Long noncoding RNA LINC01578 drives colon cancer metastasis through a positive feedback loop with the NF‐κB/YY1 axis

Metastasis accounts for poor prognosis of cancers and related deaths. Accumulating evidence has shown that long noncoding RNAs (lncRNAs) play critical roles in several types of cancer. However, which lncRNAs contribute to metastasis of colon cancer is still largely unknown. In this study, we found that lncRNA LINC01578 was correlated with metastasis and poor prognosis of colon cancer. LINC01578 was upregulated in colon cancer, associated with metastasis, advanced clinical stages, poor overall survival, disease‐specific survival, and disease‐free survival. Gain‐of‐function and loss‐of‐function assays revealed that LINC01578 enhanced colon cancer cell viability and mobility in vitro and colon cancer liver metastasis in vivo. Mechanistically, nuclear factor kappa B (NF‐κB) and Yin Yang 1 (YY1) directly bound to the LINC01578 promoter, enhanced its activity, and activated LINC01578 expression. LINC01578 was shown to be a chromatin‐bound lncRNA, which directly bound NFKBIB promoter. Furthermore, LINC01578 interacted with and recruited EZH2 to NFKBIB promoter and further repressed NFKBIB expression, thereby activating NF‐κB signaling. Through activation of NF‐κB, LINC01578 further upregulated YY1 expression. Through activation of the NF‐κB/YY1 axis, LINC01578 in turn enhanced its own promoter activity, suggesting that LINC01578 and NF‐κB/YY1 formed a positive feedback loop. Blocking NF‐κB signaling abolished the oncogenic roles of LINC01578 in colon cancer. Furthermore, the expression levels of LINC01578, NFKBIB, and YY1 were correlated in clinical tissues. Collectively, this study demonstrated that LINC01578 promoted colon cancer metastasis via forming a positive feedback loop with NF‐κB/YY1 and suggested that LINC01578 represents a potential prognostic biomarker and therapeutic target for colon cancer metastasis.

Abbreviations

ChIP

chromatin immunoprecipitation

ChIRP

chromatin isolation by RNA purification

COAD

colon adenocarcinoma

CPAT

Coding‐Potential Assessment Tool

CPC

coding potential calculator

DFS

disease‐free survival

DSS

disease‐specific survival

EdU

5‐ethynyl‐2''‐deoxyuridine

H&E

hematoxylin and eosin

HR

hazard ratio

IHC

immunohistochemistry

IKK

IκB kinase

IκB

inhibitory κB

lncRNAs

long noncoding RNAs

NC

negative control

NCBI

National Center for Biotechnology Information

NF‐κB

nuclear factor kappa B

qRT‐PCR

quantitative real‐time polymerase chain reaction

RIP

RNA immunoprecipitation

RPISeq

RNA‐Protein Interaction Prediction

TCGA

The Cancer Genome Atlas

TNF

tumor necrosis factor

TUNEL

TdT‐mediated dUTP Nick‐End Labeling

YY1

Yin Yang 1

     

Extracellular vesicles and oncogenic signaling

In recent years, extracellular vesicles (EVs) emerged as potential diagnostic and prognostic markers for cancer therapy. While the field of EV research is rapidly developing and their application as vehicles for therapeutic cargo is being tested, little is still known about the exact mechanisms of signaling specificity and cargo transfer by EVs, especially in vivo. Several signaling cascades have been found to use EVs for signaling in the tumor–stroma interaction. These include potentially oncogenic, verbatim transforming, signaling cascades such as Wnt and TGF‐β signaling, and other signaling cascades that have been tightly associated with tumor progression and metastasis, such as PD‐L1 and VEGF signaling. Multiple mechanisms of how these signaling cascades and EVs interplay to mediate these complex processes have been described, such as direct signal activation through pathway components on or in EVs or indirectly by influencing vesicle biogenesis, cargo sorting, or uptake dynamics. In this review, we summarize the current knowledge of EVs, their biogenesis, and our understanding of EV interactions with recipient cells with a focus on selected oncogenic and cancer‐associated signaling pathways. After an in‐depth look at how EVs mediate and influence signaling, we discuss potentially translatable EV functions and existing knowledge gaps.

