bluuebunny/crossref_metadata_2025 huggingface dataset:
Data from A Simple Approach to Cancer Therapy Afforded by Multivalent Pseudopeptides That Target Cell-Surface Nucleoproteins
Authors: Destouches Damien , Page Nicolas , Hamma-Kourbali Yamina , Machi Valérie , Chaloin Olivier , Frechault Sophie , Birmpas Charalampos , Katsoris Panagiotis , Beyrath Julien , Albanese Patricia , Maurer Marie , Carpentier Gilles , Strub Jean-Marc , Van Dorsselaer Alain , Muller Sylviane , Bagnard Dominique , Briand Jean Paul , Courty José
Published: March 2023

Abstract
Recent studies have implicated the involvement of cell surface forms of nucleolin in tumor growth. In this study, we investigated whether a synthetic ligand of cell-surface nucleolin known as N6L could exert antitumor activity. We found that N6L inhibits the anchorage-dependent and independent growth of tumor cell lines and that it also hampers angiogenesis. Additionally, we found that N6L is a proapoptotic molecule that increases Annexin V staining and caspase-3/7 activity in vitro and DNA fragmentation in vivo. Through affinity isolation experiments and mass-spectrometry analysis, we also identified nucleophosmin as a new N6L target. Notably, in mouse xenograft models, N6L administration inhibited human tumor growth. Biodistribution studies carried out in tumor-bearing mice indicated that following administration N6L rapidly localizes to tumor tissue, consistent with its observed antitumor effects. Our findings define N6L as a novel anticancer drug candidate warranting further investigation. Cancer Res; 71(9); 3296–305. ©2011 AACR.
Data from p53-Responsive miR-194 Inhibits Thrombospondin-1 and Promotes Angiogenesis in Colon Cancers
Authors: Sundaram Prema , Hultine Stacy , Smith Lauren M. , Dews Michael , Fox Jamie L. , Biyashev Dauren , Schelter Janell M. , Huang Qihong , Cleary Michele A. , Volpert Olga V. , Thomas-Tikhonenko Andrei
Published: March 2023

Abstract
Thrombospondin-1 (TSP-1) is an endogenous inhibitor of angiogenesis encoded by the THBS1 gene, whose promoter is activated by p53. In advanced colorectal cancers (CRC), its expression is sustained or even slightly increased despite frequent loss of p53. Here, we determined that in HCT116 CRC cells, p53 activates the THBS1 primary transcript, but fails to boost THBS1 mRNA or protein levels, implying posttranscriptional regulation by microRNAs (miRNA). In a global miRNA gain-of-function screen done in the Dicer-deficient HCT116 variant, several miRNAs negatively regulated THBS1 mRNA and protein levels, one of them being miR-194. Notably, in agreement with published data, p53 upregulated miR-194 expression in THBS1 retrovirus-transduced HCT116 cells, leading to decreased TSP-1 levels. This negative effect was mediated by a single miR-194 complementary site in the THBS1 3′-untranslated region, and its elimination resulted in TSP-1 reactivation, impaired angiogenesis in Matrigel plugs, and reduced growth of HCT116 xenografts. Conversely, transient overexpression of miR-194 in HCT116/THBS1 cells boosted Matrigel angiogenesis, and its stable overexpression in Ras-induced murine colon carcinomas increased microvascular densities and vessel sizes. Although the overall contribution of miR-194 to neoplastic growth is context dependent, p53-induced activation of this GI tract–specific miRNA during ischemia could promote angiogenesis and facilitate tissue repair. Cancer Res; 71(24); 7490–501. ©2011 AACR.
Data from Cytoplasmic CUL9/PARC Ubiquitin Ligase Is a Tumor Suppressor and Promotes p53-Dependent Apoptosis
Authors: Pei Xin-Hai , Bai Feng , Li Zhijun , Smith Matthew D. , Whitewolf Gabrielle , Jin Ran , Xiong Yue
Published: March 2023

Abstract
A wide range of cell stresses, including DNA damage, signal to p53 through posttranslational modification of p53. The cytoplasmic functions of p53 are emerging as an important constituent of role of p53 in tumor suppression. Here, we report that deletion of the Cul9 (formerly Parc) gene, which encodes an E3 ubiquitin ligase that binds to p53 and localizes in the cytoplasm, resulted in spontaneous tumor development, accelerated Eμ-Myc–induced lymphomagenesis, and rendered mice susceptible to carcinogenesis. Cul9-p53 double-mutant mice exhibited indistinguishable tumor phenotypes as p53 single-mutant mice, indicating that the function of Cul9 in tumor suppression is largely mediated by p53. Deletion of Cul9 had no significant effect on cell-cycle progression, but attenuated DNA damage–induced apoptosis. Ectopic expression of wild-type CUL9, but not a point mutant CUL9 deficient in p53 binding, promotes apoptosis. These results show CUL9 as a potential p53-activating E3 ligase in the cytoplasm. Cancer Res; 71(8); 2969–77. ©2011 AACR.
Data from Combination Therapy with HSP90 Inhibitor 17-DMAG Reconditions the Tumor Microenvironment to Improve Recruitment of Therapeutic T cells
Authors: Rao Aparna , Taylor Jennifer L. , Chi-Sabins Nina , Kawabe Mayumi , Gooding William E. , Storkus Walter J.
Published: March 2023

