Targeting G-quadruplex motifs interferes with differentiation of adipose-derived mesenchymal stem cells.

BACKGROUND: G-quadruplex (G4) motifs are nucleic acid secondary structures observed in mammalian genomes and transcriptomes able to regulate various cellular processes. Several small molecules have been developed so far to modulate G4 stability, frequently associated with anticancer activity. However, how G4 structures are regulated over homeostatic conditions is mostly unexplored. Here, we used human adipose-derived mesenchymal stem cells (ASCs) to address the role of G4 motifs during adipogenic differentiation. METHODS: Adipocyte differentiation of ASCs was investigated in the presence or absence of a well-known G4 ligand, Braco-19. Cell viability was determined by sulforhodamine B assay. Cell dimension and granularity, DNA G4 motifs and cell cycle were detected by flow cytometry. Lipid droplet accumulation was assessed by Oil Red O staining. Cell senescence was evaluated by ß-galactosidase staining. Gene expression was measured by qPCR. Protein release in the extracellular medium was quantified by ELISA. RESULTS: Braco-19 used at non-cytotoxic concentrations induced morphological changes in mature adipocytes partially restoring an undifferentiated-like status. Braco-19 reduced lipid vacuolization and PPARG, AP2, LEP and TNFA mRNA levels in terminally differentiated cells. No effect was observed in cell senescence, fibrotic markers, IL-6 and IL-8 production, while the secretion of VEGF was dose-dependently reduced. Interestingly, G4 structures were increased in differentiated adipocytes compared to their precursors. Braco-19 treatment reduced G4 content in mature adipocytes. CONCLUSIONS: Our data highlight a new role of G4 motifs as genomic structural elements related to human ASC differentiation into mature adipocytes, with potential implications in physio-pathological processes.

Low Interferon-γ Levels in Cord and Peripheral Blood of Pregnant Women Infected with SARS-CoV-2.

COVID-19 is characterized by the immune system's overreaction resulting in a 'cytokine storm', consisting in a massive release of cytokine into the bloodstream, leading to local and systemic inflammatory response. This clinical picture is further complicated in case of infection of patients with a peculiar immunological status, such as pregnancy. In this paper, we focused on Interferon-γ, which plays a pivotal immunomodulatory role in normal pregnancy and fetal development, as well as in defense against pathogens. In this study, we compared the levels of Interferon-γ and the Interferon autoantibodies of the peripheral and cord blood of pregnant women with confirmed mild COVID-19 and healthy pregnant women. The Interferon-γ was significantly lower both in the peripheral and cord blood of SARS-CoV-2-positive mothers, suggesting that infection can affect the fetal microenvironment even without severe maternal symptoms. In conclusion, further studies are needed to clarify whether lower levels of Interferon-γ due to SARS-CoV-2 infection affect the development or infection susceptibility of infants born to SARS-CoV-2-infected mothers.

Combination of dl922-947 Oncolytic Adenovirus and G-Quadruplex Binders Uncovers Improved Antitumor Activity in Breast Cancer.

G-quadruplexes (G4s) are nucleic secondary structures characterized by G-tetrads. G4 motif stabilization induces DNA damage and cancer cell death; therefore, G4-targeting small molecules are the focus of clinical investigation. DNA destabilization induced by G4 ligands might potentiate the anticancer activity of agents targeting DNA or inhibiting its repair such as oncolytic viruses. This study represents the first approach combining G4 ligands, BRACO-19 (B19), pyridostatin (PDS), and the adenovirus dl922-947 in breast cancer cells. We demonstrated that G4 binders and dl922-947 induce cytotoxicity in breast cancer cells (MDA-MB-231 and MCF-7) and at higher doses in other neoplastic cell lines of thyroid (BHT-101 cells) and prostate (PC3 cells). G4 binders induce G4 motifs distributed in the S and G2/M phases in MCF-7 cells. G4 binder/dl922-947 combination increases cell cytotoxicity and the accumulation in subG0/G1. Indeed, G4 binders favor viral entry and replication with no effect on coxsackie and adenovirus receptor. Notably, dl922-947 induces G4 motifs and its combination with PDS potentiates this effect in MCF-7 cells. The agents alone or in combination similarly enhanced cell senescence. Additionally, PDS/dl922-947 combination inactivates STING signaling in MDA-MB-231 cells. Our results suggest that G4 binder/virotherapy combination may represent a novel therapeutic anticancer approach.

