Guardians and Mediators of Metastasis: Exploring T Lymphocytes, Myeloid-Derived Suppressor Cells, and Tumor-Associated Macrophages in the Breast Cancer Microenvironment.

Breast cancers (BCs) are solid tumors composed of heterogeneous tissues consisting of cancer cells and an ever-changing tumor microenvironment (TME). The TME includes, among other non-cancer cell types, immune cells influencing the immune context of cancer tissues. In particular, the cross talk of immune cells and their interactions with cancer cells dramatically influence BC dissemination, immunoediting, and the outcomes of cancer therapies. Tumor-infiltrating lymphocytes (TILs), tumor-associated macrophages (TAMs), and myeloid-derived suppressor cells (MDSCs) represent prominent immune cell populations of breast TMEs, and they have important roles in cancer immunoescape and dissemination. Therefore, in this article we review the features of TILs, TAMs, and MDSCs in BCs. Moreover, we highlight the mechanisms by which these immune cells remodel the immune TME and lead to breast cancer metastasis.

miRNA-23a modulates sodium-hydrogen exchanger 1 expression: studies in medullary thick ascending limb of salt-induced hypertensive rats.

BACKGROUND: The kidney is the main organ in the pathophysiology of essential hypertension. Although most bicarbonate reabsorption occurs in the proximal tubule, the medullary thick ascending limb (mTAL) of the nephron also maintains acid-base balance by contributing to 25% of bicarbonate reabsorption. A crucial element in this regulation is the sodium-hydrogen exchanger 1 (NHE1), a ubiquitous membrane protein controlling intracellular pH, where proton extrusion is driven by the inward sodium flux. MicroRNA (miRNA) expression of hypertensive patients significantly differs from that of normotensive subjects. The aim of this study was to determine the functional role of miRNA alterations at the mTAL level. METHODS: By miRNA microarray analysis, we identified miRNA expression profiles in isolated mTALs from high sodium intake-induced hypertensive rats (HSD) versus their normotensive counterparts (NSD). In vitro validation was carried out in rat mTAL cells. RESULTS: Five miRNAs involved in the onset of salt-sensitive hypertension were identified, including miR-23a, which was bioinformatically predicted to target NHE1 mRNA. Data demonstrated that miRNA-23a is downregulated in the mTAL of HSD rats while NHE1 is upregulated. Consistently, transfection of an miRNA-23a mimic in an mTAL cell line, using a viral vector, resulted in NHE1 downregulation. CONCLUSION: NHE1, a protein involved in sodium reabsorption at the mTAL level and blood pressure regulation, is upregulated in our model. This was due to a downregulation of miRNA-23a. Expression levels of this miRNA are influenced by high sodium intake in the mTALs of rats. The downregulation of miRNA-23a in humans affected by essential hypertension corroborate our data and point to the potential role of miRNA-23a in the regulation of mTAL function following high salt intake.

A widespread role for SLC transmembrane transporters in resistance to cytotoxic drugs.

Solute carriers (SLCs) are the largest family of transmembrane transporters in humans and are major determinants of cellular metabolism. Several SLCs have been shown to be required for the uptake of chemical compounds into cellular systems, but systematic surveys of transporter-drug relationships in human cells are currently lacking. We performed a series of genetic screens in a haploid human cell line against 60 cytotoxic compounds representative of the chemical space populated by approved drugs. By using an SLC-focused CRISPR-Cas9 library, we identified transporters whose absence induced resistance to the drugs tested. This included dependencies involving the transporters SLC11A2/SLC16A1 for artemisinin derivatives and SLC35A2/SLC38A5 for cisplatin. The functional dependence on SLCs observed for a significant proportion of the screened compounds suggests a widespread role for SLCs in the uptake and cellular activity of cytotoxic drugs and provides an experimentally validated set of SLC-drug associations for a number of clinically relevant compounds.

The Lack of STING Impairs the MHC-I Dependent Antigen Presentation and JAK/STAT Signaling in Murine Macrophages.

STING is a transmembrane ER resident protein that was initially described as a regulator of innate immune response triggered by viral DNA and later found to be involved in a broader range of immune processes. Here, we assessed its role in the antigen presentation by generating a STING KO macrophage cell line. In the absence of STING, we observed an impaired OVA-derived SIINFEKL peptide presentation together with a decreased level of MHC-I complex on the plasma membrane, likely due to a decreased mRNA expression of ß2 m light chain as no relevant alterations of the peptide-loading complex (TAPs) were found. Moreover, JAK-STAT signaling resulted in impaired STING KO cells following OVA and LPS treatments, suggesting a dampened activation of immune response. Our data revealed a new molecular role of STING in immune mechanisms that could elucidate its role in the pathogenesis of autoimmune disorders and cancer.

