bluuebunny/crossref_metadata_2025 huggingface dataset:
Glioma Formation in Neurofibromatosis 1 Reflects Preferential Activation of K-RAS in Astrocytes
Authors: Dasgupta Biplab , Li Wen , Perry Arie , Gutmann David H.
Published: June 2022

Children with the tumor predisposition syndrome, neurofibromatosis 1 (NF1), develop optic pathway gliomas. The NF1 gene product, neurofibromin, functions as a negative regulator of RAS, such that NF1 inactivation results in RAS hyperactivation. Recent studies have highlighted the divergent biological and biochemical properties of the various RAS isoforms, which prompted us to examine the consequence of Nf1 inactivation in astrocytes on RAS isoform activation in vitro and in vivo. In this report, we show that only K-RAS is activated in Nf1−/− astrocytes and that activation of K-RAS, but not H-RAS, accounts for the proliferative advantage and abnormal actin cytoskeleton–mediated processes observed in Nf1−/− astrocytes in vitro. Moreover, dominant inhibitory K-RAS corrects these abnormalities in Nf1−/− astrocytes invitro. Lastly, we show that Nf1+/− mice with astrocyte-specific activated K-RAS expression in vivo develop optic pathway gliomas, similar to our previously reported Nf1+/− mice with astrocyte Nf1 inactivation. Collectively, our results show that K-RAS is the primary target for neurofibromin GTPase-activating protein activity in vitro and in vivo and that K-RAS activation in astrocytes recapitulates the biochemical, biological, and tumorigenic properties of neurofibromin loss.
Novel Prognostic Value of Nuclear Epidermal Growth Factor Receptor in Breast Cancer
Authors: Lo Hui-Wen , Xia Weiya , Wei Yongkun , Ali-Seyed Mohamed , Huang Shiu-Feng , Hung Mien-Chie
Published: June 2022

Epidermal growth factor receptor (EGFR) has been detected in the nucleus of cancer cells and primary tumors for decades. While localized in the nucleus, EGFR functions as a transcriptional regulator resulting in the activation of the cyclin D1 gene. Despite nuclear accumulation of EGFR is linked to increased DNA synthesis and proliferative potential, the pathological significance of nuclear EGFR, however, remains uninvestigated. Furthermore, expression of EGFR has not provided a consistent predictive value for survival of breast cancer patients. Here, we analyzed 130 breast carcinomas via immunohistochemical analyses for the levels of nuclear and non-nuclear EGFR. We found 37.7% of the cohort immunostained positively for nuclear EGFR and 6.9% with high levels of expression. Importantly, Kaplan-Meier survival analysis and log-rank test revealed a significant inverse correlation between high nuclear EGFR and overall survival (P = 0.009). Expression of nuclear EGFR correlated positively with increased levels of cyclin D1 and Ki-67, both are indicators for cell proliferation. In contrast, expression of non-nuclear EGFR did not significantly correlate with those of cyclin D1 and Ki-67 or the overall survival rate. In addition, we analyzed 37 oral squamous carcinomas for EGFR expression and found 24.3% of the cases to contain moderate/high levels of nuclear EGFR. Taken together, our findings indicate pathological significance of nuclear EGFR and may have important clinical implication.
Expression in Hematological Malignancies of a Glucocorticoid Receptor Splice Variant That Augments Glucocorticoid Receptor-mediated Effects in Transfected Cells
Authors: de Lange Pieter , Segeren Christine M. , Koper Jan W. , Wiemer Erik , Sonneveld Pieter , Brinkmann Albert O. , White Anne , Brogan Iain J. , de Jong Frank H. , Lamberts Steven W. J.
Published: June 2022

Glucocorticoids play an important role in the treatment of a number of hematological malignancies, such as multiple myeloma. The effects of glucocorticoids are mediated through the glucocorticoid receptor α, the abundance of which can be modulated by alternative splicing of the glucocorticoid receptor mRNA. Two splice variants of the glucocorticoid receptor mRNA have been described: glucocorticoid receptor β, which reportedly has a dominant negative effect on the actions of the glucocorticoid receptor α, and glucocorticoid receptor P, of which the effects are unknown. In this study, we have investigated the expression levels of these two splice variants at the mRNA level in multiple myeloma cells and in a number of other hematological tumors. Although the glucocorticoid receptor β mRNA was, if at all, expressed at very low levels, considerable amounts (up to 50% of the total glucocorticoid receptor mRNA) glucocorticoid receptor P mRNA was present in most hematological malignancies. In transient transfection studies in several cell types and in multiple myeloma cell lines, the glucocorticoid receptor P increased the activity of the glucocorticoid receptor α. These results suggest that the relative levels of the glucocorticoid receptor α and the glucocorticoid receptor P may play a role in the occurrence of glucocorticoid resistance in tumor cells during the treatment of hematological malignancies with glucocorticoids.
Antiapoptotic Effect of Serum and Glucocorticoid-Inducible Protein Kinase Is Mediated by Novel Mechanism Activating IκB Kinase
Authors: Zhang Liping , Cui Ruwen , Cheng Xiaodong , Du Jie
Published: June 2022

