bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from Intron 1–Mediated Regulation of EGFR Expression in EGFR-Dependent Malignancies Is Mediated by AP-1 and BET Proteins
Authors: Jameson Nathan M. , Ma Jianhui , Benitez Jorge , Izurieta Alejandro , Han Jee Yun , Mendez Robert , Parisian Alison , Furnari Frank
Published: April 2023

Abstract
The epidermal growth factor receptor (EGFR) is overexpressed in numerous solid tumors and is the subject of extensive therapeutic efforts. Much of the research on EGFR is focused on protein dynamics and downstream signaling; however, few studies have explored its transcriptional regulation. Here, we identified two enhancers (CE1 and CE2) present within the first intron of the EGFR gene in models of glioblastoma (GBM) and head and neck squamous cell carcinoma (HNSCC). CE1 and CE2 contain open chromatin and H3K27Ac histone marks, enhance transcription in reporter assays, and interact with the EGFR promoter. Enhancer genetic deletion by CRISPR/Cas9 significantly reduces EGFR transcript levels, with double deletion exercising an additive effect. Targeted repression of CE1 and CE2 by dCas9-KRAB demonstrates repression of transcription similar to that of genomic deletion. We identify AP-1 transcription factor family members in concert with BET bromodomain proteins as modulators of CE1 and CE2 activity in HNSCC and GBM through de novo motif identification and validate their presence. Genetic inhibition of AP-1 or pharmacologic disruption of BET/AP-1 binding results in downregulated EGFR protein and transcript levels, confirming a role for these factors in CE1 and CE2. Our results identify and characterize these novel enhancers, shedding light on the role that epigenetic mechanisms play in regulating EGFR transcription in EGFR-dependent cancers.
Implications:
We identify critical constituent enhancers present in the first intron of the EGFR gene, and provide a rationale for therapeutic targeting of EGFR intron 1 enhancers through perturbation of AP-1 and BET in EGFR-positive malignancies.
Data from Identification and Characterization of Oncogenic SOS1 Mutations in Lung Adenocarcinoma
Authors: Cai Diana , Choi Peter S. , Gelbard Maya , Meyerson Matthew
Published: April 2023

Abstract
Lung adenocarcinomas are characterized by mutations in the receptor tyrosine kinase (RTK)/Ras/Raf pathway, with up to 75% of cases containing mutations in known driver genes. However, the driver alterations in the remaining cases are yet to be determined. Recent exome sequencing analysis has identified SOS1, encoding a guanine nucleotide exchange factor, as significantly mutated in lung adenocarcinomas lacking canonical oncogenic RTK/Ras/Raf pathway mutations. Here, we demonstrate that ectopic expression of lung adenocarcinoma–derived mutants of SOS1 induces anchorage-independent cell growth in vitro and tumor formation in vivo. Biochemical experiments suggest that these mutations lead to overactivation of the Ras pathway, which can be suppressed by mutations that disrupt either the Ras-GEF or putative Rac-GEF activity of SOS1. Transcriptional profiling reveals that the expression of mutant SOS1 leads to the upregulation of MYC target genes and genes associated with Ras transformation. Furthermore, we demonstrate that an AML cancer cell line harboring a lung adenocarcinoma–associated mutant SOS1 is dependent on SOS1 for survival and is also sensitive to MEK inhibition. Our work provides experimental evidence for the role of SOS1 as an oncogene and suggests a possible therapeutic strategy to target SOS1-mutated cancers.
Implications:
This study demonstrates that SOS1 mutations found in lung adenocarcinoma are oncogenic and that MEK inhibition may be a therapeutic avenue for the treatment of SOS1-mutant cancers.
Data from BRCA1-Interacting Protein OLA1 Requires Interaction with BARD1 to Regulate Centrosome Number
Authors: Yoshino Yuki , Qi Huicheng , Fujita Hiroki , Shirota Matsuyuki , Abe Shun , Komiyama Yuhei , Shindo Kazuha , Nakayama Masahiro , Matsuzawa Ayako , Kobayashi Akihiro , Ogoh Honami , Watanabe Toshio , Ishioka Chikashi , Chiba Natsuko
Published: April 2023

Abstract
BRCA1 functions as a tumor suppressor in DNA repair and centrosome regulation. Previously, Obg-like ATPase 1 (OLA1) was shown to interact with BARD1, a heterodimer partner of BRCA1. OLA1 binds to BRCA1, BARD1, and γ-tubulin and functions in centrosome regulation. This study determined that overexpression of wild-type OLA1 (OLA1-WT) caused centrosome amplification due to centriole overduplication in mammary tissue–derived cells. Centrosome amplification induced by overexpression of the cancer-derived OLA1 mutant, which is deficient at regulating centrosome number, occurred in significantly fewer cells than in that induced by overexpression of OLA1-WT. Thus, it was hypothesized that overexpression of OLA1 with normal function efficiently induces centrosome amplification, but not that of OLA1 mutants, which are deficient at regulating centrosome number. We analyzed whether overexpression of OLA1 missense mutants of nine candidate phosphorylation residues, three residues modified with acetylation, and two ATP-binding residues caused centrosome amplification and identified five missense mutants that are deficient in the regulation of centrosome number. Three of them did not bind to BARD1. Two phosphomimetic mutations restored the binding to BARD1 and the efficient centrosome amplification by their overexpression. Knockdown and overexpression of BARD1 also caused centrosome amplification. BARD1 mutant reported in cancer failed to bind to OLA1 and rescue the BARD1 knockdown-induced centrosome amplification and reduced its centrosomal localization. Combined, these data reveal that the OLA1–BARD1 interaction is important for the regulation of centrosome number.

