Malignant Tumor Formation Research Articles

LIN28B promotes cell invasion and colorectal cancer metastasis via CLDN1 and NOTCH3.

The RNA-binding protein LIN28B is overexpressed in over 30% of patients with colorectal cancer (CRC) and is associated with poor prognosis. In the present study, we unraveled a potentially novel mechanism by which LIN28B regulates colonic epithelial cell-cell junctions and CRC metastasis. Using human CRC cells (DLD-1, Caco-2, and LoVo) with either knockdown or overexpression of LIN28B, we identified claudin 1 (CLDN1) tight junction protein as a direct downstream target and effector of LIN28B. RNA immunoprecipitation revealed that LIN28B directly binds to and posttranscriptionally regulates CLDN1 mRNA. Furthermore, using in vitro assays and a potentially novel murine model of metastatic CRC, we show that LIN28B-mediated CLDN1 expression enhances collective invasion, cell migration, and metastatic liver tumor formation. Bulk RNA sequencing of the metastatic liver tumors identified NOTCH3 as a downstream effector of the LIN28B/CLDN1 axis. Additionally, genetic and pharmacologic manipulation of NOTCH3 signaling revealed that NOTCH3 was necessary for invasion and metastatic liver tumor formation. In summary, our results suggest that LIN28B promotes invasion and liver metastasis of CRC by posttranscriptionally regulating CLDN1 and activating NOTCH3 signaling. This discovery offers a promising new therapeutic option for metastatic CRC to the liver, an area where therapeutic advancements have been relatively scarce.

Cost-effective Whole Exome Sequencing discovers pathogenic variant causing Neurofibromatosis type 1 in a family from Jammu and Kashmir, India

Neurofibromatosis type 1 (NF1) is a multisystemic hereditary disorder associated with an increased risk of benign and malignant tumor formation predominantly on the skin, bone, and peripheral nervous system. It has been reported that out of all the NF1 cases, more than 95% cases develop the disease due to heterozygous loss-of-function variants in Neurofibromin (NF1) gene. However, identification of NF1 causative variants by presently recommended method of gene-targeted Sanger sequencing is challenging and cost-intensive due to the large size of the NF1gene with 60 exons spanning about 350 kb. Further, conducting the genetic studies is difficult in low resource regions and among families with the limited financial capabilities, restricting them from availing diagnostic as well as proper disease management measures. Here, we studied a three-generation family from Jammu and Kashmir state in India, with multiple affected family members showing clinical indications of NF1. We combinedly used two applications, Whole Exome Sequencing (WES) and Sanger sequencing, for this study and discovered a nonsense variant NM_000267.3:c.2041C>T (NP_000258.1:p.Arg681Ter*) in exon 18 of NF1 gene in a cost effective manner. In silico analyses further substantiated the pathogenicity of this novel variant. The study also emphasized on the role of Next Generation Sequencing (NGS) as a cost-effective method for the discovery of pathogenic variants in disorders with known phenotypes found in large sized candidate genes. The current study is the first study based on the genetic characterization of NF1 from Jammu and Kashmir–India, highlighting the importance of the described methodology adopted for the identification and understanding of the disease in low resource region. The early diagnosis of genetic disorders would open the door to appropriate genetic counseling, reducing the disease burden in the affected families and the general population at large.

Epigenetic regulation of SMAD3 by histone methyltransferase SMYD2 promotes lung cancer metastasis

Epigenetic alterations, especially histone methylation, are key factors in cell migration and invasion in cancer metastasis. However, in lung cancer metastasis, the mechanism by which histone methylation regulates metastasis has not been fully elucidated. Here, we found that the histone methyltransferase SMYD2 is overexpressed in lung cancer and that knockdown of SMYD2 could reduce the rates of cell migration and invasion in lung cancer cell lines via direct downregulation of SMAD3 via SMYD2-mediated epigenetic regulation. Furthermore, using an in vitro epithelial-mesenchymal transition (EMT) system with a Transwell system, we generated highly invasive H1299 (In-H1299) cell lines and observed the suppression of metastatic features by SMYD2 knockdown. Finally, two types of in vivo studies revealed that the formation of metastatic tumors by shSMYD2 was significantly suppressed. Thus, we suggest that SMYD2 is a potential metastasis regulator and that the development of SMYD2-specific inhibitors may help to increase the efficacy of lung cancer treatment.

