Oxidative stress enhances the therapeutic action of a respiratory inhibitor in MYC-driven lymphoma.

MYC is a key oncogenic driver in multiple tumor types, but concomitantly endows cancer cells with a series of vulnerabilities that provide opportunities for targeted pharmacological intervention. For example, drugs that suppress mitochondrial respiration selectively kill MYC-overexpressing cells. Here, we unravel the mechanistic basis for this synthetic lethal interaction and exploit it to improve the anticancer effects of the respiratory complex I inhibitor IACS-010759. In a B-lymphoid cell line, ectopic MYC activity and treatment with IACS-010759 added up to induce oxidative stress, with consequent depletion of reduced glutathione and lethal disruption of redox homeostasis. This effect could be enhanced either with inhibitors of NADPH production through the pentose phosphate pathway, or with ascorbate (vitamin C), known to act as a pro-oxidant at high doses. In these conditions, ascorbate synergized with IACS-010759 to kill MYC-overexpressing cells in vitro and reinforced its therapeutic action against human B-cell lymphoma xenografts. Hence, complex I inhibition and high-dose ascorbate might improve the outcome of patients affected by high-grade lymphomas and potentially other MYC-driven cancers.

MYC and therapy resistance in cancer: risks and opportunities.

The MYC transcription factor, encoded by the c-MYC proto-oncogene, is activated by growth-promoting signals, and is a key regulator of biosynthetic and metabolic pathways driving cell growth and proliferation. These same processes are deregulated in MYC-driven tumors, where they become critical for cancer cell proliferation and survival. As other oncogenic insults, overexpressed MYC induces a series of cellular stresses (metabolic, oxidative, replicative, etc.) collectively known as oncogenic stress, which impact not only on tumor progression, but also on the response to therapy, with profound, multifaceted consequences on clinical outcome. On one hand, recent evidence uncovered a widespread role for MYC in therapy resistance in multiple cancer types, with either standard chemotherapeutic or targeted regimens. Reciprocally, oncogenic MYC imparts a series of molecular and metabolic dependencies to cells, thus giving rise to cancer-specific vulnerabilities that may be exploited to obtain synthetic-lethal interactions with novel anticancer drugs. Here we will review the current knowledge on the links between MYC and therapeutic responses, and will discuss possible strategies to overcome resistance through new, targeted interventions.

Polycomb group ring finger protein 6 suppresses Myc-induced lymphomagenesis.

Max is an obligate dimerization partner for the Myc transcription factors and for several repressors, such as Mnt, Mxd1-4, and Mga, collectively thought to antagonize Myc function in transcription and oncogenesis. Mga, in particular, is part of the variant Polycomb group repressive complex PRC1.6. Here, we show that ablation of the distinct PRC1.6 subunit Pcgf6-but not Mga-accelerates Myc-induced lymphomagenesis in Eµ-myc transgenic mice. Unexpectedly, however, Pcgf6 loss shows no significant impact on transcriptional profiles, in neither pre-tumoral B-cells, nor lymphomas. Altogether, these data unravel an unforeseen, Mga- and PRC1.6-independent tumor suppressor activity of Pcgf6.

Targeting mitochondrial respiration and the BCL2 family in high-grade MYC-associated B-cell lymphoma.

Multiple molecular features, such as activation of specific oncogenes (e.g., MYC, BCL2) or a variety of gene expression signatures, have been associated with disease course in diffuse large B-cell lymphoma (DLBCL), although their relationships and implications for targeted therapy remain to be fully unraveled. We report that MYC activity is closely correlated with-and most likely a driver of-gene signatures related to oxidative phosphorylation (OxPhos) in DLBCL, pointing to OxPhos enzymes, in particular mitochondrial electron transport chain (ETC) complexes, as possible therapeutic targets in high-grade MYC-associated lymphomas. In our experiments, indeed, MYC sensitized B cells to the ETC complex I inhibitor IACS-010759. Mechanistically, IACS-010759 triggered the integrated stress response (ISR) pathway, driven by the transcription factors ATF4 and CHOP, which engaged the intrinsic apoptosis pathway and lowered the apoptotic threshold in MYC-overexpressing cells. In line with these findings, the BCL2-inhibitory compound venetoclax synergized with IACS-010759 against double-hit lymphoma (DHL), a high-grade malignancy with concurrent activation of MYC and BCL2. In BCL2-negative lymphoma cells, instead, killing by IACS-010759 was potentiated by the Mcl-1 inhibitor S63845. Thus, combining an OxPhos inhibitor with select BH3-mimetic drugs provides a novel therapeutic principle against aggressive, MYC-associated DLBCL variants.