Implications: Regulation of centrosome number by BRCA1/BARD1 together with OLA1 is important for the genome integrity to prevent tumor development. Mol Cancer Res; 16(10); 1499–511. ©2018 AACR.
Data from Unraveling the Cellular Mechanism of Assembling Cholesterols for Selective Cancer Cell Death
Authors: Wang Huaimin , Feng Zhaoqianqi , Yang Cuihong , Liu Jinjian , Medina Jamie E. , Aghvami S. Ali , Dinulescu Daniela M. , Liu Jianfeng , Fraden Seth , Xu Bing
Published: April 2023

Abstract
Acquired drug resistance remains a challenge in chemotherapy. Here we show enzymatic, in situ assembling of cholesterol derivatives to act as polypharmaceuticals for selectively inducing death of cancer cells via multiple pathways and without inducing acquired drug resistance. A conjugate of tyrosine and cholesterol (TC), formed by enzyme-catalyzed dephosphorylation of phosphorylate TC, self-assembles selectively on or in cancer cells. Acting as polypharmaceuticals, the assemblies of TC augment lipid rafts, aggregate extrinsic cell death receptors (e.g., DR5, CD95, or TRAILR), modulate the expression of oncoproteins (e.g., Src and Akt), disrupt the dynamics of cytoskeletons (e.g., actin filaments or microtubules), induce endoplasmic reticulum stress, and increase the production of reactive oxygen species, thus resulting in cell death and preventing acquired drug resistance. Moreover, the assemblies inhibit the growth of platinum-resistant ovarian cancer tumor in a murine model. This work illustrates the use of instructed assembly (iA) in cellular environment to form polypharmaceuticals in situ that not only interact with multiple proteins, but also modulate membrane dynamics for developing novel anticancer therapeutics.
Implications:
As a multifaceted strategy for controlling cancer cell death, iA minimized acquired resistance of cancer cells, which is a new strategy to amplify the genetic difference between cancer and normal cells and provides a promise for overcoming drug resistance in cancer therapy.

Visual Overview: http://mcr.aacrjournals.org/content/molcanres/17/4/907/F1.large.jpg.
Data from Aurora Kinase A Promotes AR Degradation via the E3 Ligase CHIP
Authors: Sarkar Sukumar , Brautigan David L. , Larner James M.
Published: April 2023

Abstract
Reducing the levels of the androgen receptor (AR) is one of the most viable approaches to combat castration-resistant prostate cancer. Previously, we observed that proteasomal-dependent degradation of AR in response to 2-methoxyestradiol (2-ME) depends primarily on the E3 ligase C-terminus of HSP70-interacting protein (STUB1/CHIP). Here, 2-ME stimulation activates CHIP by phosphorylation via Aurora kinase A (AURKA). Aurora A kinase inhibitors and RNAi knockdown of Aurora A transcript selectively blocked CHIP phosphorylation and AR degradation. Aurora A kinase is activated by 2-ME in the S-phase as well as during mitosis, and phosphorylates CHIP at S273. Prostate cancer cells expressing an S273A mutant of CHIP have attenuated AR degradation upon 2-ME treatment compared with cells expressing wild-type CHIP, supporting the idea that CHIP phosphorylation by Aurora A activates its E3 ligase activity for the AR. These results reveal a novel 2-ME→Aurora A→CHIP→AR pathway that promotes AR degradation via the proteasome that may offer novel therapeutic opportunities for prostate cancer. Mol Cancer Res; 15(8); 1063–72. ©2017 AACR.
Data from Novel lncRNA LINC00844 Regulates Prostate Cancer Cell Migration and Invasion through AR Signaling
Authors: Lingadahalli Shreyas , Jadhao Sudhir , Sung Ying Ying , Chen Mi , Hu Lingling , Chen Xin , Cheung Edwin
Published: April 2023

Abstract
The human genome is mostly transcribed, yielding a rich repository of noncoding transcripts that are involved in a myriad of biological processes including cancer. However, how many noncoding transcripts such as long noncoding RNAs (lncRNA) function in cancer is still unclear. This study identified a novel set of clinically relevant androgen-regulated lncRNAs in prostate cancer. Among this group, LINC00844 was demonstrated to be a direct androgen-regulated target that is actively transcribed in androgen receptor (AR)–dependent prostate cancer cells. The expression of LINC00844 is higher in normal prostate compared with malignant and metastatic prostate cancer clinical specimens, and patients with low expression had a poor prognosis and significantly increased biochemical recurrence, suggesting LINC00844 functions in suppressing tumor progression and metastasis. Indeed, in vitro loss-of-function studies revealed that LINC00844 prevents prostate cancer cell migration and invasion. Moreover, findings from gene expression profiling analysis indicated that LINC00844 functions in trans, affecting global androgen-regulated gene transcription. Mechanistic evidence reveals that LINC00844 is important in facilitating AR binding to the chromatin. Finally, LINC00844 mediates its phenotypic effects in part by activating the expression of NDRG1, a crucial cancer metastasis suppressor. Collectively, LINC00844 is a novel coregulator of AR that plays a central role in the androgen transcriptional network and the development and progression of prostate cancer.
Implications:
This study highlights the function of the lncRNA, LINC00844, in regulating global AR-regulated genes in prostate cancer by modulating AR binding to chromatin.
Data from HBx-K130M/V131I Promotes Liver Cancer in Transgenic Mice via AKT/FOXO1 Signaling Pathway and Arachidonic Acid Metabolism
Authors: Chiu Amy P. , Tschida Barbara R. , Sham Tung-Ting , Lo Lilian H. , Moriarity Branden S. , Li Xiao-Xiao , Lo Regina C. , Hinton David E. , Rowlands Dewi K. , Chan Chi-On , Mok Daniel K.W. , Largaespada David A. , Warner Nadia , Keng Vincent W.
Published: April 2023

