Search

Neuroblastoma (NB) has a low frequency of recurrent mutations compared to other cancers, which hinders the development of targeted therapies and novel risk stratification strategies. Multikinase inhibitors have shown potential in treating high-risk NB, but their efficacy is likely impaired by the cancer cells’ ability to adapt to these drugs through the employment of alternative signaling pathways. Based on the expression of 48 growth factor-related genes in 1189 NB tumors, we have developed a model for NB patient survival prediction. This model discriminates between stage 4 NB tumors with favorable outcomes (>80% overall survival) and very poor outcomes (

The degradation of most intracellular proteins is a dynamic and tightly regulated process performed by proteasomes. To date, different forms of proteasomes have been identified. Currently the role of non-constitutive proteasomes (immunoproteasomes (iPs) and intermediate proteasomes (intPs)) has attracted special attention. Here, using a CRISPR-Cas9 nickase technology, four cell lines: histiocytic lymphoma, colorectal adenocarcinoma, cervix adenocarcinoma, and hepatocarcinoma were modified to express proteasomes with mCherry-tagged β5i subunit, which is a catalytic subunit of iPs and intPs. Importantly, the expression of the chimeric gene in modified cells is under the control of endogenous regulatory mechanisms and is increased following IFN-γ and/or TNF-α stimulation. Fluorescent proteasomes retain catalytic activity and are distributed within the nucleus and cytoplasm. RNAseq reveals marginal differences in gene expression profiles between the modified and wild-type cell lines. Predominant metabolic pathways and patterns of expressed receptors were identified for each cell line. Using established cell lines, we demonstrated that anti-cancer drugs Ruxolitinib, Vincristine and Gefitinib stimulated the expression of β5i-containing proteasomes, which might affect disease prognosis. Taken together, obtained cell lines can be used as a platform for real-time studies of immunoproteasome gene expression, localization of iPs and intPs, interaction of non-constitutive proteasomes with other proteins, proteasome trafficking and many other aspects of proteasome biology in living cells. Moreover, the established platform might be especially useful for fast and large-scale experiments intended to evaluate the effects of different conditions including treatment with various drugs and compounds on the proteasome pool.

Myeloid leukemia is a hematologic neoplasia characterized by a clonal proliferation of hematopoietic stem cell progenitors. Patient prognosis varies depending on the subtype of leukemia as well as eligibility for intensive treatment regimens and allogeneic stem cell transplantation. Although significant progress has been made in the therapy of patients including novel targeted treatment approaches, there is still an urgent need to optimize treatment outcome. The most common therapy is based on the use of chemotherapeutics cytarabine and anthrayclines. Here, we studied the effect of the recently synthesized marine alkaloid 3,10-dibromofascaplysin (DBF) in myeloid leukemia cells. Unsubstituted fascaplysin was early found to affect cell cycle via inhibiting CDK4/6, thus we compared the activity of DBF and other brominated derivatives with known CDK4/6 inhibitor palbociclib, which was earlier shown to be a promising candidate to treat leukemia. Unexpectedly, the effect DBF on cell cycle differs from palbociclib. In fact, DBF induced leukemic cells apoptosis and decreased the expression of genes responsible for cancer cell survival. Simultaneously, DBF was found to activate the E2F1 transcription factor. Using bioinformatical approaches we evaluated the possible molecular mechanisms, which may be associated with DBF-induced activation of E2F1. Finally, we found that DBF synergistically increase the cytotoxic effect of cytarabine in different myeloid leukemia cell lines. In conclusion, DBF is a promising drug candidate, which may be used in combinational therapeutics approaches to reduce leukemia cell growth.

Understanding the molecular basis of fibrosis, the lethal complication of COVID-19, is urgent. By the analysis of RNA-sequencing data of SARS-CoV-2-infected cells combined with data mining we identified genes involved in COVID-19 progression. To characterize their implication in the fibrosis development we established a correlation matrix based on the transcriptomic data of patients with idiopathic pulmonary fibrosis. With this method, we have identified a cluster of genes responsible for SARS-CoV-2-fibrosis including its entry receptor ACE2 and epidermal growth factor EGF. Then, we developed Vi-Fi scoring-a novel drug repurposing approach and simultaneously quantified antiviral and antifibrotic activities of the drugs based on their transcriptomic signatures. We revealed the strong dual antifibrotic and antiviral activity of EGFR/ErbB inhibitors. Before the in vitro validation, we have clustered 277 cell lines and revealed distinct COVID-19 transcriptomic signatures of the cells with similar phenotypes that defines their suitability for COVID-19 research. By ERK activity monitoring in living lung cells, we show that the drugs with predicted antifibrotic activity downregulate ERK in the host lung cells. Overall, our study provides novel insights on SARS-CoV-2 dependence on EGFR/ERK signaling and demonstrates the utility of EGFR/ErbB inhibitors for COVID-19 treatment.