Serum and glucocorticoid inducible protein kinase (SGK) plays a crucial role in promoting cell survival, but the mechanisms for this response are not clear. We show that SGK is involved in the regulation of apoptosis in breast cancer cells by modulating the transcriptional activity of nuclear transcription factor κB (NF-κB). High levels of SGK expression were observed in human breast cancer samples. When SGK was reduced the apoptotic rate increased, and increased SGK activity prevents serum withdrawal–induced apoptosis. SGK-induced cell survival was abolished by a dominant-negative form of IκB kinase β (IKKβ, K44A) or a null mutation of IKKβ in mouse embryonic fibroblast cells indicating involvement of the NF-κB pathway. Serum-induced SGK or increased expression of SGK activated NF-κB transcriptional activity, whereas small interference RNA to SGK blocked NF-κB activity. Coexpression of SGK and IKKβ significantly increased the activation of NF-κB (versus expression of IKKβ alone). Expression of dominant-negative IKKβ K44A, IκBα AA, and kinase-dead SGK (127KM) blocked the ability of SGK to stimulate NF-κB activity, suggesting that IKKβ is a target of SGK. We also show that SGK enhances the ability of IKKβ to phosphorylate endogenous IκBα in cells or recombinant glutathione S-transferase-IκBα in vitro and increases IκBα degradation; SGK physically associates with and activates IKKβ in MDA231 cells via phosphorylation of Ser181 in IKKβ. Taken together, we conclude that SGK acts as an oncogene in breast cancer cells through activation of the IKK-NF-κB pathway, thereby preventing apoptosis. Blocking SGK expression/activity represents a potential therapeutic approach for breast cancer treatment.
The Serum Glycoprotein Fetuin-A Promotes Lewis Lung Carcinoma Tumorigenesis via Adhesive-Dependent and Adhesive-Independent Mechanisms
Authors: Kundranda Madappa N. , Henderson Melodie , Carter Kathy J. , Gorden Lee , Binhazim Awadh , Ray Sanhita , Baptiste Trevor , Shokrani Masih , Leite-Browning Maria L. , Jahnen-Dechent Willi , Matrisian Lynn M. , Ochieng Josiah
Published: June 2022

Fetuin-A is a serum glycoprotein in the cystatin family associated with the regulation of soft tissue calcification. We tested the role of systemic fetuin in tumor cell growth and metastasis by injecting Lewis lung carcinoma (LLC) cells into fetuin-A null and their wild-type (WT) littermate control C57BL/6 mice via the tail vein, s.c., and intrasplenic routes. In the experimental metastasis assay, the lungs of the WT mice were filled with metastatic nodules, whereas the lungs of the fetuin-A null mutant mice were virtually free of colonies at the end of 2 weeks. Lung colonization responded to the levels of serum fetuin-A in a dose-dependent manner, as observed by the formation of half as many colonies in mice heterozygous for the fetuin-A locus compared with homozygous WT mice and restoration of lung colonization by the administration of purified fetuin-A to fetuin-A-null mice. Serum fetuin-A also influenced the growth of LLC cells injected s.c.: fetuin-A-null mice developed small s.c. tumors only after a substantial delay. Similarly, intrasplenic injection of LLC cells resulted in rapid colonization of the liver with metastasis to the lungs within 2 weeks in the WT but not fetuin-A null mice. To examine the mechanism by which fetuin-A influences LLC colonization and growth, we showed that LLC tumor cells adhere to fetuin-A in a Ca2+-dependent fashion, resulting in growth of the tumor cells. These studies support the role of fetuin-A as a major growth promoter in serum that can influence tumor establishment and growth.
Induction of Ornithine Decarboxylase Activity Is a Necessary Step for Mitogen-Activated Protein Kinase Kinase–Induced Skin Tumorigenesis
Authors: Feith David J. , Bol David K. , Carboni Joan M. , Lynch Mark J. , Sass-Kuhn Suzanne , Shoop Paula L. , Shantz Lisa M.
Published: June 2022

A transgenic mouse line overexpressing a constitutively active mutant of MEK1, a downstream effector of Ras, driven by the keratin 14 (K14) promoter, has been used to test the hypothesis that ornithine decarboxylase (ODC) induction during tumor promotion following a single initiating event [i.e., the activation of the Raf/mitogen-activated protein kinase kinase/extracellular signal-regulated kinase (Raf/MEK/ERK) pathway], is a necessary step in skin carcinogenesis. K14-MEK mice exhibit moderate hyperplasia, with spontaneous skin tumor development within 5 weeks of birth. Analysis of epidermis and dermis showed induction of MEK protein and ERK1/ERK2 phosphorylation, but no change in Akt-1, suggesting that the PI 3-kinase pathway, another pathway downstream of ras, is not activated. Examination of tumors revealed high levels of ODC protein and activity, indicating that activation of signaling cascades dependent on MEK activity is a sufficient stimulus for ODC induction. When K14-MEK mice were given α-difluoromethylornithine (DFMO), a suicide inactivator of ODC, in the drinking water from birth, there was a dramatic delay in the onset of tumor growth (∼6 weeks), and only 25% of DFMO-treated mice developed tumors by 15 weeks of age. All untreated K14-MEK mice developed tumors by 6 weeks of age. Treatment of tumor-bearing mice with DFMO reduced both tumor size and tumor number within several weeks. Tumor regression was the result of both inhibition of proliferation and increased apoptosis in tumors. The results establish ODC activation as an important component of the Raf/MEK/ERK pathway, and identify K14-MEK mice as a valuable model with which to study the regulation of ODC in ras carcinogenesis.
Loss of Heterozygosity Analysis and DNA Copy Number Measurement on 8p in Bladder Cancer Reveals Two Mechanisms of Allelic Loss
Authors: Adams Jacqui , Williams Sarah V. , Aveyard Joanne S. , Knowles Margaret A.
Published: June 2022