Abstract 4817: Protein aggregation promotes HSF1 activity enhancing cell survival during metastatic breast cancer colonization

Abstract Breast cancer is the most commonly diagnosed cancer in women and is the second leading cause of cancer-related deaths in women. Approximately 20-30% of patients will develop metastases and metastasis is responsible for greater than 90% of breast cancer deaths. Metastasis is a complex process in which the cells combat many forces to survive and spread to different areas of the body. Metastatic colonization is the rate-limiting step of metastasis and is an inefficient process in which most cells die and only a small fraction of those that survive can form metastases. We have previously shown that heat shock factor 1 (HSF1) promoted epithelial-to-mesenchymal transition (EMT) and the breast cancer stem-like population, potentially linking HSF1 to metastasis. Utilizing an HSF1 gene expression signature that assesses HSF1 transcriptional activity, we further found that patients with high HSF1 activity have significantly worse metastasis-free survival. The physiological function of HSF1 is the master regulator of the heat shock response wherein it upregulates chaperone proteins under stress conditions. Because of these functions and the fact that the process of metastatic colonization is known to involve the stem cell population and incur external stressors, we hypothesized that HSF1 may function in metastatic colonization. To test this, we subjected human breast cancer MDA- MB-231 cells with or without HSF1 knockdown to intracardiac injections in nude mice. This model injects cells directly into the circulation allowing for assessment of metastatic tumor formation in which the major barrier will be metastatic colonization. Mice receiving cells with knockdown of HSF1 had a significantly reduced metastatic burden, indicating HSF1 is necessary for the completion of metastasis and colonization. Consistent with these findings, bone metastatic tumor specimens from patients show increased HSF1 activation compared to their matched primary breast tumors. The mechanism by which HSF1 enables metastatic colonization is unknown. Metastatic colonization likely requires at least two stages that include tumor initiation (or early colonization) characterized by the seeding of a tumor followed by tumor expansion (or late colonization) characterized by rapid proliferation and an increase in tumor size. We observed increased protein amyloid aggregates that correlate with an increase in HSF1 activity during mammosphere formation, suggesting that colonization induces aggregation leading to HSF1 activation that promotes a cell survival response. Overall, my results indicate that HSF1 activity increased during breast cancer metastasis and HSF1 is necessary for colonization. Going forward, I want to understand what controls HSF1 activation during metastasis, which I hypothesize that protein aggregation is increased leading to HSF1 activation. Citation Format: Natasha Hockaden, Gabi Leriger, John Wang, Haimanti Ray, Richard Carpenter. Protein aggregation promotes HSF1 activity enhancing cell survival during metastatic breast cancer colonization. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4817.

Identification of novel peptide inhibitors for oncogenic KRAS G12D as therapeutic options using mutagenesis-based remodeling and MD simulations

The Kirsten rat sarcoma 2 viral oncogene homolog (KRAS) serves as a molecular switch, cycling between guanosine triphosphate (GTP)-bound and inactive guanosine diphosphate (GDP)-bound states. KRAS modulates numerous signal transduction pathways including the conventional RAF–MEK–ERK pathway. Mutations in the RAS genes have been linked to the formation of malignant tumors. Human malignancies typically show mutations in the Ras gene including HRAS, KRAS, and NRAS. Among all the mutations in exon 12 and exon 13 of the KRAS gene, the G12D mutation is more prevalent in pancreatic and lung cancer and accounts for around 41% of all G12 mutations, making them potential anticancer therapeutic targets. The present study is aimed at repurposing the peptide inhibitor KD2 of the KRAS G12D mutant. We employed an in-silico mutagenesis approach to design novel peptide inhibitors from the experimentally reported peptide inhibitor, and it was found that substitutions (N8W, N8I, and N8Y) might enhance the peptide’s binding affinity toward the KRAS. Molecular dynamics simulations and binding energy calculations confirmed that the newly designed peptide inhibitors are stable and that their binding affinities are stronger as compared to the wild-type peptide. The detailed analysis revealed that newly designed peptides have the potential to inhibit KRAS/Raf interaction and the oncogenic signal of the KRAS G12D mutant. Our findings strongly suggest that these peptides should be tested and clinically validated to combat the oncogenic activity of KRAS. Communicated by Ramaswamy H. Sarma