Abstract
Ineffective recognition of tumor cells by CD8+ T cells is a limitation of cancer immunotherapy. Therefore, treatment regimens that coordinately promote enhanced antitumor CD8+ T-cell activation, delivery, and target cell recognition should yield greater clinical benefit. Using an MCA205 sarcoma model, we show that in vitro treatment of tumor cells with the HSP90 inhibitor 17-DMAG results in the transient (proteasome-dependent) degradation of the HSP90 client protein EphA2 and the subsequent increased recognition of tumor cells by Type-1 anti-EphA2 CD8+ T cells. In vivo administration of 17-DMAG to tumor-bearing mice led to slowed tumor growth, enhanced/prolonged recognition of tumor cells by anti-EphA2 CD8+ T cells, reduced levels of myeloid-derived suppressor cells and regulatory T cells in the tumor microenvironment, and activation of tumor-associated vascular endothelial cells in association with elevated levels of Type-1 tumor-infiltrating lymphocytes. When combined with EphA2-specific active vaccination or the adoptive transfer of EphA2-specific CD8+ T cells, 17-DMAG cotreatment yielded a superior tumor therapeutic regimen that was capable of rendering animals free of disease. Taken together, our findings indicate that 17-DMAG functions as an immune adjuvant in the context of vaccines targeting EphA2. Cancer Res; 72(13); 3196–206. ©2012 AACR.
Data from The ΔNp63 Proteins Are Key Allies of BRCA1 in the Prevention of Basal-Like Breast Cancer
Authors: Buckley Niamh E. , Conlon Susan J. , Jirstrom Karin , Kay Elaine W. , Crawford Nyree T. , O'Grady Anthony , Sheehan Katherine , Mc Dade Simon S. , Wang Ching-Wei , McCance Dennis J. , Johnston Patrick G. , Kennedy Richard D. , Harkin D. Paul , Mullan Paul B.
Published: March 2023

Abstract
Little is known about the origin of basal-like breast cancers, an aggressive disease that is highly similar to BRCA1-mutant breast cancers. p63 family proteins that are structurally related to the p53 suppressor protein are known to function in stem cell regulation and stratified epithelia development in multiple tissues, and p63 expression may be a marker of basal-like breast cancers. Here we report that ΔNp63 isoforms of p63 are transcriptional targets for positive regulation by BRCA1. Our analyses of breast cancer tissue microarrays and BRCA1-modulated breast cancer cell lines do not support earlier reports that p63 is a marker of basal-like or BRCA1 mutant cancers. Nevertheless, we found that BRCA1 interacts with the specific p63 isoform ΔNp63γ along with transcription factor isoforms AP-2α and AP-2γ. BRCA1 required ΔNp63γ and AP-2γ to localize to an intronic enhancer region within the p63 gene to upregulate transcription of the ΔNp63 isoforms. In mammary stem/progenitor cells, siRNA-mediated knockdown of ΔNp63 expression resulted in genomic instability, increased cell proliferation, loss of DNA damage checkpoint control, and impaired growth control. Together, our findings establish that transcriptional upregulation of ΔNp63 proteins is critical for BRCA1 suppressor function and that defects in BRCA1-ΔNp63 signaling are key events in the pathogenesis of basal-like breast cancer. Cancer Res; 71(5); 1933–44. ©2011 AACR.
Data from RNA Helicase DDX5 Is a p53-Independent Target of ARF That Participates in Ribosome Biogenesis
Authors: Saporita Anthony J. , Chang Hsiang-Chun , Winkeler Crystal L. , Apicelli Anthony J. , Kladney Raleigh D. , Wang Jianbo , Townsend R. Reid , Michel Loren S. , Weber Jason D.
Published: March 2023

Abstract
The p19ARF tumor suppressor limits ribosome biogenesis and responds to hyperproliferative signals to activate the p53 checkpoint response. Although its activation of p53 has been well characterized, the role of ARF in restraining nucleolar ribosome production is poorly understood. Here we report the use of a mass spectroscopic analysis to identify protein changes within the nucleoli of Arf-deficient mouse cells. Through this approach, we discovered that ARF limited the nucleolar localization of the RNA helicase DDX5, which promotes the synthesis and maturation of rRNA, ultimately increasing ribosome output and proliferation. ARF inhibited the interaction between DDX5 and nucleophosmin (NPM), preventing association of DDX5 with the rDNA promoter and nuclear pre-ribosomes. In addition, Arf-deficient cells transformed by oncogenic RasV12 were addicted to DDX5, because reduction of DDX5 was sufficient to impair RasV12-driven colony formation in soft agar and tumor growth in mice. Taken together, our findings indicate that DDX5 is a key p53-independent target of the ARF tumor suppressor and is a novel non-oncogene participant in ribosome biogenesis. Cancer Res; 71(21); 6708–17. ©2011 AACR.
Data from Cell-Cycle Regulator Cks1 Promotes Hepatocellular Carcinoma by Supporting NF-κB–Dependent Expression of Interleukin-8
Authors: Lee Eun-Kyoung , Kim Dae-Ghon , Kim Jang-Seong , Yoon Yeup
Published: March 2023

Abstract
The cell-cycle regulator Cks1 has recently been implicated in Skp2-mediated ubiquitination of the tumor suppressor protein p27. In this article, we report that Cks1 exerts a Skp2-independent regulation of NF-κB that promotes interleukin-8 (IL-8) expression, which is critical to hepatocellular carcinoma (HCC) growth. Cks1 was upregulated frequently in human HCC tissues and cell lines. Cks1 knockdown in HCC cells elevated p27 levels and decreased tumorigenicity in a manner that was also associated with a strong downregulation of IL-8 expression. IL-8 downregulation was not phenocopied by either RNAi-mediated knockdown of Skp2 or ectopic overexpression of p27. However, attenuation of IL-8 expression itself was sufficient to blunt HCC growth. Mechanistic investigations revealed that IL-8 was controlled at a transcriptional level by Cks1 targeting of the NF-κB regulator IκBα, which led to NF-κB activation and IL-8 expression, through a p27-independent regulation of IκB kinase complex components. Collectively, our findings support the hypothesis that Cks1 supports hepatocarcinogenesis by NF-κB–mediated regulation of IL-8 expression, broadening the function of Cks1 in cancer beyond its role as a Skp2 cofactor in p27 ubiquitination. Cancer Res; 71(21); 6827–35. ©2011 AACR.
Data from An Iron Regulatory Gene Signature Predicts Outcome in Breast Cancer
Authors: Miller Lance D. , Coffman Lan G. , Chou Jeff W. , Black Michael A. , Bergh Jonas , D'Agostino Ralph , Torti Suzy V. , Torti Frank M.
Published: March 2023