IBtkα Activates the ß-Catenin-Dependent Transcription of MYC through Ubiquitylation and Proteasomal Degradation of GSK3ß in Cancerous B Cells.

The IBTK gene encodes the IBtkα protein that is a substrate receptor of E3 ubiquitin ligase, Cullin 3. We have previously reported the pro-tumorigenic activity of Ibtk in MYC-dependent B-lymphomagenesis observed in Eµ-myc transgenic mice. Here, we provide mechanistic evidence of the functional interplay between IBtkα and MYC. We show that IBtkα, albeit indirectly, activates the ß-catenin-dependent transcription of the MYC gene. Of course, IBtkα associates with GSK3ß and promotes its ubiquitylation, which is associated with proteasomal degradation. This event increases the protein level of ß-catenin, a substrate of GSK3ß, and results in the transcriptional activation of the MYC and CCND1 target genes of ß-catenin, which are involved in the control of cell division and apoptosis. In particular, we found that in Burkitt's lymphoma cells, IBtkα silencing triggered the downregulation of both MYC mRNA and protein expression, as well as a strong decrease of cell survival, mainly through the induction of apoptotic events, as assessed by using flow cytometry-based cell cycle and apoptosis analysis. Collectively, our results shed further light on the complex puzzle of IBtkα interactome and highlight IBtkα as a potential novel therapeutic target to be employed in the strategy for personalized therapy of B cell lymphoma.

Generation and Characterization of a Tumor Stromal Microenvironment and Analysis of Its Interplay with Breast Cancer Cells: An In Vitro Model to Study Breast Cancer-Associated Fibroblast Inactivation.

Breast cancer-associated fibroblasts (BCAFs), the most abundant non-cancer stromal cells of the breast tumor microenvironment (TME), dramatically sustain breast cancer (BC) progression by interacting with BC cells. BCAFs, as well as myofibroblasts, display an up regulation of activation and inflammation markers represented by α-smooth muscle actin (α-SMA) and cyclooxygenase 2 (COX-2). BCAF aggregates have been identified in the peripheral blood of metastatic BC patients. We generated an in vitro stromal model consisting of human primary BCAFs grown as monolayers or 3D cell aggregates, namely spheroids and reverted BCAFs, obtained from BCAF spheroids reverted to 2D cell adhesion growth after 216 h of 3D culture. We firstly evaluated the state of activation and inflammation and the mesenchymal status of the BCAF monolayers, BCAF spheroids and reverted BCAFs. Then, we analyzed the MCF-7 cell viability and migration following treatment with conditioned media from the different BCAF cultures. After 216 h of 3D culture, the BCAFs acquired an inactivated phenotype, associated with a significant reduction in α-SMA and COX-2 protein expression. The deactivation of the BCAF spheroids at 216 h was further confirmed by the cytostatic effect exerted by their conditioned medium on MCF-7 cells. Interestingly, the reverted BCAFs also retained a less activated phenotype as indicated by α-SMA protein expression reduction. Furthermore, the reverted BCAFs exhibited a reduced pro-tumor phenotype as indicated by the anti-migratory effect exerted by their conditioned medium on MCF-7 cells. The deactivation of BCAFs without drug treatment is possible and leads to a reduced capability of BCAFs to sustain BC progression in vitro. Consequently, this study could be a starting point to develop new therapeutic strategies targeting BCAFs and their interactions with cancer cells.

Influence of Tumor Microenvironment and Fibroblast Population Plasticity on Melanoma Growth, Therapy Resistance and Immunoescape.

Cutaneous melanoma (CM) tissue represents a network constituted by cancer cells and tumor microenvironment (TME). A key feature of CM is the high structural and cellular plasticity of TME, allowing its evolution with disease and adaptation to cancer cell and environmental alterations. In particular, during melanoma development and progression each component of TME by interacting with each other and with cancer cells is subjected to dramatic structural and cellular modifications. These alterations affect extracellular matrix (ECM) remodelling, phenotypic profile of stromal cells, cancer growth and therapeutic response. The stromal fibroblast populations of the TME include normal fibroblasts and melanoma-associated fibroblasts (MAFs) that are highly abundant and flexible cell types interacting with melanoma and stromal cells and differently influencing CM outcomes. The shift from the normal microenvironment to TME and from normal fibroblasts to MAFs deeply sustains CM growth. Hence, in this article we review the features of the normal microenvironment and TME and describe the phenotypic plasticity of normal dermal fibroblasts and MAFs, highlighting their roles in normal skin homeostasis and TME regulation. Moreover, we discuss the influence of MAFs and their secretory profiles on TME remodelling, melanoma progression, targeted therapy resistance and immunosurveillance, highlighting the cellular interactions, the signalling pathways and molecules involved in these processes.