Abbreviations

EGF(R)

epidermal growth factor (receptor)

EMT

epithelial–mesenchymal transition

EP(s)

extracellular particle(s)

EV(s)

extracellular vesicle(s)

Exo(s)

exosome(s)

Her

human epidermal growth factor receptor

ISEV

International Society for Extracellular Vesicles

MSC

mesenchymal stem cells

MV(s)

microvesicle(s)

MVB

multivesicular body

PD‐L

programmed death‐ligand

TGF‐β

transforming growth factor‐β

VEGF

vascular endothelial growth factor

Wnt

wingless and Int/wingless‐related integration site

     

Genomic and prognostic heterogeneity among RAS/BRAF V600E/TP53 co‐mutated resectable colorectal liver metastases

Hepatic resection is potentially curative for patients with colorectal liver metastases, but the treatment benefit varies. KRAS/NRAS (RAS)/TP53 co‐mutations are associated with a poor prognosis after resection, but there is large variation in patient outcome within the mutation groups, and genetic testing is currently not used to evaluate benefit from surgery. We have investigated the potential for improved prognostic stratification by combined biomarker analysis with DNA copy number aberrations (CNAs), and taking tumor heterogeneity into account. We determined the mutation status of RAS, BRAF ^V600, and TP53 in 441 liver lesions from 171 patients treated by partial hepatectomy for metastatic colorectal cancer. CNAs were profiled in 232 tumors from 67 of the patients. Mutations and high‐level amplifications of cancer‐critical genes, the latter including ERBB2 and EGFR, were predominantly homogeneous within patients. RAS/BRAF ^V600E and TP53 co‐mutations were associated with a poor patient outcome (hazard ratio, HR, 3.9, 95% confidence interval, CI, 1.3–11.1, P = 0.012) in multivariable analyses with clinicopathological variables. The genome‐wide CNA burden and intrapatient intermetastatic CNA heterogeneity varied within the mutation groups, and the CNA burden had prognostic associations in univariable analysis. Combined prognostic analyses of RAS/BRAF ^V600E/TP53 mutations and CNAs, either as a high CNA burden or high intermetastatic CNA heterogeneity, identified patients with a particularly poor outcome (co‐mutation/high CNA burden: HR 2.7, 95% CI 1.2–5.9, P = 0.013; co‐mutation/high CNA heterogeneity: HR 2.5, 95% CI 1.1–5.6, P = 0.022). In conclusion, DNA copy number profiling identified genomic and prognostic heterogeneity among patients with resectable colorectal liver metastases with co‐mutated RAS/BRAF ^V600E/TP53.

Abbreviations

5y‐CSS

five‐year cancer‐specific survival

CNA

copy number aberrations

CRC

colorectal cancer

CRLM

colorectal liver metastases

MSI

microsatellite instable

MSS

microsatellite stable

     

Targeted inhibition of ERα signaling and PIP5K1α/Akt pathways in castration‐resistant prostate cancer

Selective ERα modulator, tamoxifen, is well tolerated in a heavily pretreated castration‐resistant prostate cancer (PCa) patient cohort. However, its targeted gene network and whether expression of intratumor ERα due to androgen deprivation therapy (ADT) may play a role in PCa progression is unknown. In this study, we examined the inhibitory effect of tamoxifen on castration‐resistant PCa in vitro and in vivo. We found that tamoxifen is a potent compound that induced a high degree of apoptosis and significantly suppressed growth of xenograft tumors in mice, at a degree comparable to ISA‐2011B, an inhibitor of PIP5K1α that acts upstream of PI3K/AKT survival signaling pathway. Moreover, depletion of tumor‐associated macrophages using clodronate in combination with tamoxifen increased inhibitory effect of tamoxifen on aggressive prostate tumors. We showed that both tamoxifen and ISA‐2011B exert their on‐target effects on prostate cancer cells by targeting cyclin D1 and PIP5K1α/AKT network and the interlinked estrogen signaling. Combination treatment using tamoxifen together with ISA‐2011B resulted in tumor regression and had superior inhibitory effect compared with that of tamoxifen or ISA‐2011B alone. We have identified sets of genes that are specifically targeted by tamoxifen, ISA‐2011B or combination of both agents by RNA‐seq. We discovered that alterations in unique gene signatures, in particular estrogen‐related marker genes are associated with poor patient disease‐free survival. We further showed that ERα interacted with PIP5K1α through formation of protein complexes in the nucleus, suggesting a functional link. Our finding is the first to suggest a new therapeutic potential to inhibit or utilize the mechanisms related to ERα, PIP5K1α/AKT network, and MMP9/VEGF signaling axis, providing a strategy to treat castration‐resistant ER‐positive subtype of prostate cancer tumors with metastatic potential.