Integrated requirement of non-specific and sequence-specific DNA binding in Myc-driven transcription.

Eukaryotic transcription factors recognize specific DNA sequence motifs, but are also endowed with generic, non-specific DNA-binding activity. How these binding modes are integrated to determine select transcriptional outputs remains unresolved. We addressed this question by site-directed mutagenesis of the Myc transcription factor. Impairment of non-specific DNA backbone contacts caused pervasive loss of genome interactions and gene regulation, associated with increased intra-nuclear mobility of the Myc protein in murine cells. In contrast, a mutant lacking base-specific contacts retained DNA-binding and mobility profiles comparable to those of the wild-type protein, but failed to recognize its consensus binding motif (E-box) and could not activate Myc-target genes. Incidentally, this mutant gained weak affinity for an alternative motif, driving aberrant activation of different genes. Altogether, our data show that non-specific DNA binding is required to engage onto genomic regulatory regions; sequence recognition in turn contributes to transcriptional activation, acting at distinct levels: stabilization and positioning of Myc onto DNA, and-unexpectedly-promotion of its transcriptional activity. Hence, seemingly pervasive genome interaction profiles, as detected by ChIP-seq, actually encompass diverse DNA-binding modalities, driving defined, sequence-dependent transcriptional responses.

Low diversity or poorly explored? Mesophotic molluscs highlight undersampling in the Eastern Mediterranean.

Mesophotic assemblages are the next frontier of marine exploration in the Mediterranean Sea. Located below recreational scuba diving depths, they are difficult to access but host a diverse array of habitats structured by large invertebrate species. The Eastern Mediterranean has been much less explored than the western part of the basin and its mesophotic habitats are virtually unknown. We here describe two mesophotic (77-92 m depth) molluscan assemblages at a rocky reef and on a soft substrate off northern Israel. We record 172 species, of which 43 (25%) are first records for Israel and increase its overall marine molluscan diversity by 7%. Only five of these species have been reported in recent surveys of the nearby Lebanon, suggesting that our results are robust at a broader scale than our study area and that the reported west-to-east declining diversity gradient in the Mediterranean needs a reappraisal based on proper sampling of the eastern basin. We found only four (2%) non-indigenous species, represented by seven (0.5%) specimens. These results suggest that pristine native assemblages still thrive at this depth in Israel, in contrast to the shallow subtidal heavily affected by global warming and biological invasions, calling for strong conservation actions for these valuable but vulnerable habitats.

Revision of the Recent Alvania scabra (Philippi, 1844) complex (Mollusca, Gastropoda, Rissoidae) from the Mediterranean Sea with the description of a new species.

Herein we revise several Recent Mediterranean species of the rissoid genus Alvania Risso, 1826: Alvania scabra (Philippi, 1844), Alvania sculptilis (Monterosato, 1877), Alvania sororcula Granata-Grillo, 1877, Alvania lucinae Oberling, 1970, Alvania josefoi Oliver Templado, 2009 and Alvania scuderii Villari, 2017. They represent a rather homogeneous group of morphologically similar species, referred to as the Alvania scabra complex, which includes also some other species from the northeastern Atlantic. We designate a neotype for Rissoa scabra Philippi, 1844 and a lectotype for Rissoa oranica Pallary, 1900 to stabilize the use of the names. Alvania oranica (Pallary, 1900) is confirmed as a synonym of Alvania scabra (Philippi, 1844), and Alvania asperella (Granata-Grillo, 1877) is proposed as a synonym of Alvania sororcula (Granata-Grillo, 1877) [new synonymy]. Finally, we describe one new Mediterranean species: Alvania pizzinii Amati, Smriglio Oliverio n. sp. from Levanzo Is., Sicily.

Reactivation of Myc transcription in the mouse heart unlocks its proliferative capacity.