Many epithelial tumors show deletion of the short arm of chromosome 8 that is related to aggressive disease or adverse prognosis. In undissected samples of urothelial cell carcinoma of the bladder, at least two regions of loss of heterozygosity (LOH) were identified previously within a small region of 8p11-p12. LOH analysis on a panel of pure tumor DNA samples confirmed this and identified tumors with allelic imbalance, some with clear breakpoints in 8p12. This suggests either that these samples contained genetically distinct subclones or that breakpoints in 8p12 may confer a selective advantage without LOH. To assess the mechanism of LOH and to map breakpoints precisely, a panel of bladder cancer cell lines was examined. Microsatellite analysis of 8p markers identified regions of contiguous homozygosity that coincided with regions of LOH in tumors. Fluorescence in situ hybridization analysis was carried out on seven cell lines predicted to have 8p LOH using a chromosome 8 paint, a chromosome 8 centromeric probe, and a series of single-copy genomic probes. This revealed overall underrepresentation of 8p and overrepresentation of 8q. Several breakpoints and one interstitial deletion were identified in 8p12. Two cell lines with extensive interstitial regions of homozygosity showed no reduction in DNA copy number by fluorescence in situ hybridization analysis, indicating that, in addition to large deletions and rearrangements of 8p, small regions of interstitial LOH on 8p12 may be generated by mitotic recombination. This implicates both major DNA double-strand break repair mechanisms in the generation of 8p alterations.
The Low Molecular Weight Cyclin E Isoforms Augment Angiogenesis and Metastasis of Human Melanoma Cells In vivo
Authors: Bales Elise , Mills Lisa , Milam Nancy , McGahren-Murray Mollianne , Bandyopadhyay Debdutta , Chen Dahu , Reed Jon A. , Timchenko Nikolai , van den Oord Joost J. , Bar-Eli Menashe , Keyomarsi Khandan , Medrano Estela E.
Published: June 2022

Immunohistochemical analysis has consistently shown that cyclin E is up-regulated in human malignant melanomas in vivo. Here we analyzed such expression in more detail and show that cyclin E is overexpressed and present in low molecular weight (LMW) isoforms in metastatic melanoma and in a subset of primary invasive melanoma tumor tissues, but not in benign nevi. Human metastatic melanoma cell lines, but not normal melanocytes, also expressed the LMW cyclin E forms. The biological significance of these findings was established by showing that overexpression of two LMW cyclin E forms named cyclin E truncated 1 [cyclinE(T1)] and cyclin E truncated 2 [cyclinE(T2)] in a low tumorigenic and non-metastatic primary cutaneous melanoma cell line generated angiogenic tumors with prominent perineural invasion compared with full-length cyclin E. In addition, cyclin E(T1)– and cyclin E(T2)–expressing melanoma cells displayed a dramatic increase in the incidence and number of metastases in an experimental lung metastasis assay. Together, these results indicate that the LMW cyclin E forms are functional and likely act as regulators of human melanoma tumor progression and invasion.
Metastasis Suppression by Breast Cancer Metastasis Suppressor 1 Involves Reduction of Phosphoinositide Signaling in MDA-MB-435 Breast Carcinoma Cells
Authors: DeWald Daryll B. , Torabinejad Javad , Samant Rajeev S. , Johnston Derrick , Erin Nuray , Shope Joseph C. , Xie Yi , Welch Danny R.
Published: June 2022

Several molecules that suppress metastasis without suppressing tumorigenicity have been identified, but their mechanisms of action have not yet been determined. Many block growth at the secondary site, suggesting involvement in how cells respond to signals from the extracellular milieu. Breast cancer metastasis suppressor 1 (BRMS1)–transfected MDA-MB-435 cells were examined for modifications of phosphoinositide signaling as a potential mechanism for metastasis suppression. 435/BRMS1 cells expressed <10% of phosphatidylinositol-4, 5-bisphosphate compared with parental cells, whereas levels of the PtdIns(4)P and phosphatidylinositol-3-phosphate were unchanged. Inositol (1,4,5)-trisphosphate [Ins(1,4,5)P3] were decreased in 435/BRMS1 cells by ∼50%. Phosphatidylinositol-3,4,5-trisphosphate levels were undetectable in 435/BRMS1 cells, even when stimulated by exogenous insulin or platelet-derived growth factor. Immunofluorescence microscopy to examine cellular distribution confirmed that phosphatidylinositol-4,5-bisphosphate distribution with cells was unchanged but was uniformly decreased throughout the cell. Although the gross morphology of 435/BRMS1 cells is similar to the parent, filamentous actin was more readily apparent in 435/BRMS1. Intracellular calcium, measured using Fluo-3 and Fura-2 fluorescent calcium indicator dyes, was somewhat lower, but not statistically different in 435/BRMS1 compared with parental cell. However, when stimulated with platelet-derived growth factor, MDA-MB-435 cells, but not 435/BRMS1 cells mobilized intracellular calcium. Taken together, these results implicate signaling through phosphoinositides in the regulation of breast cancer metastasis, specifically metastasis that can be suppressed by BRMS1.
Secreted Frizzled-Related Protein 4 Is Silenced by Hypermethylation and Induces Apoptosis in β-Catenin–Deficient Human Mesothelioma Cells
Authors: He Biao , Lee Amie Y. , Dadfarmay Sina , You Liang , Xu Zhidong , Reguart Noemi , Mazieres Julien , Mikami Iwao , McCormick Frank , Jablons David M.
Published: June 2022