Abstract
Changes in iron regulation characterize the malignant state. However, the pathways that effect these changes and their specific impact on prognosis remain poorly understood. We capitalized on publicly available microarray datasets comprising 674 breast cancer cases to systematically investigate how expression of genes related to iron metabolism is linked to breast cancer prognosis. Of 61 genes involved in iron regulation, 49% were statistically significantly associated with distant metastasis-free survival. Cases were divided into test and training cohorts, and the supervised principal component method was used to stratify cases into risk groups. Optimal risk stratification was achieved with a model comprising 16 genes, which we term the iron regulatory gene signature (IRGS). Multivariable analysis revealed that the IRGS contributes information not captured by conventional prognostic indicators (HR = 1.61; 95% confidence interval: 1.16–2.24; P = 0.004). The IRGS successfully stratified homogeneously treated patients, including ER+ patients treated with tamoxifen monotherapy, both with (P = 0.006) and without (P = 0.03) lymph node metastases. To test whether multiple pathways were embedded within the IRGS, we evaluated the performance of two gene dyads with known roles in iron biology in ER+ patients treated with tamoxifen monotherapy (n = 371). For both dyads, gene combinations that minimized intracellular iron content [anti-import: TFRCLow/HFEHigh; or pro-export: SLC40A1 (ferroportin)High/HAMPLow] were associated with favorable prognosis (P < 0.005). Although the clinical utility of the IRGS will require further evaluation, its ability to both identify high-risk patients within traditionally low-risk groups and low-risk patients within high-risk groups has the potential to affect therapeutic decision making. Cancer Res; 71(21); 6728–37. ©2011 AACR.
Data from The RASSF3 Candidate Tumor Suppressor Induces Apoptosis and G1–S Cell-Cycle Arrest via p53
Authors: Kudo Takumi , Ikeda Mitsunobu , Nishikawa Misa , Yang Zeyu , Ohno Kikuo , Nakagawa Kentaro , Hata Yutaka
Published: March 2023

Abstract
RASSF3 is the smallest member of the RASSF family of proteins that function as tumor suppressors. Unlike other members of this important family, the mechanisms through which RASSF3 suppresses tumor formation remain unknown. Here, we show that RASSF3 expression induces p53-dependent apoptosis and its depletion attenuates DNA damage–induced apoptosis. We found that RASSF3-induced apoptosis depended upon p53 expression. Exogenous expression of RASSF3 induced G1–S arrest, which was also p53 dependent. In contrast, loss of RASSF3 promoted cell-cycle progression, abrogated UVB- and VP-16–induced G1–S arrest, decreased p53 protein and target gene expression, and prevented DNA repair. RASSF3 was shown to directly interact with and facilitate the ubiquitination of MDM2, the E3 ligase that targets p53 for degradation, thereby increasing p53 stabilization. Together, our findings show the tumor suppressor activity of RASSF3, which occurs through p53 stabilization and regulation of apoptosis and the cell cycle. Cancer Res; 72(11); 2901–11. ©2012 AACR.
Data from A Zebrafish Model to Study and Therapeutically Manipulate Hypoxia Signaling in Tumorigenesis
Authors: Santhakumar Kirankumar , Judson Emma C. , Elks Philip M. , McKee Sarah , Elworthy Stone , van Rooijen Ellen , Walmsley Sarah S. , Renshaw Stephen A. , Cross Simon S. , van Eeden Fredericus J.M.
Published: March 2023

Abstract
Hypoxic signaling is a central modulator of cellular physiology in cancer. Core members of oxygen-sensing pathway including the von Hippel-Lindau tumor suppressor protein (pVHL) and the hypoxia inducible factor (HIF) transcription factors have been intensively studied, but improved organismal models might speed advances for both pathobiologic understanding and therapeutic modulation. To study HIF signaling during tumorigenesis and development in zebrafish, we developed a unique in vivo reporter for hypoxia, expressing EGFP driven by prolyl hydroxylase 3 (phd3) promoter/regulatory elements. Modulation of HIF pathway in Tg(phd3::EGFP) embryos showed a specific role for hypoxic signaling in the transgene activation. Zebrafish vhl mutants display a systemic hypoxia response, reflected by strong and ubiquitous transgene expression. In contrast to human VHL patients, heterozygous Vhl mice and vhl zebrafish are not predisposed to cancer. However, upon exposure to dimethylbenzanthracene (DMBA), the vhl heterozygous fish showed an increase in the occurrence of hepatic and intestinal tumors, a subset of which exhibited strong transgene expression, suggesting loss of Vhl function in these tumor cells. Compared with control fish, DMBA-treated vhl heterozygous fish also showed an increase in proliferating cell nuclear antigen-positive renal tubules. Taken together, our findings establish Vhl as a genuine tumor suppressor in zebrafish and offer this model as a tool to noninvasively study VHL and HIF signaling during tumorigenesis and development. Cancer Res; 72(16); 4017–27. ©2012 AACR.
Data from INT6/EIF3E Interacts with ATM and Is Required for Proper Execution of the DNA Damage Response in Human Cells
Authors: Morris Christelle , Tomimatsu Nozomi , Richard Derek J. , Cluet David , Burma Sandeep , Khanna Kum Kum , Jalinot Pierre
Published: March 2023

Abstract
Altered expression of the INT6 gene, encoding the e subunit of the translational initiation factor eIF3, occurs in human breast cancers, but how INT6 relates to carcinogenesis remains unestablished. Here, we show that INT6 is involved in the DNA damage response. INT6 was required for cell survival following γ-irradiation and G2–M checkpoint control. RNA interference–mediated silencing of INT6 reduced phosphorylation of the checkpoint kinases CHK1 and CHK2 after DNA damage. In addition, INT6 silencing prevented sustained accumulation of ataxia telangiectasia mutated (ATM) at DNA damage sites in cells treated with γ-radiation or the radiomimetic drug neocarzinostatin. Mechanistically, this result could be explained by interaction of INT6 with ATM, which together with INT6 was recruited to the sites of DNA damage. Finally, INT6 silencing also reduced ubiquitylation events that promote retention of repair proteins at DNA lesions. Accordingly, accumulation of the repair factor BRCA1 was defective in the absence of INT6. Our findings reveal unexpected and striking connections of INT6 with ATM and BRCA1 and suggest that the protective action of INT6 in the onset of breast cancers relies on its involvement in the DNA damage response. Cancer Res; 72(8); 2006–16. ©2012 AACR.
Data from Nucleotide Excision Repair Factor XPC Enhances DNA Damage–Induced Apoptosis by Downregulating the Antiapoptotic Short Isoform of Caspase-2
Authors: Wang Qi-En , Han Chunhua , Zhang Bo , Sabapathy Kanaga , Wani Altaf A.
Published: March 2023