Multifunctional biomimetic nanoplatform based on photodynamic therapy and DNA repair intervention for the synergistic treatment of breast cancer.

Photodynamic therapy (PDT) is a minimally invasive and locally effective treatment method, which has been used in the clinical treatment of a variety of superficial tumors. In recent years, PDT has received extensive attention due to its induction of immunogenic cell death (ICD). However, the repair mechanism of tumor cells and low immune response limit the further development of PDT. To this end, a multifunctional biomimetic nanoplatform 4T1Mem@PGA-Ce6/Ola (MPCO) is developed to co-deliver the photosensitizer Chlorin e6 (Ce6) and Olaparib (Ola) with the function of preventing DNA repair. The nanoplatform shows efficient tumor targeting and cellular internalization properties due to cell membrane camouflage, and Ce6 and Ola produce a significant synergistic anti-tumor effect under laser irradiation. Meanwhile, the nanoplatform can also activate the cyclic guanosine monophosphate-adenosine monophosphate synthase-interferon gene stimulator signaling (cGAS-STING) pathway to produce cytokines. The damage-associated molecular patterns induced by ICD can work with these cytokines to recruit and stimulate the maturation of dendritic cells and induce the systemic anti-tumor immune response. Overall, this multifunctional biomimetic nanoplatform integrating PDT, chemotherapy, and immunotherapy is highlighted here to boost anti-tumor therapy. STATEMENT OF SIGNIFICANCE: Self-repair of DNA damage is the most important reason for the failure of primary tumor eradication and the formation of secondary and metastatic tumors. To address this issue, a multifunctional biomimetic nanoplatform 4T1Mem@PGA-Ce6/Ola (MPCO) was developed to integrate a photosensitizer Chlorine a6 and a poly (ADP-ribose) polymerase inhibitor Olaparib. With tumor targeting ability and controlled release of drugs, the MPCO was expected to enhance tumor immunogenicity and facilitate antitumor immunity through the induction of immunogenic cell death as well as the activation of the cGAS-STING pathway. This study develops a promising combination strategy against tumors and has substantial implications for the prognosis of patients with breast cancer.

Abstract B035: CXCL12 / CXCR4 signaling enhances and sustains migrating cancer stem cells via BMI1 in pancreatic ductal adenocarcinomas