Abstract
Chronic hepatitis B viral (HBV) infection remains a high underlying cause for hepatocellular carcinoma (HCC) worldwide, while the genetic mechanisms behind this remain unclear. This study elucidated the mechanisms contributing to tumor development induced by the HBV X (HBx) gene of predominantly Asian genotype B HBV and its common HBx variants. To compare the potential tumorigenic effects of K130M/V131I (Mut) and wild-type (WT) HBx on HCC, the Sleeping Beauty (SB) transposon system was used to deliver HBx Mut and WT into the livers of fumarylacetoacetate hydrolase (Fah)-deficient mice and in the context of transformation related protein 53 (Trp53) deficiency. From our results, HBx Mut had a stronger tumorigenic effect than its WT variant. Also, inflammation, necrosis, and fibrosis were evident in HBx experimental animals. Reduction of forkhead box O1 (FOXO1) with increased phosphorylation of upstream serine/threonine kinase (AKT) was detected under HBx Mut overexpression. Thus, it is proposed that HBx Mut enhances disease progression by reducing FOXO1 via phosphorylation of AKT. At the metabolomic level, HBx altered the expression of genes that participated in arachidonic acid (AA) metabolism, as a result of inflammation via accumulation of proinflammatory factors such as prostaglandins and leukotriene in liver. Taken together, the increased rate of HCC observed in chronic hepatitis B patients with K130M/V131I-mutated X protein, may be due to changes in AA metabolism and AKT/FOXO1 signaling.
Implications:
Our findings suggested that HBx-K130M/V131I–mutant variant promoted HCC progression by activating AKT/FOXO1 pathway and inducing stronger inflammation in liver via AA metabolism.
Data from p53 Maintains Baseline Expression of Multiple Tumor Suppressor Genes
Authors: Pappas Kyrie , Xu Jia , Zairis Sakellarios , Resnick-Silverman Lois , Abate Francesco , Steinbach Nicole , Ozturk Sait , Saal Lao H. , Su Tao , Cheung Pamela , Schmidt Hank , Aaronson Stuart , Hibshoosh Hanina , Manfredi James , Rabadan Raul , Parsons Ramon
Published: April 2023

Abstract
TP53 is the most commonly mutated tumor suppressor gene and its mutation drives tumorigenesis. Using ChIP-seq for p53 in the absence of acute cell stress, we found that wild-type but not mutant p53 binds and activates numerous tumor suppressor genes, including PTEN, STK11(LKB1), miR-34a, KDM6A(UTX), FOXO1, PHLDA3, and TNFRSF10B through consensus binding sites in enhancers and promoters. Depletion of p53 reduced expression of these target genes, and analysis across 18 tumor types showed that mutation of TP53 associated with reduced expression of many of these genes. Regarding PTEN, p53 activated expression of a luciferase reporter gene containing the p53-consensus site in the PTEN enhancer, and homozygous deletion of this region in cells decreased PTEN expression and increased growth and transformation. These findings show that p53 maintains expression of a team of tumor suppressor genes that may together with the stress-induced targets mediate the ability of p53 to suppress cancer development. p53 mutations selected during tumor initiation and progression, thus, inactivate multiple tumor suppressor genes in parallel, which could account for the high frequency of p53 mutations in cancer.

Implications: In this study, we investigate the activities of p53 under normal low-stress conditions and discover that p53 is capable of maintaining the expression of a group of important tumor suppressor genes at baseline, many of which are haploinsufficient, which could contribute to p53-mediated tumor suppression. Mol Cancer Res; 15(8); 1051–62. ©2017 AACR.
Data from Adaptive Resistance to EGFR-Targeted Therapy by Calcium Signaling in NSCLC Cells
Authors: Mulder Celine , Prust Nadine , van Doorn Sander , Reinecke Maria , Kuster Bernhard , van Bergen en Henegouwen Paul , Lemeer Simone
Published: April 2023

Abstract
Targeted therapies against oncogenic receptor tyrosine kinases (RTK) show promising results in the clinic. Unfortunately, despite the initial positive response, most patients develop therapeutic resistance. Most research has focused on acquired resistance occurring after an extensive time of treatment; however, the question remains as to how cells can survive an initial treatment, as early resistance to apoptosis will enable cells to develop any growth-stimulating mechanism. Here, the non–small cell lung cancer (NSCLC) PC9 cell line was used to systematically profile, by mass spectrometry, changes in the proteome, kinome, and phosphoproteome during early treatment with the EGFR inhibitor afatinib. Regardless of the response, initial drug-sensitive cells rapidly adapt to targeted therapy, and within days, cells regained the capacity to proliferate, despite persisting target inhibition. These data reveal a rapid reactivation of mTOR and MAPK signaling pathways after initial inhibition and an increase in abundance and activity of cytoskeleton and calcium signaling–related proteins. Pharmacologic inhibition of reactivated pathways resulted in increased afatinib efficacy. However more strikingly, cells that were restricted from accessing extracellular calcium were extremely sensitive to afatinib treatment. These findings were validated using three additional inhibitors tested in four different NSCLC cell lines, and the data clearly indicated a role for Ca2+ signaling during the development of adaptive resistance. From a therapeutic point of view, the increased inhibitor efficacy could limit or even prevent further resistance development.

Implications: Combined targeting of calcium signaling and RTKs may limit drug resistance and improve treatment efficacy. Mol Cancer Res; 16(11); 1773–84. ©2018 AACR.
Data from Abemaciclib Is Effective Against Pancreatic Cancer Cells and Synergizes with HuR and YAP1 Inhibition
Authors: Dhir Teena , Schultz Christopher W. , Jain Aditi , Brown Samantha Z. , Haber Alex , Goetz Austin , Xi Chunhua , Su Gloria H. , Xu Liang , Posey James , Jiang Wei , Yeo Charles J. , Golan Talia , Pishvaian Michael J. , Brody Jonathan R.
Published: April 2023