Synthesis and In Vitro Characterization of Selective Cannabinoid CB2 Receptor Agonists: Biological Evaluation against Neuroblastoma Cancer Cells.

1,8-naphthyridine-3-carboxamide structures were previously identified as a promising scaffold from which to obtain CB2R agonists with anticancer and anti-inflammatory activity. This work describes the synthesis and functional characterization of new 1,8-naphthyridin-2(1H)-one-3-carboxamides with high affinity and selectivity for CB2R. The new compounds were able to pharmacologically modulate the cAMP response without modulating CB2R-dependent ß-arrestin2 recruitment. These structures were also evaluated for their anti-cancer activity against SH-SY5Y and SK-N-BE cells. They were able to reduce the cell viability of both neuroblastoma cancer cell lines with micromolar potency (IC50 of FG158a = 11.8 µM and FG160a = 13.2 µM in SH-SY5Y cells) by a CB2R-mediated mechanism. Finally, in SH-SY5Y cells one of the newly synthesized compounds, FG158a, was able to modulate ERK1/2 expression by a CB2R-mediated effect, thus suggesting that this signaling pathway might be involved in its potential anti-cancer effect.

Humoral Response to 2-dose BNT162b2 mRNA COVID-19 Vaccination in Liver Transplant Recipients.

BACKGROUND & AIMS: In the context of the Italian severe acute respiratory syndrome coronavirus 2 vaccination program, liver transplant (LT) recipients were prioritized for vaccine administration, although the lower response to vaccines is a well-known problem in this population. We aimed to evaluate immunogenicity of BNT162b2 mRNA vaccine in LT recipients and healthy controls and to identify factors associated with negative response to vaccine. METHODS: In a cohort of adult patients with LT, we prospectively evaluated the humoral response (with anti-Spike protein IgG-LIAISON SARS-CoV-2 S1/S2-IgG chemiluminescent assay) at 1 and 3 months after 2-dose vaccination. A group of 307 vaccinated health care workers, matched by age and sex, served as controls. RESULTS: Overall, 492 LT patients were enrolled (75.41% male; median age, 64.85 years). Detectable antibodies were observed in the 75% of patients, with a median value of 73.9 AU/mL after 3 months from 2-dose vaccination. At multivariable analysis, older age (>40 years; P = .016), shorter time from liver transplantation (<5 years; P = .004), and immunosuppression with antimetabolites (P = .029) were significantly associated with non-response to vaccination. Moreover, the LT recipients showed antibody titers statistically lower than the control group (103 vs 261 AU/mL; P < .0001). Finally, in both controls and LT patients, we found a trend of inverse correlation between age and antibody titers (correlation coefficients: -0.2023 and -0.2345, respectively). CONCLUSIONS: Three months after vaccination, LT recipients showed humoral response in 75% of cases. Older age, shorter time from transplantation, and use of antimetabolites were factors associated with non-response to vaccination, and LT recipients at risk of non-response to vaccination needed to be kept under close monitoring.

Cross Talk of Macrophages with Tumor Microenvironment Cells and Modulation of Macrophages in Cancer by Virotherapy.

Cellular compartments constituting the tumor microenvironment including immune cells, fibroblasts, endothelial cells, and mesenchymal stromal/stem cells communicate with malignant cells to orchestrate a series of signals that contribute to the evolution of the tumor microenvironment. In this study, we will focus on the interplay in tumor microenvironment between macrophages and mesenchymal stem cells and macrophages and fibroblasts. In particular, cell-cell interaction and mediators secreted by these cells will be examined to explain pro/anti-tumor phenotypes induced in macrophages. Nonetheless, in the context of virotherapy, the response of macrophages as a consequence of treatment with oncolytic viruses will be analyzed regarding their polarization status and their pro/anti-tumor response.

