MicroRNA signature refine response prediction in CML

microRNAs (miRs) dysregulation have emerged as a crucial step in tumorigenesis, being related with cancer development, progression and response to treatment. In chronic myeloid leukaemia (CML), the resistance to tyrosine kinase inhibitors (TKI) is responsible for treatment failure and could be linked to changes in miRs expression. This work aimed to correlate the expression levels of 3 miRs, miR-21, miR-26b and miR-451, with response to TKI treatment in CML patients. miR-451 levels at diagnosis were significantly higher in patients with optimal response after 6 and 12 months of therapy. Conversely, patients without optimal response had highest levels of miR-21. miR-21 and miR-451 appear to be good biomarkers of response, able to predict optimal TKI responders (p < 0.05). Using the combined profile of both miRs, we create a predictive model of optimal response after one year of treatment. This study highlights the role of miR-21 and miR-451 expression levels at diagnosis in predicting which patients achieve the optimal response.info:eu-repo/semantics/publishedVersio

Genetic variants of ABC and SLC transporter genes and chronic myeloid leukaemia: impact on susceptibility and prognosis

Solute carrier (SLC) and ATP-binding cassette (ABC) transporters comprise a variety of proteins expressed on cell membranes responsible for intrusion or extrusion of substrates, respectively, including nutrients, xenobiotics, and chemotherapeutic agents. These transporters mediate the cellular disposition of tyrosine kinase inhibitors (TKIs), and their genetic variants could affect its function, potentially predisposing patients to chronic myeloid leukaemia (CML) and modulating treatment response. We explored the impact of genetic variability (single nucleotide variants—SNVs) of drug transporter genes (ABCB1, ABCG2, SLC22A1, and SLC22A5) on CML susceptibility, drug response, and BCR-ABL1 mutation status. We genotyped 10 SNVs by tetra-primers-AMRS-PCR in 198 CML patients and 404 controls, and assessed their role in CML susceptibility and prognosis. We identified five SNVs associated with CML predisposition, with some variants increasing disease risk, including TT genotype ABCB1 (rs1045642), and others showing a protective effect (GG genotype SLC22A5 rs274558). We also observed different haplotypes and genotypic profiles associated with CML predisposition. Relating to drug response impact, we found that CML patients with the CC genotype (rs2231142 ABCG2) had an increased risk of TKI resistance (six-fold). Additionally, CML patients carrying the CG genotype (rs683369 SLC22A1) presented a 4.54-fold higher risk of BCR-ABL1 mutations. Our results suggest that drug transporters’ SNVs might be involved in CML susceptibility and TKI response, and predict the risk of BCR-ABL1 mutations, highlighting the impact that SNVs could have in therapeutic selection.info:eu-repo/semantics/publishedVersio

The Hydropathy Index of the HCDR3 Region of the B-Cell Receptor Identifies Two Subgroups of IGHV-Mutated Chronic Lymphocytic Leukemia Patients With Distinct Outcome

