Targeting MYCN in Pediatric and Adult Cancers

Abstract

The deregulation of the MYC family of oncogenes, including c-MYC, MYCN and MYCL occurs in many types of cancers, and is frequently associated with a poor prognosis. The majority of functional studies have focused on c-MYC due to its broad expression profile in human cancers. The existence of highly conserved functional domains between MYCN and c-MYC suggests that MYCN participates in similar activities. MYC encodes a basic helix-loop-helix-leucine zipper (bHLH-LZ) transcription factor (TF) whose central oncogenic role in many human cancers makes it a highly desirable therapeutic target. Historically, as a TF, MYC has been regarded as "undruggable". Thus, recent efforts focus on investigating methods to indirectly target MYC to achieve anti-tumor effects. This review will primarily summarize the recent progress in understanding the function of MYCN. It will explore efforts at targeting MYCN, including strategies aimed at suppression of MYCN transcription, destabilization of MYCN protein, inhibition of MYCN transcriptional activity, repression of MYCN targets and utilization of MYCN overexpression dependent synthetic lethality.

Keywords: MYC; MYCN; Super-enhancer (SE); cancer; cofactor; pediatric cancer; transcription factor.

Figure 1

Structure and functional domains of MYCN. Three predictors of the intrinsically disordered region prediction tool PONDR are used to identify intrinsically disordered regions of MYCN (top section). Functional domains of MYCN defined by comparing c-MYC and mutagenesis assay (middle section). Examples of known MYCN protein partners and the regions of MYCN that contributed to the interaction (bottom section). Notes: Color boxes on the MYCN protein diagram: brown box, Myc homology Box (MB) I-IV; yellow box, Basic Region (BR); green box, Helix-Loop-Helix-Leucine Zipper (HLH-LZ); red box, nuclear localization signal (NLS). Gray shade box on the disorder score graph and MYCN protein diagram, regions of MYCN with relatively low disorder score. AA: amino acid.

Figure 2

Transcriptional regulation of MYCN. The schematic illustrates the presumed looping between the super-enhancer (SE) and the promoter of MYCN gene. In cancer cells, MYCN is driven by SEs that are marked by stretches of acetylated lysine 27 of histone 3 (H3K27Ac). BRD4 is a chromatin ‘reader’ that binds to acetylated lysine residues (AcK) and activates MYCN transcription. CDK7 is a TFIIH subunit that phosphorylates the carboxy-terminal domain of RNA Pol II (RNAPII) to initiate MYCN gene transcription. CDK9 is a pTEFb subunit that phosphorylates the carboxy-terminal domain of RNAPII to regulate MYCN transcriptional elongation. The enrichment or activation of these components of the transcriptional machinery in cancer cells results in aberrantly elevated transcription of MYCN (top panel). The treatment of cells with HDAC inhibitors (HDACi) inactivates MYCN SEs possibly through disrupting normal looping and depleting transcription factors (TFs) that bind to the SEs; BRD4 inhibitors (BRD4i) impact the ‘reader’ function of BRD4 to inactivate MYCN gene transcription; CDK7 inhibitors (CDK7i) and CDK9 inhibitors (CDK9i) treatment impedes the phosphorylation of RNAPII to inhibit MYCN gene transcription initiation and elongation; RA treatment inactivates MYCN transcription in a RA response element independent manner (bottom panel). Notes: circled ‘Ac’ represents H3K27Ac; circled ‘p’ represents phosphate at the RNAPII tail.

Figure 3

The regulation of MYCN protein stability. CDK1 phosphorylates MYCN at serine 62 (S62) to stabilize MYCN and prime threonine 58 (T58) for phosphorylation via GSK3β. AKT phosphorylates GSK3ß inactivating its kinase. After dephosphorylation of S62 possibly through PIN1/PP2A, MYCN is poly-ubiquitinated by the ubiquitin ligase FBXW7 and undergoes proteolytic degradation via the proteasome. AURKA binds to and stabilizes phosphorylated and poly-ubiquitinated MYCN to protect MYCN from degradation. PLK1 destabilizes FBXW7 to counteract FBXW7-mediated degradation of MYCN. The ubiquitin-specific protease HAUSP deubiquitinates MYCN to stabilize it. Thus, the treatment of cells with PI3K, AURKA, PLK1 or HAUSP inhibitors (PI3Ki, AURKAi, PLKi or HAUSPi) leads MYCN proteasomal degradation. Notes: circled ‘p’ represents phosphate; circled ‘Ub’ represents ubiquitin.

Figure 4

Targeting MYCN transcriptional activity. MYCN heterodimerizes with MAX to bind to the cis-genomic elements in DNA. MYCN interacts with cofactors WDR5 and KDM4B to activate gene transcription, while interacts with LSD1, EZH2 and HADCs to repress gene transcription through affecting chromatin status. Inhibitors such as 10058-F4, OmoMYC and MYCi361 disrupt the dimerization between MYCN and MAX to inhibit the DNA binding of MYCN. The treatment of cells with MYCN cofactor inhibitors (WDR5i, KDM4Bi, LSD1i, EZH2i or HDACi) inactivates MYCN transcriptional activity through regional epigenetic modification and/or opening or closing chromatin.

Figure 5

PROTAC strategy to directly target MYCN. The schematic illustrates the mode of action of a proteolysis targeting chimera (PROTAC) targeting MYCN. First of all, a bio-conjugatable analog of a MYCN binding ligand (such as modified 10058-F4 or MYCi361) will be conjugated to E3 ubiquitin ligase binding ligand through a linker to synthesize a MYCN PROTAC. The formation of MYCN-PROTAC-E3 ubiquitin ligase complex will result in a transfer of ubiquitin (Ub) to the lysine residues of MYCN by E2 ubiquitin-conjugating enzyme. Afterwards, the PROTAC will be released and reutilized, and the poly-ubiquitinated MYCN will undergo proteasome degradation. Notes: circled ‘Ub’ represents ubiquitin.