Biology subjects

Buechel, G.

Publications and source records attributed to Buechel, G..

6 recordsLinked to original sources

bioRxiv2026

MYC/MIZ1 suppression of lysosomal protein degradation drives immune evasion in pancreatic ductal adenocarcinoma

The MYC oncoprotein promotes immune evasion of pancreatic ductal adenocarcinoma (PDAC), but the underlying molecular mechanisms are not fully understood. Here we show that MYC protects PDAC tumors from CD4+ T cell-dependent elimination. Single cell sequencing shows that MYC suppression in tumor cells increases amino acid availability and broadly activates amino acid-responsive gene expression programs in immune cell populations. This occurs because MYC-driven uptake depletes free amino acids from tumor interstitial fluid and plasma, while MYC compromises macropinocytosis and autophagy, both of which depend on lysosomal protein degradation. MYC engages the POZ/BTB transcription factor MIZ1 to suppress lysosomal genes regulated by the TFE3/TFEB/MITF network or by free MIZ1, thereby inhibiting lysosomal protein degradation. An orthogonal genetic model enabling transient, selective inhibition of amino acid uptake in tumor cells recapitulates the effects of MYC depletion on amino acid levels in the tumor microenvironment and induces complete, CD4+ T cell-dependent tumor eradication with long-term survival. We propose that MYC-mediated, cell-autonomous disruption of lysosome function coupled to non-cell-autonomous protection from immune clearance allows MYC-low cells to benefit from MYC-high neighbors, such that intratumoral heterogeneity in MYC expression confers a selective advantage to the entire tumor. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/740267v1_ufig1.gif" ALT="Figure 1"> View larger version (53K): org.highwire.dtl.DTLVardef@140d2ccorg.highwire.dtl.DTLVardef@cf5f1borg.highwire.dtl.DTLVardef@6d132aorg.highwire.dtl.DTLVardef@10535e6_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

The MYCN/Aurora-A complex is a cyclin activating kinase for CDK12

Deregulated MYCN is a driver of aggressive pediatric and adult neuroendocrine tumors, but critical oncogenic processes downstream of MYCN remain poorly defined. In neuroblastoma, MYCN interacts with and activates the Aurora-A kinase. Here we show that Aurora-A is a CDK-activating kinase for CDK12 by phosphorylating T893 in the T-loop, thereby enhancing its kinase activity. Aurora-A-dependent activation of CDK12 controls phosphorylation of T4 of RNA polymerase and recruits transcription termination complexes, thereby preventing transcription-replication conflicts. Enhanced crosslinking and immunoprecipitation sequencing reveals that Aurora-A associates with splice sites on nascent RNA. RNA-bound Aurora-A is catalytically inactive. MYCN competes with RNA for binding to Aurora-A and displaces Aurora-A from RNA in cells, promoting its CDK12 kinase activity. Combining Aurora-A and CDK12 inhibition potently suppresses the growth of MYCN-amplified neuroblastoma cells and patient-derived xenografts. Our data demonstrate that an Aurora-A/CDK12-dependent transcription termination pathway is a critical and targetable dependency of MYCN-driven tumors.

cancer biology↗

bioRxiv2024

A trimeric USP11/USP7/TCEAL1 complex stabilizes RNAPII during early transcription to sustain oncogenic gene expression

During early transcription, RNA polymerase II (RNAPII) undergoes a series of structural transitions controlled by cyclin-dependent kinases. Whether protein ubiquitylation and proteasomal degradation affect the fate of RNAPII close to promoters is less well understood. Here we show that the deubiquitylating enzyme USP11 and its heterodimeric partner USP7 form a trimeric complex with TCEAL1, a member of the poorly understood TCEAL (TCEA/TFIIS-like) protein family. TCEAL1 shares sequence homology with the RNAPII interaction domain of the TCEA/TFIIS elongation factor, which controls the fate of backtracked RNAPII. TCEAL1 stabilizes complexes of USP11 with USP7 and with RNAPII. TCEAL1 is recruited to core promoters when transcription elongation is blocked and globally enhances the chromatin association of RNAPII during early transcription. Mechanistically, the USP11/USP7/TCEAL1 complex competes with TFIIS for binding to core promoters and protects RPB8, an essential subunit of RNAPII, from degradation, likely preventing excessive TFIIS-mediated transcript cleavage and RNAPII disassembly. In neuroblastoma and other tumors, TCEAL1-dependent genes define a TGF beta-dependent gene expression program that is characteristic for mesenchymal and invasive tumor cell types, suggesting that the USP11/USP7/TCEAL1 trimer stabilizes RNAPII during early transcription to support a critical oncogenic gene expression program (190 words).

cancer biology↗

bioRxiv2024

Targeting MYCN upregulates L1CAM tumor antigen in MYCN-dysregulated neuroblastoma to increase CAR T cell efficacy