Abbreviations

CRPC

castration‐resistant prostate cancer

DHT

dihydrotestosterone

E2

estradiol

ERα

estrogen receptor alpha

GO

gene ontology

NG‐CHM

Next‐Generation Clustered Heatmaps

PCa

prostate cancer

TMAs

tissue microarrays

     

Long non‐coding RNA RACGAP1P promotes breast cancer invasion and metastasis via miR‐345‐5p/RACGAP1‐mediated mitochondrial fission

Long non‐coding RNAs (lncRNAs) are emerging as key molecules in various cancers, yet their potential roles in the pathogenesis of breast cancer are not fully understood. Herein, using microarray analysis, we revealed that the lncRNA RACGAP1P, the pseudogene of Rac GTPase activating protein 1 (RACGAP1), was up‐regulated in breast cancer tissues. Its high expression was confirmed in 25 pairs of breast cancer tissues and 8 breast cell lines by qRT‐PCR. Subsequently, we found that RACGAP1P expression was positively correlated with lymph node metastasis, distant metastasis, TNM stage, and shorter survival time in 102 breast cancer patients. Then, in vitro and in vivo experiments were designed to investigate the biological function and regulatory mechanism of RACGAP1P in breast cancer cell lines. Overexpression of RACGAP1P in MDA‐MB‐231 and MCF7 breast cell lines increased their invasive ability and enhanced their mitochondrial fission. Conversely, inhibition of mitochondrial fission by Mdivi‐1 could reduce the invasive ability of RACGAP1P‐overexpressing cell lines. Furthermore, the promotion of mitochondrial fission by RACGAP1P depended on its competitive binding with miR‐345‐5p against its parental gene RACGAP1, leading to the activation of dynamin‐related protein 1 (Drp1). In conclusion, lncRNA RACGAP1P promotes breast cancer invasion and metastasis via miR‐345‐5p/RACGAP1 pathway‐mediated mitochondrial fission.

Abbreviations

CDS

coding sequence

ceRNAs

competitive endogenous RNAs

Drp1

dynamin‐related protein 1

FFPE

formalin‐fixed paraffin‐embedded

lncRNAs

long non‐coding RNAs

miRNAs

microRNAs

RACGAP1

Rac GTPase activating protein 1

TCGA

The Cancer Genome Atlas

     

TRAP1 enhances Warburg metabolism through modulation of PFK1 expression/activity and favors resistance to EGFR inhibitors in human colorectal carcinomas