© 2021 Rodríguez-Caballero, Fuentes Herrero, Oliva Ariza, Criado, Alcoceba, Prieto, Pérez Caro, García-Montero, González Díaz, Forconi, Sarmento-Ribeiro, Almeida and Orfao.The HCDR3 sequences of the B-cell receptor (BCR) undergo constraints in length, amino acid use, and charge during maturation of B-cell precursors and after antigen encounter, leading to BCR and antibodies with high affinity to specific antigens. Chronic lymphocytic leukemia consists of an expansion of B-cells with a mixed immature and “antigen-experienced” phenotype, with either a mutated (M-CLL) or unmutated (U-CLL) tumor BCR, associated with distinct patient outcomes. Here, we investigated the hydropathy index of the BCR of 138 CLL patients and its association with the IGHV mutational status and patient outcome. Overall, two clearly distinct subgroups of M-CLL patients emerged, based on a neutral (mean hydropathy index of -0.1) vs. negatively charged BCR (mean hydropathy index of -1.1) with molecular features closer to those of B-cell precursors and peripheral/mature B-cells, respectively. Despite that M-CLL with neutral HCDR3 did not show traits associated with a mature B-cell repertoire, important differences in IGHV gene usage of tumor cells and patient outcome were observed in this subgroup of patients once compared to both U-CLL and M-CLL with negatively charged HCDR3 sequences. Compared to M-CLL with negatively charged HCDR3 sequences, M-CLL with neutral HCDR3 sequences showed predominance of men, more advanced stages of the disease, and a greater frequency of genetic alterations—e.g., del(17p)—together with a higher rate of disease progression and shorter time to therapy (TTT), independently of other prognostic factors. Our data suggest that the hydropathy index of the HCDR3 sequences of CLL cells allows the identification of a subgroup of M-CLL with intermediate prognostic features between U-CLL and the more favorable subgroup of M-CLL with a negatively charged BCR.This work was supported by the following grants: FS/37-2017, from the Fundación Memoria D. Samuel Solórzano, Universidad de Salamanca; FIS PI17/00399-FEDER, from the Fondo de Investigación Sanitaria of Instituto de Salud Carlos III, Madrid, Spain; 0639_IDIAL_NET_3_E, from cooperative network EPINTERREG V A España Portugal (POCTEP); and ECRIN-M3, Accelerator Award Full, Cancer Research UK, Fundación Cientıfíca de la Asociación Española Contra el Cáncer (AECC), Fondazione AIRC per la Ricerca sul Cancro.

Chemical characterization and cytotoxic potential of an ellagitannin-enriched fraction from Fragaria vesca leaves

The hepatocellular carcinoma, a primary malignancy of the liver, has a very poor prognosis and a lower survival rate. Moreover, the inefficacy of conventional therapies emphasizes the importance of discovering new bioactive compounds. Several studies clearly state that plant-derived polyphenols, namely ellagitannins, have several health benefits. Fragaria vesca leaves contain high amounts of polyphenols, being especially rich in ellagitannins. Therefore, this study aimed to characterize an ellagitannin-enriched fraction (EEF) from F. vesca leaves and to unveil the anticancer potential of this fraction on human hepatocellular carcinoma cells. The analysis of EEF by HPLC-PDA-ESI/MSn allowed the detection of 12 ellagitannins. The cell viability of both EEF and crude extract was determined after 24 h of cells treatment and the halfmaximal inhibitory concentration (IC50) was evaluated. The IC50 of the EEF (113 lg/mL) was about 6 times lower than the IC50 of the crude extract (690 lg/mL). Furthermore, EEF induced cell cycle arrest at G2/M checkpoint and decreased cell proliferation in a dose-dependent way. This fraction also induced an accumulation of LC3-II protein through blockage of autophagic flux, and inhibited chymotrypsin-like activity of 26S proteasome. These results showed, for the first time, that EEF from F. vesca leaves inhibits both, autophagic and ubiquitin-proteasome system pathways, two main intracellular protein degradation systems that are targets for anticancer therapies. Additionally, a proteomic analysis allowed the identification of 914 proteins, among which 133 were modulated after cells treatment with EEF, most of them related to metabolic pathways. Overall, this study shows that the EEF from F. vesca leaves decreased cell proliferation, inhibited the proteolytic mechanisms and modulated the metabolic pathways of the cell. Additionally this study points out F. vesca as a source of valuable molecules with anticancer potential, suggesting that ellagitannins, the polyphenols identified in this fraction, could be useful in the development of new fine-tuned therapeutic strategies against carcinogenesis

Alterações fenotípicas da célula tumoral e a sua relação com a resistência à quimioterapia.