Self-assembly of bio-inspired heterochiral peptides.

Peptide hydrogels, deriving from natural protein fragments, present unique advantages as compatibility and low cost of production that allow their wide application in different fields as wound healing, cell delivery and tissue regeneration. To engineer new biomaterials, the change of the chirality of single amino acids demonstrated a powerful approach to modulate the self-assembly mechanism. Recently we unveiled that a small stretch spanning residues 268-273 in the C-terminal domain (CTD) of Nucleophosmin 1 (NPM1) is an amyloid sequence. Herein, we performed a systematic D-scan of this sequence and analyzed the structural properties of obtained peptides. The conformational and kinetic features of self-aggregates and the morphologies of derived microstructures were investigated by means of different biophysical techniques, as well as the compatibility of hydrogels was evaluated in HeLa cells. All the investigated hexapeptides formed hydrogels even if they exhibited different conformational intermediates during aggregation, and they structural featured are finely tuned by introduced chiralities.

Pharmacological Inhibition of WEE1 Potentiates the Antitumoral Effect of the dl922-947 Oncolytic Virus in Malignant Mesothelioma Cell Lines.

Malignant mesothelioma (MM) is a very aggressive asbestos-related cancer, for which no therapy proves to be effective. We have recently shown that the oncolytic adenovirus dl922-947 had antitumor effects in MM cell lines and murine xenografts. Previous studies demonstrated that dl922-947-induced host cell cycle checkpoint deregulation and consequent DNA lesions associated with the virus efficacy. However, the cellular DNA damage response (DDR) can counteract this virus action. Therefore, we assessed whether AZD1775, an inhibitor of the G2/M DNA damage checkpoint kinase WEE1, could enhance MM cell sensitivity to dl922-947. Through cell viability assays, we found that AZD1775 synergized with dl922-947 selectively in MM cell lines and increased dl922-947-induced cell death, which showed hallmarks of apoptosis (annexinV-positivity, caspase-dependency, BCL-XL decrease, chromatin condensation). Predictably, dl922-947 and/or AZD1775 activated the DDR, as indicated by increased levels of three main DDR players: phosphorylated histone H2AX (γ-H2AX), phospho-replication protein A (RPA)32, phospho-checkpoint kinase 1 (CHK1). Dl922-947 also increased inactive Tyr-15-phosphorylated cyclin-dependent kinase 1 (CDK1), a key WEE1 substrate, which is indicative of G2/M checkpoint activation. This increase in phospho-CDK1 was effectively suppressed by AZD1775, thus suggesting that this compound could, indeed, abrogate the dl922-947-induced DNA damage checkpoint in MM cells. Overall, our data suggest that the dl922-947-AZD1775 combination could be a feasible strategy against MM.

Proteostasis unbalance of nucleophosmin 1 in Acute Myeloid Leukemia: An aggregomic perspective.

The role exerted by the nucleus in the regulation of proteostasis in both health and disease is recognized of outmost importance, even though not fully understood. Many recent investigations are focused on its ability to modulate and coordinate protein quality control machineries in mammalian cells. Nucleophosmin 1 (NPM1) is one of the most abundant nucleolar proteins and its gene is mutated in ~30% of Acute Myeloid Leukemia (AML) patients. Mutations are localized in the C-terminal domain of the protein and cause cytoplasmatically delocalized and possibly aggregated forms of NPM1 (NPM1c+). Therapeutic interventions targeted on NPM1c+ are in demand and, to this end, deeper knowledge of NPM1c+ behavior in the blasts' cytosol is required. Here by means of complementary biophysical techniques we compared the conformational and aggregative behavior of the entire C-terminal domains of NPM1wt and type A NPM1c+ (bearing the most common mutation). Overall data show that only Cterm_mutA is able to form amyloid-like assemblies with fibrillar morphology and that the oligomers are toxic in human neuroblastoma SHSY cells. This study adds a novel piece of knowledge to the comprehension of the molecular roles exerted by cytoplasmatic NPM1c+ and suggests the exploitation of the amyloidogenic propensity of NPM1c+ as a new strategy for targeting AML with NPM1 mutations.