Abstract Pancreatic cancer is a fatal disease and is one of the most aggressive and metastatic malignancies worldwide. The dissemination of tumor cells is the prerequisite of metastases and correlates with a loss of epithelial differentiation and the acquisition of a migratory phenotype, a hallmark of malignant tumor progression. Migrating cancer stem cells (miCSCs) characterized by CD133+ and CXCR4+ expression play a pivotal role in malignant tumor formation, have been reported to form the invasive front of the metastasis and in silico analysis showed that both CSCs and miCSCs are significantly upregulated in PDAC. However, the regulatory pattern maintaining these CSCs and especially miCSCs in PDAC remains widely elusive. Thus, this study further helps to unravel pathways responsible for maintenance of CSC (CD133+) and miCSC (CD133+ CXCR4+) population. To identify key signaling pathways responsible for both CSCs and miCSCs maintenance, we first generated a protein-protein interaction network using STRING database. Afterwards, we validated these signaling pathway(s) involved in aiding CSC and miCSC population by performing shRNA mediated knockdown of key signaling proteins in different patient-derived pancreatic cancer cell lines. Moreover, we interrogated the involvement of the tumor-stroma crosstalk in the regulation of these pathways by co-culturing tumor cells with pancreatic stellate cells. Protein-protein interaction network incorporating relevant factors involved in EMT, stemness, as well as SHH, NF-kB and AKT signaling pathway identified a strong link between the CXCL12/CXCR4 signaling axis and BMI1. Migration assay, sphere formation assay and western blot upon shRNA mediated knockdown of either CXCR4 and/or BMI1 ascertained BMI1 as a key player downstream of the CXCL12/CXCR4 axis to mechanistically effect both EMT and stemness. Pathway focused gene expression analysis as well as ELISA and immunofluorescence for CXCL12 and actin filaments, respectively, revealed the indispensable nature of tumor-stroma crosstalk on promoting CSC and miCSC population through the chemokine CXCL12. In addition, co-culture systems revealed that particularly pancreatic stellate cells play a significant role in maintaining both CSCs and miCSCs population as well as their characteristic phenotype including but not limited to chemotherapy resistance. Taken together, our results obtained in this study established mechanistically that the CXCL12/CXCR4 signaling pathway driven by tumor-stroma crosstalk not only enhances but also maintains both CSCs and miCSCs in pancreatic ductal adenocarcinomas through BMI1 ultimately promoting metastases and therapeutic resistance. Citation Format: Kanishka Tiwary, Anton Lahusen, Syeda Inaas, Stefanie Hauff, Karolin Walter, Alexander Kleger, Thomas Seufferlein, Bruno Sainz Jr., Patrick Christian Hermann. CXCL12 / CXCR4 signaling enhances and sustains migrating cancer stem cells via BMI1 in pancreatic ductal adenocarcinomas [abstract]. In: Proceedings of the AACR Special Conference: Cancer Metastasis; 2022 Nov 14-17; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;83(2 Suppl_2):Abstract nr B035.

PHF5A facilitates the development and progression of gastric cancer through SKP2-mediated stabilization of FOS

BackgroundGastric cancer (GC) is the fifth most common cancer and the third most common cause of cancer death worldwide. Plant homeodomain (PHD)-finger domain protein PHF5A has been demonstrated to play a promoting role in a variety of cancers. This study aimed to clarify the role of PHF5A in the progression of GC and its potential mechanism of action.MethodsImmunohistochemical staining experiments were performed based on tissues from clinical GC patients to reveal PHF5A expression. A series of functional experiments in vitro and in vivo were used to clarify the role of PHF5A in GC.ResultsClinically, PHF5A was abundantly expressed in GC and existed clinical value indicating poor prognosis. In addition, GC cells with knockdown of PHF5A expression showed slowed proliferation, enhanced sensitivity to apoptosis and inhibition of migration. Mechanically, knockdown of PHF5A led to decreased protein stability of FOS, which was mediated ubiquitination of E3 ubiquitin ligase S-phase kinase-associated protein 2 (SKP2). Moreover, downregulation of FOS attenuated the promotion of PHF5A overexpression on GC cells. Consistently, Pladienolide B (PHF5A inhibitor) treatment reversed the induction of PHF5A overexpression on the malignant phenotypes and tumor formation of GC cells.ConclusionKnockdown of PHF5A inhibited the progression of GC through SKP2-mediated ubiquitination of FOS, which may be a promising candidate target with potential therapeutic value.