Acute myeloid leukemia (AML) is a genetically heterogeneous group of oncological diseases of the hematopoietic system, which are extremely difficult to treat. The development of new targeted drugs (Hylteritinib, Venetoclax) significantly improved the survival of patients, but resistance, as well as cytotoxic anti-leukemia drugs, often occurs. The search for new molecular targets for the development of effective approaches for the treatment of AML is very urgent. In blast cells of patients with AML, mutations, chromosomal rearrangements, and increased expression of a number of non-mutant genes, including transcription factor genes, are detected. The transcription factor Sp 1 binds to GC-rich regions of regulatory regions of various genes and thus controls their expression. Sp1 targets include genes responsible for proliferation, cell cycle regulation, and differentiation. In many malignant diseases, a high level of Sp1 gene expression is associated with an unfavorable prognosis, therefore, Sp1 is considered as a promising therapeutic target for cancer. In this paper, we estimated the expression levels of Sp1 in various malignant tissues. Increased Sp1 expression was detected in samples obtained from patients with AML, acute lymphoblastic leukemia, Ewing sarcoma, ovarian and kidney cancer. It is also shown that Sp1 expression correlates with the expression of genes encoding cytokine receptors and growth factors (CSF1R and IL6R), intracellular kinases (CSK, SYK, PAK1, ILK, JAK2), and transcription factor LMO2. The correlation between expression levels of Sp1 and CSF1R, SYK, Jak2 and LMO2 is also characteristic of transplanted human leukemia cells. We measured expression levels of Sp1, CSF1R, ILK, PAK1 in the cells of three transplantable lines of human leukemia and found increased levels of expression of these genes in Kasumi-1 cells. In addition, we showed that Kasumi-1 cells are most sensitive to Mitramycin, a drug that displaces Sp1 from its targets with DNA. Our data indicate the need to identify AML cells that are most sensitive to inhibition of Sp1 activity in order to assess the possibility of suppressing its activity in vivo.

Novel treatments for various types of malignant diseases are warranted. In this study, we evaluated JAK2 inhibitors (Janus kinase 2) for suppressing the growth of malignant neuroblastoma and glioblastoma cells as well as breast and non-small cell lung cancers. Neuroblastoma and glioblastoma cells are the most sensitive to the JAK2 inhibitor AG490. A study of the relative expression of receptors that can activate JAK2 suggests that cell line sensitivity to AG490 may be mediated by IL6-R, IL11-R and/or CSF1-R. AG490 enhances the effect of doxorubicin on neuroblastoma cells. Our findings suggest the possible relevance of JAK2 inhibitors for neuroblastoma therapy, especially in combination with doxorubicin.

Hallmark of neuroblastoma is an ability of this malignant tumor to undergo spontaneous regression or differentiation into benign tumor during any stage of the disease, but it is little known about mechanisms of these phenomena. We studied effect of receptor tyrosine kinase receptor KIT on expression of genes, which may be involved in tumor spontaneous regression. Downregulation of KIT expression by RNA interference in SH-SY5Y cells causes suppression of neurotrophin receptor NGFR expression that may promote the loss of sensibility of cells to nerve growth factors, also it causes upregulation of TrkA receptor expression which can stimulate cell differentiation or apoptosis in NGF dependent manner. Furthermore there is an upregulation of genes which stimulate malignant cell detection by immune system, such as genes of major histocompatibility complex HLA class I HLA-B and HLA-C, and interferon-γ receptors IFNGR1 and IFNGR2 genes. Thus KIT can mediate neuroblastoma cell sensibility to neurotrophins and immune system components--two factors directly contributing to spontaneous regression of neuroblastoma.

In this study we evaluated c-kit, VEGFA, and MYC gene expression level in seven neuroblastoma stable cell lines: SK-N-SH, SK-N-BE, SK-N-AS, SH-SY5Y, Kelly, IMR-32, and LAN-1. Expression levels of these genes can serve as diagnostic factors of cancer progression, and proteins encoded by these genes are promising targets for neuroblastoma treatment. SH-SY5Y and SK-N-AS cells have highest MYC expression and the same VEGFA expression, although SH-SY5Y has 10 times higher c-kit expression than SK-N-AS cells. Both IMR-32 and LAN-1 cells have low MYC expression level, but differ in c-kit expression, IMR-32 has significantly higher c-kit expression, than any other neuroblastoma cell line. LAN-1 on the other hand has the highest VEGFA expression. These data suggest that MYC, c-kit, and VEGFA genes can play different roles in development and progression of neuroblastoma depending on other activated molecular mechanisms in malignant cells.

The t(8;21)(q22;q22) rearrangement represents the most common chromosomal translocation in acute myeloid leukemia (AML). It results in a transcript encoding for the fusion protein AML1-ETO (AE) with transcription factor activity. AE is considered to be an attractive target for treating t(8;21) leukemia. However, AE expression alone is insufficient to cause transformation, and thus the potential of such therapy remains unclear. Several genes are deregulated in AML cells, including KIT that encodes a tyrosine kinase receptor. Here, we show that AML cells transduced with short hairpin RNA vector targeting AE mRNAs have a dramatic decrease in growth rate that is caused by induction of apoptosis and deregulation of the cell cycle. A reduction in KIT mRNA levels was also observed in AE-silenced cells, but silencing KIT expression reduced cell growth but did not induce apoptosis. Transcription profiling of cells that escape cell death revealed activation of a number of signaling pathways involved in cell survival and proliferation. In particular, we find that the extracellular signal-regulated kinase 2 (ERK2; also known as mitogen-activated protein kinase 1 (MAPK1)) protein could mediate activation of 23 out of 29 (79%) of these upregulated pathways and thus may be regarded as the key player in establishing the t(8;21)-positive leukemic cells resistant to AE suppression.