Abstract
Mutation or promoter hypermethylation of CDKN2A is found in over 90% of pancreatic ductal adenocarcinomas (PDAC) and leads to loss of function of cell-cycle inhibitors p16 (INK4A) and p14 (ARF) resulting in unchecked proliferation. The CDK4/6 inhibitor, abemaciclib, has nanomolar IC50s in PDAC cell lines and decreases growth through inhibition of phospho-Rb (pRb), G1 cell-cycle arrest, apoptosis, and the senescent phenotype detected with β-galactosidase staining and relevant mRNA elevations. Daily abemaciclib treatments in mouse PDAC xenograft studies were safe and demonstrated a 3.2-fold decrease in tumor volume compared with no treatment (P < 0.0001) accompanying a decrease in both pRb and Ki67. We determined that inhibitors of HuR (ELAVL1), a prosurvival mRNA stability factor that regulates cyclin D1, and an inhibitor of Yes-Associated Protein 1 (YAP1), a pro-oncogenic, transcriptional coactivator important for CDK6 and cyclin D1, were both synergistic with abemaciclib. Accordingly, siRNA oligonucleotides targeted against HuR, YAP1, and their common target cyclin D1, validated the synergy studies. In addition, we have seen increased sensitivity to abemaciclib in a PDAC cell line that harbors a loss of the ELAVL1 gene via CRISP-Cas9 technology. As an in vitro model for resistance, we investigated the effects of long-term abemaciclib exposure. PDAC cells chronically cultured with abemaciclib displayed a reduction in cellular growth rates (GR) and coresistance to gemcitabine and 5-fluorouracil (5-FU), but not to HuR or YAP1 inhibitors as compared with no treatment controls. We believe that our data provide compelling preclinical evidence for an abemaciclib combination–based clinical trial in patients with PDAC.
Implications:
Our data suggest that abemaciclib may be therapeutically relevant for the treatment in PDAC, especially as part of a combination regimen inhibiting YAP1 or HuR.
Data from Biomarker Assessment of HR Deficiency, Tumor BRCA1/2 Mutations, and CCNE1 Copy Number in Ovarian Cancer: Associations with Clinical Outcome Following Platinum Monotherapy
Authors: Stronach Euan A. , Paul James , Timms Kirsten M. , Hughes Elisha , Brown Krystal , Neff Christopher , Perry Michael , Gutin Alexander , El-Bahrawy Mona , Steel Jennifer H. , Liu Xinxue , Lewsley Liz-Anne , Siddiqui Nadeem , Gabra Hani , Lanchbury Jerry S. , Brown Robert
Published: April 2023

Abstract
The current study evaluated three biomarkers [homologous recombination deficiency (HRD), tumor BRCA1/2 (tBRCA) mutations, and CCNE1 copy-number variation (CNV)] in ovarian tumors from patients enrolled on the SCOTROC4 clinical trial for associations with outcome following carboplatin monotherapy. Ovarian tumors (n = 250), with high-grade serous (HGSOC) subgroup analysis (n = 179) were classified as HRD positive (HRD score ≥42 or tBRCA mutation) and as CCNE1 amplification positive (CCNE1 CNV score >2.4). Seventy-four (30%) tumors were HRD positive, including 34 (14%) with tBRCA mutations. Forty-seven (19%) were CCNE1 amplification positive, all of which were tBRCA wild-type. HRD and tBRCA, but not CCNE1 amplification, were significantly associated with CA125 complete response in the entire cohort (HRD, P = 0.00015; tBRCA P = 0.0096), and the HGSOC subgroup (HRD, P = 0.0016; tBRCA P = 0.032). HRD and lack of CCNE1 amplification were associated with improved progression-free survival (PFS) and overall survival (OS) in the full cohort and HGSOC subgroup (HRD, P = 0.00021; CCNE1 status P = 0.038). HRD remained significant for OS and PFS after adjusting for clinical factors, while CCNE1 status only remained significant for PFS. Patients with HRD-positive tumors had greater PFS and OS benefit from platinum dose intensification than HRD-negative tumors (P = 0.049 and P = 0.035, respectively). An alternative exploratory HRD score threshold (≥33 or tBRCA mutation) was also significantly associated with both PFS and OS in the HGSOC subset.

Implications: HRD, tumor BRCA1/2 mutations, and absence of CCNE1 amplification are associated with improved survival of ovarian cancer patients treated with platinum monotherapy and HRD-positive patients may benefit from platinum dose intensification. Mol Cancer Res; 16(7); 1103–11. ©2018 AACR.
Data from CDK4/6 Therapeutic Intervention and Viable Alternative to Taxanes in CRPC
Authors: Stice James P. , Wardell Suzanne E. , Norris John D. , Yllanes Alexander P. , Alley Holly M. , Haney Victoria O. , White Hannah S. , Safi Rachid , Winter Peter S. , Cocce Kimberly J. , Kishton Rigel J. , Lawrence Scott A. , Strum Jay C. , McDonnell Donald P.
Published: April 2023

Abstract
Resistance to second-generation androgen receptor (AR) antagonists and CYP17 inhibitors in patients with castration-resistant prostate cancer (CRPC) develops rapidly through reactivation of the androgen signaling axis and has been attributed to AR overexpression, production of constitutively active AR splice variants, or the selection for AR mutants with altered ligand-binding specificity. It has been established that androgens induce cell-cycle progression, in part, through upregulation of cyclin D1 (CCND1) expression and subsequent activation of cyclin-dependent kinases 4 and 6 (CDK4/6). Thus, the efficacy of the newly described CDK4/6 inhibitors (G1T28 and G1T38), docetaxel and enzalutamide, was evaluated as single agents in clinically relevant in vitro and in vivo models of hormone-sensitive and treatment-resistant prostate cancer. CDK4/6 inhibition (CDK4/6i) was as effective as docetaxel in animal models of treatment-resistant CRPC but exhibited significantly less toxicity. The in vivo effects were durable and importantly were observed in prostate cancer cells expressing wild-type AR, AR mutants, and those that have lost AR expression. CDK4/6i was also effective in prostate tumor models expressing the AR-V7 variant or the AR F876L mutation, both of which are associated with treatment resistance. Furthermore, CDK4/6i was effective in prostate cancer models where AR expression was lost. It is concluded that CDK4/6 inhibitors are a viable alternative to taxanes as therapeutic interventions in endocrine therapy–refractory CRPC.