Metabolic rewiring is a mechanism of adaptation to unfavorable environmental conditions and tumor progression. TRAP1 is an HSP90 molecular chaperone upregulated in human colorectal carcinomas (CRCs) and responsible for downregulation of oxidative phosphorylation (OXPHOS) and adaptation to metabolic stress. The mechanism by which TRAP1 regulates glycolytic metabolism and the relevance of this regulation in resistance to EGFR inhibitors were investigated in patient‐derived CRC spheres, human CRC cells, samples, and patients. A linear correlation was observed between TRAP1 levels and ¹⁸F‐fluoro‐2‐deoxy‐glucose (¹⁸F‐FDG) uptake upon PET scan or GLUT1 expression in human CRCs. Consistently, TRAP1 enhances GLUT1 expression, glucose uptake, and lactate production and downregulates OXPHOS in CRC patient‐derived spheroids and cell lines. Mechanistically, TRAP1 maximizes lactate production to balance low OXPHOS through the regulation of the glycolytic enzyme phosphofructokinase‐1 (PFK1); this depends on the interaction between TRAP1 and PFK1, which favors PFK1 glycolytic activity and prevents its ubiquitination/degradation. By contrast, TRAP1/PFK1 interaction is lost in conditions of enhanced OXPHOS, which results in loss of TRAP1 regulation of PFK1 activity and lactate production. Notably, TRAP1 regulation of glycolysis is involved in resistance of RAS‐wild‐type CRCs to EGFR monoclonals. Indeed, either TRAP1 upregulation or high glycolytic metabolism impairs cetuximab activity in vitro, whereas TRAP1 targeting and/or inhibition of glycolytic pathway enhances cell response to cetuximab. Finally, a linear correlation between ¹⁸F‐FDG PET uptake and poor response to cetuximab in first‐line therapy in human metastatic CRCs was observed. These results suggest that TRAP1 is a key determinant of CRC metabolic rewiring and favors resistance to EGFR inhibitors through regulation of glycolytic metabolism.

Abbreviations

¹⁸F‐FDG

¹⁸F‐fluoro‐2‐deoxy‐glucose

2‐DG

2‐deoxy‐glucose

BRAF

v‐raf murine sarcoma viral oncogene homolog B

CRC

human colorectal carcinoma

ECAR

extracellular acidification rate

EGFR

epidermal growth factor receptor

GLUT1

glucose transporter member 1

HKII

hexokinase II

HSP90

heat‐shock protein 90

MCT4

solute carrier family 16 (monocarboxylic acid transporters), member 3

OCR

oxygen consumption rates

OXPHOS

oxidative phosphorylation

PET

positron emission tomography

PFK1

phosphofructokinase 1

RAS

Kirsten rat sarcoma viral oncogene homolog

SUVmax

maximum standardized uptake value

TRAP1

TNF receptor‐associated protein 1

     

Deubiquitinase OTUD6A promotes proliferation of cancer cells via regulating Drp1 stability and mitochondrial fission

Dynamin‐related protein 1 (Drp1) is a cytosolic protein responsible for mitochondrial fission and is essential in the initiation and development of several human diseases, including cancer. However, the regulation of Drp1, especially of its ubiquitination, remains unclear. In this study, we report that the ovarian tumor‐associated protease deubiquitinase 6A (OTUD6A) deubiquitylates and stabilizes Drp1, thereby facilitating regulation of mitochondrial morphology and tumorigenesis. OTUD6A is upregulated in human patients with colorectal cancer. The depletion of OTUD6A leads to lower Drp1 levels and suppressed mitochondrial fission, and the affected cells are consequently less prone to tumorigenesis. Conversely, the overexpression of OTUD6A increases Drp1 levels and its protein half‐life and enhances cancer cell growth. Therefore, our results reveal a novel upstream protein of Drp1, and its role in tumorigenesis that is played, in part, through the activation of mitochondrial fission mediated by Drp1.

Abbreviations

Drp1

dynamin‐related protein 1

DUBs

deubiquitinases

His‐ub

His‐ubiquitin

IB

immunoblot

IHC

immunohistochemistry

IP

immunoprecipitation

MARCH5

membrane‐associated ring‐CH‐type finger 5

Mfn1

mitofusin 1

Mfn2

mitofusin 2

NIK

NF‐κB‐inducing kinase

Opa1

optic atrophy 1

OTUs

ovarian tumor‐associated proteases

OTUD6A

ovarian tumor‐associated protease deubiquitinase 6A

     

BAP1 suppresses prostate cancer progression by deubiquitinating and stabilizing PTEN