Bisphenol A exposure inhibits vascular smooth muscle cell responses: Involvement of proliferation, migration, and invasion

Previous studies have associated bisphenol A (BPA) with malignant tumor formation, infertility, and atherosclerosis in vitro and in vivo. However, the precise mechanisms through which BPA affects the cardiovascular system under normal conditions remain unclear. Therefore, this study investigated the biological mechanisms through which BPA affects the responses of aortic vascular smooth muscle cells (VSMCs). BPA treatment inhibited the proliferative activity of VSMCs and induced G2/M-phase cell cycle arrest via stimulation of the ATM-CHK2-Cdc25C-p21WAF1-Cdc2 cascade in VSMCs. Furthermore, BPA treatment upregulated the phosphorylation of mitogen-activated protein kinase (MAPK) pathways such as ERK, JNK, and p38 MAPK in VSMCs. However, the phosphorylation level of AKT was down-regulated by BPA treatment. Additionally, the phosphorylation of ERK, JNK, and p38 MAPK was suppressed when the cells were treated with their respective inhibitors (U0126, SP600125, and SB203580). BPA suppressed MMP-9 activity by reducing the binding activity of AP-1, Sp-1, and NF-κB, thus inhibiting the invasive and migratory ability of VSMCs. These data demonstrate that BPA interferes with the proliferation, migration, and invasion capacities of VSMCs. Therefore, our findings suggest that overexposure to BPA can lead to cardiovascular damage due to dysregulated VSMC responses.

Viral Carcinogenesis: A Concise Review

Abstract As per the International Classification of Diseases, oral cancer is a subtype of head-and-neck cancer which develops in the tongue, lips, salivary glands, floor of the mouth, gingiva, buccal surfaces, oropharynx, and other intraoral areas. It is commonly presumed at least 5 to 10 mutations in different genes cause the transformation of normal cells to malignant. They may be triggered on by exposure to a single agent or a combination of agents, random errors in genetic replication, or errors in DNA repair procedures. Oncogenic viruses are a serious threat to humans, farm animals, and pets. These viruses cause cell transformation, uncontrollable cell growth, and the formation of malignant tumors. This review summarizes on the role of such viruses in carcinogenesis, antiviral therapy, and therapeutic vaccines.

RAB11A Promotes Cell Malignant Progression and Tumor Formation of Prostate Cancer via Activating FAK/AKT Signaling Pathway.

RAB11A, a member of the GTPase family, acts as a regulator in diverse cancers development. The dysregulation of the FAK/AKT signaling pathway is mainly related to tumorigenesis. This study aimed to investigate the possible effect of RAB11A in prostate cancer and further explore the potential mechanisms. In this study, we illustrated the tumor-promoting effects of RAB11A based on in vivo and in vitro experiments. RAB11A expression was upregulated in prostate cancer cells. RAB11A knockdown decreased the prostate cancer cell proliferation, migration, and invasion. RAB11A also induced the epithelial-mesenchymal transition. PF562271 suppressed the malignant characteristics of prostate cancer cells caused by RAB11A knockdown. Furthermore, the interference of RAB11A reduced the tumor growth and the protein levels of p-FAK/FAK and p-AKT/AKT in vivo. RAB11A promotes cell malignant progression and tumor formation in prostate cancer via activating FAK/AKT signaling pathway.

Poly(ADP-ribosyl)ating enzymes cooperate to coordinate development

The transcriptome is subject to rapid and massive changes during the transition between developmental stages. These changes require tight control to avoid the undesired reactivation of gene expression that is only important for previous developmental stages and, if unchecked during transition between developmental stages, could lead to anarchic proliferation and formation of malignant tumors. In this context, the involvement of chromatin factors is important since they can directly regulate the expression of multiple genes at the same time. Poly(ADP-ribose) enzymes, involved in several processes from DNA repair to transcription regulation, might play a role in this regulation. Here, we report that PARP-1 and PARG cooperate to temporally regulate the gene expression profile during the larval/pupa transition. PARP-1 and PARG are both essential in repressing the expression of genes coding for digestive enzymes and larval cuticle proteins, while PARG positively regulate the expression of defense response genes. These results suggest a cooperative coordination between PARP-1 and PARG that specifically maintains the integrity of expression profile between developmental stages.