The secreted frizzled-related proteins (SFRPs) function as negative regulators of Wnt signaling and have important implications in tumorigenesis. Frequent promoter hypermethylation of SFRPs has been identified in human cancer. Restoration of SFRP function attenuates Wnt signaling and induces apoptosis in a variety of cancer types. Wnt signaling is known to inhibit apoptosis through activation of β-catenin/Tcf–mediated transcription. Recently, we identified aberrant Wnt activation as a result of Dishevelled overexpression in malignant mesothelioma. Here, we report that silencing of SFRP4 is correlated with promoter hypermethylation in β-catenin–deficient mesothelioma cell lines. Reexpression of SFRP4 in these β-catenin–deficient mesothelioma cell lines blocks Wnt signaling, induces apoptosis, and suppresses growth. Conversely, knocking down SFRP4 by small interfering RNA in cell lines expressing both SFRP4 and β-catenin stimulates Wnt signaling, promotes cell growth, and inhibits chemodrug-induced apoptosis. Our results suggest that methylation silencing of SFRP4 may play an important role in aberrant Wnt activation in mesothelioma even in the absence of β-catenin. Our data also suggest that β-catenin–independent noncanonical pathway(s) may be involved in the apoptotic inhibition caused by activation of Wnt signaling.
Homeobox Msx1 Interacts with p53 Tumor Suppressor and Inhibits Tumor Growth by Inducing Apoptosis
Authors: Park Kyoungsook , Kim Kwangbae , Rho Seung Bae , Choi Kyusam , Kim Dojin , Oh Sun-Hee , Park Jinhee , Lee Seung-Hoon , Lee Je-Ho
Published: June 2022

The stability of wild-type p53 is critical for its apoptotic function. In some cancers, wild-type p53 is inactivated by interaction with viral and cellular proteins, and restoration of its activity has therapeutic potential. Here, we identify homeobox Msx1 as a p53-interacting protein and show its novel function as a p53 regulator. Overexpression of homeobox Msx1 induced apoptosis of cancer cells harboring nonfunctional wild-type p53 and suppressed growth of human tumor xenografts in nude mice. The homeodomain of Msx1 functions as a protein-protein interacting motif rather than a DNA-binding domain and is essential for stabilization, nuclear accumulation, and apoptotic function of wild-type p53. The identification of a novel function of Msx1 as a p53 regulator may open new avenues for developing improved molecular therapies for tumors with a nonmutational p53 inactivation mechanism.
Suppression of Melanotroph Carcinogenesis Leads to Accelerated Progression of Pituitary Anterior Lobe Tumors and Medullary Thyroid Carcinomas in Rb+/− Mice
Authors: Zhou Zongxiang , Flesken-Nikitin Andrea , Levine Corinna G. , Shmidt Elena N. , Eng Jessica P. , Nikitina Ekaterina Yu. , Spencer David M. , Nikitin Alexander Yu.
Published: June 2022

Mice with a single copy of the retinoblastoma gene (Rb+/−) develop a syndrome of multiple neuroendocrine neoplasia. They usually succumb to fast-growing, Rb-deficient melanotroph tumors of the pituitary intermediate lobe, which are extremely rare in humans. Thus, full assessment of Rb role in other, more relevant to human pathology, neoplasms is complicated. To prevent melanotroph neoplasia while preserving spontaneous carcinogenesis in other types of cells, we have prepared transgenic mice in which 770-bp fragment of pro-opiomelanocortin promoter directs expression of the human RB gene to melanotrophs (TgPOMC-RB). In three independent lines, transgenic mice crossed to Rb+/− background are devoid of melanotroph tumors but develop the usual spectrum of other neoplasms. Interestingly, abrogation of melanotroph carcinogenesis results in accelerated progression of pituitary anterior lobe tumors and medullary thyroid carcinomas. A combination of immunologic tests, cell culture studies, and tumorigenicity assays indicates that α-melanocyte–stimulating hormone, which is overproduced by melanotroph tumors, attenuates neoplastic progression by decreasing cell proliferation and inducing apoptosis. Taken together, we show that cell lineage–specific complementation of Rb function can be successfully used for refining available models of stochastic carcinogenesis and identify α-melanocyte–stimulating hormone as a potential attenuating factor during progression of neuroendocrine neoplasms.
Growth Stimulation of COX-2–Negative Pancreatic Cancer by a Selective COX-2 Inhibitor
Authors: Eibl Guido , Takata Yasunori , Boros Laszlo G. , Liu Joey , Okada Yuji , Reber Howard A. , Hines Oscar J.
Published: June 2022

Cyclooxygenase 2 (COX-2) inhibitors are promising antiangiogenic agents in several preclinical models. The aim of the present study was to evaluate the effect of selective COX-2 inhibitors on vascular endothelial growth factor (VEGF) production in vitro and angiogenesis and growth of pancreatic cancer in vivo, focusing on putative differences between COX-2–negative and COX-2–positive tumors. VEGF production and angiogenesis in vitro were determined by ELISA and endothelial cell migration assay. To determine whether the effect of COX-2 inhibitors was mediated by peroxisome proliferator–activated receptor γ (PPAR-γ), we used a dominant-negative PPAR-γ and a pharmacologic inhibitor. In vitro findings were validated in a pancreatic cancer animal model. Microvessel density was assessed by CD31 immunostaining. Intratumoral prostaglandin and VEGF levels were measured by mass spectroscopy and ELISA. Selective COX-2 inhibitors had a concentration-dependent effect on VEGF production in vitro. Higher concentrations increased VEGF levels and stimulated angiogenesis by activating PPAR-γ. In vivo, nimesulide increased VEGF production by cancer cells in COX-2–positive and COX-2–negative pancreatic tumors. In COX-2–negative pancreatic cancer, this effect was associated with an increase in angiogenesis and growth. In COX-2–positive pancreatic cancer, the nimesulide-induced increase of VEGF production by the cancer cells was offset by a decrease in VEGF production by the nonmalignant cell types leading to reduced tumor angiogenesis and growth. Selective COX-2 inhibitors had opposite effects on growth and angiogenesis in pancreatic cancer depending on COX-2 expression. These findings imply that assessing the COX-2 profile of the pancreatic tumor is mandatory before initiating therapy with a selective COX-2 inhibitor.
Expression-Based Discovery of Variation in the Human Glutathione S-Transferase M3 Promoter and Functional Analysis in a Glioma Cell Line Using Allele-Specific Chromatin Immunoprecipitation
Authors: Liu Xuemei , Campbell Michelle R. , Pittman Gary S. , Faulkner Eric C. , Watson Mary A. , Bell Douglas A.
Published: June 2022

