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1.
Chem Sci ; 12(38): 12587-12599, 2021 Oct 06.
Artigo em Inglês | MEDLINE | ID: mdl-34703544

RESUMO

Oxaliplatin is a very potent platinum(ii) drug which is frequently used in poly-chemotherapy schemes against advanced colorectal cancer. However, its benefit is limited by severe adverse effects as well as resistance development. Based on their higher tolerability, platinum(iv) prodrugs came into focus of interest. However, comparable to their platinum(ii) counterparts they lack tumor specificity and are frequently prematurely activated in the blood circulation. With the aim to exploit the enhanced albumin consumption and accumulation in the malignant tissue, we have recently developed a new albumin-targeted prodrug, which supposed to release oxaliplatin in a highly tumor-specific manner. In more detail, we designed a platinum(iv) complex containing two maleimide moieties in the axial position (KP2156), which allows selective binding to the cysteine 34. In the present study, diverse cell biological and analytical tools such as laser ablation inductively-coupled plasma mass spectrometry (LA-ICP-MS), isotope labeling, and nano-scale secondary ion mass spectrometry (NanoSIMS) were employed to better understand the in vivo distribution and activation process of KP2156 (in comparison to free oxaliplatin and a non-albumin-binding succinimide analogue). KP2156 forms very stable albumin adducts in the bloodstream resulting in a superior pharmacological profile, such as distinctly prolonged terminal excretion half-life and enhanced effective platinum dose (measured by ICP-MS). The albumin-bound drug is accumulating in the malignant tissue, where it enters the cancer cells via clathrin- and caveolin-dependent endocytosis, and is activated by reduction to release oxaliplatin. This results in profound, long-lasting anticancer activity of KP2156 against CT26 colon cancer tumors in vivo based on cell cycle arrest and apoptotic cell death. Summarizing, albumin-binding of platinum(iv) complexes potently enhances the efficacy of oxaliplatin therapy and should be further developed towards clinical phase I trials.

2.
Eur J Clin Pharmacol ; 77(10): 1473-1484, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-33969434

RESUMO

PURPOSE: AT04A and AT06A are two AFFITOPE® peptide vaccine candidates being developed for the treatment of hypercholesterolemia by inducing proprotein convertase subtilisin/kexin type 9 (PCSK9)-specific antibodies. This study aimed to investigate safety, tolerability, antibody development, and reduction of low-density lipoprotein cholesterol (LDLc) following four subcutaneous immunizations. METHODS: This phase I, single-blind, randomized, placebo-controlled study was conducted in a total of 72 healthy subjects with a mean fasting LDLc level at baseline of 117.1 mg/dL (range 77-196 mg/dL). Each cohort enrolled 24 subjects to receive three priming immunizations at weeks 0, 4, and 8 and to receive a single booster immunization at week 60 of either AT04A, AT06A, or placebo. In addition to safety (primary objective), the antigenic peptide- and PCSK9-specific antibody response and the impact on LDLc were evaluated over a period of 90 weeks. RESULTS: The most common systemic treatment-related adverse events (AEs) reported were fatigue, headache, and myalgia in 75% of subjects in the AT06A group and 58% and 46% of subjects in the placebo and AT04A groups, respectively. Injection site reactions (ISR) representing 63% of all treatment-emergent adverse events (TEAEs), were transient and mostly of mild or moderate intensity and rarely severe (3%). Both active treatments triggered a robust, long-lasting antibody response towards the antigenic peptides used for immunization that optimally cross-reacted with the target epitope on PCSK9. In the AT04A group, a reduction in serum LDLc was observed with a mean peak reduction of 11.2% and 13.3% from baseline compared to placebo at week 20 and 70 respectively, and over the whole study period, the mean LDLc reduction for the AT04A group vs. placebo was -7.2% (95% CI [-10.4 to -3.9], P < 0.0001). In this group, PCSK9 target epitope titers above 50 were associated with clinically relevant LDLc reductions with an individual maximal decrease of 39%. CONCLUSIONS: Although both AT04A and AT06 were safe and immunogenic, only AT04A demonstrated significant LDLc-lowering activity, justifying further development. TRIAL REGISTRATION: EudraCT: 2015-001719-11. ClinicalTrials.gov Identifier: NCT02508896.

3.
Cancer Chemother Pharmacol ; 88(2): 203-209, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33907881

RESUMO

PURPOSE: As critical parameter after extravasation of cytotoxic vesicants, anthracyclines were determined in removed tissue from patients requiring surgical intervention due to tissue necrosis. We monitored their distribution within the affected lesion to establish a possible dose-toxicity relation. METHODS: From six patients scheduled for surgery, removed tissue flaps were systematically analysed by HPLC (epirubicin: 5 subjects; doxorubicin: 1 subject). RESULTS: After extravasation, tissue concentrations were highly variable with an individual anthracycline distribution pattern ranging from a few nanograms up to 17 µg per 100 mg tissue, which indicated a substantial difference in tissue sensitivity among patients. The resection borders coincided with the extension of the erythema and guided the surgical intervention after demarcation of the lesion, which occurred usually 2 or 3 weeks after extravasation. At that time, drug was hardly detected at the resection borders. Wound drains were negative for the extravasated drugs while showing a time profile of vascular growth factors and inflammatory cytokines, which was highly similar to routine surgery. In all six patients, surgical debridement with immediate wound closure led to healing within approximately 2 weeks, when therapy was resumed in all patients with reasonable time delay. CONCLUSION: Surgical intervention after demarcation of the extravasation lesion allows for almost uninterrupted continuation of treatment independent of the amount of extravasated anthracycline. As even minor amounts of the vesicants may trigger tissue necrosis, preventive measures merit the highest priority.


