Identification Of MYC Research Articles

Luminescence complementation technology for the identification of MYC:TRRAP inhibitors.

Mechanism-based targeted therapies have exhibited remarkable success in treating otherwise untreatable or unresectable cancers. Novel targeted therapies that correct dysregulated transcriptional programs in cancer are an unmet medical need. The transcription factor MYC is the most frequently amplified gene in human cancer and is overexpressed because of mutations in an array of oncogenic signaling pathways. The fact that many cancer cells cannot survive without MYC – a phenomenon termed “MYC addiction” – provides a compelling case for the development of MYC-specific targeted therapies. We propose a new strategy to inhibit MYC function by disrupting its essential interaction with TRRAP using small molecules. To achieve our goal, we developed a platform using luminescence complementation for identifying small molecules as inhibitors of the MYC:TRRAP interaction. Here we present validation of this assay by measuring the disruption of TRRAP binding caused by substitutions to the invariant and essential MYC homology 2 region of MYC.

Abstract PS5-07: A retrospective analysis of association of MYC and RAD21 amplification with final overall survival (OS) data in the phase 3 EMBRACA study with talazoparib

Abstract Background: Loss-of-function mutations in genes encoding components of the homologous recombination DNA damage response (DDR) machinery, notably BRCA1/2, are associated with tumor sensitivity to poly(ADP-ribose) polymerase inhibitors (PARPi). Little is known about the potential contribution of tumor alterations in non-DDR genes to modulating sensitivity to PARP inhibitors in the clinic. In EMBRACA, the PARPi TALA improved progression-free survival (PFS) (HR [95% CI] 0.54 [0.41-0.71], P<0.001) vs chemotherapy (CT) in germline BRCA1/2-mutated HER2-negative locally advanced/metastatic breast cancer. Methods: Baseline tumor tissue from 308 pts (71%; intent-to-treat) was tested by FoundationOne®. To support exploratory identification of tumor gene alterations associated with best vs worst outcomes, pts were mapped into 2 groups: [1] BEST: Pts in TALA arm with OS ≥ 30 mo and duration of treatment ≥ 24 mo, Pts in CT arm with OS ≥ 30 mo; [2] WORST: Pts in TALA or CT arm with a PFS event (PD by Independent Radiological Facility or death) ≤ 12 wks. 2 pts had short PFS but long OS and were initially mapped to both groups but then assigned to BEST and excluded from WORST. Results: Exploratory heat map visualizations of known/likely pathogenic genetic alterations defined by the FoundationOne® gene panel were used to assess differences in genetic alterations between BEST and WORST in TNBC and non-TNBC subpopulations, leading to the identification of MYC and RAD21 as commonly altered genes of high interest. Based on these results showing imbalances in tumor amplification events between BEST and WORST outcome pts, Cox regression analysis was used to explore potential associations of MYC or RAD21 amplification with OS across the evaluable ITT population. In TNBC pts receiving TALA, OS was shorter with MYC amplification than without [HR (95% CI) 1.877 (1.102, 3.196)]. No such association was evident in TNBC pts receiving CT [HR (95% CI) 0.706 (0.303, 1.643)]. In contrast, for non-TNBC pts receiving TALA, no association with OS was evident for MYC amplification versus without [HR (95% CI) 0.603 (0.310, 1.173)], while for pts receiving CT OS was shorter with MYC amplification than without [HR (95% CI) 1.917 (1.037, 3.544)]. RAD21 amplification status was not associated with OS in either TNBC or non-TNBC patients for either treatment arm, although it should be noted that two RAD21 subgroups were very small (n=7 and 11; others had n ≥ 21). Conclusions: Based on these exploratory, retrospective analyses MYC amplification was associated with shorter OS in TNBC pts receiving TALA. This may reflect the role of MYC in positive regulation of genes involved in homologous recombination such as RAD51 (Carey et al, Cancer Res 2018;78(3):742-757). MYC and RAD21 are both located at 8q24 and frequently coamplified in breast cancer (eg, Annunziato et al, Nat Commun. 2019;10(1):397), hence the lack of association of RAD21 amplification with outcome in TNBC suggests that the association of MYC amplification with shorter OS seen in pts receiving TALA may be gene-specific. Further investigation is warranted. Funding: Pfizer TNBCNon-TNBCTALACTTALACTGene AlterationBest n=10Worst n=15Best n=7Worst n=16Best n=17Worst n=11Best n=20Worst n=11MYC amplification0 (0%)7 (47%)2 (29%)6 (38%)5 (29%)1 (9%)3 (15%)5 (46%)RAD21 amplification1 (10%)2 (13%)3 (43%)5 (31%)5 (29%)1 (9%)7 (35%)4 (36%) Citation Format: Johannes Ettl, A. Douglas Laird, Joanne L. Blum, Hope S. Rugo, Sara A. Hurvitz, Miguel Martín, Henri Roché, Young-Hyuck Im, Annabel Goodwin, Rinat Yerushalmi, Tiziana Usari, Silvana Lanzalone, Akos Czibere, Julia Hopkins, Lee A. Albacker, Lida A. Mina, Jennifer K. Litton. A retrospective analysis of association of MYC and RAD21 amplification with final overall survival (OS) data in the phase 3 EMBRACA study with talazoparib [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS5-07.