Tumor-Associated Macrophage Status in Cancer Treatment.

Tumor-associated macrophages (TAMs) represent the most abundant innate immune cells in tumors. TAMs, exhibiting anti-inflammatory phenotype, are key players in cancer progression, metastasis and resistance to therapy. A high TAM infiltration is generally associated with poor prognosis, but macrophages are highly plastic cells that can adopt either proinflammatory/antitumor or anti-inflammatory/protumor features in response to tumor microenvironment stimuli. In the context of cancer therapy, many anticancer therapeutics, apart from their direct effect on tumor cells, display different effects on TAM activation status and density. In this review, we aim to evaluate the indirect effects of anticancer therapies in the modulation of TAM phenotypes and pro/antitumor activity.

Virotherapy: From single agents to combinatorial treatments.

Virotherpay is emerging as a promising strategy against cancer, and three oncolytic viruses (OVs) have gained approval in different countries for the treatment of several cancer types. Beyond the capability to selectively infect, replicate and lyse cancer cells, OVs act through a multitude of events, including modification of the tumour micro/macro-environment as well as a complex modulation of the anti-tumour immune response by activation of danger signals and immunogenic cell death pathways. Most OVs show limited effects, depending on the viral platform and the interactions with the host. OVs used as monotherapy only in a minority of patients elicited a full response. Better outcomes were obtained using OVs in combination with other treatments, such as immune therapy or chemotherapy, suggesting that the full potential of OVs can be unleashed in combination with other treatment modalities. Here, we report the main described combination of OVs with conventional chemotherapeutic agents: platinum salts, mitotic inhibitors, anthracyclines and other antibiotics, anti-metabolites, alkylating agents and topoisomerase inhibitors. Additionally, our work provides an overview of OV combination with targeted therapies: histone deacetylase inhibitors, kinase inhibitors, monoclonal antibodies, inhibitors of DNA repair, inhibitors of the proteasome complex and statins that demonstrated enhanced OV anti-neoplastic activity. Although further studies are required to assess the best combinations to translate the results in the clinic, it is clear that combined therapies, acting with complementary mechanisms of action might be useful to target cancer lesions resistant to currently available treatments.

Reply to Comment on "Malfitano, A.M. et al. Virotherapy as a Potential Therapeutic Approach for the Treatment of Aggressive Thyroid Cancer" Cancers 2019, 11, 1532.

Our work is focused on the future clinical use of oncolytic viruses (OVs) for the treatment of aggressive thyroid carcinomas. Therefore, we provide a brief description of the overall use of OVs in the clinic. Rigvir is among the few OVs that have already been used for the treatment of patients, and studies describing its effects have been briefly commented and cited in our text [1]. [...].

G-Quadruplex Binders Induce Immunogenic Cell Death Markers in Aggressive Breast Cancer Cells.

BACKGROUND: DNA G-quadruplex (G4) structures represent potential anti-cancer targets. In this study, we compared the effect of two G4-targeting compounds, C066-3108 and the gold standard BRACO-19. METHODS: In breast and prostate cancer cells, cytotoxicity induced by both molecules was measured by a sulforhodamine B assay. In breast cancer cells, cycle, apoptosis, the formation of G4 structures, calreticulin and high mobility group box 1 (HMGB1), as well as T cell activation, were analyzed by flow cytometry and adenosine triphosphate (ATP) by luminescence. RESULTS: Both ligands inhibited cell survival and induced DNA damage. In MCF-7 cells, G4 ligands increased the subG0/G1 phase of the cell cycle inducing apoptosis and reduced intracellular ATP. In untreated MCF-7 cells, we observed a slight presence of G4 structures associated with the G2/M phase. In MDA-MB231 cells, G4 ligands decreased the G1 and enhanced the G2/M phase. We observed a decrease of intracellular ATP, calreticulin cell surface exposure and an increase of HMGB1, accompanied by T cell activation. Both compounds induced G4 structure formation in the subG0/G1 phase. CONCLUSIONS: Our data report similar effects for both compounds and the first evidence that G4 ligands induce the release of danger signals associated with immunogenic cell death and induction of T cell activation.