It is unclear why some tissues are refractory to the mitogenic effects of the oncogene Myc. Here we show that Myc activation induces rapid transcriptional responses followed by proliferation in some, but not all, organs. Despite such disparities in proliferative response, Myc is bound to DNA at open elements in responsive (liver) and non-responsive (heart) tissues, but fails to induce a robust transcriptional and proliferative response in the heart. Using heart as an exemplar of a non-responsive tissue, we show that Myc-driven transcription is re-engaged in mature cardiomyocytes by elevating levels of the positive transcription elongation factor (P-TEFb), instating a large proliferative response. Hence, P-TEFb activity is a key limiting determinant of whether the heart is permissive for Myc transcriptional activation. These data provide a greater understanding of how Myc transcriptional activity is determined and indicate modification of P-TEFb levels could be utilised to drive regeneration of adult cardiomyocytes for the treatment of heart myopathies.

Cooperation Between MYC and ß-Catenin in Liver Tumorigenesis Requires Yap/Taz.

BACKGROUND AND AIMS: Activation of MYC and catenin beta-1 (CTNNB1, encoding ß-catenin) can co-occur in liver cancer, but how these oncogenes cooperate in tumorigenesis remains unclear. APPROACH AND RESULTS: We generated a mouse model allowing conditional activation of MYC and WNT/ß-catenin signaling (through either ß-catenin activation or loss of APC - adenomatous polyposis coli) upon expression of CRE recombinase in the liver and monitored their effects on hepatocyte proliferation, apoptosis, gene expression profiles, and tumorigenesis. Activation of WNT/ß-catenin signaling strongly accelerated MYC-driven carcinogenesis in the liver. Both pathways also cooperated in promoting cellular transformation in vitro, demonstrating their cell-autonomous action. Short-term induction of MYC and ß-catenin in hepatocytes, followed by RNA-sequencing profiling, allowed the identification of a "Myc/ß-catenin signature," composed of a discrete set of Myc-activated genes whose expression increased in the presence of active ß-catenin. Notably, this signature enriched for targets of Yes-associated protein (Yap) and transcriptional coactivator with PDZ-binding motif (Taz), two transcriptional coactivators known to be activated by WNT/ß-catenin signaling and to cooperate with MYC in mitogenic activation and liver transformation. Consistent with these regulatory connections, Yap/Taz accumulated upon Myc/ß-catenin activation and were required not only for the ensuing proliferative response, but also for tumor cell growth and survival. Finally, the Myc/ß-catenin signature was enriched in a subset of human hepatocellular carcinomas characterized by comparatively poor prognosis. CONCLUSIONS: Myc and ß-catenin show a strong cooperative action in liver carcinogenesis, with Yap and Taz serving as mediators of this effect. These findings warrant efforts toward therapeutic targeting of Yap/Taz in aggressive liver tumors marked by elevated Myc/ß-catenin activity.

An early Myc-dependent transcriptional program orchestrates cell growth during B-cell activation.

Upon activation, lymphocytes exit quiescence and undergo substantial increases in cell size, accompanied by activation of energy-producing and anabolic pathways, widespread chromatin decompaction, and elevated transcriptional activity. These changes depend upon prior induction of the Myc transcription factor, but how Myc controls them remains unclear. We addressed this issue by profiling the response to LPS stimulation in wild-type and c-myc-deleted primary mouse B-cells. Myc is rapidly induced, becomes detectable on virtually all active promoters and enhancers, but has no direct impact on global transcriptional activity. Instead, Myc contributes to the swift up- and down-regulation of several hundred genes, including many known regulators of the aforementioned cellular processes. Myc-activated promoters are enriched for E-box consensus motifs, bind Myc at the highest levels, and show enhanced RNA Polymerase II recruitment, the opposite being true at down-regulated loci. Remarkably, the Myc-dependent signature identified in activated B-cells is also enriched in Myc-driven B-cell lymphomas: hence, besides modulation of new cancer-specific programs, the oncogenic action of Myc may largely rely on sustained deregulation of its normal physiological targets.

MYC in Germinal Center-derived lymphomas: Mechanisms and therapeutic opportunities.