Identification of MYC as an antinecroptotic protein that stifles RIPK1–RIPK3 complex formation

The underlying mechanism of necroptosis in relation to cancer is still unclear. Here, MYC, a potent oncogene, is an antinecroptotic factor that directly suppresses the formation of the RIPK1-RIPK3 complex. Gene set enrichment analyses reveal that the MYC pathway is the most prominently down-regulated signaling pathway during necroptosis. Depletion or deletion of MYC promotes the RIPK1-RIPK3 interaction, thereby stabilizing the RIPK1 and RIPK3 proteins and facilitating necroptosis. Interestingly, MYC binds to RIPK3 in the cytoplasm and inhibits the interaction between RIPK1 and RIPK3 in vitro. Furthermore, MYC-nick, a truncated form that is mainly localized in the cytoplasm, prevented TNF-induced necroptosis. Finally, down-regulation of MYC enhances necroptosis in leukemia cells and suppresses tumor growth in a xenograft model upon treatment with birinapant and emricasan. MYC-mediated suppression of necroptosis is a mechanism of necroptosis resistance in cancer, and approaches targeting MYC to induce necroptosis represent an attractive therapeutic strategy for cancer.

A practical approach to FISH testing for MYC rearrangements and brief review of MYC in aggressive B-cell lymphomas

Aggressive B-cell lymphomas, including the WHO diagnoses of diffuse large B-cell lymphoma, high-grade B-cell lymphoma with MYC and BCL2 and/or BCL6 rearrangements, high-grade B-cell lymphoma, not otherwise specified, and Burkitt lymphoma, together account for approximately 40% of B-cell non-Hodgkin lymphomas. Identification of MYC, BCL2, and BCL6 rearrangements in these neoplasms is critical for diagnostic, prognostic, and therapeutic purposes. Herein, we address technical issues surrounding identification of these genetic abnormalities, discuss their diagnostic, prognostic, and therapeutic implications, and present an algorithm for use of interphase FISH in the work-up of aggressive B-cell lymphomas. To maximize sensitivity while still limiting cost, it is recommended that interphase FISH be performed in all B-cell lymphomas with large cell or high-grade morphology using both IGH/MYC dual-color dual-fusion (D-FISH) and MYC break-apart probes (BAP) as an initial probe set, followed by BCL2 BAP (or IGH/BCL2 D-FISH) and BCL6 BAP if a MYC rearrangement is identified. In pediatric patients or aggressive B-cell lymphomas with Burkitt-like morphology, a complete analysis at the outset using BAP probes for MYC, BCL2 (or IGH/BCL2 D-FISH), and BCL6 as well as D-FISH probes for IGH/MYC, IGK/MYC, and IGL/MYC is recommended.