Abstract
XPC protein is a critical DNA damage recognition factor in nucleotide excision repair for which genetic deficiency confers a predisposition to cancer. In this study, we show that XPC has a function that is independent of its canonical function in DNA repair, potentially altering the interpretation of how XPC deficiency leads to heightened cancer susceptibility. XPC enhances apoptosis induced by DNA damage in a p53 nullizygous background, acting downstream of mitochondrial permeabilization and upstream of caspase-9 activation in the DNA damage–induced apoptosis cascade. We found that deficiency in XPC upregulated production of the short isoform of caspase-2 (casp-2S). This upregulation occurred at both protein and mRNA levels through repression of the caspase-2 promoter by XPC protein. Targeted RNAi-mediated downregulation of casp-2S–enhanced UV-induced apoptosis as well as activation of caspase-9 and caspase-6 in XPC-deficient cells, but not in XPC-proficient cells. In addition, XPC overexpression in various p53-deficient cancer cells resistant to cisplatin improved their sensitivity to cisplatin-induced apoptosis. Given that casp-2S functions as an antiapoptotic protein, our findings suggest that XPC enhances DNA damage–induced apoptosis through inhibition of casp-2S transcription. Together, these findings offer a mechanistic foundation to overcome the resistance of highly prevalent p53-deficient tumors to cell death induced by DNA-damaging therapeutic agents, by targeting strategies that inhibit the expression or function of casp-2S. Cancer Res; 72(3); 666–75. ©2011 AACR.
Data from The Unfolded Protein Response Induces the Angiogenic Switch in Human Tumor Cells through the PERK/ATF4 Pathway
Authors: Wang Yugang , Alam Goleeta N. , Ning Yu , Visioli Fernanda , Dong Zhihong , Nör Jacques E. , Polverini Peter J.
Published: March 2023

Abstract
Neovascularization is a limiting factor in tumor growth and progression. It is well known that changes in the tumor microenvironment, such as hypoxia and glucose deprivation (GD), can induce VEGF production. However, the mechanism linking GD to tumor growth and angiogenesis is unclear. We hypothesize that GD induces the angiogenic switch in tumors through activation of the unfolded protein response (UPR). We report that UPR activation in human tumors results in elevated expression of proangiogenic mediators and a concomitant decrease in angiogenesis inhibitors. cDNA microarray results showed that GD-induced UPR activation promoted upregulation of a number of proangiogenic mediators (VEGF, FGF-2, IL-6, etc.) and downregulation of several angiogenic inhibitors (THBS1, CXCL14, and CXCL10). In vitro studies revealed that partially blocking UPR signaling by silencing protein kinase RNA–like ER kinase (PERK) or activating transcription factor 4 (ATF4) significantly reduced the production of angiogenesis mediators induced by GD. However, suppressing the alpha subunit of hypoxia-inducible factors had no effect on this process. Chromatin immunoprecipitation (ChIP) confirmed binding of ATF4 to a regulatory site in the VEGF gene. In vivo results confirmed that knockdown of PERK in tumor cells slows down tumor growth and decreases tumor blood vessel density. Collectively, these results show that the PERK/ATF4 arm of UPR mediates the angiogenic switch and is a potential target for antiangiogenic cancer therapy. Cancer Res; 72(20); 5396–406. ©2012 AACR.
Data from Iodide Transporter NIS Regulates Cancer Cell Motility and Invasiveness by Interacting with the Rho Guanine Nucleotide Exchange Factor LARG
Authors: Lacoste Claire , Hervé Julie , Bou Nader Myriam , Dos Santos Alexandre , Moniaux Nicolas , Valogne Yannick , Montjean Rodrick , Dorseuil Olivier , Samuel Didier , Cassio Doris , Portulano Carla , Carrasco Nancy , Bréchot Christian , Faivre Jamila
Published: March 2023

Abstract
A number of solute carrier (SLC) proteins are subject to changes in expression and activity during carcinogenesis. Whether these changes play a role in carcinogenesis is unclear, except for some nutrients and ion carriers whose deregulation ensures the necessary reprogramming of energy metabolism in cancer cells. In this study, we investigated the functional role in tumor progression of the sodium/iodide symporter (NIS; aka SLC5A5), which is upregulated and mislocalized in many human carcinomas. Notably, we found that NIS enhanced cell migration and invasion without ion transport being involved. These functions were mediated by NIS binding to leukemia-associated RhoA guanine exchange factor, a Rho guanine exchange factor that activates the small GTPase RhoA. Sequestering NIS in intracellular organelles or impairing its targeting to the cell surface (as observed in many cancers) led to a further increase in cell motility and invasiveness. In sum, our results established NIS as a carrier protein that interacts with a major cell signaling hub to facilitate tumor cell locomotion and invasion. Cancer Res; 72(21); 5505–15. ©2012 AACR.
Data from Systemic Delivery of Salmonella typhimurium Transformed with IDO shRNA Enhances Intratumoral Vector Colonization and Suppresses Tumor Growth
Authors: Blache Céline A. , Manuel Edwin R. , Kaltcheva Teodora I. , Wong Andrea N. , Ellenhorn Joshua D.I. , Blazar Bruce R. , Diamond Don J.
Published: March 2023

