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Oligomerization of antibody fragments via modification with polyethylene glycol (pegylation) may alter their function and properties, leading to a multivalent interaction of the resulting constructs with the target antigen. In a recent study, we generated pegylated monomers and multimers of scFv fragments of GD2-specific antibodies using maleimide-thiol chemistry. Multimerization enhanced the antigen-binding properties and demonstrated a more efficient tumor uptake in a syngeneic GD2-positive mouse cancer model compared to monomeric antibody fragments, thereby providing a rationale for improving the therapeutic characteristics of GD2-specific antibody fragments. In this work, we obtained pegylated conjugates of scFv fragments of GD2-specific antibodies with maytansinoids DM1 or DM4 using tetravalent PEG-maleimide (PEG4). The protein products from the two-stage thiol-maleimide reaction resolved by gel electrophoresis indicated that pegylated scFv fragments constituted the predominant part of the protein bands, and most of the scFv formed pegylated monomers and dimers. The conjugates retained the ability to bind ganglioside GD2 comparable to that of the parental scFv fragment and to specifically interact with GD2-positive cells. Both induced significant inhibitory effects in the GD2-positive B78-D14 cell line, in contrast to the GD2-negative B16 cell line. The decrease in the B78-D14 cell viability when treated with scFv-PEG4-DM4 was more prominent than that for scFv-PEG4-DM1, and was characterized by a twofold lower half-maximal inhibitory concentration (IC50). Unlike the parental scFv fragment, the product of scFv and PEG4 conjugation (scFv-PEG4), consisting predominantly of pegylated scFv multimers and monomers, induced direct cell death in the GD2-positive B78-D14 cells. However, the potency of scFv-PEG4 was low in the selected concentration range, thus demonstrating that the cytotoxic effect of DM1 and DM4 within the antibody fragment-drug conjugates was primary. The suggested approach may contribute to development of novel configurations of antibody fragment-drug conjugates for cancer treatment.
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Liver diseases, characterized by high morbidity and mortality, represent a substantial medical problem globally. The current therapeutic approaches are mainly aimed at reducing symptoms and slowing down the progression of the diseases. Organ transplantation remains the only effective treatment method in cases of severe liver pathology. In this regard, the development of new effective approaches aimed at stimulating liver regeneration, both by activation of the organ's own resources or by different therapeutic agents that trigger regeneration, does not cease to be relevant. To date, many systematic reviews and meta-analyses have been published confirming the effectiveness of mesenchymal stromal cell (MSC) transplantation in the treatment of liver diseases of various severities and etiologies. However, despite the successful use of MSCs in clinical practice and the promising therapeutic results in animal models of liver diseases, the mechanisms of their protective and regenerative action remain poorly understood. Specifically, data about the molecular agents produced by these cells and mediating their therapeutic action are fragmentary and often contradictory. Since MSCs or MSC-like cells are found in all tissues and organs, it is likely that many key intercellular interactions within the tissue niches are dependent on MSCs. In this context, it is essential to understand the mechanisms underlying communication between MSCs and differentiated parenchymal cells of each particular tissue. This is important both from the perspective of basic science and for the development of therapeutic approaches involving the modulation of the activity of resident MSCs. With regard to the liver, the research is concentrated on the intercommunication between MSCs and hepatocytes under normal conditions and during the development of the pathological process. The goals of this review were to identify the key factors mediating the crosstalk between MSCs and hepatocytes and determine the possible mechanisms of interaction of the two cell types under normal and stressful conditions. The analysis of the hepatocyte-MSC interaction showed that MSCs carry out chaperone-like functions, including the synthesis of the supportive extracellular matrix proteins; prevention of apoptosis, pyroptosis, and ferroptosis; support of regeneration; elimination of lipotoxicity and ER stress; promotion of antioxidant effects; and donation of mitochondria. The underlying mechanisms suggest very close interdependence, including even direct cytoplasm and organelle exchange.
