NK cells with adhesion defects and reduced cytotoxic functions are associated with a poor prognosis in Multiple Myeloma.

The promising results obtained with immunotherapeutic approaches for multiple myeloma (MM) call for a better stratification of patients based on immune components. The most pressing being cytotoxic lymphocytes such as Natural Killer (NK) cells that are mandatory for MM surveillance and therapy. In this study, we performed a single cell RNA sequencing analysis of NK cells from 10 MM patients and 10 age/sex matched healthy donors (HD) that revealed important transcriptomic changes in NK cell landscape affecting both the bone marrow and peripheral blood compartment. The frequency of mature cytotoxic "CD56dim" NK cell subsets was reduced in MM patients at the advantage of late-stage NK cell subsets expressing NFB and IFN-I inflammatory signatures. These NK cell subsets accumulating in MM patients were characterized by a low CD16 and CD226 expression and poor cytotoxic functions. MM CD16/CD226Lo NK cells also had adhesion defects with reduced LFA-1 integrin activation and actin polymerization that may account for their limited effector functions in vitro. Finally, analysis of BM infiltrating NK cells in a retrospective cohort of 177 MM patients from the IFM 2009 trial demonstrated that a high frequency of NK cells and their low CD16 and CD226 expression were associated with a shorter overall survival. Thus, CD16/CD226Lo NK cells with reduced effector functions accumulate along MM development and negatively impact patients' clinical outcome. Given the growing interest in harnessing NK cells to treat myeloma, this improved knowledge around MM-associated NK cell dysfunction will stimulate the development of more efficient immunotherapeutic drugs against MM.

Between hope and reality: treatment of genetic diseases through nucleic acid-based drugs.

Rare diseases (RD) affect a small number of people compared to the general population and are mostly genetic in origin. The first clinical signs often appear at birth or in childhood, and patients endure high levels of pain and progressive loss of autonomy frequently associated with short life expectancy. Until recently, the low prevalence of RD and the gatekeeping delay in their diagnosis have long hampered research. The era of nucleic acid (NA)-based therapies has revolutionized the landscape of RD treatment and new hopes arise with the perspectives of disease-modifying drugs development as some NA-based therapies are now entering the clinical stage. Herein, we review NA-based drugs that were approved and are currently under investigation for the treatment of RD. We also discuss the recent structural improvements of NA-based therapeutics and delivery system, which overcome the main limitations in their market expansion and the current approaches that are developed to address the endosomal escape issue. We finally open the discussion on the ethical and societal issues that raise this new technology in terms of regulatory approval and sustainability of production.

TCTP regulates genotoxic stress and tumorigenicity via intercellular vesicular signaling.

Oncogenic intercellular signaling is regulated by extracellular vesicles (EVs), but the underlying mechanisms remain mostly unclear. Since TCTP (translationally controlled tumor protein) is an EV component, we investigated whether it has a role in genotoxic stress signaling and malignant transformation. By generating a Tctp-inducible knockout mouse model (Tctp-/f-), we report that Tctp is required for genotoxic stress-induced apoptosis signaling via small EVs (sEVs). Human breast cancer cells knocked-down for TCTP show impaired spontaneous EV secretion, thereby reducing sEV-dependent malignant growth. Since Trp53-/- mice are prone to tumor formation, we derived tumor cells from Trp53-/-;Tctp-/f- double mutant mice and describe a drastic decrease in tumori-genicity with concomitant decrease in sEV secretion and content. Remarkably, Trp53-/-;Tctp-/f- mice show highly prolonged survival. Treatment of Trp53-/- mice with sertraline, which inhibits TCTP function, increases their survival. Mechanistically, TCTP binds DDX3, recruiting RNAs, including miRNAs, to sEVs. Our findings establish TCTP as an essential protagonist in the regulation of sEV-signaling in the context of apoptosis and tumorigenicity.

Castration-Resistant Prostate Cancer: From Uncovered Resistance Mechanisms to Current Treatments.

Prostate cancer (PC) is the second most common cancer in men worldwide. Despite recent advances in diagnosis and treatment, castration-resistant prostate cancer (CRPC) remains a significant medical challenge. Prostate cancer cells can develop mechanisms to resist androgen deprivation therapy, such as AR overexpression, AR mutations, alterations in AR coregulators, increased steroidogenic signaling pathways, outlaw pathways, and bypass pathways. Various treatment options for CRPC exist, including androgen deprivation therapy, chemotherapy, immunotherapy, localized or systemic therapeutic radiation, and PARP inhibitors. However, more research is needed to combat CRPC effectively. Further investigation into the underlying mechanisms of the disease and the development of new therapeutic strategies will be crucial in improving patient outcomes. The present work summarizes the current knowledge regarding the underlying mechanisms that promote CRPC, including both AR-dependent and independent pathways. Additionally, we provide an overview of the currently approved therapeutic options for CRPC, with special emphasis on chemotherapy, radiation therapy, immunotherapy, PARP inhibitors, and potential combination strategies.