Discovery and functional evaluation of biologically significant regulatory single nucleotide polymorphisms (SNP) in carcinogen metabolism genes is a difficult challenge because the phenotypic consequences may be both transient and subtle. We have used a gene expression screening approach to identify a functional regulatory SNP in glutathione S-transferase M3 (GSTM3). Anttila et al. proposed that variation in GSTM3 expression was affected by exposure to cigarette smoke and inheritance of the GSTM1-null genotype. To investigate the mechanism of GSTM3 expression was affected by exposure to cigarette smoke and inheritance of the GSTM1-null genotype. To investigate the mechanism of GSTM3 expression variation, we measured GSTM3 expression in lymphoblast cells from a human Centre d'Etude du Polymorphisme Humain family and observed a low expression phenotype. Promoter sequencing revealed two novel GSTM3 promoter SNPs: A/C and A/G SNPs, 63 and 783 bp upstream of the codon 1 start site, respectively. In this pedigree, the two children homozygous for the −63C/C genotype had 8-fold lower GSTM3 expression relative to the two children with the −63A/A genotype, with no association between A−783G SNP and GSTM3 expression. Further evaluation using genotyped glioma cell lines and with luciferase reporter constructs showed that the −63C allele was associated with lower GSTM3 expression (P < 0.0001 and P < 0.003). RNA pol II chromatin immunoprecipitation was combined with quantitative probed-based allelic discrimination genotyping to provide direct evidence of a 9-fold reduced RNA pol II binding capacity for the −63C allele. These results show that the GSTM3 −63C allele strongly affects gene expression in human cell lines and suggests that individuals who carry the low expression allele may be deficient in glutathione transferase catalyzed biological functions.
Abstract 6298: The novel long noncoding RNA lncNeur promotes neuroblastoma by up-regulating AURKA and Myc expression
Authors: Wu Jing , Liu Pei Yan , Cheung Belamy , Marshall Glenn , Dinger Marcel , Liu Tao
Published: June 2022

Background: Neuroblastoma is the most common solid tumor in early childhood, and accounts for approximately 15% of childhood cancer death. We have recently analyzed RNA sequencing data from 493 human neuroblastoma tissues, and identified the long noncoding RNA lincNeur as one of the transcripts, high expression of which most considerably correlated with poor prognosis in both MYCN oncogene-amplified and MYCN non-amplified neuroblastoma patients.

Aims: To determine the tumorigenic role of lincNeur in neuroblastoma, and to identify the mechanism of action through which lincNeur exerts oncogenic function, also to screen for small molecule compound inhibitors of lincNeur.

Methods and results: We performed RNA-binding protein pull-down assay and RNA-immunoprecipitation assay. RALY was identified and validated as one of the binding proteins of lincNeur. We also identified down-stream targets of lincNeur via genome-wide differential gene expression analysis. E2F1 and AURKA were identified as among the transcripts significantly down-regulated after lincNeur knockdown. RT-PCR and western blot analysis showed that lincNeur knockdown reduced E2F1 and AURKA mRNA and protein expression, and reduced N-Myc and c-Myc protein but not mRNA expression. In addition, lincNeur or RALY knockdown reduced, while over-expression of lincNeur or RALY promoted, neuroblastoma cell proliferation. We also demonstrated lincNeur knockdown significantly suppressed RALY protein binding to E2F1 and AURKA mRNAs. Mouse xenograft model experiment showed that lincNeur knockdown suppressed neuroblastoma tumor progression in vivo. Importantly, we have confirmed the binding site between lincNeur and RALY, and are screening small molecule compound libraries for inhibitors of lincNeur and RALY interaction for neuroblastoma therapy.

Conclusions: The novel long noncoding RNA lincNeur promotes neuroblastoma by interacting with RALY to up-regulate E2F1 and AURKA mRNA and protein, as well as N-Myc and c-Myc protein expression. Small molecule compound inhibitors of lincNeur and RALY interaction is a promising therapeutic approach.

Citation Format: Jing Wu, Pei Yan Liu, Belamy Cheung, Glenn Marshall, Marcel Dinger, Tao Liu. The novel long noncoding RNA lncNeur promotes neuroblastoma by up-regulating AURKA and Myc expression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6298.
Abstract PL01-05: Genomic medicine meets cancer interception
Authors: Domchek Susan M.
Published: June 2022

The increased identification of individuals with an elevated risk of developing cancer based on inherited genetic susceptibility allows for targeted cancer interception opportunities. Understanding the molecular basis for cancer susceptibility creates the potential to disrupt the oncogenic process even after it has started. Individuals with pathogenic variants in BRCA1 and BRCA2 are at increased risk for the development of breast, ovarian, pancreatic, and prostate cancers. Work over the past decade has allowed for an understanding of the cells of origin of BRCA1 - related breast cancer and has led to a Phase III study of denosumab vs placebo to examine the impact of denosumab on breast cancer risk. The adjuvant study of the PARP inhibitor olaparib in high risk BRCA-associated breast cancer demonstrated an improvement in event free survival along with a favorable safety profile, paving the way for further work in the prevention space. Immunologic approaches have led to clinical trials using vaccines which will include healthy BRCA1/2 carriers. As our molecular understanding of BRCA haploinsufficiency improves, so too will our options for cancer interception. All of this work could lead to a halo effect with impact on sporadic cancer interception.