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Hepatopatias , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais , Animais , Hepatócitos/metabolismo , Hepatopatias/metabolismo , Células-Tronco Mesenquimais/metabolismo , HomeostaseRESUMO
Ganglioside GD2 is a well-established target expressed on multiple solid tumors, many of which are characterized by low treatment efficiency. Antibody-drug conjugates (ADCs) have demonstrated marked success in a number of solid tumors, and GD2-directed drug conjugates may also hold strong therapeutic potential. In a recent study, we showed that ADCs based on the approved antibody dinutuximab and the drugs monomethyl auristatin E (MMAE) or F (MMAF) manifested potent and selective cytotoxicity in a panel of tumor cell lines and strongly inhibited solid tumor growth in GD2-positive mouse cancer models. Here, we employed two different GD2-binding moieties-minibodies and scFv fragments that carry variable antibody domains identical to those of dinutuximab, and site-directly conjugated them to MMAE or MMAF by thiol-maleimide chemistry with drug-to-antibody ratios (DAR) of 2 and 1, respectively. Specific binding of the antibody fragment-drug conjugates (FDCs) to GD2 was confirmed in direct ELISA, flow cytometry, and confocal microscopy. Selective cytotoxic and cytostatic effects of the conjugates were observed in GD2-positive but not GD2-negative neuroblastoma and melanoma cell lines. Minibody-based FDCs demonstrated more pronounced cytotoxic effects and stronger antigen binding compared to scFv-based FDCs. The developed molecules may offer considerable practical benefit, since antibody fragment-drug conjugates are capable of enhancing therapeutic efficacy of ADCs by improving their pharmacokinetic characteristics and reducing side effects.
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Antineoplásicos , Imunoconjugados , Neuroblastoma , Animais , Camundongos , Fragmentos de Imunoglobulinas , Linhagem Celular Tumoral , Antineoplásicos/farmacologia , Imunoconjugados/uso terapêutico , Neuroblastoma/patologia , Modelos Animais de Doenças , Gangliosídeos/metabolismoRESUMO
Over the past two decades, mesenchymal stem cells (MSCs) have shown promising therapeutic effects both in preclinical studies (in animal models of a wide range of diseases) and in clinical trials. However, the efficacy of MSC-based therapy is not always predictable. Moreover, despite the large number of studies, the mechanisms underlying the regenerative potential of MSCs are not fully elucidated. Recently, it has been reliably established that transplanted MSCs can undergo rapid apoptosis and clearance from the recipient's body, still exhibiting therapeutic effects, especially those associated with their immunosuppressive/immunomodulating properties. The mechanisms underlying these effects can be mediated by the efferocytosis of apoptotic MSCs by host phagocytic cells. In this concise review, we briefly describe three types of MSC-generated extracellular vesicles, through which their therapeutic functions can potentially be carried out; we focused on reviewing recent data on apoptotic MSCs and MSC-derived apoptotic bodies (MSC-ApoBDs), their functions, and the mechanisms of their therapeutic effects.
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Mesenchymal stem cells (MSCs) have a pronounced therapeutic potential in various pathological conditions. Though therapeutic effects of MSC transplantation have been studied for a long time, the underlying mechanisms are still not clear. It has been shown that transplanted MSCs are rapidly eliminated, presumably by apoptosis. As the mechanisms of MSC apoptosis are not fully understood, in the present work we analyzed MSC sensitivity to Fas-induced apoptosis using MSCs isolated from the biopsies of liver fibrosis patients (L-MSCs). The level of cell death was analyzed by flow cytometry in the propidium iodide test. The luminescent ATP assay was used to measure cellular ATP levels; and the mitochondrial membrane potential was assessed using the potential-dependent dye JC-1. We found that human L-MSCs were resistant to Fas-induced cell death over a wide range of FasL and anti-Fas mAb concentrations. At the same time, intrinsic death signal inducers CoCl2 and staurosporine caused apoptosis of L-MSCs in a dose-dependent manner. Despite the absence of Fas-induced cell death treatment of L-MSCs with low concentrations of FasL or anti-Fas mAb resulted in a cellular ATP level decrease, while high concentrations of the inducers caused a decline of the mitochondrial membrane potential. Pre-incubation of L-MSCs with the pro-inflammatory cytokine TNF-α did not promote L-MSC cell death. Our data indicate that human L-MSCs have increased resistance to receptor-mediated cell death even under inflammatory conditions.