Abstract B007: LIN28B promotes collective cell invasion and colorectal cancer metastasis via a novel CLDN1 and NOTCH3 axis

Abstract Colorectal cancer (CRC) is the third most commonly diagnosed cancer and the second most common cause of cancer death worldwide. The 5-year survival rate is 90% in patients with localized tumors, but the survival rate drastically decreases to 14% in patients with distant metastasis. Therefore, it is imperative to elucidate the molecular mechanisms underlying CRC metastasis. The RNA-binding protein LIN28B is overexpressed in over 30% of patients with CRC and is associated with poor prognosis. Indeed, our previous work revealed that LIN28B promotes liver metastasis in a subcutaneous xenograft model of CRC as well as a portal vein injection model. In the present study, we investigated the mechanism by which LIN28B promotes colorectal tumor progression and metastasis. To assess biological changes induced by LIN28B, we established CRC cells (DLD-1 and LoVo) with high expression of LIN28B (LIN28Bhigh). LIN28B overexpression upregulated Claudin-1 (CLDN1), a protein that functions as a major constituent of the tight junction complexes. RNA immunoprecipitation revealed that LIN28B directly binds to and stabilizes CLDN1 at a post-transcriptional level. Knockdown of CLDN1 expression in LIN28Bhigh cells suppresses cell aggregation, wound healing rate, and collective invasion. Using a mouse model of metastatic CRC, we reveal that knockdown of CLDN1 inhibits liver metastasis of LIN28Bhigh CRC cells. RNA-sequencing of metastatic liver tumors from LIN28Bhigh cells showed that NOTCH3 works downstream of the LIN28B-CLDN1 axis to induce cell aggregation, collective invasion, and metastatic liver tumor formation. Inhibition of Notch signaling by DAPT, an inhibitor of the γ-secretase complex, significantly reduced metastatic liver tumors in vivo. Taken together, our results indicate that LIN28B promotes cell aggregation, invasion, and liver metastasis in CRC through post-transcriptional induction of CLDN1 and upregulation of NOTCH3. Development of new therapies that target LIN28B-CLDN1-NOTCH3 axis may provide an effective strategy for stage 4 CRC. This work was supported by 9R01CA277795-22. Citation Format: Alice E. Shin, Kensuke Sugiura, Yasunori Masuike, Kensuke Suzuki, Christopher J. Lengner, Anil K. Rustgi. LIN28B promotes collective cell invasion and colorectal cancer metastasis via a novel CLDN1 and NOTCH3 axis [abstract]. In: Proceedings of the AACR Special Conference on Colorectal Cancer; 2022 Oct 1-4; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;82(23 Suppl_1):Abstract nr B007.

Effects of β2 Integrins on Osteoclasts, Macrophages, Chondrocytes, and Synovial Fibroblasts in Osteoarthritis.

β2 integrins are transmembrane receptors that exist widely in human immune cells and participate in pathological processes such as chronic inflammation, thrombosis, and malignant tumor formation. They mainly mediate intercellular adhesion, coordinate the ingestion of extracellular matrix components, and regulate cytoskeleton formation, thereby regulating cell signaling. Osteoarthritis (OA) is a chronic joint disease that causes joint pain and increases disease burden; it has a high prevalence among populations worldwide. Previous studies have reported that β2 integrins are overexpressed in OA and may play an essential role in the occurrence of OA. The important roles of β2 integrins in the maturation and differentiation of osteoclasts, the regulation of bone homeostasis, and the polarization and migration of macrophages have also been reported. The present review aims to highlight the role of β2 integrins in OA pathogenesis and outline their potential for serving as therapeutic targets.