Assuntos
Antibióticos Antineoplásicos/efeitos adversos , Antibióticos Antineoplásicos/farmacocinética , Epirubicina/efeitos adversos , Epirubicina/farmacocinética , Distribuição Tecidual/fisiologia , Idoso , Antraciclinas/efeitos adversos , Antraciclinas/farmacocinética , Antraciclinas/uso terapêutico , Antineoplásicos/efeitos adversos , Antineoplásicos/farmacocinética , Antineoplásicos/uso terapêutico , Citocinas/metabolismo , Doxorrubicina/efeitos adversos , Doxorrubicina/farmacocinética , Doxorrubicina/uso terapêutico , Epirubicina/uso terapêutico , Feminino , Humanos , Inflamação/metabolismo , Masculino , Pessoa de Meia-Idade , Necrose/induzido quimicamente , Necrose/metabolismo , Dermatopatias/tratamento farmacológico , Dermatopatias/metabolismo , Retalhos Cirúrgicos/patologia , Cicatrização/efeitos dos fármacos
4.
Wien Klin Wochenschr ; 133(1-2): 32-40, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-33296026

RESUMO

BACKGROUND: Advanced therapy-refractory parotid gland carcinomas have a poor prognosis with limited therapy options. We used molecular profiling to offer molecular guided therapies to patients with advanced metastatic parotid gland malignancies. METHODS: In this retrospective analysis we describe the molecular profiling of ten patients diagnosed with therapy-refractory metastatic parotid gland malignancies. RESULTS: We identified seven genetic aberrations in five patients: two mutations in CDKN2A and one mutation in APC, ATM, TP53, SMARCB1 and FGFR1, respectively. No mutations were detected in five patients. The IHC demonstrated frequent expressions of EGFR and p­mTOR, as well as PTEN in eight patients. For four fifths (n = 8) of the patients, a targeted therapy was suggested. Eventually, three patients received the targeted therapy recommendation and one patient achieved stable disease for 14 months. CONCLUSION: A total of eight therapy recommendations were provided. Based on our observations, molecular-guided therapies may be a feasible treatment approach for this rare disease entity.


Assuntos
Carcinoma , Glândula Parótida , Adulto , Humanos , Terapia de Alvo Molecular , Medicina de Precisão , Estudos Retrospectivos
5.
J Pers Med ; 10(4)2020 Dec 11.
Artigo em Inglês | MEDLINE | ID: mdl-33322358

RESUMO

In this analysis, we examined the efficacy, feasibility, and limitations of molecular-based targeted therapies in heavily pretreated metastatic colorectal cancer (mCRC) patients after failure of all standard treatments. In this single-center, real-world retrospective analysis of our platform for precision medicine, we mapped the molecular profiles of 60 mCRC patients. Tumor samples of the patients were analyzed using next-generation sequencing panels of mutation hotspots, microsatellite instability testing, and immunohistochemistry. All profiles were reviewed by a multidisciplinary team to provide a targeted treatment recommendation after consensus discussion. In total, we detected 166 mutations in 53 patients. The five most frequently found mutations were TP53, KRAS, APC, PIK3CA, and PTEN. In 28 cases (47% of all patients), a molecularly targeted therapy could be recommended. Eventually, 12 patients (20%) received the recommended therapy. Six patients (10%) had a clinical benefit. The median time to treatment failure was 3.1 months. Our study demonstrates the feasibility and applicability of using targeted therapies in daily clinical practice for heavily pretreated mCRC patients. This could be used as a targeted treatment option in half of the patients.

6.
Cancers (Basel) ; 12(11)2020 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-33203166

RESUMO

Recurrent/metastatic (R/M) head and neck cancers bear a poor prognosis. In this analysis, we examined the efficacy and the outcome of targeted therapy recommendations based on the patients' molecular tumor portrait after failure of all standard therapy options. In this single-center, real-world retrospective analysis of our platform for precision medicine, we analyzed the molecular profile of 50 patients diagnosed with R/M head and neck cancer. Tumor samples of the patients were examined using next-generation sequencing panels of mutation hotspots, microsatellite instability (MSI) testing, and immunohistochemistry (IHC). In 31 cases (62.0% of all patients), a molecular-driven targeted therapy approach was recommended. Eventually, 14 patients (28%) received the suggested targeted therapy. Six of fourteen patients (43%) achieved stable disease conditions and four patients (29%) experienced a progressive disease. The median time to treatment failure was 2.8 months. Therapy recommendations were significantly more often issued for men (p = 0.037) than for women. This analysis demonstrated that precision medicine provided the basis for molecular-driven therapy recommendations in over half of the patients with advanced therapy refractory head and neck cancers, with significantly more therapy recommendations for men. Our analysis showed that although precision medicine approaches are implementable and feasible for the management of recurrent/metastatic head and neck cancers in daily clinical routine, there are major limitations and challenges that have to be overcome.