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Glioma is the most common type of primary CNS tumor, composed of cells that resemble normal glial cells. Recent genetic studies have provided insight into the inter-tumoral heterogeneity of gliomas, resulting in the updated 2021 WHO classification of gliomas. Thorough understanding of inter-tumoral heterogeneity has already improved the prognosis and treatment outcomes of some types of gliomas. Currently, the challenge for researchers is to study the intratumoral cell heterogeneity of newly defined glioma subtypes. Cancer stem cells (CSCs) present in gliomas and many other tumors are an example of intratumoral heterogeneity of great importance. In this review, we discuss the modern concept of glioma stem cells and recent single-cell sequencing-driven progress in the research of intratumoral glioma cell heterogeneity. The particular emphasis was placed on the recently revealed variations of the cell composition of the subtypes of the adult-type diffuse gliomas, including astrocytoma, oligodendroglioma and glioblastoma. The novel data explain the inconsistencies in earlier glioma stem cell research and also provide insight into the development of more effective targeted therapy and the cell-based immunotherapy of gliomas. Separate sections are devoted to the description of single-cell sequencing approach and its role in the development of cell-based immunotherapies for glioma.
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Astrocitoma , Glioblastoma , Glioma , Oligodendroglioma , Humanos , Glioma/genética , Glioma/terapia , Glioma/patologia , Oligodendroglioma/patologia , Glioblastoma/patologia , Astrocitoma/patologia , Células-Tronco Neoplásicas/patologiaRESUMO
CD133 is an extensively studied marker of the most malignant tumor cell population, designated as cancer stem cells (CSCs). However, the function of this glycoprotein and its involvement in cell regulatory cascades are still poorly understood. Here we show a positive correlation between the level of CD133 plasma membrane expression and the proliferative activity of cells of the Caco-2, HT-29, and HUH7 cancer cell lines. Despite a substantial difference in the proliferative activities of cell populations with different levels of CD133 expression, transcriptomic and proteomic profiling revealed only minor distinctions between them. Nonetheless, a further in silico assessment of the differentially expressed transcripts and proteins revealed 16 proteins that could be involved in the regulation of CD133 expression; these were assigned ranks reflecting the apparent extent of their involvement. Among them, the TRIM28 transcription factor had the highest rank. The prominent role of TRIM28 in CD133 expression modulation was confirmed experimentally in the Caco2 cell line clones: the knockout, though not the knockdown, of the TRIM28 gene downregulated CD133. These results for the first time highlight an important role of the TRIM28 transcription factor in the regulation of CD133-associated cancer cell heterogeneity.
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Antígeno AC133/genética , Células-Tronco Neoplásicas/citologia , Proteína 28 com Motivo Tripartido/metabolismo , Antígeno AC133/metabolismo , Células CACO-2 , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Fenótipo , Proteômica , Fatores de Transcrição/metabolismoRESUMO
Antigen-binding fragments of antibodies specific to the tumor-associated ganglioside GD2 are well poised to play a substantial role in modern GD2-targeted cancer therapies, however, rapid elimination from the body and reduced affinity compared to full-length antibodies limit their therapeutic potential. In this study, scFv fragments of GD2-specific antibodies 14.18 were produced in a mammalian expression system that specifically bind to ganglioside GD2, followed by site-directed pegylation to generate mono-, di-, and tetra-scFv fragments. Fractionated pegylated dimers and tetramers of scFv fragments showed significant increase of the binding to GD2 which was not accompanied by cross-reactivity with other gangliosides. Pegylated multimeric di-scFvs and tetra-scFvs exhibited cytotoxic effects in GD2-positive tumor cells, while their circulation time in blood significantly increased compared with monomeric antibody fragments. We also demonstrated a more efficient tumor uptake of the multimers in a syngeneic GD2-positive mouse cancer model. The findings of this study provide the rationale for improving therapeutic characteristics of GD2-specific antibody fragments by multimerization and propose a strategy to generate such molecules. On the basis of multimeric antibody fragments, bispecific antibodies and conjugates with cytotoxic drugs or radioactive isotopes may be developed that will possess improved pharmacokinetic and pharmacodynamic properties.