Abstract
Generating antitumor responses through the inhibition of tumor-derived immune suppression represents a promising strategy in the development of cancer immunotherapeutics. Here, we present a strategy incorporating delivery of the bacterium Salmonella typhimurium (ST), naturally tropic for the hypoxic tumor environment, transformed with a small hairpin RNA (shRNA) plasmid against the immunosuppressive molecule indoleamine 2,3-dioxygenase 1 (shIDO). When systemically delivered into mice, shIDO silences host IDO expression and leads to massive intratumoral cell death that is associated with significant tumor infiltration by polymorphonuclear neutrophils (PMN). shIDO-ST treatment causes tumor cell death independently of host IDO and adaptive immunity, which may have important implications for use in immunosuppressed patients with cancer. Furthermore, shIDO-ST treatment increases reactive oxygen species (ROS) produced by infiltrating PMNs and, conversely, PMN immunodepletion abrogates tumor control. Silencing of host IDO significantly enhances S. typhimurium colonization, suggesting that IDO expression within the tumor controls the immune response to S. typhimurium. In summary, we present a novel approach to cancer treatment that involves the specific silencing of tumor-derived IDO that allows for the recruitment of ROS-producing PMNs, which may act primarily to clear S. typhimurium infection, but in the process also induces apoptosis of surrounding tumor tissue resulting in a vigorous antitumor effect. Cancer Res; 72(24); 6447–56. ©2012 AACR.
Data from Interleukin-10 Ablation Promotes Tumor Development, Growth, and Metastasis
Authors: Tanikawa Takashi , Wilke Cailin Moira , Kryczek Ilona , Chen Grace Y. , Kao John , Núñez Gabriel , Zou Weiping
Published: March 2023

Abstract
Interleukin-10 (IL-10) is a broadly acting immune inhibitory cytokine that is generally thought to support tumor growth. Here we challenge this view with evidence that genetic ablation of IL-10 in the mouse significantly heightens sensitivity to chemical carcinogenesis, growth of transplanted tumors, and formation of metastases. Tumor growth in IL-10–deficient (IL-10−/−) mice was associated with an increased level of myeloid-derived suppressor cells (MDSC) and CD4+Foxp3+ regulatory T (Treg) cells in both the tumor microenvironment and the tumor-draining lymph nodes. IL-10−/− MDSCs express high levels of MHC and IL-1, and they efficiently induced formation of Treg cells. IL-1 signaling blockade reduced tumor growth mediated by IL-10 deficiency, associated with a partial rescue of tumor infiltration and function of effector T cells and a decrease in tumor angiogenesis and tumor infiltration by Treg cells. Taken together, our findings establish that endogenous IL-10 inhibits inflammatory cytokine production and hampers the development of Treg cells and MDSCs, two key components of the immunosuppressive tumor microenvironment, thereby inhibiting tumor development, growth, and metastasis. Cancer Res; 72(2); 420–9. ©2011 AACR.
Data from NF-κB Hyperactivation in Tumor Tissues Allows Tumor-Selective Reprogramming of the Chemokine Microenvironment to Enhance the Recruitment of Cytolytic T Effector Cells
Authors: Muthuswamy Ravikumar , Berk Erik , Junecko Beth Fallert , Zeh Herbert J. , Zureikat Amer H. , Normolle Daniel , Luong The Minh , Reinhart Todd A. , Bartlett David L. , Kalinski Pawel
Published: March 2023

Abstract
Tumor infiltration with effector CD8+ T cells (Teff) predicts longer recurrence-free survival in many types of human cancer, illustrating the broad significance of Teff for effective immunosurveillance. Colorectal tumors with reduced accumulation of Teff express low levels of Teff-attracting chemokines such as CXCL10/IP10 and CCL5/RANTES. In this study, we investigated the feasibility of enhancing tumor production of Teff-attracting chemokines as a cancer therapeutic strategy using a tissue explant culture system to analyze chemokine induction in intact tumor tissues. In different tumor explants, we observed highly heterogeneous responses to IFNα or poly-I:C (a TLR3 ligand) when they were applied individually. In contrast, a combination of IFNα and poly-I:C uniformly enhanced the production of CXCL10 and CCL5 in all tumor lesions. Moreover, these effects could be optimized by the further addition of COX inhibitors. Applying this triple combination also uniformly suppressed the production of CCL22/MDC, a chemokine associated with infiltration of T regulatory cells (Treg). The Teff-enhancing effects of this treatment occurred selectively in tumor tissues, as compared with tissues derived from tumor margins. These effects relied on the increased propensity of tumor-associated cells (mostly fibroblasts and infiltrating inflammatory cells) to hyperactivate NF-κB and produce Teff-attracting chemokines in response to treatment, resulting in an enhanced ability of the treated tumors to attract Teff cells and reduced ability to attract Treg cells. Together, our findings suggest the feasibility of exploiting NF-κB hyperactivation in the tumor microenvironment to selectively enhance Teff entry into colon tumors. Cancer Res; 72(15); 3735–43. ©2012 AACR.
Data from Genetic Determinants for Promoter Hypermethylation in the Lungs of Smokers: A Candidate Gene-Based Study
Authors: Leng Shuguang , Stidley Christine A. , Liu Yushi , Edlund Christopher K. , Willink Randall P. , Han Younghun , Landi Maria Teresa , Thun Michael , Picchi Maria A. , Bruse Shannon E. , Crowell Richard E. , Van Den Berg David , Caporaso Neil E. , Amos Christopher I. , Siegfried Jill M. , Tesfaigzi Yohannes , Gilliland Frank D. , Belinsky Steven A.
Published: March 2023