7.
J Pers Med ; 10(4)2020 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-33114048

RESUMO

Advanced therapy-refractory solid tumors bear a dismal prognosis and constitute a major challenge in offering effective treatment strategies. In this real-world retrospective analysis of our precision medicine platform MONDTI, we describe the molecular profile of 554 patients diagnosed with 17 different types of advanced solid tumors after failure of all standard treatment options. In 304 cases (54.9% of all patients), a molecular-driven targeted therapy approach could be recommended, with a recommendation rate above 50% in 12 tumor entities. The three highest rates for therapy recommendation per tumor classification were observed in urologic malignancies (90.0%), mesothelioma (78.6%), and male reproductive cancers (71.4%). Tumor type (p = 0.46), expression of p-mTOR (p = 0.011), expression of EGFR (p = 0.046), and expression of PD-L1 (p = 0.023) had a significant impact on the targeted therapy recommendation rate. Therapy recommendations were significantly more often issued for men (p = 0.015) due to gender-specific differences in the molecular profiles of patients with head and neck cancer and malignant mesothelioma. This analysis demonstrates that precision medicine was feasible and provided the basis for molecular-driven therapy recommendations in patients with advanced therapy refractory solid tumors.

8.
Ther Adv Med Oncol ; 12: 1758835920938611, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32699558

RESUMO

Background: Metastatic pancreatic ductal adenocarcinoma (mPDAC) bears a dismal prognosis due to the limited activity of systemic chemotherapy. In our platform for precision medicine, we aim to offer molecular-guided treatments to patients without further standard therapy options. Methods: In this single center, real-world retrospective analysis of our platform, we describe the molecular-based therapy approaches used in all 50 patients diagnosed with therapy-refractory mPDAC. A molecular portrait of the tumor specimens was created by next-generation sequencing, immunohistochemistry (IHC), microsatellite instability (MSI) testing, and fluorescence in situ hybridization. Results: In total, we detected 123 mutations in 50 patients. The five most frequent mutations were KRAS (n = 40; 80%), TP53 (n = 29; 58%), CDKN2A (n = 8; 16%), SMAD4 (n = 4; 8%), and NOTCH1 (n = 4; 8%), which together accounted for 40.2% of all mutations. Two patients had gene fusions, namely, TBL1XR1-PIK3CA and EIF3E-RSPO2. IHC detected expression of EGFR, phosphorylated mTOR, and PTEN in 36 (72%), 33 (66%), and 17 patients (34%), respectively. For 14 (28%) of the 50 patients, a targeted therapy was suggested based on the identified molecular targets. The recommended treatments included the mTOR inhibitor everolimus (n = 3), pembrolizumab (n = 3), palbociclib (n = 2), nintedanib (n = 2), and cetuximab, crizotinib, tamoxifen, and the combination of lapatinib and trastuzumab, in one patient each.Finally, five patients received the recommended therapy. Four patients died due to disease progression before radiological assessment. One patient was treated with nintedanib and achieved stable disease for 6 months. Conclusion: Based on our observations, precision medicine approaches are feasible and implementable in clinical routine and may provide molecular-based therapy recommendations for mPDAC.

9.
Oncologist ; 25(7): e1060-e1069, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32369643

RESUMO

INTRODUCTION: Advanced gynecologic cancers have a poor prognosis and constitute a major challenge for adequate treatment strategies. By analyzing and targeting molecular alterations, molecular guided treatments may be a viable option for the treatment of advanced gynecologic cancers. PATIENTS AND METHODS: In this single-center, real-world retrospective analysis of our platform for precision cancer medicine (PCM), we describe the molecular profiling of 72 patients diagnosed with different types of advanced gynecologic malignancies. Tumor samples of the patients were examined by next-generation sequencing panel and immunohistochemistry (IHC). RESULTS: In total, we identified 209 genetic aberrations in 72 patients. The ten most frequent alterations were TP53 (n = 42, 20%), KRAS (n = 14, 6.6%), PIK3CA (n = 11, 5.2%), PIK3R1 (n = 9, 4.3%), ATR (n = 8, 3.8%), PTEN (n = 8, 3.8%), BRCA1 (n = 6, 2.8%), NF1 (n = 4, 1.9%), NOTCH1 (n = 4, 1.9%), and POLE (n = 4, 1.9%), which account for more than half of all molecular alterations (52.6%). In 21 (29.1%) patients only one mutation could be detected, and 44 (61.1%) patients had more than one mutation. No molecular alterations were detected in seven (9.7%) patients. IHC detected expression of phosphorylated mammalian target of rapamycin and epidermal growth factor receptor in 58 (80.6%) and 53 (73.6%) patients, respectively. In over two thirds (n = 49, 68.1%), a targeted therapy was suggested, based on the identified genetic aberrations. The most frequently recommended specific treatment was the combination of everolimus with exemestane (n = 18, 25 %). CONCLUSION: Based on our observations, it seems that PCM might be a feasible approach for advanced gynecologic cancers with limited treatment options. IMPLICATIONS FOR PRACTICE: Nowadays molecular profiling of advanced gynecologic malignancies is feasible in the clinical routine. A molecular portrait should be done for every patient with an advanced therapy-refractory gynecologic malignancy to offer molecular-based treatment concepts.