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Antineoplásicos Imunológicos , Gangliosídeos/antagonistas & inibidores , Neoplasias Experimentais , Polietilenoglicóis/química , Anticorpos de Cadeia Única , Animais , Antineoplásicos Imunológicos/química , Antineoplásicos Imunológicos/farmacocinética , Antineoplásicos Imunológicos/farmacologia , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Neoplasias Experimentais/sangue , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/patologia , Anticorpos de Cadeia Única/química , Anticorpos de Cadeia Única/farmacocinética , Anticorpos de Cadeia Única/farmacologiaRESUMO
BACKGROUND: Ganglioside GD2 is expressed on plasma membranes of various types of malignant cells. One of the most promising approaches for cancer immunotherapy is the treatment with monoclonal antibodies recognizing tumor-associated markers such as ganglioside GD2. It is considered that major mechanisms of anticancer activity of anti-GD2 antibodies are complement-dependent cytotoxicity and/or antibody-mediated cellular cytotoxicity. At the same time, several studies suggested that anti-GD2 antibodies are capable of direct induction of cell death of number of tumor cell lines, but it has not been investigated in details. In this study we investigated the functional role of ganglioside GD2 in the induction of cell death of multiple tumor cell lines by using GD2-specific monoclonal antibodies. METHODS: Expression of GD2 on different tumor cell lines was analyzed by flow cytometry using anti-GD2 antibodies. By using HPTLC followed by densitometric analysis we measured the amount of ganglioside GD2 in total ganglioside fractions isolated from tumor cell lines. An MTT assay was performed to assess viability of GD2-positive and -negative tumor cell lines treated with anti-GD2 mAbs. Cross-reactivity of anti-GD2 mAbs with other gangliosides or other surface molecules was investigated by ELISA and flow cytometry. Inhibition of GD2 expression was achieved by using of inhibitor for ganglioside synthesis PDMP and/or siRNA for GM2/GD2 and GD3 synthases. RESULTS: Anti-GD2 mAbs effectively induced non-classical cell death that combined features of both apoptosis and necrosis in GD2-positive tumor cells and did not affect GD2-negative tumors. Anti-GD2 mAbs directly induced cell death, which included alteration of mitochondrial membrane potential, induction of apoptotic volume decrease and cell membrane permeability. This cytotoxic effect was mediated exclusively by specific binding of anti-GD2 antibodies with ganglioside GD2 but not with other molecules. Moreover, the level of GD2 expression correlated with susceptibility of tumor cell lines to cytotoxic effect of anti-GD2 antibodies. CONCLUSIONS: Results of this study demonstrate that anti-GD2 antibodies not only passively bind to the surface of tumor cells but also directly induce rapid cell death after the incubation with GD2-positive tumor cells. These results suggest a new role of GD2 as a receptor that actively transduces death signal in malignant cells.