Abstract
The detection of tumor suppressor gene promoter methylation in sputum-derived exfoliated cells predicts early lung cancer. Here, we identified genetic determinants for this epigenetic process and examined their biologic effects on gene regulation. A two-stage approach involving discovery and replication was used to assess the association between promoter hypermethylation of a 12-gene panel and common variation in 40 genes involved in carcinogen metabolism, regulation of methylation, and DNA damage response in members of the Lovelace Smokers Cohort (N = 1,434). Molecular validation of three identified variants was conducted using primary bronchial epithelial cells. Association of study-wide significance (P < 8.2 × 10−5) was identified for rs1641511, rs3730859, and rs1883264 in TP53, LIG1, and BIK, respectively. These single-nucleotide polymorphisms (SNP) were significantly associated with altered expression of the corresponding genes in primary bronchial epithelial cells. In addition, rs3730859 in LIG1 was also moderately associated with increased risk for lung cancer among Caucasian smokers. Together, our findings suggest that genetic variation in DNA replication and apoptosis pathways impacts the propensity for gene promoter hypermethylation in the aerodigestive tract of smokers. The incorporation of genetic biomarkers for gene promoter hypermethylation with clinical and somatic markers may improve risk assessment models for lung cancer. Cancer Res; 72(3); 707–15. ©2011 AACR.
Data from Hepatocyte–Stellate Cell Cross-Talk in the Liver Engenders a Permissive Inflammatory Microenvironment That Drives Progression in Hepatocellular Carcinoma
Authors: Coulouarn Cédric , Corlu Anne , Glaise Denise , Guénon Isabelle , Thorgeirsson Snorri S. , Clément Bruno
Published: March 2023

Abstract
Many solid malignant tumors arise on a background of inflamed and/or fibrotic tissues, features that are found in more than 80% hepatocellular carcinomas (HCC). Activated hepatic stellate cells (HSC) play a critical role in fibrogenesis associated with HCC onset and progression, yet their functional impact on hepatocyte fate remains largely unexplored. Here, we used a coculture model to investigate the cross-talk between hepatocytes (human hepatoma cells) and activated human HSCs. Unsupervised genome-wide expression profiling showed that hepatocyte–HSC cross-talk is bidirectional and results in the deregulation of functionally relevant gene networks. Notably, coculturing increased the expression of proinflammatory cytokines and modified the phenotype of hepatocytes toward motile cells. Hepatocyte–HSC cross-talk also generated a permissive proangiogenic microenvironment, particularly by inducing VEGFA and matrix metalloproteinase (MMP)9 expression in HSCs. An integrative genomic analysis revealed that the expression of genes associated with hepatocyte–HSC cross-talk correlated with HCC progression in mice and was predictive of a poor prognosis and metastasis propensity in human HCCs. Interestingly, the effects of cross-talk on migration and angiogenesis were reversed by the histone deacetylase inhibitor trichostatin A. Our findings, therefore, indicate that the cross-talk between hepatoma cells and activated HSCs is an important feature of HCC progression, which may be targeted by epigenetic modulation. Cancer Res; 72(10); 2533–42. ©2012 AACR.
Data from Mre11-Dependent Degradation of Stalled DNA Replication Forks Is Prevented by BRCA2 and PARP1
Authors: Ying Songmin , Hamdy Freddie C. , Helleday Thomas
Published: March 2023

Abstract
PARP inhibitors are currently being used in clinical trials to treat BRCA1- or BRCA2-defective tumors, based on the synthetic lethal interaction between PARP1 and BRCA1/2-mediated homologous recombination (HR). However, the molecular mechanisms that drive this synthetic lethality remain unclear. Here, we show increased levels of Mre11, a key component of MRN (Mre11-Rad50-Nbs1) complex that plays a role in the restart of stalled replication forks and enhanced resection at stalled replication forks in BRCA2-deficient cells. BRCA2-deficient cells also showed hypersensitivity to the Mre11 inhibitor mirin. Interestingly, PARP1 activity was required to protect stalled forks from Mre11-dependent degradation. Resistance to PARP inhibition in BRCA2-mutant cells led to reduced levels of Mre11 foci and also rescued their sensitivity to mirin. Taken together, our findings not only show that Mre11 activity is required for the survival of BRCA2 mutant cells but also elucidate roles for both the BRCA2 and PARP1 proteins in protecting stalled replication forks, which offers insight into the molecular mechanisms of the synthetic lethality between BRCA2 and PARP1. Cancer Res; 72(11); 2814–21. ©2012 AACR.
Data from Chronic Autophagy Is a Cellular Adaptation to Tumor Acidic pH Microenvironments
Authors: Wojtkowiak Jonathan W. , Rothberg Jennifer M. , Kumar Virendra , Schramm Karla J. , Haller Edward , Proemsey Joshua B. , Lloyd Mark C. , Sloane Bonnie F. , Gillies Robert J.
Published: March 2023

Abstract
Tumor cell survival relies upon adaptation to the acidic conditions of the tumor microenvironment. To investigate potential acidosis survival mechanisms, we examined the effect of low pH (6.7) on human breast carcinoma cells. Acute low pH exposure reduced proliferation rate, induced a G1 cell cycle arrest, and increased cytoplasmic vacuolization. Gene expression analysis revealed elevated levels of ATG5 and BNIP3 in acid-conditioned cells, suggesting cells exposed to low pH may utilize autophagy as a survival mechanism. In support of this hypothesis, we found that acute low pH stimulated autophagy as defined by an increase in LC3-positive punctate vesicles, double-membrane vacuoles, and decreased phosphorylation of AKT and ribosomal protein S6. Notably, cells exposed to low pH for approximately 3 months restored their proliferative capacity while maintaining the cytoplasmic vacuolated phenotype. Although autophagy is typically transient, elevated autophagy markers were maintained chronically in low pH conditioned cells as visualized by increased protein expression of LC3-II and double-membrane vacuoles. Furthermore, these cells exhibited elevated sensitivity to PI3K-class III inhibition by 3-methyladenine. In mouse tumors, LC3 expression was reduced by systemic treatment with sodium bicarbonate, which raises intratumoral pH. Taken together, these results argue that acidic conditions in the tumor microenvironment promote autophagy, and that chronic autophagy occurs as a survival adaptation in this setting. Cancer Res; 72(16); 3938–47. ©2012 AACR.
Data from ZNF217 Is a Marker of Poor Prognosis in Breast Cancer That Drives Epithelial–Mesenchymal Transition and Invasion
Authors: Vendrell Julie A. , Thollet Aurélie , Nguyen Nhan T. , Ghayad Sandra E. , Vinot Stéphanie , Bièche Ivan , Grisard Evelyne , Josserand Véronique , Coll Jean-Luc , Roux Pierre , Corbo Laura , Treilleux Isabelle , Rimokh Ruth , Cohen Pascale A.
Published: March 2023