Assuntos
Neoplasias dos Genitais Femininos , Medicina de Precisão , Feminino , Neoplasias dos Genitais Femininos/tratamento farmacológico , Neoplasias dos Genitais Femininos/genética , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Terapia de Alvo Molecular , Mutação , Estudos Retrospectivos
10.
Thorac Cancer ; 11(7): 1979-1988, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32438515

RESUMO

BACKGROUND: Malignant mesothelioma is an aggressive cancer and has a poor prognosis. Here, we analyzed the feasibility, molecular and gender aspects of targeted therapy recommendations for malignant mesothelioma based on the individual molecular tumor profile. METHODS: In this single-center, real-world retrospective analysis of our platform for precision medicine, we evaluated the molecular profiling of malignant mesothelioma in 14 patients, including nine men and five women. Tumor samples of the patients were examined with a 50 gene next-generation sequencing (NGS) panel, immunohistochemistry, and fluorescence in situ hybridization, to detect possible molecular aberrations which may be targeted by off-label therapy custom-tailored to the individual patient. RESULTS: In total, we identified 11 mutations in six of the 14 patients, including BAP1, FANCA, NF1, NF2, PD-L1, RAD52D, SETD2, SRC, and TP53. No mutation was detected in eight of the 14 patients. Targeted therapy was recommended for 11 out of the 14 patients. All recommendations were mainly based on the molecular characteristics determined by immunohistochemistry. Targeted therapy recommendations were significantly more often for men than women due to gender-specific differences in PDGFRα expression. Eventually, four patients received the targeted therapy, of whom one patient subsequently achieved stable disease. CONCLUSIONS: Our observations suggest that a molecular-guided treatment approach is feasible for the management of advanced malignant mesothelioma. Our analysis revealed gender specific differences in PDGFRα expression that should be further evaluated in clinical trials.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Biomarcadores Tumorais/genética , Mesotelioma Maligno/tratamento farmacológico , Mutação , Recidiva Local de Neoplasia/tratamento farmacológico , Seleção de Pacientes , Neoplasias Peritoneais/tratamento farmacológico , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Mesotelioma Maligno/genética , Mesotelioma Maligno/patologia , Pessoa de Meia-Idade , Terapia de Alvo Molecular , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/patologia , Neoplasias Peritoneais/genética , Neoplasias Peritoneais/secundário , Medicina de Precisão , Prognóstico , Estudos Retrospectivos , Caracteres Sexuais , Taxa de Sobrevida
11.
Cell Death Dis ; 10(10): 696, 2019 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-31541075

RESUMO

Medulloblastoma, an embryonal tumor of the cerebellum/fourth ventricle, is one of the most frequent malignant brain tumors in children. Although genetic variants are increasingly used in treatment stratification, survival of high-risk patients, characterized by leptomeningeal dissemination, TP53 mutation or MYC amplification, is still poor. FOXM1, a proliferation-specific oncogenic transcription factor, is deregulated in various solid tumors, including medulloblastoma, and triggers cellular proliferation, migration and genomic instability. In tissue samples obtained from medulloblastoma patients, the significant upregulation of FOXM1 was associated with a loss of its putative regulating microRNA, miR-4521. To understand the underlying mechanism, we investigated the effect of miR-4521 on the expression of the transcription factor FOXM1 in medulloblastoma cell lines. Transfection of this microRNA reduced proliferation and invasion of several medulloblastoma cell lines and induced programmed cell death through activation of caspase 3/7. Further, downstream targets of FOXM1 such as PLK1 and cyclin B1 were significantly reduced thus affecting the cell cycle progression in medulloblastoma cell lines. In conclusion, a restoration of miRNA-4521 may selectively suppress the pathophysiological effect of aberrant FOXM1 expression and serve as a targeted approach for medulloblastoma therapy.