The rearrangement of immunoglobulin loci during the germinal center reaction is associated with an increased risk of chromosomal translocations that activate oncogenes such as MYC, BCL2 or BCL6, thus contributing to the development of B-cell lymphomas. MYC and BCL2 activation are initiating events in Burkitt's (BL) and Follicular Lymphoma (FL), respectively, but can occur at later stages in other subtypes such as Diffuse Large-B Cell Lymphoma (DLBCL). MYC can also be activated during the progression of FL to the transformed stage. Thus, either DLBCL or FL can give rise to aggressive double-hit lymphomas (DHL) with concurrent activation of MYC and BCL2. Research over the last three decades has improved our understanding of the functions of these oncogenes and the basis for their cooperative action in lymphomagenesis. MYC, in particular, is a transcription factor that contributes to cell activation, growth and proliferation, while concomitantly sensitizing cells to apoptosis, the latter being blocked by BCL2. Here, we review our current knowledge about the role of MYC in germinal center B-cells and lymphomas, discuss MYC-induced dependencies that can sensitize cancer cells to select pharmacological inhibitors, and illustrate their therapeutic potential in aggressive lymphomas-and in particular in DHL, in combination with BCL2 inhibitors.

p53 Loss in Breast Cancer Leads to Myc Activation, Increased Cell Plasticity, and Expression of a Mitotic Signature with Prognostic Value.

Loss of p53 function is invariably associated with cancer. Its role in tumor growth was recently linked to its effects on cancer stem cells (CSCs), although the underlying molecular mechanisms remain unknown. Here, we show that c-myc is a transcriptional target of p53 in mammary stem cells (MaSCs) and is activated in breast tumors as a consequence of p53 loss. Constitutive Myc expression in normal mammary cells leads to increased frequency of MaSC symmetric divisions, extended MaSC replicative-potential, and MaSC-reprogramming of progenitors, whereas Myc activation in breast cancer is necessary and sufficient to maintain the expanding pool of CSCs. Concomitant p53 loss and Myc activation trigger the expression of 189 mitotic genes, which identify patients at high risk of mortality and relapse, independently of other risk factors. Altogether, deregulation of the p53:Myc axis in mammary tumors increases CSC content and plasticity and is a critical determinant of tumor growth and clinical aggressiveness.

Catalogue of the primary types of marine molluscan taxa described by Tommaso Allery Di Maria, Marquis of Monterosato, deposited in the Museo Civico di Zoologia, Roma.

We have compiled a complete list of new marine molluscan taxa introduced by Tommaso Allery Di Maria, Marquis of Monterosato (1841-1927). The dates of publication of every single work have been checked against available evidence, and an updated bibliography is also presented. Finally, the type material of all marine taxa expected to be in the collection Monterosato (presently preserved in the Museo Civico di Zoologia in Rome) has been searched in the main collection, and all retrieved specimens have been catalogued. A large majority of the material has been found, representative specimens of each taxon have been illustrated, and remarks on nomenclature and taxonomy have been provided yielding 42 new synonymies, 46 nominal taxa rediscovered, and 6 new combinations.

Therapeutic synergy between tigecycline and venetoclax in a preclinical model of MYC/BCL2 double-hit B cell lymphoma.

High-grade B cell lymphomas with concurrent activation of the MYC and BCL2 oncogenes, also known as double-hit lymphomas (DHL), show dismal prognosis with current therapies. MYC activation sensitizes cells to inhibition of mitochondrial translation by the antibiotic tigecycline, and treatment with this compound provides a therapeutic window in a mouse model of MYC-driven lymphoma. We now addressed the utility of this antibiotic for treatment of DHL. BCL2 activation in mouse Eµ-myc lymphomas antagonized tigecycline-induced cell death, which was specifically restored by combined treatment with the BCL2 inhibitor venetoclax. In line with these findings, tigecycline and two related antibiotics, tetracycline and doxycycline, synergized with venetoclax in killing human MYC/BCL2 DHL cells. Treatment of mice engrafted with either DHL cell lines or a patient-derived xenograft revealed strong antitumoral effects of the tigecycline/venetoclax combination, including long-term tumor eradication with one of the cell lines. This drug combination also had the potential to cooperate with rituximab, a component of current front-line regimens. Venetoclax and tigecycline are currently in the clinic with distinct indications: Our preclinical results warrant the repurposing of these drugs for combinatorial treatment of DHL.

Transcriptional integration of mitogenic and mechanical signals by Myc and YAP.