Molecular T-Cell Repertoire Analysis as Source of Prognostic and Predictive Biomarkers for Checkpoint Blockade Immunotherapy.

The T cells are key players of the response to checkpoint blockade immunotherapy (CBI) and monitoring the strength and specificity of antitumor T-cell reactivity remains a crucial but elusive component of precision immunotherapy. The entire assembly of T-cell receptor (TCR) sequences accounts for antigen specificity and strength of the T-cell immune response. The TCR repertoire hence represents a "footprint" of the conditions faced by T cells that dynamically evolves according to the challenges that arise for the immune system, such as tumor neo-antigenic load. Hence, TCR repertoire analysis is becoming increasingly important to comprehensively understand the nature of a successful antitumor T-cell response, and to improve the success and safety of current CBI.

IBTK Haploinsufficiency Affects the Tumor Microenvironment of Myc-Driven Lymphoma in E-myc Mice.

The tumor microenvironment is a dynamic and interactive supporting network of various components, including blood vessels, cytokines, chemokines, and immune cells, which sustain the tumor cell's survival and growth. Murine models of lymphoma are useful to study tumor biology, the microenvironment, and mechanisms of response to therapy. Lymphomas are heterogeneous hematologic malignancies, and the complex microenvironment from which they arise and their multifaceted genetic basis represents a challenge for the generation and use of an appropriate murine model. So, it is important to choose the correct methodology. Recently, we supported the first evidence on the pro-oncogenic action of IBTK in Myc-driven B cell lymphomagenesis in mice, inhibiting apoptosis in the pre-cancerous stage. We used the transgenic Eµ-myc mouse model of non-Hodgkin's lymphoma and Ibtk hemizygous mice to evaluate the tumor development of Myc-driven lymphoma. Here, we report that the allelic loss of Ibtk alters the immunophenotype of Myc-driven B cell lymphomas, increasing the rate of pre-B cells and affecting the tumor microenvironment in Eµ-myc mice. In particular, we observed enhanced tumor angiogenesis, increasing pro-angiogenic and lymphangiogenic factors, such as VEGF, MMP-9, CCL2, and VEGFD, and a significant recruitment of tumor-associated macrophages in lymphomas of Ibtk+/- Eµ-myc compared to Ibtk+/+ Eµ-myc mice. In summary, these results indicate that IBTK haploinsufficiency promotes Myc tumor development by modifying the tumor microenvironment.

Insights about MYC and Apoptosis in B-Lymphomagenesis: An Update from Murine Models.

The balance between cell survival and cell death represents an essential part of human tissue homeostasis, while altered apoptosis contributes to several pathologies and can affect the treatment efficacy. Impaired apoptosis is one of the main cancer hallmarks and some types of lymphomas harbor mutations that directly affect key regulators of cell death (such as BCL-2 family members). The development of novel techniques in the field of immunology and new animal models has greatly accelerated our understanding of oncogenic mechanisms in MYC-associated lymphomas. Mouse models are a powerful tool to reveal multiple genes implicated in the genesis of lymphoma and are extensively used to clarify the molecular mechanism of lymphoma, validating the gene function. Key features of MYC-induced apoptosis will be discussed here along with more recent studies on MYC direct and indirect interactors, including their cooperative action in lymphomagenesis. We review our current knowledge about the role of MYC-induced apoptosis in B-cell malignancies, discussing the transcriptional regulation network of MYC and regulatory feedback action of miRs during MYC-driven lymphomagenesis. More importantly, the finding of new modulators of apoptosis now enabling researchers to translate the discoveries that have been made in the laboratory into clinical practice to positively impact human health.

Revisiting Bacterial Ubiquitin Ligase Effectors: Weapons for Host Exploitation.