Assuntos
Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Proteína Forkhead Box M1/biossíntese , Meduloblastoma/genética , Meduloblastoma/metabolismo , MicroRNAs/metabolismo , Regiões 3' não Traduzidas , Adolescente , Neoplasias Encefálicas/patologia , Pontos de Checagem do Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Proliferação de Células/fisiologia , Criança , Pré-Escolar , Regulação para Baixo , Feminino , Proteína Forkhead Box M1/genética , Proteína Forkhead Box M1/metabolismo , Humanos , Lactente , Masculino , Meduloblastoma/patologia , MicroRNAs/biossíntese , MicroRNAs/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Esferoides Celulares , Regulação para Cima
12.
Clin Cancer Res ; 25(9): 2737-2744, 2019 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-30647078

RESUMO

PURPOSE: Brain metastases (BMs) are a rare but devastating condition in estrogen receptor (ER)-positive metastatic breast cancer (MBC). Although endocrine therapy (ET) is the mainstay of treatment in this disease subtype, only case reports have been published concerning the activity of ET in BMs henceforth. Therefore, we aimed to systematically investigate the impact of ET after diagnosis of BM on outcome and clinical course of disease in patients with ER-positive MBC. EXPERIMENTAL DESIGN: Patient characteristics, detailed information about BMs including diagnosis-specific graded prognostic assessment class (DS-GPA), and clinical outcome were obtained by retrospective chart review for all patients treated for ER-positive breast cancer BMs between 1990 and 2017 at an academic care center. Overall survival (OS) was measured as the interval from diagnosis of BM until death or last date of follow-up. RESULTS: Overall, 198 patients [female: 195/198 (98.5%); male: 3/198 (1.5%)] with ER-positive breast cancer BMs were available for this analysis. Eighty-eight of 198 patients (44.4%) received ET after diagnosis of BM including aromatase inhibitors (AIs; letrozole, anastrozole, exemestane), tamoxifen, and fulvestrant. Median OS was significantly longer in patients receiving ET after diagnosis of BM compared with patients who did not (15 vs. 4 months, P < 0.001; log-rank test). No significant difference in terms of OS was observed between patients receiving AIs, tamoxifen, or fulvestrant. In patients with concomitant leptomeningeal carcinomatosis (LC), ET prolonged median OS significantly as well (7 vs. 3 months, P = 0.012; log-rank test). In a multivariate analysis including DS-GPA and ET, only treatment with ET after diagnosis of BM (HR, 0.69; 95% confidence interval, 0.48-0.99; P = 0.046) was associated with prognosis (Cox regression model). CONCLUSIONS: Continuing ET after BM diagnosis was associated with a significantly prolonged OS in this large single-center cohort. No substantial differences between substances were observed. These findings should be validated in a prospective cohort.


Assuntos
Inibidores da Aromatase/uso terapêutico , Neoplasias Encefálicas/mortalidade , Neoplasias da Mama/mortalidade , Carcinoma Ductal de Mama/mortalidade , Carcinoma Lobular/mortalidade , Continuidade da Assistência ao Paciente , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/secundário , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/patologia , Carcinoma Ductal de Mama/tratamento farmacológico , Carcinoma Ductal de Mama/patologia , Carcinoma Lobular/tratamento farmacológico , Carcinoma Lobular/patologia , Feminino , Seguimentos , Humanos , Pessoa de Meia-Idade , Prognóstico , Estudos Retrospectivos , Taxa de Sobrevida , Adulto Jovem
13.
Anal Chem ; 90(22): 13178-13182, 2018 11 20.
Artigo em Inglês | MEDLINE | ID: mdl-30383359

RESUMO

Cancer cells communicate with the whole organism via extracellular vesicles (EVs), which propagate molecular information in support of the malignant phenotype. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) was employed for protein profiling of EVs derived from CCL-228 as the primary colon tumor, the lymph node metastasis CCL-227, and subclones resistant to 5, 25, and 125 µM 5-fluorouracil (FU). EVs were harvested from cell culture supernatant by ultracentrifugation to serve as a model for circulating cancer cell-derived biomarker carriers from body fluids (i.e., liquid biopsy). Protein mass spectra were recorded using standard MALDI matrixes (e.g., CHCA, sinapinic acid) in the range m/ z 2000-20000 on different MALDI-TOF-MS systems and subjected to multivariate data analysis . By using hierarchical clustering, PCA and PLS-DA, discriminatory protein patterns of the EVs from the different cell populations were obtained. Peaks in the range  m/ z 2000-6500 and m/ z 5500-15500 were found to be unique to EVs and the cells, respectively. This clearly demonstrates the differential expression of proteins in EVs as the result of an increasing chemoresistance of their parent cells. The sensitivity of the MALDI-MS based assay was in the low µg/mL (≈1.2-5 × 1010 particles/mL) range. Consequently, our MALDI-MS protein profiling approach shows the potential to serve as novel tool for minimally invasive cancer diagnostics and chemotherapy monitoring in the future, e.g., for early detection of therapy resistance without biopsy.