Mouse tumor susceptibility genes identify drug combinations for multiple myeloma

Long-term genetic studies utilizing backcross and congenic strain analyses coupled with positional cloning strategies and functional studies identified Cdkn2a, Mtor, and Mndal as mouse plasmacytoma susceptibility/resistance genes. Tumor incidence data in congenic strains carrying the resistance alleles of Cdkn2a and Mtor led us to hypothesize that drug combinations affecting these pathways are likely to have an additive, if not synergistic effect in inhibiting tumor cell growth. Traditional and novel systems-level genomic approaches were used to assess combination activity, disease specificity, and clinical potential of a drug combination involving rapamycin/everolimus, an Mtor inhibitor, with entinostat, an histone deacetylase inhibitor. The combination synergistically repressed oncogenic MYC and activated the Cdkn2a tumor suppressor. The identification of MYC as a primary upstream regulator led to the identification of small molecule binders of the G-quadruplex structure that forms in the NHEIII region of the MYC promoter. These studies highlight the importance of identifying drug combinations which simultaneously upregulate tumor suppressors and downregulate oncogenes.

Abstract 528: Identification of MYC as a novel repressor of miR-29 in pancreatic ductal adenocarcinoma

Abstract Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignant disease with a dismal 5-year survival rate of 8%. The lack of effective therapies reinforces the need to better understand molecular mechanisms in PDAC. In our previous work, we found that downregulation of miR-29 is a common phenomenon of PDAC, and its restored expression had a potent tumor-suppressive effect in pancreatic cancer cells. However, the mechanism associated with the loss of miR-29 in PDAC has yet to be elucidated. In our initial work, using PDAC cell lines, we found that miR-29 is downregulated independent of KRAS. In an effort to understand the transcriptional factors mediating miR-29 repression, we selected several transcription factors that have a role in PDAC pathogenesis and binding sites in the miR-29 promoter region. siRNA mediated knockdown (KD) of three candidate transcription factors, SMAD4, MYC, and Gli3 revealed that the KD of MYC resulted in the highest gain of miR-29 expression (2-fold increase). Furthermore, MYC nuclear localization negatively correlates with miR-29 expression in various pancreatic cancer cell lines, and treatment of pancreatic cancer cells with small-molecule MYC inhibitor resulted in a dosage-dependent increase of miR-29 expression. miR-29a/b1 locus located on human chromosome 7 has two canonical MYC binding motifs upstream of the miR-29 transcription start site, which were functionally validated via luciferase assay. Finally, autophagy has been shown to be upregulated by MYC and is known to mitigate stressors of the integrated stress response (ISR) pathway in cancer. In our previous work, we showed that miR-29 blocks autophagy in pancreatic cancer cells. Consistently, we found miR-29 overexpression led to decreased cancer cell viability in combination with gemcitabine and induced the ISR as indicated by increased phospho-eIF2α. Taken together, our results demonstrate that MYC plays a key role in the regulation of miR-29 expression in pancreatic cancer. Furthermore, our data indicate the potential use of miR-29 as a novel therapeutic agent for PDAC, and these results could help form the basis for future studies to develop a more targeted approach of miR-29 therapeutic administration. Citation Format: Jason Kwon, Kayla Quirin, Tricia Factora, Janaiah Kota. Identification of MYC as a novel repressor of miR-29 in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 528.

Cell-of-Origin Assignment in Diffuse Large B-Cell Lymphoma Determined By Gene Expression in Formalin-Fixed Paraffin-Embedded Tissue Has Prognostic Significance Independent of IPI and MYC/BCL2 Immunohistochemistry