Deubiquitinase BAP1 is an important tumor suppressor in several malignancies, but its functions and critical substrates in prostate cancer (PCa) remain unclear. Here, we report that the mRNA and protein expression levels of BAP1 are downregulated in clinical PCa specimens. BAP1 can physically bind to and deubiquitinate PTEN, which inhibits the ubiquitination‐mediated degradation of PTEN and thus stabilizes PTEN protein. Ectopically expressed BAP1 in PCa cells increases PTEN protein level and subsequently inhibits the AKT signaling pathway, thus suppressing PCa progression. Conversely, knockdown of BAP1 in PCa cells leads to the decrease in PTEN protein level and the activation of the Akt signaling pathway, therefore promoting malignant transformation and cancer metastasis. However, these can be reversed by the re‐expression of PTEN. More importantly, we found that BAP1 protein level positively correlates with PTEN in a substantial fraction of human cancers. These findings demonstrate that BAP1 is an important deubiquitinase of PTEN for its stability and the BAP1‐PTEN signaling axis plays a crucial role in tumor suppression.

Abbreviations

BAP1

the BRCA1‐associated protein 1

DUBs

deubiquitinases

GEO

Gene Expression Omnibus

IP

immunoprecipitation

KEGG

the Kyoto Encyclopedia of Genes and Genomes

PCa

prostate cancer

PTEN

phosphatase and tensin homolog deleted on chromosome 10

qRT–PCR

quantitative real‐time polymerase chain reaction

TCGA

the Cancer Genome Atlas

USP

the ubiquitin–proteasome system

VM

vasculogenic mimicry

     

Downregulation of miR‐326 and its host gene β‐arrestin1 induces pro‐survival activity of E2F1 and promotes medulloblastoma growth

Persistent mortality rates of medulloblastoma (MB) and severe side effects of the current therapies require the definition of the molecular mechanisms that contribute to tumor progression. Using cultured MB cancer stem cells and xenograft tumors generated in mice, we show that low expression of miR‐326 and its host gene β‐arrestin1 (ARRB1) promotes tumor growth enhancing the E2F1 pro‐survival function. Our models revealed that miR‐326 and ARRB1 are controlled by a bivalent domain, since the H3K27me3 repressive mark is found at their regulatory region together with the activation‐associated H3K4me3 mark. High levels of EZH2, a feature of MB, are responsible for the presence of H3K27me3. Ectopic expression of miR‐326 and ARRB1 provides hints into how their low levels regulate E2F1 activity. MiR‐326 targets E2F1 mRNA, thereby reducing its protein levels; ARRB1, triggering E2F1 acetylation, reverses its function into pro‐apoptotic activity. Similar to miR‐326 and ARRB1 overexpression, we also show that EZH2 inhibition restores miR‐326/ARRB1 expression, limiting E2F1 pro‐proliferative activity. Our results reveal a new regulatory molecular axis critical for MB progression.

Abbreviations

ARRB1

β‐arrestin1

BTC

bulk tumor cell

CSCs

cancer stem cells

EZH2

enhancer of zeste homolog 2

GCP

granule cell progenitors

MB

medulloblastoma

OFC

oncosphere‐forming cell

     

TGFβ promotes YAP‐dependent AXL induction in mesenchymal‐type lung cancer cells

The acquisition of chemoresistance remains a major cause of cancer mortality due to the limited accessibility of targeted or immune therapies. However, given that severe alterations of molecular features during epithelial‐to‐mesenchymal transition (EMT) lead to acquired chemoresistance, emerging studies have focused on identifying targetable drivers associated with acquired chemoresistance. Particularly, AXL, a key receptor tyrosine kinase that confers resistance against targets and chemotherapeutics, is highly expressed in mesenchymal cancer cells. However, the underlying mechanism of AXL induction in mesenchymal cancer cells is poorly understood. Our study revealed that the YAP signature, which was highly enriched in mesenchymal‐type lung cancer, was closely correlated to AXL expression in 181 lung cancer cell lines. Moreover, using isogenic lung cancer cell pairs, we also found that doxorubicin treatment induced YAP nuclear translocation in mesenchymal‐type lung cancer cells to induce AXL expression. Additionally, the concurrent activation of TGFβ signaling coordinated YAP‐dependent AXL expression through SMAD4. These data suggest that crosstalk between YAP and the TGFβ/SMAD axis upon treatment with chemotherapeutics might be a promising target to improve chemosensitivity in mesenchymal‐type lung cancer.