Implications: The preclinical efficacy of CDK4/6 monotherapy observed here suggests the need for near-term clinical studies of these agents in advanced prostate cancer. Mol Cancer Res; 15(6); 660–9. ©2017 AACR.
Data from Protein Kinase D–Dependent Downregulation of Immediate Early Genes through Class IIA Histone Deacetylases in Acute Lymphoblastic Leukemia
Authors: Sun Guangyan , Shvab Anna , Leclerc Guy J. , Li Bin , Beckedorff Felipe , Shiekhattar Ramin , Barredo Julio C.
Published: April 2023

Abstract
Acute lymphoblastic leukemia (ALL) is a leading cause of cancer-related death in children and adolescents, and cure rates for relapsed/refractory ALL remain dismal, highlighting the need for novel targeted therapies. To identify genome-wide metabolic-stress regulated genes, we used RNA-sequencing in ALL cells treated with AICAR, an AMPK activator. RNA-sequencing identified the immediate early genes (IEGs) as a subset of genes downregulated by AICAR. We show that AICAR-induced IEGs downregulation was blocked by an adenosine uptake inhibitor indicating AICAR was responsible for IEGs reprogramming. Using pharmacologic and genetic models we established this mechanism was AMPK-independent. Further investigations using kinase assays, PKD/PKC inhibitors and rescue experiments, demonstrated that AICAR directly inhibited PKD kinase activity and identified PKD as responsible for IEGs downregulation. Mechanistically, PKD inhibition suppressed phosphorylation and nuclear export of class IIa HDACs, which lowered histone H3 acetylation and decreased NFκB(p65) recruitment to IEGs promoters. Finally, PKD inhibition induced apoptosis via DUSP1/DUSP6 downregulation eliciting a DNA damage response. More importantly, ALL patient cells exhibited the same PKD-HDACs-IEGs–mediated mechanism. As proof of principle of the therapeutic potential of targeting PKD, we established the in vivo relevance of our findings using an NSG ALL mouse model. In conclusion, we identified a previously unreported PKD-dependent survival mechanism in response to AICAR-induced cellular stress in ALL through regulation of DUSPs and IEGs' expression.
Implications:
PKD mediates early transcriptional responses in ALL cells as an adaptive survival mechanism to overcome cellular stress.
Data from Meta-Analysis and Systematic Review of the Genomics of Mucosal Melanoma
Authors: Broit Natasa , Johansson Peter A. , Rodgers Chloe B. , Walpole Sebastian T. , Newell Felicity , Hayward Nicholas K. , Pritchard Antonia L.
Published: April 2023

Abstract
Mucosal melanoma is a rare subtype of melanoma. To date, there has been no comprehensive systematic collation and statistical analysis of the aberrations and aggregated frequency of driver events across multiple studies. Published studies using whole genome, whole exome, targeted gene panel, or individual gene sequencing were identified. Datasets from these studies were collated to summarize mutations, structural variants, and regions of copy-number alteration. Studies using next-generation sequencing were divided into the “main” cohort (n = 173; fresh-frozen samples), “validation” cohort (n = 48; formalin-fixed, paraffin-embedded samples) and a second “validation” cohort comprised 104 tumors sequenced using a targeted panel. Studies assessing mutations in BRAF, KIT, and NRAS were summarized to assess hotspot mutations. Statistical analysis of the main cohort variant data revealed KIT, NF1, BRAF, NRAS, SF3B1, and SPRED1 as significantly mutated genes. ATRX and SF3B1 mutations occurred more commonly in lower anatomy melanomas and CTNNB1 in the upper anatomy. NF1, PTEN, CDKN2A, SPRED1, ATM, CHEK2, and ARID1B were commonly affected by chromosomal copy loss, while TERT, KIT, BRAF, YAP1, CDK4, CCND1, GAB2, MDM2, SKP2, and MITF were commonly amplified. Further notable genomic alterations occurring at lower frequencies indicated commonality of signaling networks in tumorigenesis, including MAPK, PI3K, Notch, Wnt/β-catenin, cell cycle, DNA repair, and telomere maintenance pathways. This analysis identified genomic aberrations that provide some insight to the way in which specific pathways may be disrupted.
Implications:
Our analysis has shown that mucosal melanomas have a diverse range of genomic alterations in several biological pathways.
Data from Proteomic Analysis of Src Family Kinase Phosphorylation States in Cancer Cells Suggests Deregulation of the Unique Domain
Authors: Ruiz-Saenz Ana , Zahedi Farima , Peterson Elliott , Yoo Ashley , Dreyer Courtney A. , Spassov Danislav S. , Oses-Prieto Juan , Burlingame Alma , Moasser Mark M.
Published: April 2023

Abstract
The Src family kinases (SFK) are homologs of retroviral oncogenes, earning them the label of proto-oncogenes. Their functions are influenced by positive and negative regulatory tyrosine phosphorylation events and inhibitory and activating intramolecular and extramolecular interactions. This regulation is disrupted in their viral oncogene counterparts. However, in contrast to most other proto-oncogenes, the genetic alteration of these genes does not seem to occur in human tumors and how and whether their functions are altered in human cancers remain to be determined. To look for proteomic-level alterations, we took a more granular look at the activation states of SFKs based on their two known regulatory tyrosine phosphorylations, but found no significant differences in their activity states when comparing immortalized epithelial cells with cancer cells. SFKs are known to have other less well-studied phosphorylations, particularly within their unstructured N-terminal unique domains (UD), although their role in cancers has not been explored. In comparing panels of epithelial cells with cancer cells, we found a decrease in S17 phosphorylation in the UD of Src in cancer cells. Dephosphorylated S17 favors the dimerization of Src that is mediated through the UD and suggests increased Src dimerization in cancers. These data highlight the important role of the UD of Src and suggest that a deeper understanding of proteomic-level alterations of the unstructured UD of SFKs may provide considerable insights into how SFKs are deregulated in cancers.
Implications:
This work highlights the role of the N-terminal UD of Src kinases in regulating their signaling functions and possibly in their deregulation in human cancers.
Data from Palbociclib Renders Human Papilloma Virus–Negative Head and Neck Squamous Cell Carcinoma Vulnerable to the Senolytic Agent Navitoclax
Authors: Gadsden Nicholas J. , Fulcher Cory D. , Li Daniel , Shrivastava Nitisha , Thomas Carlos , Segall Jeffrey E. , Prystowsky Michael B. , Schlecht Nicolas F. , Gavathiotis Evripidis , Ow Thomas J.
Published: April 2023