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Gangliosídeos/biossíntese , Imunoterapia , Melanoma/imunologia , Transdução de Sinais/genética , Anticorpos Monoclonais/uso terapêutico , Apoptose/imunologia , Linhagem Celular Tumoral , Gangliosídeos/antagonistas & inibidores , Gangliosídeos/metabolismo , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Melanoma/metabolismo , Melanoma/patologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Potencial da Membrana Mitocondrial/imunologia , Transdução de Sinais/imunologiaRESUMO
Non-alcoholic steatohepatitis (NASH) is a serious public health issue associated with the obesity pandemic. Obesity is the main risk factor for the non-alcoholic fatty liver disease (NAFLD), which progresses to NASH and then to end-stage liver disease. Currently, there are no specific pharmacotherapies of NAFLD/NASH approved by the FDA or other national regulatory bodies and the treatment includes lifestyle adjustment and medicines for improving lipid metabolism, enhancing sensitivity to insulin, balancing oxidation, and counteracting fibrosis. Accordingly, further basic research and development of new therapeutic approaches are greatly needed. Mesenchymal stem cells (MSCs) and MSC-derived extracellular vesicles prevent induced hepatocyte death in vitro and attenuate NASH symptoms in animal models of the disease. They interact with hepatocytes directly, but also target other liver cells, including Kupffer cells and macrophages recruited from the blood flow. This review provides an update on the pathogenesis of NAFLD/NASH and the key role of macrophages in the development of the disease. We examine in detail the mechanisms of the cross-talk between the MSCs and the macrophages, which are likely to be among the key targets of MSCs and their derivatives in the course of NAFLD/NASH cell therapy.
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BACKGROUND: Both ganglioside GD2-targeted immunotherapy and antibody-drug conjugates (ADCs) have demonstrated clinical success as solid tumor therapies in recent years, yet no research has been carried out to develop anti-GD2 ADCs against solid tumors. This is the first study to analyze cytotoxic activity of clinically relevant anti-GD2 ADCs in a wide panel of cell lines with varying GD2 expression and their effects in mouse models of GD2-positive solid cancer. METHODS: Anti-GD2 ADCs were generated based on the GD2-specific antibody ch14.18 approved for the treatment of neuroblastoma and commonly used drugs monomethyl auristatin E (MMAE) or F (MMAF), conjugated via a cleavable linker by thiol-maleimide chemistry. The antibody was produced in a mammalian expression system, and its specific binding to GD2 was analyzed. Antigen-binding properties and biodistribution of the ADCs in mice were studied in comparison with the parent antibody. Cytotoxic effects of the ADCs were evaluated in a wide panel of GD2-positive and GD2-negative tumor cell lines of neuroblastoma, glioma, sarcoma, melanoma, and breast cancer. Their antitumor effects were studied in the B78-D14 melanoma and EL-4 lymphoma syngeneic mouse models. RESULTS: The ch14.18-MMAE and ch14.18-MMAF ADCs retained antigen-binding properties of the parent antibody. Direct dependence of the cytotoxic effect on the level of GD2 expression was observed in cell lines of different origin for both ADCs, with IC50 below 1 nM for the cells with high GD2 expression and no cytotoxic effect for GD2-negative cells. Within the analyzed cell lines, ch14.18-MMAF was more effective in the cells overexpressing GD2, while ch14.18-MMAE had more prominent activity in the cells expressing low GD2 levels. The ADCs had a similar biodistribution profile in the B78-D14 melanoma model compared with the parent antibody, reaching 7.7% ID/g in the tumor at 48 hours postinjection. The average tumor size in groups treated with ch14.18-MMAE or ch14.18-MMAF was 2.6 times and 3.8 times smaller, respectively, compared with the control group. Antitumor effects of the anti-GD2 ADCs were also confirmed in the EL-4 lymphoma model. CONCLUSION: These findings validate the potential of ADCs targeting ganglioside GD2 in treating multiple GD2-expressing solid tumors.