Citation Format: Susan M. Domchek. Genomic medicine meets cancer interception [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr PL01-05.
Abstract PL04-03: Germline and microbiome variants impact immunotherapy efficacy through modulation of myeloid cells
Authors: Gajewski Thomas F.
Published: June 2022

Most cancers express tumor antigens that can be recognized by T cells of the host. The fact that cancers become clinically relevant and grow nonetheless implies that immune escape must occur to allow cancer outgrowth. We have observed two major phenotypes of human melanoma metastases based on gene expression profiling and confirmatory assays. One subgroup of patients has a T cell-inflamed phenotype that includes expression of chemokines, T cell markers, and a type I IFN signature. In contrast, the other major subset lacks this phenotype and appears to display immune “exclusion”. Factors that influence the degree of spontaneous immune infiltration are being investigated, as sources of inter-patient heterogeneity. These include tumor cell-intrinsic oncogenic events, the composition of the gut microbiota, and polymorphisms in immune regulatory genes. We now know that each of these dimensions can be functionally important. The first tumor cell-intrinsic oncogenic pathway identified that mediates immune exclusion is the Wnt/β-catenin pathway. Tumors with active β-catenin fail to recruit Batf3-lineage dendritic cells into the tumor site. Regarding the commensal microbiota, mouse models identified commensal Bifidobacterium as one key component that augments spontaneous anti-tumor immunity and increases efficacy of anti-PD-L1 therapy in vivo. Similar analyses in human cancer patients revealed bacteria sequences enriched in anti-PD-1 responders, and also bacteria sequences enriched in non-responders. Fecal transfer into germ-free mice has confirmed a causal role for the gut microbiota in regulating immunotherapy efficacy. Recent experiments have revealed that one major mechanism by which gut microbes impact on distant ani-tumor immunity is through modulation of immune-regulatory myeloid cells, ie the M1/M2 ratio and MDSCs. Regarding germline variants, our first identified SNP connected to immune cell infiltration is in the PKCδ gene. Loss of function variants are associated with greater immune cell infiltration. PKCδ knockout hosts show improved immune-mediated tumor control and anti-PD-L1 efficacy, but with comparable T cell priming. However, activated T cell accumulation in the tumor microenvironment increases overtime, which is associated with a shift from M2 to M1. Myeloid cell-specific PKCδKO mice using LysM-Cre Tg mice recapitulate the phenotype. Thus, in each of these 3 instances, tumor and host factors impact on anti-tumor immunity by modulating myeloid cell participation—Batf3-DCs, MDSCs, and M1/M2 cells.

Citation Format: Thomas F. Gajewski. Germline and microbiome variants impact immunotherapy efficacy through modulation of myeloid cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr PL04-03.
Abstract SY03-02: Extrachromosomal DNA (ecDNA): Cancer's dynamic circular genome
Authors: Mischel Paul S.
Published: June 2022

Human genes are arranged on 23 pairs of chromosomes, but in cancer, tumor-promoting genes and regulatory elements can free themselves from chromosomes and relocate to circular, extrachromosomal pieces of DNA (ecDNA). ecDNA poses one of the greatest challenges for cancer patients, affecting children and adults and women and men, with a wide variety of cancer types. Although ecDNA was first observed more than 50 years ago, its critical importance has only recently come to light. ecDNA, because of its non-chromosomal inheritance, drives high-copy number oncogene amplification and enables tumors to evolve their genomes rapidly, contributing to treatment resistance and shorter survival for patients. Furthermore, the circular ecDNA architecture fundamentally alters gene regulation and transcription. In this talk, I will discuss recent collaborative discoveries that highlight how the higher order, spatial organization of ecDNA, and its mechanism of inheritance, contributes to aggressive tumorigenesis, accelerated evolution, and treatment resistance.

Citation Format: Paul S. Mischel. Extrachromosomal DNA (ecDNA): Cancer's dynamic circular genome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr SY03-02.
Abstract SY23-03: Mesenchymal lineage heterogeneity as a determinant of matrix composition and tumor progression
Authors: Sherman Mara H.
Published: June 2022

The mechanisms underlying evolution of tumor-associated stroma remain poorly understood. In solid tumors featuring a prominent stromal reaction, an improved understanding of the functions and origins of abundant stromal cell types may facilitate the development of new and effective therapies. Pancreatic ductal adenocarcinoma (PDAC) is the quintessence of a fibro-inflammatory malignancy, with 50-90% of tumor volume occupied by a dense, desmoplastic stroma. Cancer-associated fibroblasts (CAFs) are the key cell type which drives the stromal reaction in PDAC, and recent reports suggest that stromal CAFs represent a heterogeneous population of cells from diverse origins, potentially including cell types which support and others which suppress tumor growth. Pancreatic stellate cells (PSCs) are lipid-storing cells in healthy pancreas which can transdifferentiate to an activated CAF phenotype. PSCs have been suggested as the predominant source of fibroblasts in the PDAC tumor microenvironment. However, proper lineage tracing studies have never been performed, such that the relative contribution and specific functions of PSCs in the tumor microenvironment are unknown. We have developed a novel mouse model to track PSC differentiation and function during pancreatic tumor progression in vivo. This model revealed that PSC-derived CAFs in fact give rise to a numerically minor but functionally significant subset of PDAC CAFs, and may represent a viable therapeutic target. We adapted our mouse model to enable targeted ablation of PSCs and derivative CAFs within their host tissue for the first time. Functional studies in this model reveal non-redundant functions for PSC-derived CAFs in shaping the PDAC microenvironment, and highlight the significance of mesenchymal lineage heterogeneity for pancreatic tumorigenesis. Further, we find that tumor genotype with respect to p53 status dictates stromal evolutionary routes with respect to cell of origin, a finding that may extend to additional cancer cell-intrinsic genomic alterations. Together, these findings shed light on mechanisms that shape the tumor microenvironment during pancreatic cancer progression, and may hold relevance in additional solid tumor types.