The molecularly defined cell-of-origin (COO) subtypes of diffuse large B-cell lymphoma (DLBCL), activated B-cell like (ABC) and germinal center B-cell like (GCB), have distinct underlying biology and outcomes in the R-CHOP treatment era. The recent advent of therapeutic agents that show subtype-specific activity has made the robust assignment of COO increasingly important for identifying patients that will respond to these agents. Although it has become common practice to use widely applicable immunohistochemistry-based algorithms to assign COO, these methods are reported to have widely variable concordance with COO as determined by gene expression profiling (GEP). Furthermore, using these IHC-based algorithms, the relationship between COO and treatment outcome to R-CHOP has been very inconsistent. We recently described an accurate and robust gene expression based assay for determining COO applicable to formalin-fixed paraffin-embedded (FFPE) biopsies. Here we sought to determine the prognostic significance of COO assigned using this assay and the relationship with other established prognostic factors.The Lymph2Cx assay, a 20 gene assay based on NanoString technology, was applied, as previously described (Scott et al Blood 2014;123:1214-17), to RNA extracted from the diagnostic FFPE biopsies (tumor content ≥ 60%) of 274 patients with de novo DLBCL. The median follow-up of living patients within this cohort, uniformly treated with R-CHOP at the BC Cancer Agency (Vancouver, Canada) was 6.1 years (range 1.2 – 13.2). Tissue microarrays of duplicate 0.6mm cores were stained for MYC and BCL2 (DAKO 124) to assess the proportion of tumor cells that expressed these proteins. Identification of MYC+/BCL2+ tumors (as described in Johnson et al J Clin Oncol 2012; 30: 3452-9; cut-offs: MYC ≥ 40%, BCL2 ≥ 50%) was performed independently by two expert hematopathologists (AM and PF) with consensus on discordant cases reached with a third hematopathologist (RDG).GEP data of sufficient quality was obtained from 271/274 (99%) of the biopsies, assigning the following COOs: 90 (33%) ABC, 153 (56%) GCB and 28 (10%) were unclassifiable. In comparison with the GCB group, the ABC group had a greater proportion of patients with stage III/IV disease (63% vs 45%, P=0.01) and higher IPI scores (P=0.03). The ABC subtype was associated with significantly inferior outcomes compared with the GCB group (Figure 1). This association was independent of IPI groupings (low (0-1), intermediate (2-3) and high (4-5)) in multivariate analyses. The prognostic significance of the COO was most evident in the intermediate IPI group (5 year time-to-progression (TTP) ABC 45% vs GCB 71%; log-rank P=0.001, HR 2.7 (95% CI 1.6 – 6.3)).It has been proposed that the prognostic power of COO is largely attributable to the greater proportion of MYC+BCL2+ cases within the ABC subtype (Hu et al Blood 2013; 121: 4021-31). We confirm that the proportion of tumors that are MYC+/BCL2+ by IHC is significantly higher in the ABC vs GCB subtype (57% vs 21%, P<0.001). However, multivariate analyses demonstrated that COO was significantly associated with TTP and disease specific survival, independent of MYC/BCL2 IHC. The significant association between MYC+/BCL2+ IHC and inferior outcomes was only observed in the GCB group, while the association between COO and treatment outcomes was observed only in the non-MYC+/BCL2+ group (Figure 2). When multivariate analyses were performed including the IPI groupings, MYC/BCL2 IHC and COO as variables, only the IPI and COO were significantly associated with TTP.We have previously shown the accuracy of the Lymph2Cx assay at assigning COO, establishing its potential as a predictive biomarker. This large study of R-CHOP treated patients demonstrates that this assay separates patients into groups with significantly different outcomes, recapitulating the results of COO assigned using GEP on frozen tissue (Lenz et al N Engl J Med 2008; 359: 2313-23). Furthermore, the prognostic significance is independent of established and recently described prognostic factors providing additional support for its use in clinical trials and, ultimately, routine clinical practice. [Display omitted] [Display omitted] DisclosuresScott:NIH: Patents & Royalties. Wright:NIH: Patents & Royalties. Staudt:NIH: Patents & Royalties. Connors:NIH: Patents & Royalties. Rimsza:NIH: Patents & Royalties. Gascoyne:NIH: Patents & Royalties; Celgene: Consultancy.

Genetic Modeling of PIM Proteins in Cancer: Proviral Tagging and Cooperation with Oncogenes, Tumor Suppressor Genes, and Carcinogens.