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Antineoplásicos , Imunoconjugados , Melanoma , Neuroblastoma , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Gangliosídeos , Imunoconjugados/farmacologia , Imunoconjugados/uso terapêutico , Mamíferos , Camundongos , Distribuição TecidualRESUMO
Animal model studies and first clinical trials have demonstrated the safety and efficacy of the mesenchymal stem cells' (MSCs) transplantation in stroke. Intra-arterial (IA) administration looks especially promising, since it provides targeted cell delivery to the ischemic brain, is highly effective, and can be safe as long as the infusion is conducted appropriately. However, wider clinical application of the IA MSCs transplantation will only be possible after a better understanding of the mechanism of their therapeutic action is achieved. On the way to achieve this goal, the study of transplanted cells' fate and their interactions with the blood-brain barrier (BBB) structures could be one of the key factors. In this review, we analyze the available data concerning one of the most important aspects of the transplanted MSCs' action-the ability of cells to cross the blood-brain barrier (BBB) in vitro and in vivo after IA administration into animals with experimental stroke. The collected data show that some of the transplanted MSCs temporarily attach to the walls of the cerebral vessels and then return to the bloodstream or penetrate the BBB and either undergo homing in the perivascular space or penetrate deeper into the parenchyma. Transmigration across the BBB is not necessary for the induction of therapeutic effects, which can be incited through a paracrine mechanism even by cells located inside the blood vessels.
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Barreira Hematoencefálica/patologia , Terapia Baseada em Transplante de Células e Tecidos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Acidente Vascular Cerebral/terapia , Animais , Ensaios Clínicos como Assunto , Humanos , Injeções Intra-ArteriaisRESUMO
The tumor-associated ganglioside GD2 represents an attractive target for cancer immunotherapy. GD2-positive tumors are more responsive to such targeted therapy, and new methods are needed for the screening of GD2 molecular tumor phenotypes. In this work, we built a gene expression-based binary classifier predicting the GD2-positive tumor phenotypes. To this end, we compared RNA sequencing data from human tumor biopsy material from experimental samples and public databases as well as from GD2-positive and GD2-negative cancer cell lines, for expression levels of genes encoding enzymes involved in ganglioside biosynthesis. We identified a 2-gene expression signature combining ganglioside synthase genes ST8SIA1 and B4GALNT1 that serves as a more efficient predictor of GD2-positive phenotype (Matthews Correlation Coefficient (MCC) 0.32, 0.88, and 0.98 in three independent comparisons) compared to the individual ganglioside biosynthesis genes (MCC 0.02-0.32, 0.1-0.75, and 0.04-1 for the same independent comparisons). No individual gene showed a higher MCC score than the expression signature MCC score in two or more comparisons. Our diagnostic approach can hopefully be applied for pan-cancer prediction of GD2 phenotypes using gene expression data.
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Chronic liver diseases constitute a significant economic, social, and biomedical burden. Among commonly adopted approaches, only organ transplantation can radically help patients with end-stage liver pathologies. Cell therapy with hepatocytes as a treatment for chronic liver disease has demonstrated promising results. However, quality human hepatocytes are in short supply. Stem/progenitor cells capable of differentiating into functionally active hepatocytes provide an attractive alternative approach to cell therapy for liver diseases, as well as to liver-tissue engineering, drug screening, and basic research. The application of methods generally used to isolate mesenchymal stem cells (MSCs) and maintain them in culture to human liver tissue provides cells, designated here as liver MSCs. They have much in common with MSCs from other tissues, but differ in two aspects-expression of a range of hepatocyte-specific genes and, possibly, inherent commitment to hepatogenic differentiation. The aim of this review is to analyze data regarding liver MSCs, probably another type of liver stem/progenitor cells different from hepatic stellate cells or so-called hepatic progenitor cells. The review presents an analysis of the phenotypic characteristics of liver MSCs, their differentiation and therapeutic potential, methods for isolating these cells from human liver, and discusses issues of their origin and heterogeneity. Human liver MSCs are a fascinating object of fundamental research with a potential for important practical applications.