Um dos problemas da oncologia médica é o desenvolvimento de resistência natural ou adquirida, das células neoplásicas aos fármacos anticancerígenos. Os mecanismos envolvidos na sindrome de resistência a múltiplos fármacos (MDR) estão ainda mal esclarecidos. O objectivo deste trabalho é avaliar a contribuição da persistência do stresse oxidativo, da mitocôndria, das alterações membranares e da expressão das proteinas envolvidas na regulação da morte celular por apoptose, na patogenia, e no desenvolvimento de recidiva e de resistência à quimioterapia na leucemia. Observámos uma diminuição das defesas antioxidantes, particularmente das defesas de natureza enzimática, superóxido dismutase e peroxidase do glutatião, nos doentes com leucemia aguda, e das defesas de natureza não enzimática, glutatião reduzido e vitamina E, nos doentes com leucemia crónica. Encontrámos também um aumento da produção de peróxidos intracelulares, diminuição da actividade da cadeia respiratória mitocondrial, nomeadamente do complexo IV, diminuição da desidrogenase do lactato e ainda um aumento da expressão de Bcl-2 e diminuição da expressão de Bax e de Fas. Estes dados poderão relacionar-se com a elevada capacidade proliferativa das células, na leucemia linfoblástica aguda, e/ou com a resistência à morte celular, na leucemia linfocítica crónica. Por outro lado, os doentes com leucemia linfoblástica aguda em recidiva apresentam aumento da expressão de glicoproteína P de superfície, da razão fosfolípido/proteína e fosfolípido/colesterol, e do conteúdo em triacilglicerídeos. Os doentes com leucemia linfoblástica aguda, de sobrevida mais curta e recidiva mais precoce, são os que apresentam níveis de expressão de Bcl-2 mais elevados e/ou de Bax mais baixos, ou níveis mais elevados de GSH, o que poderá contribuir para essa recidiva e, provavelmente, para a resistência à quimioterapia. Verificou-se também que a citotoxicidade induzida por fármacos anticancerígenos em células de leucemia humana em cultura é mediada por morte celular apoptótica, para a qual pode contribuir o aumento da expressão de Bax e de Fas, bem como o aumento da produção de peróxidos, e a alteração da actividade da cadeia respiratória mitocondrial. Nas células resistentes aos fármacos anticancerígenos, observámos, além do aumento da expressão de glicoproteina P, de Bcl-2 e da razão Bcl-2/Bax, uma diminuição da expressão de Bax e de Fas e um aumento dos níveis de glutatião reduzido, o que pode contribuir para a resistência à quimioterapia, podendo a disfunção mitocondrial ter um papel determinante nos mecanismos de resistência. Um conhecimento mais aprofundado sobre a participação do stresse oxidativo, do envolvimento da mitocôndria e das proteínas reguladoras da apoptose nos mecanismos de resistência à morte celular, pode permitir uma avaliação prognóstica e melhor abordagem terapêutica do doente com leucemia

Apoptosis and (in) Pain&mdash;Potential Clinical Implications

The deregulation of apoptosis is involved in the development of several pathologies, and recent evidence suggests that apoptosis may be involved in chronic pain, namely in neuropathic pain. Neuropathic pain is a chronic pain state caused by primary damage or dysfunction of the nervous system; however, the details of the molecular mechanisms have not yet been fully elucidated. Recently, it was found that nerve endings contain transient receptor potential (TRP) channels that sense and detect signals released by injured tissues and respond to these damage signals. TRP channels are similar to the voltage-gated potassium channels or nucleotide-gated channels that participate in calcium and magnesium homeostasis. TRP channels allowing calcium to penetrate into nerve terminals can activate apoptosis, leading to nerve terminal destruction. Further, some TRPs are activated by acid and reactive oxygen species (ROS). ROS are mainly produced in the mitochondrial respiratory chain, and an increase in ROS production and/or a decrease in the antioxidant network may induce oxidative stress (OS). Depending on the OS levels, they can promote cellular proliferation and/or cell degeneration or death. Previous studies have indicated that proinflammatory cytokines, such as tumor necrosis factor-&alpha; (TNF-&alpha;), play an important role in the peripheral mediation of neuropathic pain. This article aims to perform a review of the involvement of apoptosis in pain, particularly the role of OS and neuroinflammation, and the clinical relevance of this knowledge. The potential discovery of new biomarkers and therapeutic targets can result in the development of more effective and targeted drugs to treat chronic pain, namely neuropathic pain. Highlights: Oxidative stress and neuroinflammation can activate cell signaling pathways that can lead to nerve terminal destruction by apoptosis. These could constitute potential new pain biomarkers and targets for therapy in neuropathic pain