Assuntos
Biomarcadores Tumorais/análise , Resistencia a Medicamentos Antineoplásicos/fisiologia , Vesículas Extracelulares/química , Proteínas de Neoplasias/análise , Proteômica/métodos , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Humanos , Limite de Detecção , Proteínas de Neoplasias/metabolismo , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos
14.
ESC Heart Fail ; 5(6): 1176-1183, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30230713

RESUMO

AIMS: Cardiac repair has steered clinical attention and remains an unmet need, because available regenerative therapies lack robust mechanistic evidence. Pressure-controlled intermittent coronary sinus occlusion (PICSO), known to induce angiogenetic and vasoactive molecules as well as to reduce regional ischemia, may activate endogenous regenerative processes in failing myocardium. We aimed to investigate the effects of PICSO in patients with advanced heart failure undergoing cardiac resynchronization therapy. METHODS AND RESULTS: Eight out of 32 patients were treated with PICSO, and the remainder served as controls. After electrode testing including left ventricular leads, PICSO was performed for 20 min. To test immediate molecular responses, in both patient groups, coronary venous blood samples were taken at baseline and after 20 min, the time required for the intervention. Sera were tested for microRNAs and growth factors. To test the ability of up-regulated soluble factors on cell proliferation and expression of transcription factors [e.g. Krüppel-like factor 4 (KLF-4)], sera were co-cultured with human cardiomyocytes and fibroblasts. As compared with controls, significant differential expression (differences between pre-values and post-values in relation to both patient cohorts) of microRNA patterns associated with cardiac development was observed with PICSO. Importantly, miR-143 (P < 0.048) and miR-145 (P < 0,047) increased, both targeting a network of transcription factors (including KLF-4) that promote differentiation and repress proliferation of vascular smooth muscle cells. Additionally, an increase of miR-19b (P < 0.019) known to alleviate endothelial cell apoptosis was found, whereas disadvantageous miR-320b (P < 0.023) suspect to impair expression of c-myc, normally provoking cell cycle re-entry in post-mitotic myocytes and miR-25 (P < 0.023), decreased, a target of anti-miR application to improve contractility in the failing heart. Co-cultured post-PICSO sera significantly increased cellular proliferation both in fibroblasts (P < 0.001) and adult cardiomycytes (P < 0.004) sampled from a transplant recipient as compared with controls. Adult cardiomyocytes showed a seven-fold increase of the transcription factor KLF-4 protein when co-cultured with treated sera as compared with controls. CONCLUSIONS: Here, we show for the first time that PICSO, a trans-coronary sinus catheter intervention, is associated with an increase in morphogens secreted into cardiac veins, normally present during cardiac development, and a significant induction of cell proliferation. Present findings support the notion that epigenetic modifications, that is, haemodynamic stimuli on venous vascular cells, may reverse myocardial deterioration. Further investigations are needed to decipher the maze of complex interacting molecular pathways in failing myocardium and the potential role of PICSO to reinitiate developmental processes to prevent further myocardial decay eventually reaching clinical significance.


Assuntos
Oclusão com Balão/métodos , Cateterismo Cardíaco/métodos , Circulação Coronária/fisiologia , Seio Coronário/fisiopatologia , Vasos Coronários/fisiopatologia , Insuficiência Cardíaca/terapia , Idoso , Biomarcadores/sangue , Feminino , Insuficiência Cardíaca/sangue , Insuficiência Cardíaca/fisiopatologia , Humanos , Masculino , Pessoa de Meia-Idade , Pressão
15.
Sci Rep ; 8(1): 124, 2018 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-29317666

RESUMO

There are no  dose-finding trials available for rituximab that could guide dosing in non-malignant diseases. We hypothesized that currently used doses (≥375 mg/m2) exceed several hundred-fold the half-maximal effective dose, which is most sensitive for detecting putative differences between biosimilars and important for dose finding. In an open label, exploratory trial healthy volunteers received single infusions of rituximab at doses of 0.1, 0.3 or 1.0 mg/m2. Subsequently, in a double-blind, randomized trial healthy volunteers received single infusions of two rituximab products at doses of 0.1 and 0.3 mg/m2. In the exploratory trial rituximab transiently depleted CD20+ cells by a mean 68% (range: 57-95%), 74% (55-82%) and 97% (94-100%) immediately after the infusion of 0.1 (n = 4), 0.3 (n = 4) and 1 mg/m2 (n = 8), respectively. In the randomized trial CD20+ cells decreased by a mean 48% (25-84%) - 55% (26-85%) and 81 (67-89%) - 87% (77-96%) after infusion of 0.1 mg/m2 (n = 12) or 0.3 mg/m2 (n = 8 proposed biosimilar, n = 4 reference product) of the proposed biosimilar or the reference product, respectively. It is important to understand that in healthy volunteers <1% of the authorized rituximab doses depletes almost all circulating B lymphocytes. Thus, for non-malignant diseases alternative, more cost-effective dosing regimens seem plausible, but require clinical testing. (EudraCT-No. 2010-023781-45; EudraCT-No. 2013-001077-24).