New 2,4-bis[(substituted-aminomethyl)phenyl]phenylquinazoline and 2,4-bis[(substituted-aminomethyl)phenyl]phenylquinoline Derivatives: Synthesis and Biological Evaluation as Novel Anticancer Agents by Targeting G-Quadruplex.

The syntheses of novel 2,4-bis[(substituted-aminomethyl)phenyl]phenylquinazolines 12 and 2,4-bis[(substituted-aminomethyl)phenyl]phenylquinolines 13 are reported here in six steps starting from various halogeno-quinazoline-2,4-(1H,3H)-diones or substituted anilines. The antiproliferative activities of the products were determined in vitro against a panel of breast (MCF-7 and MDA-MB-231), human adherent cervical (HeLa and SiHa), and ovarian (A2780) cell lines. Disubstituted 6- and 7-phenyl-bis(3-dimethylaminopropyl)aminomethylphenyl-quinazolines 12b, 12f, and 12i displayed the most interesting antiproliferative activities against six human cancer cell lines. In the series of quinoline derivatives, 6-phenyl-bis(3-dimethylaminopropyl)aminomethylphenylquinoline 13a proved to be the most active. G-quadruplexes (G4) stacked non-canonical nucleic acid structures found in specific G-rich DNA, or RNA sequences in the human genome are considered as potential targets for the development of anticancer agents. Then, as small aza-organic heterocyclic derivatives are well known to target and stabilize G4 structures, their ability to bind G4 structures have been determined through FRET melting, circular dichroism, and native mass spectrometry assays. Finally, telomerase inhibition ability has been also assessed using the MCF-7 cell line.

MYC and Twist1 cooperate to drive metastasis by eliciting crosstalk between cancer and innate immunity.

Metastasis is a major cause of cancer mortality. We generated an autochthonous transgenic mouse model whereby conditional expression of MYC and Twist1 enables hepatocellular carcinoma (HCC) to metastasize in >90% of mice. MYC and Twist1 cooperate and their sustained expression is required to elicit a transcriptional program associated with the activation of innate immunity, through secretion of a cytokinome that elicits recruitment and polarization of tumor associated macrophages (TAMs). Systemic treatment with Ccl2 and Il13 induced MYC-HCCs to metastasize; whereas, blockade of Ccl2 and Il13 abrogated MYC/Twist1-HCC metastasis. Further, in 33 human cancers (n = 9502) MYC and TWIST1 predict poor survival (p=4.3×10-10), CCL2/IL13 expression (p<10-109) and TAM infiltration (p<10-96). Finally, in the plasma of patients with HCC (n = 25) but not cirrhosis (n = 10), CCL2 and IL13 were increased and IL13 predicted invasive tumors. Therefore, MYC and TWIST1 generally appear to cooperate in human cancer to elicit a cytokinome that enables metastasis through crosstalk between cancer and immune microenvironment.

Lipid nanoparticles that deliver IL-12 messenger RNA suppress tumorigenesis in MYC oncogene-driven hepatocellular carcinoma.

Interleukin-12 (IL-12) is a promising candidate for cancer immunotherapy because of its ability to activate a number of host immune subsets that recognize and destroy cancer cells. We found that human hepatocellular carcinoma (HCC) patients with higher than median levels of IL-12 have significantly favorable clinical outcomes. Here, we report that a messenger RNA (mRNA) lipid nanoparticle delivering IL-12 (IL-12-LNP) slows down the progression of MYC oncogene-driven HCC. IL-12-LNP was well distributed within the HCC tumor and was not associated with significant animal toxicity. Treatment with IL-12-LNP significantly reduced liver tumor burden measured by dynamic magnetic resonance imaging (MRI), and increased survival of MYC-induced HCC transgenic mice in comparison to control mice. Importantly, IL-12-LNP exhibited no effect on transgenic MYC levels confirming that its therapeutic efficacy was not related to the downregulation of a driver oncogene. IL-12-LNP elicited marked infiltration of activated CD44+ CD3+ CD4+ T helper cells into the tumor, and increased the production of Interferon γ (IFNγ). Collectively, our findings suggest that IL-12-LNP administration may be an effective immunotherapy against HCC.