Mammalian cells must integrate environmental cues to determine coherent physiological responses. The transcription factors Myc and YAP-TEAD act downstream from mitogenic signals, with the latter responding also to mechanical cues. Here, we show that these factors coordinately regulate genes required for cell proliferation. Activation of Myc led to extensive association with its genomic targets, most of which were prebound by TEAD. At these loci, recruitment of YAP was Myc-dependent and led to full transcriptional activation. This cooperation was critical for cell cycle entry, organ growth, and tumorigenesis. Thus, Myc and YAP-TEAD integrate mitogenic and mechanical cues at the transcriptional level to provide multifactorial control of cell proliferation.

Integrative analysis of RNA polymerase II and transcriptional dynamics upon MYC activation.

Overexpression of the MYC transcription factor causes its widespread interaction with regulatory elements in the genome but leads to the up- and down-regulation of discrete sets of genes. The molecular determinants of these selective transcriptional responses remain elusive. Here, we present an integrated time-course analysis of transcription and mRNA dynamics following MYC activation in proliferating mouse fibroblasts, based on chromatin immunoprecipitation, metabolic labeling of newly synthesized RNA, extensive sequencing, and mathematical modeling. Transcriptional activation correlated with the highest increases in MYC binding at promoters. Repression followed a reciprocal scenario, with the lowest gains in MYC binding. Altogether, the relative abundance (henceforth, "share") of MYC at promoters was the strongest predictor of transcriptional responses in diverse cell types, predominating over MYC's association with the corepressor ZBTB17 (also known as MIZ1). MYC activation elicited immediate loading of RNA polymerase II (RNAPII) at activated promoters, followed by increases in pause-release, while repressed promoters showed opposite effects. Gains and losses in RNAPII loading were proportional to the changes in the MYC share, suggesting that repression by MYC may be partly indirect, owing to competition for limiting amounts of RNAPII. Secondary to the changes in RNAPII loading, the dynamics of elongation and pre-mRNA processing were also rapidly altered at MYC regulated genes, leading to the transient accumulation of partially or aberrantly processed mRNAs. Altogether, our results shed light on how overexpressed MYC alters the various phases of the RNAPII cycle and the resulting transcriptional response.

FunChIP: an R/Bioconductor package for functional classification of ChIP-seq shapes.

SUMMARY: Chromatin Immunoprecipitation followed by sequencing (ChIP-seq) generates local accumulations of sequencing reads on the genome ("peaks"), which correspond to specific protein-DNA interactions or chromatin modifications. Peaks are detected by considering their total area above a background signal, usually neglecting their shapes, which instead may convey additional biological information. We present FunChIP, an R/Bioconductor package for clustering peaks according to a functional representation of their shapes: after approximating their profiles with cubic B-splines, FunChIP minimizes their functional distance and classifies the peaks applying a k-mean alignment and clustering algorithm. The whole pipeline is user-friendly and provides visualization functions for a quick inspection of the results. An application to the transcription factor Myc in 3T9 murine fibroblasts shows that clusters of peaks with different shapes are associated with different genomic locations and different transcriptional regulatory activity. AVAILABILITY AND IMPLEMENTATION: The package is implemented in R and is available under Artistic Licence 2.0 from the Bioconductor website (http://bioconductor.org/packages/FunChIP). CONTACT: marco.morelli@iit.it. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Opposing macrophage polarization programs show extensive epigenomic and transcriptional cross-talk.

Stimulation of macrophages with interferon-γ (IFN-γ) and interleukin 4 (IL-4) triggers distinct and opposing activation programs. During mixed infections or cancer, macrophages are often exposed to both cytokines, but how these two programs influence each other remains unclear. We found that IFN-γ and IL-4 mutually inhibited the epigenomic and transcriptional changes induced by each cytokine alone. Computational and functional analyses revealed the genomic bases for gene-specific cross-repression. For instance, while binding motifs for the transcription factors STAT1 and IRF1 were associated with robust and IL-4-resistant responses to IFN-γ, their coexistence with binding sites for auxiliary transcription factors such as AP-1 generated vulnerability to IL-4-mediated inhibition. These data provide a core mechanistic framework for the integration of signals that control macrophage activation in complex environmental conditions.