Abbreviations

AUC

area under the curve

AXL

AXL receptor tyrosine kinase

BCL2

B‐cell lymphoma 2

CTD2

cancer target discovery and development

CTGF

connective tissue growth factor

DEG

differentially expressed genes

DOXO

doxorubicin

EMT

epithelial–mesenchymal transition

Eto

etoposide

FDA

Food and Drug Administration

ITGB3

integrin beta‐3

MAPK

mitogen‐activated protein kinase

MMP2

matrix metalloproteinase‐2

MMP9

matrix metalloproteinase‐9

mRNA

messenger RNA

NF‐κB

nuclear factor kappa‐light‐chain‐enhancer of activated B cells

SBE

SMAD binding element

SERPINE1

serpin family E member 1

siRNA

small interfering RNA

ssGSEA

single‐sample gene set enrichment analysis

TCGA

The Cancer Genome Atlas

TGFβ

transforming growth factor beta

YAP

Yes‐associated protein

YAP8SA

mutants of inhibitory phosphorylation site at eight serine to Alanine of YAP

ZEB1

zinc finger E‐box binding homeobox 1

ZEB2

zinc finger E‐box‐binding homeobox 2

     

Downregulation of Glutamine Synthetase,not glutaminolysis,is responsible for glutamine addiction in Notch1‐driven acute lymphoblastic leukemia

The cellular receptor Notch1 is a central regulator of T‐cell development, and as a consequence, Notch1 pathway appears upregulated in > 65% of the cases of T‐cell acute lymphoblastic leukemia (T‐ALL). However, strategies targeting Notch1 signaling render only modest results in the clinic due to treatment resistance and severe side effects. While many investigations reported the different aspects of tumor cell growth and leukemia progression controlled by Notch1, less is known regarding the modifications of cellular metabolism induced by Notch1 upregulation in T‐ALL. Previously, glutaminolysis inhibition has been proposed to synergize with anti‐Notch therapies in T‐ALL models. In this work, we report that Notch1 upregulation in T‐ALL induced a change in the metabolism of the important amino acid glutamine, preventing glutamine synthesis through the downregulation of glutamine synthetase (GS). Downregulation of GS was responsible for glutamine addiction in Notch1‐driven T‐ALL both in vitro and in vivo. Our results also confirmed an increase in glutaminolysis mediated by Notch1. Increased glutaminolysis resulted in the activation of the mammalian target of rapamycin complex 1 (mTORC1) pathway, a central controller of cell growth. However, glutaminolysis did not play any role in Notch1‐induced glutamine addiction. Finally, the combined treatment targeting mTORC1 and limiting glutamine availability had a synergistic effect to induce apoptosis and to prevent Notch1‐driven leukemia progression. Our results placed glutamine limitation and mTORC1 inhibition as a potential therapy against Notch1‐driven leukemia.

Abbreviations

7‐AAD

7‐Aminoactinomycin D

BPTES

bis‐2‐(5‐phenylacetamido‐1,2,4‐thiadiazol‐2‐yl)ethyl sulfide

DON

diazo‐5‐oxo‐L‐norleucine

ECAR

extracellular acidification rate

GDH

glutamate dehydrogenase

GLS

glutaminase

GS

glutamine synthetase

GSI

γ‐secretase inhibitor

MSO

L‐methionine sulfoximine

mTORC1

mammalian target of rapamycin complex 1

NICD

Notch intracellular domain

PI

propidium iodide

RAP

rapamycin

T‐ALL

T‐cell acute lymphoblastic leukemia

TCA

tricarboxylic acid

αKG

α‐ketoglutarate

     

A cancer‐associated CDKN1B mutation induces p27 phosphorylation on a novel residue: a new mechanism for tumor suppressor loss‐of‐function