The MYC oncogene is a global regulator of the immune response.

The MYC proto-oncogene is a gene product that coordinates the transcriptional regulation of a multitude of genes that are essential to cellular programs required for normal as well as neoplastic cellular growth and proliferation, including cell cycle, self-renewal, survival, cell growth, metabolism, protein and ribosomal biogenesis, and differentiation. Here, we propose that MYC regulates these programs in a manner that is coordinated with a global influence on the host immune response. MYC had been presumed to contribute to tumorigenesis through tumor cell-intrinsic influences. More recently, MYC expression in tumor cells has been shown to regulate the tumor microenvironment through effects on both innate and adaptive immune effector cells and immune regulatory cytokines. Then, MYC was shown to regulate the expression of the immune checkpoint gene products CD47 and programmed death-ligand 1. Similarly, other oncogenes, which are known to modulate MYC, have been shown to regulate immune checkpoints. Hence, MYC may generally prevent highly proliferative cells from eliciting an immune response. MYC-driven neoplastic cells have coopted this mechanism to bypass immune detection. Thus, MYC inactivation can restore the immune response against a tumor. MYC-induced tumors may be particularly sensitive to immuno-oncology therapeutic interventions.

Anti-miR-17 therapy delays tumorigenesis in MYC-driven hepatocellular carcinoma (HCC).

Hepatocellular carcinoma (HCC) remains a significant clinical challenge with few therapeutic options. Genomic amplification and/or overexpression of the MYC oncogene is a common molecular event in HCC, thus making it an attractive target for drug therapy. Unfortunately, currently there are no direct drug therapies against MYC. As an alternative strategy, microRNAs regulated by MYC may be downstream targets for therapeutic blockade. MiR-17 family is a microRNA family transcriptionally regulated by MYC and it is commonly overexpressed in human HCCs. In this study, we performed systemic delivery of a novel lipid nanoparticle (LNP) encapsulating an anti-miR-17 oligonucleotide in a conditional transgenic mouse model of MYC driven HCC. Treatment with anti-miR-17 in vivo, but not with a control anti-miRNA, resulted in significant de-repression of direct targets of miR-17, robust apoptosis, decreased proliferation and led to delayed tumorigenesis in MYC-driven HCCs. Global gene expression profiling revealed engagement of miR-17 target genes and inhibition of key transcriptional programs of MYC, including cell cycle progression and proliferation. Hence, anti-miR-17 is an effective therapy for MYC-driven HCC.

TCTP Has a Crucial Role in the Different Stages of Prostate Cancer Malignant Progression.

Prostate cancer (PC) is the second most common cause of cancer-related mortality in men in the western world after lung cancer. Many patients are not candidates for resection given the advanced stage of their cancer. The primary treatment for advanced PC is the castration therapy which supresses the production of androgens, hormone that promotes PC growth. Despite the efficiency of the castration therapy, most patients develop castration resistant disease which remains uncurable. Clearly, novel approaches are required to effectively treat castration resistant PC (CRPC). New strategies that identify the molecular mechanisms by which PC becomes resistant to conventional therapies may enable the identification of novel therapeutic targets that could improve clinical outcome. Recent studies have demonstrated the implication of TCTP's over-expression in PC and CRPC, and its role in resistance to treatment. TCTP's interaction with p53 and their negative feedback loop regulation have also been described to be causal for PC progression and invasion. A novel nanotherapy that inhibits TCTP has been developed as a new therapeutical strategy in CRPC. This chapter will highlight the role of TCTP as new therapeutic target in PC, in particular, therapy-resistant advanced PC and report the development of novel nanotherapy against TCTP that restore treatment-sensitivity in CRPC that deserve to be tested in clinical trial.

Lipid-oligonucleotide conjugates improve cellular uptake and efficiency of TCTP-antisense in castration-resistant prostate cancer.

Translationally controlled tumor protein (TCTP) has been implicated in a plethora of important cellular processes related to cell growth, cell cycle progression, malignant transformation and inhibition of apoptosis. Therefore, TCTP is now recognized as a potential therapeutic target in several cancers including prostate, breast and lung cancers. We previously showed that TCTP is overexpressed in castration-resistant prostate cancer (CRPC), and it has been implicated resistance to treatment. Recently, we developed TCTP antisense oligonucleotides (ASOs) to inhibit TCTP expression. However, the intracellular delivery and silencing activity of these oligonucleotides remains a challenge, and depend on the use of transfection agents and delivery systems. Here we show that lipid-modified ASO (LASOs) has improved penetration and efficiency in inhibiting TCTP expression in the absence of additional transfection agents, both in vitro and in vivo. Transfection with TCTP-LASO led to rapid and prolonged internalization via macropinocytosis, TCTP downregulation and significant decreased cell viability. We also show that lipid-modification led to delayed tumor progression in CRPC xenografts models, with no significant toxic effects observed.