Protein ubiquitylation plays a central role in eukaryotic cell physiology. It is involved in several regulatory processes, ranging from protein folding or degradation, subcellular localization of proteins, vesicular trafficking and endocytosis to DNA repair, cell cycle, innate immunity, autophagy, and apoptosis. As such, it is reasonable that pathogens have developed a way to exploit such a crucial system to enhance their virulence against the host. Hence, bacteria have evolved a wide range of effectors capable of mimicking the main players of the eukaryotic ubiquitin system, in particular ubiquitin ligases, by interfering with host physiology. Here, we give an overview of this topic and, in particular, we detail and discuss the mechanisms developed by pathogenic bacteria to hijack the host ubiquitination system for their own benefit.

Monitoring multiple myeloma by idiotype-specific peptide binders of tumor-derived exosomes.

Tumor-derived exosomes (TDEs) play a pivotal role in tumor establishment and progression, and are emerging biomarkers for tumor diagnosis in personalized medicine. To date, there is a lack of efficient technology platforms for exosome isolation and characterization. Multiple myeloma (MM) is an incurable B-cell malignancy due to the rapid development of drug-resistance. MM-released exosomes express the immunoglobulin B-cell receptor (Ig-BCR) of the tumor B-cells, which can be targeted by Idiotype-binding peptides (Id-peptides). In this study, we analyzed the production of MM-released exosomes in the murine 5T33MM multiple myeloma model as biomarkers of tumor growth. To this end, we selected Id-peptides by screening a phage display library using as bait the Ig-BCR expressed by 5T33MM cells. By FACS, the FITC-conjugated Id-peptides detected the MM-released exosomes in the serum of 5T33MM-engrafted mice, levels of which are correlated with tumor progression at an earlier time point compared to serum paraprotein. These results indicate that Id-peptide-based recognition of MM-released exosomes may represent a very sensitive diagnostic approach for clinical evaluation of disease progression.

CRL3IBTK Regulates the Tumor Suppressor Pdcd4 through Ubiquitylation Coupled to Proteasomal Degradation.

The human inhibitor of Bruton's tyrosine kinase isoform α (IBtkα) is a BTB protein encoded by the IBTK gene, which maps to chromosomal locus 6q14.1, a mutational hot spot in lymphoproliferative disorders. Here, we demonstrate that IBtkα forms a CRL3(IBTK) complex promoting its self-ubiquitylation. We identified the tumor suppressor Pdcd4 as IBtkα interactor and ubiquitylation substrate of CRL3(IBTK) for proteasomal degradation. Serum-induced degradation of Pdcd4 required both IBtkα and Cul3, indicating that CRL3(IBTK) regulated the Pdcd4 stability in serum signaling. By promoting Pdcd4 degradation, IBtkα counteracted the suppressive effect of Pdcd4 on translation of reporter luciferase mRNAs with stem-loop structured or unstructured 5'-UTR. IBtkα depletion by RNAi caused Pdcd4 accumulation and decreased the translation of Bcl-xL mRNA, a well known target of Pdcd4 repression. By characterizing CRL3(IBTK) as a novel ubiquitin ligase, this study provides new insights into regulatory mechanisms of cellular pathways, such as the Pdcd4-dependent translation of mRNAs.

Single-cell real-time imaging of transgene expression upon lipofection.

Here we address the process of lipofection by quantifying the expression of a genetically-encoded fluorescent reporter at the single-cell level, and in real-time, by confocal imaging in live cells. The Lipofectamine gold-standard formulation is compared to the alternative promising DC-Chol/DOPE formulation. In both cases, we report that only dividing cells are able to produce a detectable amount of the fluorescent reporter protein. Notably, by measuring fluorescence over time in each pair of daughter cells, we find that Lipofectamine-based transfection statistically yields a remarkably higher degree of "symmetry" in protein expression between daughter cells as compared to DC-Chol/DOPE. A model is envisioned in which the degree of symmetry of protein expression is linked to the number of bioavailable DNA copies within the cell before nuclear breakdown. Reported results open new perspectives for the understanding of the lipofection mechanism and define a new experimental platform for the quantitative comparison of transfection reagents.

Selective Enzymatic Transformation to Aldehydes in vivo by Fungal Carboxylate Reductase from Neurospora crassa.

The enzymatic reduction of carboxylic acids is in its infancy with only a handful of biocatalysts available to this end. We have increased the spectrum of carboxylate-reducing enzymes (CARs) with the sequence of a fungal CAR from Neurospora crassa OR74A (NcCAR). NcCAR was efficiently expressed in E. coli using an autoinduction protocol at low temperature. It was purified and characterized in vitro, revealing a broad substrate acceptance, a pH optimum at pH 5.5-6.0, a Tm of 45 °C and inhibition by the co-product pyrophosphate which can be alleviated by the addition of pyrophosphatase. The synthetic utility of NcCAR was demonstrated in a whole-cell biotransformation using the Escherichia coli K-12 MG1655 RARE strain in order to suppress overreduction to undesired alcohol. The fragrance compound piperonal was prepared from piperonylic acid (30 mM) on gram scale in 92 % isolated yield in >98% purity. This corresponds to a productivity of 1.5 g/L/h.