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Fígado/citologia , Fígado/fisiologia , Células-Tronco Mesenquimais/fisiologia , Tecido Adiposo/citologia , Adulto , Diferenciação Celular/fisiologia , Doença Hepática Terminal/patologia , Doença Hepática Terminal/fisiopatologia , Doença Hepática Terminal/terapia , Hepatócitos/fisiologia , Humanos , Imunomodulação/fisiologia , Transplante de Células-Tronco MesenquimaisRESUMO
Monoclonal antibodies (mAbs) are an important class of therapeutic agents approved for the therapy of many types of malignancies. However, in certain cases applications of conventional mAbs have several limitations in anticancer immunotherapy. These limitations include insufficient efficacy and adverse effects. The antigen-binding fragments of antibodies have a considerable potential to overcome the disadvantages of conventional mAbs, such as poor penetration into solid tumors and Fc-mediated bystander activation of the immune system. Fragments of antibodies retain antigen specificity and part of functional properties of conventional mAbs and at the same time have much better penetration into the tumors and a greatly reduced level of adverse effects. Recent advantages in antibody engineering allowed to produce different types of antibody fragments with improved structure and properties for efficient elimination of tumor cells. These molecules opened up new perspectives for anticancer therapy. Here, we will overview the structural features of the various types of antibody fragments and their applications for anticancer therapy as separate molecules and as part of complex conjugates or structures. Mechanisms of antitumor action of antibody fragments as well as their advantages and disadvantages for clinical application will be discussed in this review.
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Produtos Biológicos/uso terapêutico , Fragmentos de Imunoglobulinas/uso terapêutico , Neoplasias/tratamento farmacológico , Antígenos/imunologia , Humanos , Fragmentos de Imunoglobulinas/imunologiaRESUMO
Neuroblastoma is a pediatric solid cancer of heterogeneous clinical behavior. The unique features of this type of cancer frequently hamper the process of determining clinical presentation and predicting therapy effectiveness. The tumor can spontaneously regress without treatment or actively develop and give rise to metastases despite aggressive multimodal therapy. In recent years, immunotherapy has become one of the most promising approaches to the treatment of neuroblastoma. Still, only one drug for targeted immunotherapy of neuroblastoma, chimeric monoclonal GD2-specific antibodies, is used in the clinic today, and its application has significant limitations. In this regard, the development of effective and safe GD2-targeted immunotherapies and analysis of other potential molecular targets for the treatment of neuroblastoma represents an important and topical task. The review summarizes biological characteristics of the origin and development of neuroblastoma and outlines molecular markers of neuroblastoma and modern immunotherapy approaches directed towards these markers.
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Imunoterapia/métodos , Neuroblastoma/patologia , Neuroblastoma/terapia , HumanosRESUMO
Sequential courses of anticancer target therapy lead to selection of drug-resistant cells, which results in continuous decrease of clinical response. Here we present a new approach for predicting effective combinations of target drugs, which act in a synergistic manner. Synergistic combinations of drugs may prevent or postpone acquired resistance, thus increasing treatment efficiency. We cultured human ovarian carcinoma SKOV-3 and neuroblastoma NGP-127 cancer cell lines in the presence of Tyrosine Kinase Inhibitors (Pazopanib, Sorafenib, and Sunitinib) and Rapalogues (Temsirolimus and Everolimus) for four months and obtained cell lines demonstrating increased drug resistance. We investigated gene expression profiles of intact and resistant cells by microarrays and analyzed alterations in 378 cancer-related signaling pathways using the bioinformatical platform Oncobox. This revealed numerous pathways linked with development of drug resistant phenotypes. Our approach is based on targeting proteins involved in as many as possible signaling pathways upregulated in resistant cells. We tested 13 combinations of drugs and/or selective inhibitors predicted by Oncobox and 10 random combinations. Synergy scores for Oncobox predictions were significantly higher than for randomly selected drug combinations. Thus, the proposed approach significantly outperforms random selection of drugs and can be adopted to enhance discovery of new synergistic combinations of anticancer target drugs.