MYC: Master Regulator of Immune Privilege.

Cancers are often initiated by genetic events that activate proto-oncogenes or inactivate tumor-suppressor genes. These events are also crucial for sustained tumor cell proliferation and survival, a phenomenon described as oncogene addiction. In addition to this cell-intrinsic role, recent evidence indicates that oncogenes also directly regulate immune responses, leading to immunosuppression. Expression of many oncogenes or loss of tumor suppressors induces the expression of immune checkpoints that regulate the immune response, such as PD-L1. We discuss here how oncogenes, and in particular MYC, suppress immune surveillance, and how oncogene-targeted therapies may restore the immune response against tumors.

Salmonella Typhimurium utilizes a T6SS-mediated antibacterial weapon to establish in the host gut.

The mammalian gastrointestinal tract is colonized by a high-density polymicrobial community where bacteria compete for niches and resources. One key competition strategy includes cell contact-dependent mechanisms of interbacterial antagonism, such as the type VI secretion system (T6SS), a multiprotein needle-like apparatus that injects effector proteins into prokaryotic and/or eukaryotic target cells. However, the contribution of T6SS antibacterial activity during pathogen invasion of the gut has not been demonstrated. We report that successful establishment in the gut by the enteropathogenic bacterium Salmonella enterica serovar Typhimurium requires a T6SS encoded within Salmonella pathogenicity island-6 (SPI-6). In an in vitro setting, we demonstrate that bile salts increase SPI-6 antibacterial activity and that S Typhimurium kills commensal bacteria in a T6SS-dependent manner. Furthermore, we provide evidence that one of the two T6SS nanotube subunits, Hcp1, is required for killing Klebsiella oxytoca in vitro and that this activity is mediated by the specific interaction of Hcp1 with the antibacterial amidase Tae4. Finally, we show that K. oxytoca is killed in the host gut in an Hcp1-dependent manner and that the T6SS antibacterial activity is essential for Salmonella to establish infection within the host gut. Our findings provide an example of pathogen T6SS-dependent killing of commensal bacteria as a mechanism to successfully colonize the host gut.

MYC regulates the antitumor immune response through CD47 and PD-L1.

The MYC oncogene codes for a transcription factor that is overexpressed in many human cancers. Here we show that MYC regulates the expression of two immune checkpoint proteins on the tumor cell surface: the innate immune regulator CD47 (cluster of differentiation 47) and the adaptive immune checkpoint PD-L1 (programmed death-ligand 1). Suppression of MYC in mouse tumors and human tumor cells caused a reduction in the levels of CD47 and PD-L1 messenger RNA and protein. MYC was found to bind directly to the promoters of the Cd47 and Pd-l1 genes. MYC inactivation in mouse tumors down-regulated CD47 and PD-L1 expression and enhanced the antitumor immune response. In contrast, when MYC was inactivated in tumors with enforced expression of CD47 or PD-L1, the immune response was suppressed, and tumors continued to grow. Thus, MYC appears to initiate and maintain tumorigenesis, in part, through the modulation of immune regulatory molecules.

The functional landscape of Hsp27 reveals new cellular processes such as DNA repair and alternative splicing and proposes novel anticancer targets.

Previously, we identified the stress-induced chaperone, Hsp27, as highly overexpressed in castration-resistant prostate cancer and developed an Hsp27 inhibitor (OGX-427) currently tested in phase I/II clinical trials as a chemosensitizing agent in different cancers. To better understand the Hsp27 poorly-defined cytoprotective functions in cancers and increase the OGX-427 pharmacological safety, we established the Hsp27-protein interaction network using a yeast two-hybrid approach and identified 226 interaction partners. As an example, we showed that targeting Hsp27 interaction with TCTP, a partner protein identified in our screen increases therapy sensitivity, opening a new promising field of research for therapeutic approaches that could decrease or abolish toxicity for normal cells. Results of an in-depth bioinformatics network analysis allying the Hsp27 interaction map into the human interactome underlined the multifunctional character of this protein. We identified interactions of Hsp27 with proteins involved in eight well known functions previously related to Hsp27 and uncovered 17 potential new ones, such as DNA repair and RNA splicing. Validation of Hsp27 involvement in both processes in human prostate cancer cells supports our system biology-predicted functions and provides new insights into Hsp27 roles in cancer cells.