Combination of elacridar with imatinib modulates resistance associated with drug efflux transporters in chronic myeloid leukemia

Multidrug resistance (MDR) development has emerged as a complication that compromises the success of several chemotherapeutic agents. In chronic myeloid leukemia (CML), imatinib resistance has been associated with changes in BCR-ABL1 and intracellular drug concentration, controlled by SLC and ABC transporters. We evaluate the therapeutic potential of a P-glycoprotein and BCRP inhibitor, elacridar, in sensitive (K562 and LAMA-84) and imatinib-resistant (K562-RC and K562-RD) CML cell lines as monotherapy and combined with imatinib. Cell viability was analyzed by resazurin assay. Drug transporter activity, cell death, cell proliferation rate, and cell cycle distribution were analyzed by flow cytometry. Both resistant models presented an increased activity of BCRP and P-gP compared to K562 cells. Elacridar as monotherapy did not reach IC50 in any CML models but activated apoptosis without cytostatic effect. Nevertheless, the association of elacridar (250 nM) with imatinib overcomes resistance, re-sensitizing K562-RC and K562-RD cells with five and ten times lower imatinib concentrations, respectively. Drug combination induced apoptosis with increased cleaved-caspases-3, cleaved-PARP and DNA damage, reduced cell proliferation rate, and arrested CML cells in the S phase. These data suggest that elacridar combined with imatinib might represent a new therapeutic option for overcoming TKI resistance involving efflux transporters. View Full-Textinfo:eu-repo/semantics/publishedVersio

Modelos experimentais em oncologia: O contributo da cultura de células para o conhecimento da biologia do cancro Experimental models in oncology: Contribution of cell culture on understanding the biology of cancer

A cultura de tecidos surgiu no século xx (Harrison, 1907) para estudar o comportamento das células animais em ambiente homeostático e em situações de stress. A capacidade de estudar as células a nível molecular relaciona-se com a forma como as células podem crescer e ser manipuladas em laboratório. A cultura de células in vitro permitiu estudar crescimento, diferenciação e morte celular e efectuar manipulações genéticas necessárias ao perfeito conhecimento da estrutura e funções dos genes. A cultura de células estaminais humanas veio colmatar algumas limitações inerentes aos restantes modelos de cultura. Ao que parece, as células estaminais cancerígenas mantêm-se quiescentes nos locais metastáticos até serem activadas por sinais apropriados do microambiente. Vários estudos revelaram que diferentes tipos de cancros podem surgir da transformação maligna de células estaminais. A eliminação destas células progenitoras tumorais é essencial para o desenvolvimento de novas abordagens terapêuticas mais eficazes em cancros agressivos. Por outro lado, a utilização de células dendríticas modificadas em cultura poderá contribuir para a produção de uma potencial vacina terapêutica eficaz para obter a regressão tumoral.In the beginning of the 20th century, tissue culture was started with the aim of studying the behaviour of animal cells in normal and stress conditions. The cell study at molecular level depends on their capacity of growing and how they can be manipulated in laboratory. In vitro cell culture allows us the possibility of studying biological key processes, such as growth, differentiation and cell death, and also to do genetic manipulations essential to the knowledge of structure and genes function. Human stem cells culture provides strategies to circumvent other models’ deficiencies. It seems that cancer stem cells remain quiescent until activation by appropriated micro-environmental stimulation. Several studies reveal that different cancer types could be due to stem cell malignant transformations. Removal of these cells is essential to the development of more effective cancer therapies for advanced disease. On the other hand, dendritic cells modified in culture may be used as a therapeutic vaccine in order to induce tumour withraw