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Acquired resistance to chemotherapy and radiation therapy is one of the major obstacles decreasing efficiency of treatment of the oncologic diseases. In this study, on the two cell lines (ovarian carcinoma SKOV-3 and neuroblastoma NGP-127), we modeled acquired resistance to five target anticancer drugs. The cells were grown on gradually increasing concentrations of the clinically relevant tyrosine kinase inhibitors (TKIs) Sorafenib, Pazopanib and Sunitinib, and rapalogs Everolimus and Temsirolimus, for 20 weeks. After 20 weeks of culturing, the half-inhibitory concentrations (IC50) increased by 25 - 186% for the particular combinations of the drugs and cell types. We next subjected cells to 10 Gy irradiation, a dose frequently used in clinical radiation therapy. For the SKOV-3, but not NGP-127 cells, for the TKIs Sorafenib, Pazopanib and Sunitinib, we noticed statistically significant increase in capacity to repair radiation-induced DNA double strand breaks compared to naïve control cells not previously treated with TKIs. These peculiarities were linked with the increased activation of ATM DNA repair pathway in the TKI-treated SKOV-3, but not NGP-127 cells. Our results provide a new cell culture model for studying anti-cancer therapy efficiency and evidence that there may be a tissue-specific radioresistance emerging as a side effect of treatment with TKIs.
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The emerging field of regenerative medicine offers innovative methods of cell therapy and tissue/organ engineering as a novel approach to liver disease treatment. The ultimate scientific foundation of both cell therapy of liver diseases and liver tissue and organ engineering is delivered by the in-depth studies of the cellular and molecular mechanisms of liver regeneration. The cellular mechanisms of the homeostatic and injury-induced liver regeneration are unique. Restoration of the mass of liver parenchyma is achieved by compensatory hypertrophy and hyperplasia of the differentiated parenchymal cells, hepatocytes, while expansion and differentiation of the resident stem/progenitor cells play a minor or negligible role. Participation of blood-borne cells of the bone marrow origin in liver parenchyma regeneration has been proven but does not exceed 1-2% of newly formed hepatocytes. Liver regeneration is activated spontaneously after injury and can be further stimulated by cell therapy with hepatocytes, hematopoietic stem cells, or mesenchymal stem cells. Further studies aimed at improving the outcomes of cell therapy of liver diseases are underway. In case of liver failure, transplantation of engineered liver can become the best option in the foreseeable future. Engineering of a transplantable liver or its major part is an enormous challenge, but rapid progress in induced pluripotency, tissue engineering, and bioprinting research shows that it may be doable.
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Terapia Baseada em Transplante de Células e Tecidos , Hepatopatias/genética , Regeneração Hepática , Engenharia Tecidual , Diferenciação Celular/genética , Hepatócitos/patologia , Humanos , Fígado/patologia , Hepatopatias/patologia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologiaRESUMO
In vivo tracking of transplanted mesenchymal stem cells (MSCs) migration and homing is vital for understanding the mechanisms of beneficial effects of MSCs transplantation in animal models of diseases and in clinical trials. Transplanted cells can be labeled with superparamagnetic iron oxide (SPIO) particles and visualized in vivo using a number of iron sensitive MRI techniques. However, the applicability of those techniques for SPIO-labeled MSCs tracking in live brain has not been sufficiently investigated. The goal of this study was to estimate the efficiency of various MRI techniques of SPIO-labeled cell tracing in the brain. To achieve that goal, the precision and specificity of T2WI, T2*WI and SWI (Susceptibility-Weighted Imaging) techniques of SPIO-labeled MSCs tracing in vitro and in live rat brain were for the first time compared in the same experiment. We have shown that SWI presents the most sensitive pulse sequence for SPIO-labeled MSCs MR visualization. After intracerebral administration due to limitations caused by local micro-hemorrhages the visualization threshold was 102 cells, while after intra-arterial transplantation SWI permitted detection of several cells or even single cells. There is just one publication claiming detection of individual SPIO-labeled MSCs in live brain, while the other state much lower sensitivity, describe detection of different cell types or high resolution tracing of MSCs in other tissues. This study confirms the possibility of single cell tracing in live brain and outlines the necessary conditions. SWI is a method convenient for the detection of single SPIO labeled MSCs and small groups of SPIO labeled MSCs in brain tissue and can be appropriate for monitoring migration and homing of transplanted cells in basic and translational neuroscience.