Citation Format: Mara H. Sherman. Mesenchymal lineage heterogeneity as a determinant of matrix composition and tumor progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr SY23-03.
Abstract SY31-04: Uncovering targetable dependencies of latent brain metastatic cells
Authors: Malladi Srinivas
Published: June 2022

Uncovering Targetable Dependencies of Latent Brain Metastatic Cells

Srinivas Malladi, Department of Pathology, Harold C. Simmons Comprehensive Cancer Center, UT Southwestern Medical Center, Dallas, TX, 75390, USA

ABSTRACT

HER2+ breast cancer patients presenting with brain metastases have poor survival outcomes. Metachronous brain metastases or relapse are observed in approximately fifty percent of HER2+ breast cancer patients considered disease free after a variable length of time post primary diagnosis and treatment. Disseminated latent residual cells that survive current therapies are responsible for these relapses or metachronous metastasis. How disseminated tumor cells survive as latent/dormant entities for extended period before initiating metachronous metastasis is poorly understood. Moreover, the basis for disparate metastatic fitness among disseminated tumor cells of similar oncotype within a distal organ is unknown and vital for better clinical management. Through a phenotypic screen in mice, we have isolated HER2+ latent (Lat) and brain metastatic (BM) cells.Transcriptomic and metabolic profiles of these brain-tropic breast cancer cells identified actionable dependencies and unique therapeutic opportunities. Brain-tropic Lat and metastatic cells have distinct metabolic profiles that determine their metastatic fitness. Both latent and metastatic cells survive in equilibrium with innate immune surveillance, oxidize glutamine and maintain cellular redox homeostasis through the anionic amino acid transporter xCT. Likewise, xCT expression was significantly higher in matched metachronous brain metastatic samples compared to primary tumors from HER2+ breast cancer patients. Genetic or pharmacological blockade of xCT eradicates residual disease and brain metastatic relapse in these preclinical models. Lactate secreted by aggressive metastatic cells limits innate immune surveillance and triggers overt metastasis. Furthermore, transcriptomic analysis identified augmented AXL expression. Depleting or inhibiting AXL in these HER2 targeted therapy resistant brain-tropic cells attenuates metastatic incidence. In sum, by investigating phenotypically distinct brain-tropic latent and metastatic cells, we uncovered the impact of metabolic diversity and adaptations on metastatic fitness and identified targetable vulnerabilities.

Citation Format: Srinivas Malladi. Uncovering targetable dependencies of latent brain metastatic cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr SY31-04.
Abstract SY39-01: The DNA damageome and cancer evolution
Authors: Rosenberg Susan
Published: June 2022

DNA damage provokes mutations, genome instability, cancer and biological aging and results from external agents or endogenous cellular processes. We identified networks of unrelated proteins of diverse functions that promote DNA damage when overproduced: the DNA “damageome” proteins (DDPs). We found a large overproduction DDP network in Escherichia coli and find that the proteins promote mutagenesis and genome instability by various mechanisms—three of which we identify. Their 284 human homologs are over-represented among known cancer drivers, and their expression in tumors predicts heavy mutagenesis and poor prognosis. When overproduced in human cells, many of the human homologs promote DNA damage and mutation, with DNA-damaging mechanisms like those in E. coli. As the earliest upstream provocateurs of cancer evolution, the proteins of DNA damageome networks may indicate that there are evolution-accelerating functions of many human known and newly implicated cancer-promoting proteins.

Citation Format: Susan Rosenberg. The DNA damageome and cancer evolution [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr SY39-01.
Abstract SY39-03: Lung cancer evolution and its prognostic impact
Authors: Jamal-Hanjani Mariam
Published: June 2022

Lung cancer is the leading cause of cancer-associated mortality worldwide, and genomic intratumour heterogeneity as a result of cancer evolution has been linked to therapeutic resistance and poor outcomes. TRACERx is a prospective longitudinal cancer evolution programme which aims to investigate the relationship between intratumour heterogeneity and clinical outcomes, and to map the evolutionary pathways adopted in lung cancer from early to late stage disease. Here we present our findings from mutational, copy number and mutational signature analyses in over 420 tumours, and provide insights into the shared patterns of tumour evolution and their impact on prognosis. We demonstrate a significant association between somatic copy number alteration (SCNA) intratumour heterogeneity, defined as the percentage of subclonal SCNAs as previously reported, and poor outcomes. Furthermore, we find that subclonal genome doubling, a driver of SCNA intratumour heterogeneity, is associated with poor outcome independent of clinical variables, such as tumour stage.