IBTK Differently Modulates Gene Expression and RNA Splicing in HeLa and K562 Cells.

The IBTK gene encodes the major protein isoform IBTKα that was recently characterized as substrate receptor of Cul3-dependent E3 ligase, regulating ubiquitination coupled to proteasomal degradation of Pdcd4, an inhibitor of translation. Due to the presence of Ankyrin-BTB-RCC1 domains that mediate several protein-protein interactions, IBTKα could exert expanded regulatory roles, including interaction with transcription regulators. To verify the effects of IBTKα on gene expression, we analyzed HeLa and K562 cell transcriptomes by RNA-Sequencing before and after IBTK knock-down by shRNA transduction. In HeLa cells, 1285 (2.03%) of 63,128 mapped transcripts were differentially expressed in IBTK-shRNA-transduced cells, as compared to cells treated with control-shRNA, with 587 upregulated (45.7%) and 698 downregulated (54.3%) RNAs. In K562 cells, 1959 (3.1%) of 63128 mapped RNAs were differentially expressed in IBTK-shRNA-transduced cells, including 1053 upregulated (53.7%) and 906 downregulated (46.3%). Only 137 transcripts (0.22%) were commonly deregulated by IBTK silencing in both HeLa and K562 cells, indicating that most IBTKα effects on gene expression are cell type-specific. Based on gene ontology classification, the genes responsive to IBTK are involved in different biological processes, including in particular chromatin and nucleosomal organization, gene expression regulation, and cellular traffic and migration. In addition, IBTK RNA interference affected RNA maturation in both cell lines, as shown by the evidence of alternative 3'- and 5'-splicing, mutually exclusive exons, retained introns, and skipped exons. Altogether, these results indicate that IBTK differently modulates gene expression and RNA splicing in HeLa and K562 cells, demonstrating a novel biological role of this protein.

B-cell receptor-guided delivery of peptide-siRNA complex for B-cell lymphoma therapy.

BACKGROUND: Despite the clinical response of conventional anticancer therapy, including chemotherapeutic treatments, radiation therapy and corticosteroids, tumorigenic B-cell lymphomas show an incomplete response to clinical practices that result in a minimal residual disease (MRD) where few residual neoplastic cells undetected in vivo, replenish the cancer cell reservoir. This scenario, which is also shared with other cancer diseases, requires the development of strategies to advance in novel, selective targeting toward the tumorigenic cells that survive to the anticancer agents. METHODS: Here, we have taken advantage of the therapeutic properties of an idiotype specific peptide (pA20-36) that bind specifically to murine B-lymphoma cells in the setting of an anti cancer strategy, based on the selected delivery of electrostatic-based complex, peptide-siRNA. To this end, two engineered, arginine rich, peptides that included the pA20-36 targeting sequence were designed to bind fluorescent-labelled siRNA. One peptide presented 9 Arg at the C-terminal of pA20-36 whereas the other included 5 Arg at the N- and C-terminus, respectively. RESULTS: Compared to the control and random peptide-siRNA complexes, both pA20-36-siRNA complexes were endowed with the selective delivering of fluorescent-labelled siRNA toward the A20 murine B-cell lymphoma, as evaluated by cytofluorimetry and confocal microscopy, whereas fluorescent-labelled siRNA alone was not internalized in the selected cells. Compared to peptide controls, the use of the modified pA20-36 peptides complexed with siRNA anti-GAPDH and anti-Bcl2 showed a down-regulation in the expression levels of the corresponding genes. CONCLUSIONS: Peptide-siRNA complex can be suitable tool for both selective peptide-driven cell targeting and gene silencing. In this setting, the improvement of this strategy is expected to provide a safe and non-invasive approach for the delivery of therapeutic molecules.

Early molecular and behavioral response to lipopolysaccharide in the WAG/Rij rat model of absence epilepsy and depressive-like behavior, involves interplay between AMPK, AKT/mTOR pathways and neuroinflammatory cytokine release.