Abstract
The Krüppel-like zinc finger protein ZNF217 is a candidate oncogene in breast cancer. In this study, we showed that high levels of expression of ZNF217 mRNA are associated with poor prognosis and the development of metastases in breast cancer. Overexpression of ZNF217 in breast cancer cells stimulated migration and invasion in vitro and promoted the development of spontaneous lung or node metastases in mice in vivo. ZNF217 also promoted epithelial–mesenchymal transition (EMT) in human mammary epithelial cells, and the TGF-β–activated Smad signaling pathway was identified as a major driver of ZNF217-induced EMT. In addition, a TGF-β autocrine loop sustained activation of the TGF-β pathway in ZNF217-overexpressing mammary epithelial cells, most likely because of ZNF217-mediated direct upregulation of TGFB2 or TGFB3. Inhibition of the TGF-β pathway led to the reversal of ZNF217-mediated EMT. Together, our findings indicate that ZNF217 mRNA expression may represent a novel prognostic biomarker in breast cancer. Therapeutic targeting of ZNF217 of the TGF-β signaling pathway may benefit the subset of patients whose tumors express high levels of ZNF217. Cancer Res; 72(14); 3593–606. ©2012 AACR.
Data from CD24 Is an Effector of HIF-1–Driven Primary Tumor Growth and Metastasis
Authors: Thomas Shibu , Harding Michael A. , Smith Steven C. , Overdevest Jonathan B. , Nitz Matthew D. , Frierson Henry F. , Tomlins Scott A. , Kristiansen Glen , Theodorescu Dan
Published: March 2023

Abstract
Hypoxia drives malignant progression in part by promoting accumulation of the oncogenic transcription factor hypoxia inducible factor–1α (HIF-1α) in tumor cells. Tumor aggressiveness also relates to elevation of the cancer stem cell–associated membrane protein CD24, which has been causally implicated in tumor formation and metastasis in experimental models. Here, we link these two elements by showing that hypoxia induces CD24 expression through a functional hypoxia responsive element in the CD24 promoter. HIF-1α overexpression induced CD24 mRNA and protein under normoxic conditions, with this effect traced to a recruitment of endogenous HIF-1α to the CD24 promoter. Short hairpin RNA–mediated attenuation of HIF-1α or CD24 expression reduced cancer cell survival in vitro and in vivo at the levels of primary and metastatic tumor growth. CD24 overexpression in HIF-1α–depleted cancer cells rescued this decrease, whereas HIF-1α overexpression in CD24-depleted cells did not. Analysis of clinical tumor specimens revealed a correlation between HIF-1α and CD24 levels and an association of their coexpression to decreased patient survival. Our results establish a mechanistic linkage between 2 critically important molecules in cancer, identifying CD24 as a critical HIF-1α transcriptional target and biologic effector, strengthening the rationale to target CD24 for cancer therapy. Cancer Res; 72(21); 5600–12. ©2012 AACR.
Data from Prohibitin Attenuates Colitis-Associated Tumorigenesis in Mice by Modulating p53 and STAT3 Apoptotic Responses
Authors: Kathiria Arwa S. , Neumann William L. , Rhees Jennifer , Hotchkiss Erin , Cheng Yulan , Genta Robert M. , Meltzer Stephen J. , Souza Rhonda F. , Theiss Arianne L.
Published: March 2023

Abstract
Although inflammatory bowel disease is associated with higher risk of colorectal cancer, the precise pathogenic mechanisms underlying this association are not completely understood. Prohibitin 1 (PHB), a protein implicated in the regulation of proliferation, apoptosis, and transcription, is decreased in intestinal inflammation. In this study, we have established a key function for PHB in mediating colitis-associated cancer. Wild-type and transgenic (Tg) mice specifically overexpressing PHB in intestinal epithelial cells were subjected to a classical two-stage protocol of colitis-associated carcinogenesis. In addition, wild-type and p53 null human cell models were used to assess PHB interaction with STAT3 and p53. Wild-type mice exhibited decreased mucosal PHB protein expression during colitis-associated carcinogenesis. Tg mice exhibited decreased susceptibility in a manner associated with increased apoptosis, p53, Bax, and Bad expression plus decreased Bcl-xL and Bcl-2 expression. PHB overexpression in wild-type but not p53 null human cells increased expression of Bax, Bad, and caspase-3 cleavage. In wild-type p53 cells, PHB overexpression decreased basal and interleukin-6-induced STAT3 activation and expression of the STAT3 responsive genes Bcl-xL and Bcl-2. PHB coimmunoprecipitated with phospho-STAT3 in addition to p53 in cultured cell lysates and colon mucosa. This is the first study to show interaction between PHB and STAT3 in vivo. In summary, our findings suggest that PHB protects against colitis-associated cancer by modulating p53- and STAT3-mediated apoptosis. Modulation of PHB expression in intestinal epithelial cells may offer a potential therapeutic approach to prevent colitis-associated carcinogenesis. Cancer Res; 72(22); 5778–89. ©2012 AACR.
Data from In Vivo Imaging of Drug-Induced Mitochondrial Outer Membrane Permeabilization at Single-Cell Resolution
Authors: Earley Sarah , Vinegoni Claudio , Dunham Joshua , Gorbatov Rostic , Feruglio Paolo Fumene , Weissleder Ralph
Published: March 2023