The PIM proteins, which were initially discovered as proviral insertion sites in Moloney-murine leukemia virus infection, are a family of highly homologous serine/threonine kinases that have been reported to be overexpressed in hematological malignancies and solid tumors. The PIM proteins have also been associated with metastasis and overall treatment responses and implicated in the regulation of apoptosis, metabolism, the cell cycle, and homing and migration, which makes these proteins interesting targets for anti-cancer drug discovery. The use of retroviral insertional mutagenesis and refined approaches such as complementation tagging has allowed the identification of myc, pim, and a third group of genes (including bmi1 and gfi1) as complementing genes in lymphomagenesis. Moreover, mouse modeling of human cancer has provided an understanding of the molecular pathways that are involved in tumor initiation and progression at the physiological level. In particular, genetically modified mice have allowed researchers to further elucidate the role of each of the Pim isoforms in various tumor types. PIM kinases have been identified as weak oncogenes because experimental overexpression in lymphoid tissue, prostate, and liver induces tumors at a relatively low incidence and with a long latency. However, very strong synergistic tumorigenicity between Pim1/2 and c-Myc and other oncogenes has been observed in lymphoid tissues. Mouse models have also been used to study whether the inhibition of specific PIM isoforms is required to prevent carcinogen-induced sarcomas, indicating that the absence of Pim2 and Pim3 greatly reduces sarcoma growth and bone invasion; the extent of this effect is similar to that observed in the absence of all three isoforms. This review will summarize some of the animal models that have been used to understand the isoform-specific contribution of PIM kinases to tumorigenesis.

Oncogenic Mechanisms in Burkitt Lymphoma

Burkitt lymphoma is a germinal center B-cell-derived cancer that was instrumental in the identification of MYC as an important human oncogene more than three decades ago. Recently, new genomics technologies have uncovered several additional oncogenic mechanisms that cooperate with MYC to create this highly aggressive cancer. The transcription factor TCF-3 is central to Burkitt lymphoma pathogenesis. TCF-3 is rendered constitutively active in Burkitt lymphoma by two related mechanisms: (1) somatic mutations that inactivate its negative regulator ID3, and (2) somatic mutations in TCF-3 that block the ability of ID3 to bind and interfere with its activity as a transcription factor. TCF-3 is also a master regulator of normal germinal center B-cell differentiation. Within the germinal center, TCF-3 up-regulates genes that are characteristically expressed in the rapidly dividing centroblasts, the putative cell of origin for Burkitt lymphoma, while repressing genes expressed in the less proliferative centrocytes. TCF-3 promotes antigen-independent (tonic) B-cell-receptor signaling in Burkitt lymphoma by transactivating immunoglobulin heavy- and light-chain genes while repressing PTPN6, which encodes the phosphatase SHP-1, a negative regulator of B-cell-receptor signaling. Tonic B-cell-receptor signaling sustains Burkitt lymphoma survival by engaging the PI3 kinase pathway. In addition, TCF-3 promotes cell-cycle progression by transactivating CCND3, encoding a D-type cyclin that regulates the G1-S phase transition. Additionally, CCND3 accumulates oncogenic mutations that stabilize cyclin D3 protein expression and drive proliferation. These new insights into Burkitt lymphoma pathogenesis suggest new therapeutic strategies, which are sorely needed in developing regions of the world where this cancer is endemic.

MYC and MiR: vicious circle

MYC and MiR: vicious circle

The Transcriptional Repressor dMnt Is a Regulator of Growth in Drosophila melanogaster

The Myc-Max-Mad/Mnt network of transcription factors has been implicated in oncogenesis and the regulation of proliferation in vertebrate cells. The identification of Myc and Max homologs in Drosophila melanogaster has demonstrated a critical role for dMyc in cell growth control. In this report, we identify and characterize the third member of this network, dMnt, the sole fly homolog of the mammalian Mnt and Mad family of transcriptional repressors. dMnt possesses two regions characteristic of Mad and Mnt proteins: a basic helix-loop-helix-zipper domain, through which it dimerizes with dMax to form a sequence-specific DNA binding complex, and a Sin-interacting domain, which mediates interaction with the dSin3 corepressor. Using the upstream activation sequence/GAL4 system, we show that expression of dMnt results in an inhibition of cellular growth and proliferation. Furthermore, we have generated a dMnt null allele, which results in flies with larger cells, increased weight, and decreased life span compared to wild-type flies. Our results demonstrate that dMnt is a transcriptional repressor that regulates D. melanogaster body size.