TCTP as therapeutic target in cancers.

The translationally controlled tumor protein (TCTP) is a highly conserved protein present in eukaryotic organisms. This protein, located both in the cytoplasmic and the nucleus, is expressed in various tissues and is regulated in response to a wide range of extracellular stimuli. TCTP interacts with itself and other protein including MCL1 and p53. TCTP has been shown to play an important role in physiological events, such as cell proliferation, cell death and immune responses but also in stress response and tumor reversion. Moreover, TCTP expression is associated with malignancy and chemoresistance. In this review, we will evaluate pathways regulated by TCTP and current inhibitory strategy to target TCTP in cancerous diseases.

Hsp27 as a therapeutic target in cancers.

Heat shock protein 27 (Hsp27), induced by heat shock, environmental and pathophysiological stressors, is a multidimensional protein that acts as a protein chaperone and an antioxidant. This protein plays a major role in the inhibition of apoptosis and actin cytoskeletal remodeling. This stress-activated protein is up-regulated in many cancers and is associated with poor prognosis as well as treatment resistance by protecting cells from therapeutic agent that normally induces apoptosis. This review highlights the most recent findings and role of Hsp27 in cancer and the different strategies to target and inhibit Hsp27 for clinical purposes.

Targeting TCTP as a new therapeutic strategy in castration-resistant prostate cancer.

Heat shock protein 27 (Hsp27) is highly overexpressed in castration-resistant prostate cancer (CRPC) and an antisense inhibitor (OGX-427) is currently in phase II clinical trials. In order to understand mechanisms of action of Hsp27 and find new therapeutic targets specific of CRPC, we screened for Hsp27 client proteins. Here, we report that translationally controlled tumor protein (TCTP) is a new Hsp27 client protein involved in Hsp27 cytoprotection. We found that TCTP expression is absent or weak in normal prostate cells, moderately expressed in 18.5% of treatment naive PC, and becomes uniformly and strongly expressed in 75% of CRPC. To define TCTP function, we developed and worldwide patented a TCTP antisense oligonucleotide (ASO). Interestingly, we found that CRPC progression correlates with TCTP overexpression and loss of P53. TCTP knockdown restored P53 expression and function, suggesting that castration-sensitivity is directly linked to P53 expression. Collectively, these findings provide a new Hsp27 cytoprotection mechanism in CRPC, and preclinical proof-of-concept that combining ASO-mediated TCTP knockdown with castration and/or docetaxel therapy could serve as a novel strategy to treat CRPC, with no or little toxicity for normal prostate cells.

TP53INP1 as new therapeutic target in castration-resistant prostate cancer.

BACKGROUND: Prostate cancer (PC) is one of the most common malignancies in industrialized countries, and the second leading cause of cancer-related death in the United States. We recently showed that over-expression of tumor protein 53-induced nuclear protein 1 (TP53INP1), a cell stress response protein, is a worse prognostic factor in PC, particularly predictive of biological cancer relapse. Moreover, treatment of castration-sensitive (CS) LNCaP tumor cells with a TP53INP1 antisense oligonucleotide (TP53INP1 ASO) inhibits proliferation and induces apoptosis. The aim of this study was to investigate variations of TP53INP1 expression in PC during androgen withdrawal therapy and in castration-resistant prostate cancer (CRPC). METHODS: Quantitative measurements of immunohistochemical expression of TP53INP1 using high-throughput densitometry, assessed on digitized microscopic tissue micro-array images were correlated with hormone therapy (HT) status in human PC. Northern blot analysis of TP53INP1 after castration was performed in LNCaP xenograft. Treatment of CR C4-2 tumor cells in vitro with TP53INP1 ASO was analyzed. We also analyzed the effect of TP53INP1 ASO treatment in vivo on tumor xenograft growth. RESULTS: TP53INP1 protein expression decreases during HT and increases after HT in human CRPC. TP53INP1 mRNA increases significantly in CR tumors of LNCaP xenograft. Moreover, treatment of CR C4-2 cells with TP53INP1 ASO downregulates TP53INP1 protein level, inhibits proliferation, and induces apoptosis. Finally, in vivo, TP53INP1 ASO treatment significantly inhibits the tumoral progression of CR C4-2 xenograft and enhances docetaxel cytotoxicity. CONCLUSIONS: These results suggest that TP53INP1 could be considered as a relevant-specific target for molecular therapy of CRPC.