CDKN1B haploinsufficiency promotes the development of several human cancers. The gene encodes p27^Kip1, a protein playing pivotal roles in the control of growth, differentiation, cytoskeleton dynamics, and cytokinesis. CDKN1B haploinsufficiency has been associated with chromosomal or gene aberrations. However, very few data exist on the mechanisms by which CDKN1B missense mutations facilitate carcinogenesis. Here, we report a functional study on a cancer‐associated germinal p27^Kip1 variant, namely glycine9‐>arginine‐p27^Kip1 (G9R‐p27^Kip1) identified in a parathyroid adenoma. We unexpectedly found that G9R‐p27^Kip1 lacks the major tumor suppressor activities of p27^Kip1 including its antiproliferative and pro‐apoptotic functions. In addition, G9R‐p27^Kip1 transfection in cell lines induces the formation of more numerous and larger spheres when compared to wild‐type p27^Kip1‐transfected cells. We demonstrated that the mutation creates a consensus sequence for basophilic kinases causing a massive phosphorylation of G9R‐p27^Kip1 on S12, a residue normally never found modified in p27^Kip1. The novel S12 phosphorylation appears responsible for the loss of function of G9R‐p27^Kip1 since S12AG9R‐p27^Kip1 recovers most of the p27^Kip1 tumor suppressor activities. In addition, the expression of the phosphomimetic S12D‐p27^Kip1 recapitulates G9R‐p27^Kip1 properties. Mechanistically, S12 phosphorylation enhances the nuclear localization of the mutant protein and also reduces its cyclin‐dependent kinase (CDK)2/CDK1 inhibition activity. To our knowledge, this is the first reported case of quantitative phosphorylation of a p27^Kip1 variant on a physiologically unmodified residue associated with the loss of several tumor suppressor activities. In addition, our findings demonstrate that haploinsufficiency might be due to unpredictable post‐translational modifications due to generation of novel consensus sequences by cancer‐associated missense mutations.

Abbreviations

1D/WB

monodimensional western blotting

2D/WB

two‐dimensional western blotting

CDK

cyclin‐dependent kinase

CHX

cycloheximide

G9R‐p27

glycine9‐>arginine‐p27

IUPs

intrinsically unstructured proteins

mAbs

monoclonal antibodies

MEN

multiple endocrine neoplasia

MENX

multiple endocrine neoplasia X

PTMs

post‐translational modifications

rAbs

rabbit antibodies

TSG

tumor suppressor gene

wt‐p27

wild‐type p27

     

HRD1 inhibits fatty acid oxidation and tumorigenesis by ubiquitinating CPT2 in triple‐negative breast cancer

Dependence on glutamine and acceleration of fatty acid oxidation (FAO) are both metabolic characteristics of triple‐negative breast cancer (TNBC). With the rapid growth of tumors, accelerated glutamine catabolism depletes local glutamine, resulting in glutamine deficiency. Studies have shown that the use of alternative energy sources, such as fatty acids, enables tumor cells to continue to proliferate rapidly in a glutamine‐deficient microenvironment. However, the detailed mechanisms behind this metabolic change are still unclear. Herein, we identified HRD1 as a regulatory protein for FAO that specifically inhibits TNBC cell proliferation under glutamine‐deficient conditions. Furthermore, we observed that HRD1 expression is significantly downregulated under glutamine deprivation and HRD1 directly ubiquitinates and stabilizes CPT2 through K48‐linked ubiquitination. In addition, the inhibition of CPT2 expression dramatically suppresses TNBC cell proliferation mediated by HRD1 knockdown in vitro and in vivo. Finally, we found that the glutaminase inhibitor CB839 significantly inhibited TNBC cell tumor growth, but not in the HRD1 knock‐downed TNBC cells. These findings provide an invaluable insight into HRD1 as a regulator of lipid metabolism and have important implications for TNBC therapeutic targeting.

Abbreviations

CPT1A

carnitine palmitoyltransferase 1A

CPT2

carnitine palmitoyltransferase 2

FAO

fatty acid oxidation

GLS

glutaminase

HRD1

HMG‐CoA reductase degradation protein 1

LC‐MS

liquid chromatography mass spectrometry

TNBC

triple‐negative breast cancer