Abstract
Observing drug responses in the tumor microenvironment in vivo can be technically challenging. As a result, cellular responses to molecularly targeted cancer drugs are often studied in cell culture, which does not accurately represent the behavior of cancer cells growing in vivo. Using high-resolution microscopy and fluorescently labeled genetic reporters for apoptosis, we developed an approach to visualize drug-induced cell death at single-cell resolution in vivo. Stable expression of the mitochondrial intermembrane protein IMS-RP was established in human breast and pancreatic cancer cells. Image analysis was then used to quantify release of IMS-RP into the cytoplasm upon apoptosis and irreversible mitochondrial permeabilization. Both breast and pancreatic cancer cells showed higher basal apoptotic rates in vivo than in culture. To study drug-induced apoptosis, we exposed tumor cells to navitoclax (ABT-263), an inhibitor of Bcl-2, Bcl-xL, and Bcl-w, both in vitro and in vivo. Although the tumors responded to Bcl-2 inhibition in vivo, inducing apoptosis in around 20% of cancer cells, the observed response was much higher in cell culture. Together, our findings show an imaging technique that can be used to directly visualize cell death within the tumor microenvironment in response to drug treatment. Cancer Res; 72(12); 2949–56. ©2012 AACR.
Data from MLK3 Regulates Paxillin Phosphorylation in Chemokine-Mediated Breast Cancer Cell Migration and Invasion to Drive Metastasis
Authors: Chen Jian , Gallo Kathleen A.
Published: March 2023

Abstract
MLK3 kinase activates multiple mitogen-activated protein kinases and plays a critical role in cancer cell migration and invasion. In the tumor microenvironment, prometastatic factors drive breast cancer invasion and metastasis, but their associated signaling pathways are not well-known. Here, we provide evidence that MLK3 is required for chemokine (CXCL12)-induced invasion of basal breast cancer cells. We found that MLK3 induced robust phosphorylation of the focal adhesion scaffold paxillin on Ser 178 and Tyr 118, which was blocked by silencing or inhibition of MLK3-JNK. Silencing or inhibition of MLK3, inhibition of JNK, or expression of paxillin S178A all led to enhanced Rho activity, indicating that the MLK3–JNK–paxillin axis limits Rho activity to promote focal adhesion turnover and migration. Consistent with this, MLK3 silencing increased focal adhesions and stress fibers in breast cancer cells. MLK3 silencing also decreased the formation of breast cancer lung metastases in vivo, and breast cancer cells derived from mouse lung metastases showed enhanced Ser 178 paxillin phosphorylation. Taken together, our findings suggest that the MLK3–JNK–paxillin signaling axis may represent a potential therapeutic target and/or prognostic marker in breast cancer metastasis. Cancer Res; 72(16); 4130–40. ©2012 AACR.
Data from Global Quantitative Phosphoproteome Analysis of Human Tumor Xenografts Treated with a CD44 Antagonist
Authors: Weigand Stefan , Herting Frank , Maisel Daniela , Nopora Adam , Voss Edgar , Schaab Christoph , Klammer Martin , Tebbe Andreas
Published: March 2023

Abstract
The cell surface glycoprotein CD44 plays an important role in the development and progression of various tumor types. RG7356 is a humanized antibody targeting the constant region of CD44 that shows antitumor efficacy in mice implanted with CD44-expressing tumors such as MDA-MB-231 breast cancer cells. CD44 receptor seems to function as the main receptor for hyaluronic acid and osteopontin, serving as coreceptor for growth factor pathways like cMet, EGFR, HER-2, and VEGFR and by cytoskeletal modulation via ERM and Rho kinase signaling. To assess the direct impact of RG7356 binding to the CD44 receptor, a global mass spectrometry–based phosphoproteomics approach was applied to freshly isolated MDA-MB-231 tumor xenografts. Results from a global phosphoproteomics screen were further corroborated by Western blot and ELISA analyses of tumor lysates from CD44-expressing tumors. Short-term treatment of tumor-bearing mice with RG7356 resulted in modifications of the MAPK pathway in the responsive model, although no effects on downstream phosphorylation were observed in a nonresponsive xenograft model. Taken together, our approach augments the value of other high throughput techniques to identify biomarkers for clinical development of targeted agents. Cancer Res; 72(17); 4329–39. ©2012 AACR.
Data from Alternate Splicing of the p53 Inhibitor HDMX Offers a Superior Prognostic Biomarker than p53 Mutation in Human Cancer
Authors: Lenos Kristiaan , Grawenda Anna M. , Lodder Kirsten , Kuijjer Marieke L. , Teunisse Amina F.A.S. , Repapi Emmanouela , Grochola Lukasz F. , Bartel Frank , Hogendoorn Pancras C.W. , Wuerl Peter , Taubert Helge , Cleton-Jansen Anne-Marie , Bond Gareth L. , Jochemsen Aart G.
Published: March 2023

Abstract
Conventional high-grade osteosarcoma is the most common primary bone malignancy. Although altered expression of the p53 inhibitor HDMX (Mdmx/Mdm4) is associated with cancer risk, progression, and outcome in other tumor types, little is known about its role in osteosarcoma. High expression of the Hdmx splice variant HDMX-S relative to the full-length transcript (the HDMX-S/HDMX-FL ratio) correlates with reduced HDMX protein expression, faster progression, and poorer survival in several cancers. Here, we show that the HDMX-S/HDMX-FL ratio positively correlates with less HDMX protein expression, faster metastatic progression, and a trend to worse overall survival in osteosarcomas. We found that the HDMX-S/HDMX-FL ratio associated with common somatic genetic lesions connected with p53 inhibition, such as p53 mutation and HDM2 overexpression in osteosarcoma cell lines. Interestingly, this finding was not limited to osteosarcomas as we observed similar associations in breast cancer and a variety of other cancer cell lines, as well as in tumors from patients with soft tissue sarcoma. The HDMX-S/HDMX-FL ratio better defined patients with sarcoma with worse survival rates than p53 mutational status. We propose a novel role for alternative splicing of HDMX, whereby it serves as a mechanism by which HDMX protein levels are reduced in cancer cells that have already inhibited p53 activity. Alternative splicing of HDMX could, therefore, serve as a more effective biomarker for p53 pathway attenuation in cancers than p53 gene mutation. Cancer Res; 72(16); 4074–84. ©2012 AACR.