The mammalian target of rapamycin (mTOR) pathway has been recently indicated as a suitable drug target for the prevention of epileptogenesis. The mTOR pathway is known for its involvement in the control of the immune system. Since neuroinflammation is recognized as a major contributor to epileptogenesis, we wished to examine whether the neuroprotective effects of mTOR modulation could involve a suppression of the neuroinflammatory process in epileptic brain. We have investigated the early molecular mechanisms involved in the effects of intracerebral administration of the lipopolysaccharide (LPS) in the WAG/Rij rat model of absence epilepsy, in relation to seizure generation and depressive-like behavior; we also tested whether the effects of LPS could be modulated by treatment with rapamycin (RAP), a specific mTOR inhibitor. We determined, in specific rat brain areas, levels of p-mTOR/p-p70S6K and also p-AKT/p-AMPK as downstream or upstream indicators of mTOR activity and tested the effects of LPS and RAP co-administration. Changes in the brain levels of pro-inflammatory cytokines IL-1ß and TNF-α and their relative mRNA expression levels were measured, and the involvement of nuclear factor-κB (NF-κB) was also examined in vitro. We confirmed that RAP inhibits the aggravation of absence seizures and depressive-like/sickness behavior induced by LPS in the WAG/Rij rats through the activation of mTOR and show that this effect is correlated with the ability of RAP to dampen and delay LPS increases in neuroinflammatory cytokines IL-1ß and TNF-α, most likely through inhibition of the activation of NF-κB. Our results suggest that such a mechanism could contribute to the antiseizure, antiepileptogenic and behavioral effects of RAP and further highlight the potential therapeutic usefulness of mTOR inhibition in the management of human epilepsy and other neurological disorders. Furthermore, we show that LPS-dependent neuroinflammatory effects are also mediated by a complex interplay between AKT, AMPK and mTOR with specificity to selective brain areas. In conclusion, neuroinflammation appears to be a highly coordinated phenomenon, where timing of intervention may be carefully evaluated in order to identify the best suitable target.

Human immunodeficiency virus-1 Tat activates NF-κB via physical interaction with IκB-α and p65.

Nuclear factor (NF)-κB is a master regulator of pro-inflammatory genes and is upregulated in human immunodeficiency virus 1 (HIV-1) infection. Mechanisms underlying the NF-κB deregulation by HIV-1 are relevant for immune dysfunction in AIDS. We report that in single round HIV-1 infection, or single-pulse PMA stimulation, the HIV-1 Tat transactivator activated NF-κB by hijacking the inhibitor IκB-α and by preventing the repressor binding to the NF-κB complex. Moreover, Tat associated with the p65 subunit of NF-κB and increased the p65 DNA-binding affinity and transcriptional activity. The arginine- and cysteine-rich domains of Tat were required for IκB-α and p65 association, respectively, and for sustaining the NF-κB activity. Among an array of NF-κB-responsive genes, Tat mostly activated the MIP-1α expression in a p65-dependent manner, and bound to the MIP-1α NF-κB enhancer thus promoting the recruitment of p65 with displacement of IκB-α; similar findings were obtained for the NF-κB-responsive genes CSF3, LTA, NFKBIA and TLR2. Our results support a novel mechanism of NF-κB activation via physical interaction of Tat with IκB-α and p65, and may contribute to further insights into the deregulation of the inflammatory response by HIV-1.

Enhanced pro-apoptotic activity of rituximab through IBTK silencing in non-Hodgkin lymphoma B-cells.

Rituximab is a commonly used chemotherapeutic drug for patients with aggressive lymphomas, such as non-Hodgkin's lymphoma (NHL). Currently, the combination of Rituximab and chemotherapy (R-CHOP) stands as the most prevalent first-line therapy for NHL. Nevertheless, the development of new therapeutic approaches remains imperative. An increasing body of evidence highlights a novel role for IBTK in tumorigenesis and cancer growth. In this study, we aim to broaden our understanding of IBTK's function in B-lymphoma, with a particular focus on its impact on the expression of the oncogene MYC. Here, we assessed the effects of combining Rituximab with IBTK silencing on cell viability through cell cycle analysis and Annexin V assays in vitro. Furthermore, we leveraged the transplantability of Eµ-myc lymphomas to investigate whether the inhibition of IBTK could elicit anti-tumor effects in the treatment of lymphomas in vivo. Our data suggests that IBTK silencing may serve as an effective anti-tumor agent for aggressive B-Lymphomas, underscoring its role in promoting apoptosis when used in combination with Rituximab, both in in vitro and in vivo settings.