Assuntos
Antineoplásicos Imunológicos/administração & dosagem , Medicamentos Biossimilares/administração & dosagem , Voluntários Saudáveis , Rituximab/administração & dosagem , Adulto , Antineoplásicos Imunológicos/efeitos adversos , Antineoplásicos Imunológicos/farmacocinética , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Linfócitos B/metabolismo , Medicamentos Biossimilares/efeitos adversos , Medicamentos Biossimilares/farmacocinética , Monitoramento de Medicamentos , Feminino , Humanos , Contagem de Linfócitos , Masculino , Pessoa de Meia-Idade , Projetos Piloto , Projetos de Pesquisa , Rituximab/efeitos adversos , Rituximab/farmacocinética , Fatores de Tempo , Adulto Jovem
16.
Oncotarget ; 8(41): 71206-71222, 2017 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-29050357

RESUMO

Soon after microRNAs entered the stage as novel regulators of gene expression, they were found to regulate -and to be regulated by- the development, progression and aggressiveness of virtually all human types of cancer. Therefore, miRNAs in general harbor a huge potential as diagnostic and prognostic markers as well as potential therapeutic targets in cancer. The miR-17-92 cluster was found to be overexpressed in many human cancers and to promote unrestrained cell growth, and has therefore been termed onco-miR-1. In addition, its expression is often dysregulated in many other diseases. MiR-17-5p, its most prominent member, is an essential regulator of fundamental cellular processes like proliferation, autophagy and apoptosis, and its deficiency is neonatally lethal in the mouse. Many cancer types are associated with elevated miR-17-5p expression, and the degree of overexpression might correlate with cancer aggressiveness and responsiveness to chemotherapeutics - suggesting miR-17-5p to be an alarm signal. Liver, gastric or colorectal cancers are examples where miR-17-5p has been observed exclusively as an oncogene, while, in other cancer types, like breast, prostate and lung cancer, the role of miR-17-5p is not as clear-cut, and it might also act as tumor-suppressor. However, in all cancer types studied so far, miR-17-5p has been found at elevated levels in the circulation. In this review, we therefore recapitulate the current state of knowledge about miR-17-5p in the context of cancer, and suggest that elevated miR-17-5p levels in the plasma might be a sensitive and early alarm signal for cancer ('alarmiR'), albeit not a specific alarm for a specific type of tumor.

17.
Food Chem Toxicol ; 108(Pt A): 53-62, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28716444

RESUMO

Cellular growth inhibition exerted by thiosemicarbazones is mainly attributed to down-regulation of ribonucleotide reductase (RNR) activity, with RNR being responsible for the rate-limiting step of de novo DNA synthesis. In this study, we investigated the antineoplastic effects of three newly synthesized thiosemicarbazone derivatives, thiazolyl hydrazones, in human HL-60 promyelocytic leukemia cells. The cytotoxicity of compounds alone and in combination with arabinofuranosylcytosine (AraC) was determined by growth inhibition assays. Effects on deoxyribonucleoside triphosphate (dNTP) concentrations were quantified by HPLC, and the incorporation of radio-labeled 14C-cytidine into nascent DNA was measured using a beta counter. Cell cycle distribution was analyzed by FACS, and protein levels of RNR subunits and checkpoint kinases were evaluated by Western blotting. VG12, VG19, and VG22 dose-dependently decreased intracellular dNTP concentrations, impaired cell cycle progression and, consequently, inhibited the growth of HL-60 cells. VG19 also lowered the protein levels of RNR subunits R1 and R2 and significantly diminished the incorporation of radio-labeled 14C-cytidine, being equivalent to an inhibition of DNA synthesis. Combination of thiazolyl hydrazones with AraC synergistically potentiated the antiproliferative effects seen with each drug alone and might therefore improve conventional chemotherapeutic regimens for the treatment of human malignancies such as acute promyelocytic or chronic myelogenous leukemia.


Assuntos
Citarabina/farmacocinética , Regulação para Baixo/efeitos dos fármacos , Hidrazonas/farmacocinética , Ribonucleotídeo Redutases/antagonistas & inibidores , Tiossemicarbazonas/farmacocinética , Ciclo Celular/efeitos dos fármacos , Citarabina/administração & dosagem , Citarabina/farmacologia , Sinergismo Farmacológico , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Células HL-60 , Humanos , Hidrazonas/administração & dosagem , Hidrazonas/química , Hidrazonas/farmacologia , Estrutura Molecular , Ribonucleotídeo Redutases/genética , Ribonucleotídeo Redutases/metabolismo , Tiossemicarbazonas/administração & dosagem , Tiossemicarbazonas/química , Tiossemicarbazonas/farmacologia
18.
Oncotarget ; 8(17): 29519-29534, 2017 Apr 25.
Artigo em Inglês | MEDLINE | ID: mdl-28392501