The Oncolytic Virus dl922-947 Triggers Immunogenic Cell Death in Mesothelioma and Reduces Xenograft Growth.

Background: Malignant pleural mesothelioma (MPM) is an aggressive cancer associated with asbestos exposure that urgently requires effective therapeutic strategies. Current treatments are unable to increase significantly patient survival, which is often limited to <1 year from diagnosis. Virotherapy, based on the use of oncolytic viruses that exert anti-cancer effects by direct cell lysis and through the induction of anti-tumor immune response, represents an alternative therapeutic option for rare tumors with limited life expectancy. In this study, we propose the use of the adenovirus dl922-947, engineered to allow selective replication in cancer cells, to counteract MPM. Methods: We performed a thorough preclinical assessment of dl922-947 effects in a set of MPM cell lines and xenografts. Cytotoxicity of dl922-947 alone and in combination assays was evaluated by sulforhodamine B assay. Cell cycle, calreticulin expression, and high mobility group box protein 1 (HMGB1) secretion were determined by flow cytometry, whereas ATP content was determined by a luminescence-based bioassay. The modulation of angiogenic factors in MPM-infected cells was evaluated through ELISA. Results: We found that dl922-947 infection exhibits cytotoxic effects in MPM cell lines, affecting cell viability, cell cycle progression, and regulating main hallmarks of immunogenic cell death inducing calreticulin surface exposure, HMGB1 and ATP release. Our results also suggest that dl922-947 may affect angiogenic signals by regulation of VEGF-A and IL-8 secretion. Furthermore, dl922-947 shows anti-tumor efficacy in murine xenograft models reducing tumor growth and enhancing survival. Finally, the combination with cisplatin potentiated the cytotoxic effect of dl922-947. Conclusions: Overall our data identify virotherapy, based on the use of dl922-947, as a new possible therapeutic strategy against MPM, which could be used alone, in combination with standard chemotherapy drugs, as shown here, or other approaches also aimed at enhancing the antitumoral immune response elicited by the virus.

Platinum(II) O,S Complexes Inhibit the Aggregation of Amyloid Model Systems.

Platinum(II) complexes with different cinnamic acid derivatives as ligands were investigated for their ability to inhibit the aggregation process of amyloid systems derived from Aß, Yeast Prion Protein Sup35p and the C-terminal domain of nucleophosmin 1. Thioflavin T binding assays and circular dichroism data indicate that these compounds strongly inhibit the aggregation of investigated peptides exhibiting IC50 values in the micromolar range. MS analysis confirms the formation of adducts between peptides and Pt(II) complexes that are also able to reduce amyloid cytotoxicity in human SH-SY5Y neuroblastoma cells. Overall data suggests that bidentate ligands based on ß-hydroxy dithiocinnamic esters can be used to develop platinum or platinoid compounds with anti-amyloid aggregation properties.

Structural insights into amyloid structures of the C-terminal region of nucleophosmin 1 in type A mutation of acute myeloid leukemia.