Abstract
We demonstrate that inhibition of cyclin-dependent kinases 4/6 (CDK4/6) leads to senescence in human papillomavirus (HPV)–negative (−) head and neck squamous cell carcinoma (HNSCC), but not in HPV-positive (+) HNSCC. The BCL-2 family inhibitor, navitoclax, has been shown to eliminate senescent cells effectively. We evaluated the efficacy of combining palbociclib and navitoclax in HPV− HNSCC. Three HPV− HNSCC cell lines (CAL27, HN31, and PCI15B) and three HPV+ HNSCC cell lines (UPCI-SCC-090, UPCI-SCC-154, and UM-SCC-47) were treated with palbociclib. Treatment drove reduced expression of phosphorylated Rb (p-Rb) and phenotypic evidence of senescence in all HPV− cell lines, whereas HPV+ cell lines did not display a consistent response by Rb or p-Rb and did not exhibit morphologic changes of senescence in response to palbociclib. In addition, treatment of HPV− cells with palbociclib increased both β-galactosidase protein expression and BCL-xL protein expression compared with untreated controls in HPV− cells. Co-expression of β-galactosidase and BCL-xL occurred consistently, indicating elevated BCL-xL expression in senescent cells. Combining palbociclib with navitoclax led to decreased HPV− HNSCC cell survival and led to increased apoptosis levels in HPV− cell lines compared with each agent given alone.
Implications:
This work exploits a key genomic hallmark of HPV− HNSCC (CDKN2A disruption) using palbociclib to induce BCL-xL–dependent senescence, which subsequently causes the cancer cells to be vulnerable to the senolytic agent, navitoclax.
Data from EMT-Derived Alterations in Glutamine Metabolism Sensitize Mesenchymal Breast Cells to mTOR Inhibition
Authors: Karvelsson Sigurdur Trausti , Sigurdsson Arnar , Seip Kotryna , Grinde Maria Tunset , Wang Qiong , Johannsson Freyr , Mælandsmo Gunhild Mari , Moestue Siver Andreas , Rolfsson Ottar , Halldorsson Skarphedinn
Published: April 2023

Abstract
Epithelial-to-mesenchymal transition (EMT) is a fundamental developmental process with strong implications in cancer progression. Understanding the metabolic alterations associated with EMT may open new avenues of treatment and prevention. Here we used 13C carbon analogs of glucose and glutamine to examine differences in their utilization within central carbon and lipid metabolism following EMT in breast epithelial cell lines. We found that there are inherent differences in metabolic profiles before and after EMT. We observed EMT-dependent re-routing of the TCA-cycle, characterized by increased mitochondrial IDH2-mediated reductive carboxylation of glutamine to lipid biosynthesis with a concomitant lowering of glycolytic rates and glutamine-dependent glutathione (GSH) generation. Using weighted correlation network analysis, we identified cancer drugs whose efficacy against the NCI-60 Human Tumor Cell Line panel is significantly associated with GSH abundance and confirmed these in vitro. We report that EMT-linked alterations in GSH synthesis modulate the sensitivity of breast epithelial cells to mTOR inhibitors.
Implications:
EMT in breast cells causes an increased demand for glutamine for fatty acid biosynthesis, altering its contribution to glutathione biosynthesis, which sensitizes the cells to mTOR inhibitors.
Data from The Ubiquitin-Specific Peptidase USP18 Promotes Lipolysis, Fatty Acid Oxidation, and Lung Cancer Growth
Authors: Liu Xi , Lu Yun , Chen Zibo , Liu Xiuxia , Hu Weiguo , Zheng Lin , Chen Yulong , Kurie Jonathan M. , Shi Mi , Mustachio Lisa Maria , Adresson Thorkell , Fox Stephen , Roszik Jason , Kawakami Masanori , Freemantle Sarah J. , Dmitrovsky Ethan
Published: April 2023

Abstract
Ubiquitin specific peptidase 18 (USP18), previously known as UBP43, is the IFN-stimulated gene 15 (ISG15) deconjugase. USP18 removes ISG15 from substrate proteins. This study reports that USP18-null mice (vs. wild-type mice) exhibited lower lipolysis rates, altered fat to body weight ratios, and cold sensitivity. USP18 is a regulator of lipid and fatty acid metabolism. Prior work established that USP18 promotes lung tumorigenesis. We sought to learn whether this occurs through altered lipid and fatty acid metabolism. Loss of USP18 repressed adipose triglyceride lipase (ATGL) expression; gain of USP18 expression upregulated ATGL in lung cancer cells. The E1-like ubiquitin activating enzyme promoted ISG15 conjugation of ATGL and destabilization. Immunoprecipitation assays confirmed that ISG15 covalently conjugates to ATGL. Protein expression of thermogenic regulators was examined in brown fat of USP18-null versus wild-type mice. Uncoupling protein 1 (UCP1) was repressed in USP18-null fat. Gain of USP18 expression augmented UCP1 protein via reduced ubiquitination. Gain of UCP1 expression in lung cancer cell lines enhanced cellular proliferation. UCP1 knockdown inhibited proliferation. Beta-hydroxybutyrate colorimetric assays performed after gain of UCP1 expression revealed increased cellular fatty acid beta-oxidation, augmenting fatty acid beta-oxidation in Seahorse assays. Combined USP18, ATGL, and UCP1 profiles were interrogated in The Cancer Genome Atlas. Intriguingly, lung cancers with increased USP18, ATGL, and UCP1 expression had an unfavorable survival. These findings reveal that USP18 is a pharmacologic target that controls fatty acid metabolism.
Implications:
USP18 is an antineoplastic target that affects lung cancer fatty acid metabolism.
Data from A Novel Therapeutic Strategy Targeting the Mesenchymal Phenotype of Malignant Pleural Mesothelioma by Suppressing LSD1
Authors: Wirawan Aditya , Tajima Ken , Takahashi Fumiyuki , Mitsuishi Yoichiro , Winardi Wira , Hidayat Moulid , Hayakawa Daisuke , Matsumoto Naohisa , Izumi Kenta , Asao Tetsuhiko , Ko Ryo , Shimada Naoko , Takamochi Kazuya , Suzuki Kenji , Abe Masaaki , Hino Okio , Sekido Yoshitaka , Takahashi Kazuhisa
Published: April 2023