BackgroundCurrent treatment protocols have only limited success in pediatric patients with neuroblastomas harboring amplifications of the central oncogene, MYCN. Adoptive T cell therapy presents an innovative strategy to improve cure rates. However, L1CAM-targeting CAR T cells achieved only limited response against refractory/relapsed neuroblastoma in an ongoing phase I trial to date. Here, we investigate how oncogenic MYCN levels influence tumor cell response to CAR T cells, as one possible factor limiting success in trials. MethodsHigh MYCN levels were induced in SK-N-AS cells harboring the normal diploid MYCN complement using a tetracycline-inducible system. The inducible MYCN cell model or MYCN-amplified neuroblastoma cell lines were cocultured with L1CAM-CAR T cells. CAR T cell effector function was assessed via activation marker expression (flow cytometry), cytokine release and tumor cytotoxicity (biophotonic signal assessment). The cell model was characterized using RNA sequencing, and our data compared to publicly available RNA and proteomic data sets from neuroblastomas. ChIP-sequencing data was used to determine transcriptional L1CAM regulation by MYCN using public data sets. Synergism between CAR T cells and the MLN8237 AURKA inhibitor, which indirectly inhibits MYCN activity, was assessed in vitro using the Bliss model and in vivo in an immunocompromised mouse model. ResultsInducing high MYCN levels in the neuroblastoma cell model reduced L1CAM expression and, consequently, L1CAM-CAR T cell effector function (activation, cytokine release and cytotoxicity) in vitro. Primary neuroblastomas possessing high MYCN levels expressed lower levels of both the L1CAM transcript and L1CAM tumor antigen. Indirectly inhibiting MYCN via AURKA using MLN8237 treatment restored L1CAM expression on tumor cells in vitro and restored L1CAM-CAR T cell effector function. Combining MLN8237 and L1CAM-CAR T cell treatment synergistically increased neuroblastoma-directed killing in MYCN-overexpressing cells in vitro and in vivo concomitant with severe in vivo toxicity. ConclusionWe shed new light on a primary resistance mechanism in MYCN-driven neuroblastoma against L1CAM-CAR T cells via target antigen downregulation. These data suggest that combining L1CAM-CAR T cell therapy with pharmacological MYCN inhibition may benefit patients with high-risk neuroblastomas harboring MYCN amplifications.

immunology↗

bioRxiv2023

Direct RNA-binding by MYCN mediates feedback from RNA processing to transcription control

The MYCN oncoprotein broadly binds active promoters in a heterodimer with its partner protein MAX. MYCN also interacts with the nuclear exosome, a 3-5 exoribonuclease complex, suggesting a function in RNA metabolism. Here we show that MYCN forms stable high molecular weight complexes with the exosome and multiple RNA-binding proteins. In cells, MYCN binds to thousands of intronic RNAs; recombinant MYCN directly binds RNA via a short, highly conserved sequence termed MYCBoxI. Perturbing exosome function results in global re-localization of MYCN from promoters to intronic RNAs. At promoters, MYCN is then replaced by the MNT(MXD6) repressor protein, which inhibits MYCN-dependent transcription. MYCN promotes the degradation of its bound introns via the nuclear exosome targeting (NEXT) complex. Our data demonstrate that MYCN is an RNA-binding protein that regulates nascent transcript turnover and show that competition between its RNA- and DNA-bound states links the dynamics of the MYCN/MAX/MXD network to mRNA processing.

cancer biology↗

bioRxiv2022

Selenocysteine metabolism is a targetable vulnerability in MYCN-amplified cancers

Understanding the operational molecular, and metabolic networks that determine the balance between pro- and anti-ferroptotic regulatory pathways could unravel unique vulnerabilities to be exploited for cancer therapy. Here we identify the selenoprotein P (SELENOP) receptor, LRP8, as a key determinant protecting MYCN-amplified neuroblastoma cells from ferroptosis in vitro and in orthotopic neuroblastoma mouse models. Specifically, the exquisite dependency on LRP8-mediated selenocysteine import is caused by the failure of MYCN-amplified cells to efficiently utilize alternative forms of selenium/selenocysteine based uptake necessary for selenoprotein biosynthesis. Increased activity of one of such transporters, SLC7A11, in MYCN-amplified cells leads to cysteine overload, progressive mitochondrial decline and impaired proliferation. These data reveal in LRP8 a targetable, and specific vulnerability of MYCN-amplified neuroblastoma cells and disclose a yet-unaccounted mechanism for selective ferroptosis induction that has the potential to become an important therapeutic entry point for MYCN-amplified neuroblastoma. Statement of significanceGiven the largely unsuccessful repurposing of adult oncology drugs for the treatment of neuroblastoma, our discoveries pave the way for novel ferroptosis based strategies for this entity. Specifically, targeting of LRP8 may offer novel therapeutic and safer opportunities for a number of pediatric malignancies and MYCN driven cancers.

cancer biology↗