Acute myeloid leukemia (AML) is a clinically and a molecularly heterogeneous disease characterized by the accumulation of undifferentiated and uncontrolled proliferation of hematopoietic progenitor cells. The sub-group named "AML with gene mutations" includes mutations in nucleophosmin (NPM1) assumed as a distinct leukemic entity. NPM1 is an abundant multifunctional protein belonging to the nucleoplasmin family of nuclear chaperones. AML mutated protein is translocated into the cytoplasm (NPM1c+) retaining all functional domains except the loss of a unique NoLs (nucleolar localization signal) at the C-term domain (CTD) and the subsequent disruption of a three helix bundle as tertiary structure. The oligomeric state of NPM1 is of outmost importance for its biological roles and our previous studies linked an aggregation propensity of distinct regions of CTD to leukomogenic potentials of AML mutations. Here we investigated a polypeptide spanning the third and second helices of the bundle of type A mutated CTD. By a combination of several techniques, we ascertained the amyloid character of the aggregates and of fibrils resulting from a self-recognition mechanism. Further amyloid assemblies resulted cytoxic in MTT assay strengthening a new idea of a therapeutic strategy in AML consisting in the self-degradation of mutated NPM1.

Potential involvement of neutrophils in human thyroid cancer.

BACKGROUND: Neutrophil functions have long been regarded as limited to acute inflammation and the defense against microbes. The role(s) of neutrophils in cancer remain poorly understood. Neutrophils infiltrate tumors and are key effector cells in the orchestration of inflammatory responses. Thyroid cancer (TC) is the most recurrent endocrine malignant tumor and is responsible for 70% of deaths due to endocrine cancers. No studies are so far available on the role of neutrophils in TC. OBJECTIVE: Our purpose was to study the involvement of tumor-associated neutrophils in TC. METHODS: Highly purified human neutrophils (>99%) from healthy donors were stimulated in vitro with conditioned media derived from TC cell lines TPC1 and 8505c (TC-CMs). Neutrophil functions (e.g., chemotaxis, activation, plasticity, survival, gene expression, and protein release) were evaluated. RESULTS: TC-derived soluble factors promoted neutrophil chemotaxis and survival. Neutrophil chemotaxis toward a TC-CM was mediated, at least in part, by CXCL8/IL-8, and survival was mediated by granulocyte-macrophage colony-stimulating factor (GM-CSF). In addition, each TC-CM induced morphological changes and activation of neutrophils (e.g., CD11b and CD66b upregulation and CD62L shedding) and modified neutrophils' kinetic properties. Furthermore, each TC-CM induced production of reactive oxygen species, expression of proinflammatory and angiogenic mediators (CXCL8/IL-8, VEGF-A, and TNF-α), and a release of matrix metalloproteinase 9 (MMP-9). Moreover, in TC patients, tumor-associated neutrophils correlated with larger tumor size. CONCLUSIONS: TC cell lines produce soluble factors able to "educate" neutrophils toward an activated functional state. These data will advance the understanding of the molecular and cellular mechanisms of innate immunity in TC.

Lipopolysaccharide-Elicited TSLPR Expression Enriches a Functionally Discrete Subset of Human CD14+ CD1c+ Monocytes.

Thymic stromal lymphopoietin (TSLP) is a cytokine produced mainly by epithelial cells in response to inflammatory or microbial stimuli and binds to the TSLP receptor (TSLPR) complex, a heterodimer composed of TSLPR and IL-7 receptor α (CD127). TSLP activates multiple immune cell subsets expressing the TSLPR complex and plays a role in several models of disease. Although human monocytes express TSLPR and CD127 mRNAs in response to the TLR4 agonist LPS, their responsiveness to TSLP is poorly defined. We demonstrate that TSLP enhances human CD14+ monocyte CCL17 production in response to LPS and IL-4. Surprisingly, only a subset of CD14+ CD16- monocytes, TSLPR+ monocytes (TSLPR+ mono), expresses TSLPR complex upon LPS stimulation in an NF-κB- and p38-dependent manner. Phenotypic, functional, and transcriptomic analysis revealed specific features of TSLPR+ mono, including higher CCL17 and IL-10 production and increased expression of genes with important immune functions (i.e., GAS6, ALOX15B, FCGR2B, LAIR1). Strikingly, TSLPR+ mono express higher levels of the dendritic cell marker CD1c. This evidence led us to identify a subset of peripheral blood CD14+ CD1c+ cells that expresses the highest levels of TSLPR upon LPS stimulation. The translational relevance of these findings is highlighted by the higher expression of TSLPR and CD127 mRNAs in monocytes isolated from patients with Gram-negative sepsis compared with healthy control subjects. Our results emphasize a phenotypic and functional heterogeneity in an apparently homogeneous population of human CD14+ CD16- monocytes and prompt further ontogenetic and functional analysis of CD14+ CD1c+ and LPS-activated CD14+ CD1c+ TSLPR+ mono.