Abstract
Malignant pleural mesothelioma (MPM) is a highly aggressive tumor that has a low overall survival; however, no significant treatment advances have been made in the past 15 years. Large-scale molecular studies have identified a poor prognostic subset of MPM linked to the epithelial–mesenchymal transition (EMT) that may contribute toward resistance to chemotherapy, suggesting that EMT could be targeted to treat patients with MPM. Previously, we reported that histone modifiers regulating EMT could be therapeutic targets; therefore, in this study, we investigated whether targeting lysine-specific demethylase 1 (LSD1/KDM1), a histone-modifying enzyme responsible for demethylating histone H3 lysine 4 and lysine 9, could represent a novel therapeutic strategy for MPM. We suppressed LSD1 and investigated the EMT phenotype using EMT marker expression and wound-healing assay; and chemosensitivity using apoptosis assay. We found that suppressing LSD1 induces an epithelial phenotype in sarcomatoid MPM cells, while attenuating the mesenchymal phenotype sensitized MPM cells to cisplatin-induced apoptosis. Subsequent genome-wide identification, comprehensive microarray analysis, and Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) to assess genome-wide changes in chromatin accessibility suggested that LSD1 directly regulates milk fat globulin protein E8 (MFGE8), an integrin ligand that is involved in the FAK pathway. Furthermore, we found that LSD1 regulates the mesenchymal phenotype and apoptosis by activating the FAK–AKT–GSK3β pathway via a positive feedback loop involving MFGE8 and Snail expression, thereby leading to cisplatin resistance.
Implications:
This study suggests that LSD1 regulates the mesenchymal phenotype and apoptosis, and that LSD1 inhibitors could be combined with the cisplatin as a novel therapy for patients with MPM.
Data from Comprehensive Mutational Analysis of the BRCA1-Associated DNA Helicase and Tumor-Suppressor FANCJ/BACH1/BRIP1
Authors: Calvo Jennifer A. , Fritchman Briana , Hernandez Desiree , Persky Nicole S. , Johannessen Cory M. , Piccioni Federica , Kelch Brian A. , Cantor Sharon B.
Published: April 2023

Abstract
FANCJ (BRIP1/BACH1) is a hereditary breast and ovarian cancer (HBOC) gene encoding a DNA helicase. Similar to HBOC genes, BRCA1 and BRCA2, FANCJ is critical for processing DNA inter-strand crosslinks (ICL) induced by chemotherapeutics, such as cisplatin. Consequently, cells deficient in FANCJ or its catalytic activity are sensitive to ICL-inducing agents. Unfortunately, the majority of FANCJ clinical mutations remain uncharacterized, limiting therapeutic opportunities to effectively use cisplatin to treat tumors with mutated FANCJ. Here, we sought to perform a comprehensive screen to identify FANCJ loss-of-function (LOF) mutations. We developed a FANCJ lentivirus mutation library representing approximately 450 patient–derived FANCJ nonsense and missense mutations to introduce FANCJ mutants into FANCJ knockout (K/O) HeLa cells. We performed a high-throughput screen to identify FANCJ LOF mutants that, as compared with wild-type FANCJ, fail to robustly restore resistance to ICL-inducing agents, cisplatin or mitomycin C (MMC). On the basis of the failure to confer resistance to either cisplatin or MMC, we identified 26 missense and 25 nonsense LOF mutations. Nonsense mutations elucidated a relationship between location of truncation and ICL sensitivity, as the majority of nonsense mutations before amino acid 860 confer ICL sensitivity. Further validation of a subset of LOF mutations confirmed the ability of the screen to identify FANCJ mutations unable to confer ICL resistance. Finally, mapping the location of LOF mutations to a new homology model provides additional functional information.
Implications:
We identify 51 FANCJ LOF mutations, providing important classification of FANCJ mutations that will afford additional therapeutic strategies for affected patients.
Data from G9a Promotes Invasion and Metastasis of Non–Small Cell Lung Cancer through Enhancing Focal Adhesion Kinase Activation via NF-κB Signaling Pathway
Authors: Sun Ting , Zhang Keqiang , Pangeni Rajendra P. , Wu Jun , Li Wendong , Du Yong , Guo Yuming , Chaurasiya Shyambabu , Arvanitis Leonidas , Raz Dan J.
Published: April 2023

Abstract
Potential roles of euchromatic histone methyltransferase 2 (EHMT2 or G9a) in invasion and metastasis are not well understood in non–small cell lung cancer (NSCLC). Here, we investigated the effect and underlying mechanisms of G9a and therapeutic implications of targeting G9a in the invasion and metastasis of NSCLC. Overexpression of G9a significantly enhanced in vitro proliferation and invasion, while knockdown of G9a drastically suppressed in vivo growth and metastasis of A549 and H1299 NSCLC cells. Knockdown or inhibition of G9a significantly decreased the expression of focal adhesion kinase (FAK) protein and activation of FAK pathway. In addition, defactinib, a potent FAK inhibitor, partially abolished the G9a-enhanced invasion in these NSCLC cells. Furthermore, targeting G9a was found to suppress NF-κB transcriptional activity in NSCLC cells through stabilizing NF-κB inhibitor alpha (IκBα), while an NF-κB inhibitor Parthenilide partially abolished the G9a-enhanced FAK activation, which suggests that G9a-enhanced invasion and activation of FAK is mediated by elevated NF-κB activity. Notably, a strong positive correlation between the IHC staining of G9a and phosphorylated FAK proteins was identified in H1299 xenografts and 159 cases of NSCLC tissues (R = 0.408).
Implications:
The findings of this study strongly demonstrate that G9a may promote invasion and metastasis of NSCLC cells by enhancing FAK signaling pathway via elevating NF-κB transcriptional activity, indicating potential significance and therapeutic implications of these pathways in the invasion and metastasis of NSCLCs that overexpress G9a protein.
Data from Inflammation-Induced Metastatic Colonization of the Lung Is Facilitated by Hepatocyte Growth Factor-Secreting Monocyte-Derived Macrophages
Authors: Arif Arif A. , Huang Yu-Hsuan , Freeman Spencer A. , Atif Jawairia , Dean Pamela , Lai Jacqueline C.Y. , Blanchet Marie-Renee , Wiegand Kimberly C. , McNagny Kelly M. , Underhill T. Michael , Gold Michael R. , Johnson Pauline , Roskelley Calvin D.
Published: April 2023