Citation Format: Mariam Jamal-Hanjani. Lung cancer evolution and its prognostic impact [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr SY39-03.
Impaired Antitumor Immune Response inMYCN-amplified Neuroblastoma Is Associated with Lack of CCL2 Secretion and Poor Dendritic Cell Recruitment
Authors: Kacher Jamila , Manches Olivier , Aspord Caroline , Sartelet Hervé , Chaperot Laurence
Published: June 2022

In neuroblastoma, MYCN amplification is associated with sparse immune infiltrate and poor prognosis. Dendritic cells (DC) are crucial immune sentinels but their involvement in neuroblastoma pathogenesis is poorly understood. We observed that the migration of monocytes, myeloid and plasmacytoid DC induced by MYCN-nonamplified neuroblastoma supernatants was abrogated by the addition of anti-CCL2 antibodies, demonstrating the involvement of the CCR2/CCL2 axis in their recruitment by these tumors. Using public RNA sequencing and microarray datasets, we describe lower level of expression of CCL2 in MYCN-amplified neuroblastoma tumors, and we propose a working model for T-cell recruitment in neuroblastoma tumors in which CCL2 produced by neuroblastoma cells initiates the recruitment of monocytes, myeloid and plasmacytoid DCs. Among these cells, the CD1c+ subset may recruit T cells by means of CCL19/CCL22 secretion. In vitro, supernatants from DCs cocultured with neuroblastoma cell lines and activated contain CCL22 and CCL19, and are chemotactic for both CD4+ and CD8+ T cells. We also looked at immunomodulation induced by neuroblastoma cell lines, and found MYCN-nonamplified neuroblastoma cell lines were able to create a microenvironment where DC activation is enhanced. Overall, our findings highlight a major role for CCL2/CCR2 axis in monocytes, myeloid and plasmacytoid cells recruitment toward MYCN-nonamplified neuroblastoma, allowing further immune cell recruitment, and show that these tumors present a microenvironment that can favor DC responses.

Significance:

In MYCN-nonamplified neuroblastoma, CCL2 produced by neuroblastoma cells induces the recruitment of antigen-presenting cells (DCs and monocytes/macrophages), allowing infiltration by T cells, in link with CCL19 and CCL22 production, hence favoring immune responses.
Paediatric Dialysis at a Tertiary Hospital in South-West Nigeria: A 4-Year Report
Authors: Ademola Adebowale D. , Asinobi Adanze O. , Alao Michael A. , Wade Andrew W.
Published: June 2022

Introduction: Dialysis is potentially lifesaving in children with acute kidney injury (AKI) or chronic kidney disease (CKD), but availability is limited in low-income countries and lower-middle-income countries (LMICs). Methods: In the present study, we perform a 4-year study of patients who received peritoneal dialysis (PD) or haemodialysis (HD) at the Paediatric Nephrology Unit of the University College Hospital Ibadan, Nigeria. Subgroup analysis was performed on patients with sepsis or malaria AKI who underwent HD or PD for predictors of in-hospital mortality. Results: A total of 167 children aged 7 days to 18 years, median 7 (interquartile range 3–12) years, (60.5% males) were studied. In total, 129 (77.2%) had AKI, while 38 had CKD. Regarding AKI, 83 children (64.3%) received HD only, 42 underwent PD only, while 4 underwent both HD and PD. Malaria AKI was treated with HD in 43 (51.8%) or PD in 8 (10.5%), while sepsis AKI was treated with HD in 20 (21.4%) or PD in 33 (78.6%). Mortality in AKI was 16.3% overall, 10.8% in children on HD only, and 26.2% in children on PD only. Patients with sepsis AKI had higher mortality compared to patients with malaria AKI (RR 7.96 [1.70–37.37]). Subgroup analysis showed that age, diagnosis, and dialysis modality were not independent risk factors for mortality. The aetiology of CKD was glomerulonephritis in 26 (68.4%): treatment was HD in 36 and PD in 2 with mortality being 26.3%. Conclusions: PD for AKI showed relatively good outcomes in a LMIC. However, funding and support for a formal dialysis program for the management of AKI and CKD are needed.
Fibrillary Glomerulonephritis, DNAJB9, and the Unfolded Protein Response
Authors: Andeen Nicole K. , Kung Vanderlene L. , Robertson Josh , Gurley Susan B. , Avasare Rupali S. , Sitaraman Sneha
Published: June 2022

Background: Fibrillary glomerulonephritis (FGN) is found in approximately 1% of native kidney biopsies and was traditionally defined by glomerular deposition of fibrils larger than amyloid (12–24 nm diameter) composed of polyclonal IgG. Recent identification of DNAJB9 as a sensitive and specific marker of FGN has revolutionized FGN diagnosis and opened new avenues to studying FGN pathogenesis. In this review, we synthesize recent literature to provide an updated appraisal of the clinical and pathologic features of FGN, discuss diagnostic challenges and pitfalls, and propose molecular models of disease in light of DNAJB9. Summary: DNAJB9 tissue assays, paraffin immunofluorescence studies, and IgG subclass testing demonstrate that FGN is distinct from other glomerular diseases with organized deposits and highlight FGN morphologic variants. Additionally, these newer techniques show that FGN is only rarely monoclonal, and patients with monoclonal FGN usually do not have a monoclonal gammopathy. DNAJB9 mutation does not appear to affect the genetic architecture of FGN; however, the accumulation of DNAJB9 in FGN deposits suggests that disease is driven, at least in part, by proteins involved in the unfolded protein response. Treatments for FGN remain empiric, with some encouraging data suggesting that rituximab-based therapy is effective and that transplantation is a good option for patients progressing to ESKD. Key Messages: DNAJB9 aids in distinguishing FGN from other glomerular diseases with organized deposits. Further investigations into the role of DNAJB9 in FGN pathogenesis are necessary to better understand disease initiation and progression and to ultimately develop targeted therapies.