , L-Lactato Deshidrogenasa/metabolismo , Masculino , Melanoma/patología , Persona de Mediana Edad , Pronóstico , Neoplasias Cutáneas/patología , Análisis de Supervivencia , Resultado del TratamientoRESUMEN
PURPOSE: To identify baseline peripheral blood biomarkers associated with clinical outcome following ipilimumab treatment in advanced melanoma patients. EXPERIMENTAL DESIGN: Frequencies of myeloid-derived suppressor cells (MDSC) and regulatory T cells (Treg), serum lactate dehydrogenase (LDH), routine blood counts, and clinical characteristics were assessed in 209 patients. Endpoints were overall survival (OS) and best overall response. Statistical calculations were done by Kaplan-Meier and Cox regression analysis, including calibration and discrimination by C-statistics. RESULTS: Low baseline LDH, absolute monocyte counts (AMC), Lin(-)CD14(+)HLA-DR(-/low)-MDSC frequencies, and high absolute eosinophil counts (AEC), relative lymphocyte counts (RLC), and CD4(+)CD25(+)FoxP3(+)-Treg frequencies were significantly associated with better survival, and were considered in a combination model. Patients (43.5%) presenting with the best biomarker signature had a 30% response rate and median survival of 16 months. In contrast, patients with the worst biomarkers (27.5%) had only a 3% response rate and median survival of 4 months. The occurrence of adverse events correlated with neither baseline biomarker signatures nor the clinical benefit of ipilimumab. In another model, limited to the routine parameters LDH, AMC, AEC, and RLC, the number of favorable factors (4 vs. 3 vs. 2-0) was also associated with OS (P < 0.001 for all pairwise comparisons) in the main study and additionally in an independent validation cohort. CONCLUSIONS: A baseline signature of low LDH, AMC, and MDSCs as well as high AEC, Tregs, and RLC is associated with favorable outcome following ipilimumab. Prospective investigation of the predictive impact of these markers following ipilimumab and other treatments, e.g., PD-1 antibodies, is warranted. Clin Cancer Res; 22(12); 2908-18. ©2016 AACR.
Asunto(s)
Antineoplásicos Inmunológicos/uso terapéutico , Biomarcadores de Tumor/sangre , Ipilimumab/uso terapéutico , Melanoma/tratamiento farmacológico , Neoplasias Cutáneas/tratamiento farmacológico , Anciano , Antineoplásicos Inmunológicos/efectos adversos , Recuento de Eritrocitos , Femenino , Humanos , Ipilimumab/efectos adversos , L-Lactato Deshidrogenasa/sangre , Recuento de Linfocitos , Masculino , Melanoma/mortalidad , Melanoma/patología , Células Supresoras de Origen Mieloide/citología , Neoplasias Cutáneas/mortalidad , Neoplasias Cutáneas/patología , Linfocitos T Reguladores/citología , Resultado del TratamientoRESUMEN
Human γδ T-cells possess regulatory and cytotoxic capabilities, and could potentially influence the efficacy of immunotherapies. We analysed the frequencies of peripheral γδ T-cells, including their most prominent subsets (Vδ1+ and Vδ2+ cells) and differentiation states in 109 melanoma patients and 109 healthy controls. We additionally analysed the impact of γδ T-cells on overall survival (OS) calculated from the first dose of ipilimumab in melanoma patients. Higher median frequencies of Vδ1+ cells and lower median frequencies of Vδ2+ cells were identified in patients compared to healthy subjects (Vδ1+: 30% versus 15%, Vδ2+: 39% versus 64%, both p < 0.001). Patients with higher frequencies of Vδ1+ cells (≥30%) had poorer OS (p = 0.043) and a Vδ1+ differentiation signature dominated by late-differentiated phenotypes. In contrast, higher frequencies of Vδ2+ cells (≥39%) were associated with longer survival (p = 0.031) independent of the M category or lactate dehydrogenase level. Patients with decreasing frequencies of Vδ2+ cells under ipilimumab treatment had worse OS and a lower rate of clinical benefit than patients without such decreases. Therefore, we suggest frequencies of both Vδ1+ and Vδ2+ cells as candidate biomarkers for outcome in melanoma patients following ipilimumab. Further studies are needed to validate these results and to clarify whether they represent prognostic associations or whether γδ T-cells are specifically and/or functionally linked to the mode of action of ipilimumab.
Asunto(s)
Anticuerpos Monoclonales/uso terapéutico , Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/sangre , Inmunoterapia/métodos , Melanoma/tratamiento farmacológico , Melanoma/inmunología , Receptores de Antígenos de Linfocitos T gamma-delta/sangre , Subgrupos de Linfocitos T/citología , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Diferenciación Celular , Femenino , Citometría de Flujo , Humanos , Ipilimumab , Masculino , Persona de Mediana Edad , Pronóstico , Subgrupos de Linfocitos T/inmunologíaRESUMEN
The melanoma-associated antigens Melan-A and NY-ESO-1 stimulate different T-cell responses in late-stage melanoma patients. Either CD4+ or CD8+ T-cell reactivity against NY-ESO-1 was associated with better prognosis, but for Melan-A, only CD8+ but not CD4+ T-cell responses were associated with longer survival.
RESUMEN
The percentage of circulating CD14+CD11b+HLA-DR-/low myeloid-derived suppressor cells (MDSCs) inversely correlates with survival among advanced melanoma patients. High levels of MDSCs are associated with the absence of T lymphocytes specific for melanoma-derived antigens, implying a causal and clinically relevant interaction between these cell subsets. MDSCs might therefore represent prognostic markers as well as targets for the development of novel therapeutic interventions against melanoma.
RESUMEN
PURPOSE: To analyze the prognostic relevance and relative impact of circulating myeloid-derived suppressor cells (MDSC) and regulatory T cells (Treg) compared with functional tumor antigen-specific T cells in patients with melanoma with distant metastasis. EXPERIMENTAL DESIGN: The percentage of CD14(+)CD11b(+)HLA-DR(-/low) MDSCs, CD4(+)CD25(+)FoxP3(+) Tregs, and the presence of NY-ESO-1- or Melan-A-specific T cells was analyzed in 94 patients and validated in an additional cohort of 39 patients by flow cytometry. Univariate survival differences were calculated according to Kaplan-Meier and log-rank tests. Multivariate analyses were performed using Cox regression models. RESULTS: NY-ESO-1-specific T cells, the M-category, and the frequency of MDSCs were associated with survival. The absence of NY-ESO-1-specific T cells and the M-category M1c independently increased the risk of death. In a second Cox model not considering results on antigen-specific T cells, a frequency of >11% MDSCs showed independent impact. Its association with survival was confirmed in the additional patient cohort. Median survival of patients with a lower frequency of MDSCs was 13 months versus 8 months for others (P < 0.001, combined cohorts). We observed a strong correlation between high levels of MDSCs and the absence of melanoma antigen-specific T cells implying a causal and clinically relevant interaction. No prognostic impact was observed for Tregs. CONCLUSIONS: Circulating CD14(+)CD11b(+)HLA-DR(-/low) MDSCs have a negative impact on survival and inversely correlate with the presence of functional antigen-specific T cells in patients with advanced melanoma. Our findings provide a rationale to investigate MDSC-depleting strategies in the therapeutic setting especially in combination with vaccination or T-cell transfer approaches.