GM-CSF and IL-3 Modulate Human Monocyte TNF-α Production and Renewal in In Vitro Models of Trained Immunity.

GM-CSF and IL-3 are hematopoietic cytokines that also modulate the effector functions of several immune cell subsets. In particular, GM-CSF and IL-3 exert a significant control on monocyte and macrophage effector functions, as assessed in experimental models of inflammatory and autoimmune diseases and also in human studies. Here, we sought to investigate the mechanisms and the extent to which GM-CSF and IL-3 modulate the pro-inflammatory, LPS-mediated, activation of human CD14+ monocytes taking into account the new concept of trained immunity (i.e., the priming stimulus modulates the response to subsequent stimuli mainly by inducing chromatin remodeling and increased transcription at relevant genetic loci). We demonstrate that GM-CSF and IL-3 priming enhances TNF-α production upon subsequent LPS stimulation (short-term model of trained immunity) in a p38- and SIRT2-dependent manner without increasing TNF primary transcript levels (a more direct measure of transcription), thus supporting a posttranscriptional regulation of TNF-α in primed monocytes. GM-CSF and IL-3 priming followed by 6 days of resting also results in increased TNF-α production upon LPS stimulation (long-term model of trained immunity). In this case, however, GM-CSF and IL-3 priming induces a c-Myc-dependent monocyte renewal and increase in cell number that is in turn responsible for heightened TNF-α production. Overall, our results provide insights to understand the biology of monocytes in health and disease conditions in which the hematopoietic cytokines GM-CSF and IL-3 play a role and also extend our knowledge of the cellular and molecular mechanisms of trained immunity.

The oncolytic virus dl922-947 reduces IL-8/CXCL8 and MCP-1/CCL2 expression and impairs angiogenesis and macrophage infiltration in anaplastic thyroid carcinoma.

Anaplastic thyroid carcinoma (ATC) is one of the most aggressive human solid tumor and current treatments are ineffective in increasing patients' survival. Thus, the development of new therapeutic approaches for ATC is needed. We have previously shown that the oncolytic adenovirus dl922-947 induces ATC cell death in vitro and tumor regression in vivo. However, the impact of dl922-947 on the pro-tumorigenic ATC microenvironment is still unknown. Since viruses are able to regulate cytokine and chemokine production from infected cells, we sought to investigate whether dl922-947 virotherapy has such effect on ATC cells, thereby modulating ATC microenvironment. dl922-947 decreased IL-8/CXCL8 and MCP-1/CCL2 production by the ATC cell lines 8505-c and BHT101-5. These results correlated with dl922-947-mediated reduction of NF-κB p65 binding to IL8 promoter in 8505-c and BHT101-5 cells and CCL2 promoter in 8505-c cells. IL-8 stimulates cancer cell proliferation, survival and invasion, and also angiogenesis. dl922-947-mediated reduction of IL-8 impaired ATC cell motility in vitro and ATC-induced angiogenesis in vitro and in vivo. We also show that dl922-947-mediated reduction of the monocyte-attracting chemokine CCL2 decreased monocyte chemotaxis in vitro and tumor macrophage density in vivo. Interestingly, dl922-947 treatment induced the switch of tumor macrophages toward a pro-inflammatory M1 phenotype, likely by increasing the expression of the pro-inflammatory cytokine interferon-γ. Altogether, we demonstrate that dl922-947 treatment re-shape the pro-tumorigenic ATC microenvironment by modulating cancer-cell intrinsic factors and the immune response. An in-depth knowledge of dl922-947-mediated effects on ATC microenvironment may help to refine ATC virotherapy in the context of cancer immunotherapy.