RESUMO

In a continuous and mutual exchange of information, cancer cells are invariably exposed to microenvironment transformation. This continuous alteration of the genetic, molecular and cellular peritumoral stroma background has become as critical as the management of primary tumor progression events in cancer cells. The communication between stroma and tumor cells within the extracellular matrix is one of the triggers in colon and liver carcinogenesis. All non- codingRNAs including long non-coding RNAs, microRNAs and ultraconserved genes play a critical role in almost all cancers and are responsible for the modulation of the tumor microenvironment in several malignant processes such as initiation, progression and dissemination. This review details the involvement of non codingRNAs in the evolution of human colorectal carcinoma and hepatocellular carcinoma in relationship with the microenvironment. Recent research has shown that a considerable number of dysregulated non- codingRNAs could be valuable diagnostic and prognostic biomarkers in cancer. Therefore, more in-depth knowledge of the role non- codingRNAs play in stroma-tumor communication and of the complex regulatory mechanisms between ultraconserved genes and microRNAs supports the validation of future effective therapeutic targets in patients suffering from hepatocellular and colorectal carcinoma, two distinctive entities which share quite a lot common non-coding RNAs.


Assuntos
Carcinoma Hepatocelular/genética , Neoplasias Colorretais/genética , Neoplasias Hepáticas/genética , RNA Longo não Codificante/metabolismo , Carcinoma Hepatocelular/patologia , Neoplasias Colorretais/patologia , Progressão da Doença , Humanos , Neoplasias Hepáticas/patologia , Microambiente Tumoral
19.
Cell Mol Life Sci ; 74(10): 1907-1921, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28013338

RESUMO

Retraction of mesenchymal stromal cells supports the invasion of colorectal cancer cells (CRC) into the adjacent compartment. CRC-secreted 12(S)-HETE enhances the retraction of cancer-associated fibroblasts (CAFs) and therefore, 12(S)-HETE may enforce invasivity of CRC. Understanding the mechanisms of metastatic CRC is crucial for successful intervention. Therefore, we studied pro-invasive contributions of stromal cells in physiologically relevant three-dimensional in vitro assays consisting of CRC spheroids, CAFs, extracellular matrix and endothelial cells, as well as in reductionist models. In order to elucidate how CAFs support CRC invasion, tumour spheroid-induced CAF retraction and free intracellular Ca2+ levels were measured and pharmacological- or siRNA-based inhibition of selected signalling cascades was performed. CRC spheroids caused the retraction of CAFs, generating entry gates in the adjacent surrogate stroma. The responsible trigger factor 12(S)-HETE provoked a signal, which was transduced by PLC, IP3, free intracellular Ca2+, Ca2+-calmodulin-kinase-II, RHO/ROCK and MYLK which led to the activation of myosin light chain 2, and subsequent CAF mobility. RHO activity was observed downstream as well as upstream of Ca2+ release. Thus, Ca2+ signalling served as central signal amplifier. Treatment with the FDA-approved drugs carbamazepine, cinnarizine, nifedipine and bepridil HCl, which reportedly interfere with cellular calcium availability, inhibited CAF-retraction. The elucidation of signalling pathways and identification of approved inhibitory drugs warrant development of intervention strategies targeting tumour-stroma interaction.


Assuntos
Ácido 12-Hidroxi-5,8,10,14-Eicosatetraenoico/metabolismo , Fibroblastos Associados a Câncer/patologia , Colo/patologia , Neoplasias Colorretais/patologia , Reto/patologia , Transdução de Sinais , Cálcio/metabolismo , Fibroblastos Associados a Câncer/metabolismo , Miosinas Cardíacas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Colo/metabolismo , Neoplasias Colorretais/metabolismo , Humanos , Cadeias Leves de Miosina/metabolismo , Invasividade Neoplásica/patologia , Reto/metabolismo , Quinases Associadas a rho/metabolismo
20.
Biomol Concepts ; 7(5-6): 311-319, 2016 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-27837593

RESUMO

In the last decade, microRNAs (miRs or miRNAs) became of great interest in cancer research due to their multifunctional and active regulation in a variety of vital cellular processes. In this review, we discuss the miR-200 family, which is composed of five members (miR-141, miR-200a/200b/200c and miR-429). Although being among the best investigated miRNAs in the field, there are still many open issues. Here, we describe the potential role of miR-200 as prognostic and/or predictive biomarker, its influence on motility and cell migration as well as its role in epithelial to mesenchymal transition (EMT) and metastasis formation in different tumour types. Recent studies also demonstrated the influence of miR-200 on drug resistance and described a correlation between miR-200 expression levels and overall survival of patients. Despite intense research in this field, the full role of the miR-200 family in cancer progression and metastasis is not completely understood and seems to differ between different tumour types and different cellular backgrounds. To elucidate these differences further, a finer characterisation of the role of the individual miRNA-200 family members is currently under investigation.


Assuntos
MicroRNAs/genética , Família Multigênica , Progressão da Doença , Resistencia a Medicamentos Antineoplásicos/genética , Transição Epitelial-Mesenquimal/genética , Fator de Iniciação 3 em Eucariotos , Regulação Neoplásica da Expressão Gênica , Humanos , Metástase Neoplásica , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/mortalidade , Neoplasias/patologia , Prognóstico , Interferência de RNA
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