Abstract
A rate-limiting step for circulating tumor cells to colonize distant organ sites is their ability to locate a microenvironmental niche that supports their survival and growth. This can be achieved by features intrinsic to the tumor cells and/or by the conditioning of a “premetastatic” niche. To determine if pulmonary inflammation promotes the latter, we initiated models for inflammatory asthma, hypersensitivity pneumonitis, or bleomycin-induced sterile inflammation before introducing tumor cells with low metastatic potential into the circulation. All types of inflammation increased the end-stage metastatic burden of the lungs 14 days after tumor cell inoculation without overtly affecting tumor extravasation. Instead, the number and size of early micrometastatic lesions found within the interstitial tissues 96 hours after tumor cell inoculation were increased in the inflamed lungs, coincident with increased tumor cell survival and the presence of nearby inflammation-induced monocyte-derived macrophages (MoDM; CD11b+CD11c+). Remarkably, the adoptive transfer of these MoDM was sufficient to increase lung metastasis in the absence of inflammation. These inflammation-induced MoDM secrete a number of growth factors and cytokines, one of which is hepatocyte growth factor (HGF), that augmented tumor cell survival under conditions of stress in vitro. Importantly, blocking HGF signaling with the cMET inhibitor capmatinib abolished inflammation-induced early micrometastatic lesion formation in vivo. These findings indicate that inflammation-induced MoDM and HGF in particular increase the efficiency of early metastatic colonization in the lung by locally preconditioning the microenvironment.
Implications:
Inflammation preconditions the distant site microenvironment to increase the metastatic potential of tumor cells that arrive there.
Data from Cytidine Deaminase APOBEC3A Regulates PD-L1 Expression in Cancer Cells in a JNK/c-JUN-Dependent Manner
Authors: Zhao Kailiang , Zhang Qiang , Flanagan Sheryl A. , Lang Xueting , Jiang Long , Parsels Leslie A. , Parsels Joshua D. , Zou Weiping , Lawrence Theodore S. , Buisson Rémi , Green Michael D. , Morgan Meredith A.
Published: April 2023

Abstract
Programmed death-ligand 1 (PD-L1) promotes tumor immune evasion by engaging the PD-1 receptor and inhibiting T-cell activity. While the regulation of PD-L1 expression is not fully understood, its expression is associated with tumor mutational burden and response to immune checkpoint therapy. Here, we report that Apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3A (APOBEC3A) is an important regulator of PD-L1 expression. Using an APOBEC3A inducible expression system as well as siRNA against endogenous APOBEC3A, we found that APOBEC3A regulates PD-L1 mRNA and protein levels as well as PD-L1 cell surface expression in cancer. Mechanistically, APOBEC3A-induced PD-L1 expression was dependent on APOBEC3A catalytic activity as catalytically dead APOBEC3A mutant (E72A) failed to induce PD-L1 expression. Furthermore, APOBEC3A-induced PD-L1 expression was dependent on replication-associated DNA damage and JNK/c-JUN signaling but not interferon signaling. In addition, we confirmed the relevance of these finding in patient tumors as APOBEC3A expression and mutational signature correlated with PD-L1 expression in multiple patient cancer types. These data provide a novel link between APOBEC3A, its DNA mutagenic activity and PD-L1-mediated antitumoral immunity. This work nominates APOBEC3A as a mechanism of immune evasion and a potential biomarker for the therapeutic efficacy of immune checkpoint blockade.
Implications:
APOBEC3A catalytic activity induces replication-associated DNA damage to promote PD-L1 expression implying that APOBEC3A-driven mutagenesis represents both a mechanism of tumor immune evasion and a therapeutically targetable vulnerability in cancer cells.
Data from SHOC2 Is a Critical Modulator of Sensitivity to EGFR–TKIs in Non–Small Cell Lung Cancer Cells
Authors: Terai Hideki , Hamamoto Junko , Emoto Katsura , Masuda Takeshi , Manabe Tadashi , Kuronuma Satoshi , Kobayashi Keigo , Masuzawa Keita , Ikemura Shinnosuke , Nakayama Sohei , Kawada Ichiro , Suzuki Yusuke , Takeuchi Osamu , Suzuki Yukio , Ohtsuki Sumio , Yasuda Hiroyuki , Soejima Kenzo , Fukunaga Koichi
Published: April 2023

Abstract
EGFR mutation-positive patients with non–small cell lung cancer (NSCLC) respond well to treatment with EGFR–tyrosine kinase inhibitors (EGFR–TKI); however, treatment with EGFR–TKIs is not curative, owing to the presence of residual cancer cells with intrinsic or acquired resistance to this class of drugs. Additional treatment targets that may enhance the efficacy of EGFR–TKIs remain elusive. Using a CRISPR/Cas9-based screen, we identified the leucine-rich repeat scaffold protein SHOC2 as a key modulator of sensitivity to EGFR–TKI treatment. On the basis of in vitro assays, we demonstrated that SHOC2 expression levels strongly correlate with the sensitivity to EGFR–TKIs and that SHOC2 affects the sensitivity to EGFR–TKIs in NSCLC cells via SHOC2/MRAS/PP1c and SHOC2/SCRIB signaling. The potential SHOC2 inhibitor celastrol phenocopied SHOC2 depletion. In addition, we confirmed that SHOC2 expression levels were important for the sensitivity to EGFR–TKIs in vivo. Furthermore, IHC showed the accumulation of cancer cells that express high levels of SHOC2 in lung cancer tissues obtained from patients with NSCLC who experienced acquired resistance to EGFR–TKIs. These data indicate that SHOC2 may be a therapeutic target for patients with NSCLC or a biomarker to predict sensitivity to EGFR–TKI therapy in EGFR mutation-positive patients with NSCLC. Our findings may help improve treatment strategies for patients with NSCLC harboring EGFR mutations.
Implications:
This study showed that SHOC2 works as a modulator of sensitivity to EGFR–TKIs and the expression levels of SHOC2 can be used as a biomarker for sensitivity to EGFR–TKIs.