Von Hippel–Lindau (VHL) disease and VHL-associated tumors in Indian subjects: VHL gene testing in a resource constraint setting

BackgroundVon Hippel–Lindau (VHL) syndrome is a familial cancer syndrome caused by mutations in VHL gene. It is characterized by the formation of benign and malignant tumors like retinal angioma, cerebellar hemangioblastoma, spinal hemangioblastoma, renal cell carcinoma, pheochromocytoma, pancreatic and renal cysts, and endolymphatic sac tumors. Germline mutations in VHL gene have also been reported in isolated VHL-associated tumors. VHL gene is a small gene with 3 coding exons and can be easily tested even in a resource constraint setting.ObjectiveTo describe clinical presentation and estimate the diagnostic yield of in VHL and VHL-associated tumors.MethodsThis is a descriptive study in a hospital setting. Here, we describe the clinical and molecular data of 69 patients with suspected VHL or having VHL-associated tumors. Sanger sequencing of coding sequences and conserved splice sites of VHL gene were done in all patients. Multiplex ligation-dependent probe amplification (MLPA) of VHL gene to detect large deletions/duplications was performed for 18 patients with no pathogenic sequence variations.ResultsAmong tumor types at presentation, pheochromocytoma was seen in 49% (34/69), hemangioblastoma was seen in 30% (21/69), and renal cell carcinoma was seen in 7% (5/69). Rest had other tumors like paraganglioma, endolymphatic sac papillary tumors, cerebellar astrocytoma and pancreatic cyst. Seven patients (10%) had more than one tumor at the time of diagnosis. Pathogenic variations in VHL gene were identified in 31probands by Sanger sequencing; 18 were missense, 2 nonsense and 2 small indels. A heterozygous deletion of exon 3 was detected by MLPA in one patient among 18 patients for whom MLPA was done. Overall, the molecular yield was 46% cases (32/69). Family history was present in 7 mutation positive cases (22%). Overall, 11 families (16%) opted for pre-symptomatic mutation testing in the family.ConclusionsMutation testing is indicated in VHL and VHL-associated tumors. The testing facility is easy and can be adopted easily in developing countries like India. The yield is good, and with fairly high incidence of familial cases, molecular testing can help in pre-symptomatic testing and surveillance.

Establishment of a Prognostic Model for Hepatocellular Carcinoma Based on Bioinformatics and the Role of NR6A1 in the Progression of HCC.

Background and AimsGenerally acceptable prognostic models for hepatocellular carcinoma (HCC) are not available. This study aimed to establish a prognostic model for HCC by identifying immune-related differentially expressed genes (IR-DEGs) and to investigate the potential role of NR6A1 in the progression of HCC.MethodsBioinformatics analysis using The Cancer Genome Atlas and ImmPort databases was used to identify IR-DEGs. Lasso Cox regression and multivariate Cox regression analysis were used to establish a prognostic model of HCC. Kaplan-Meier analysis and the receiver operating characteristic (ROC) curves were used to evaluate the performance of the prognostic model, which was further verified in the International Cancer Genome Consortium (ICGC) database. Gene set enrichment analysis was used to explore the potential pathways of NR6A1. Cell counting kit 8, colony formation, wound healing, and Transwell migration assays using Huh7 cells, and tumor formation models in nude mice were conducted.ResultsA prognostic model established based on ten identified IR-DEGs including HSPA4, FABP6, MAPT, NDRG1, APLN, IL17D, LHB, SPP1, GLP1R, and NR6A1, effectively predicted the prognosis of HCC patients, was confirmed by the ROC curves and verified in ICGC database. NR6A1 expression was significantly up-regulated in HCC patients, and NR6A1 was significantly associated with a low survival rate. Gene set enrichment analysis showed the enrichment of cell cycle, mTOR, WNT, and ERBB signaling pathways in patients with high NR6A1 expression. NR6A1 promoted cell proliferation, invasiveness, migration, and malignant tumor formation and growth in vitro and in vivo.ConclusionsAn effective prognostic model for HCC, based on a novel signature of 10 immune-related genes, was established. NR6A1 was up-regulated in HCC and was associated with a poor prognosis of HCC. NR6A1 promoted cell proliferation, migration, and growth of HCC, most likely through the cell cycle, mTOR, WNT, and ERBB signaling pathways.

Role of microRNA/lncRNA Intertwined With the Wnt/β-Catenin Axis in Regulating the Pathogenesis of Triple-Negative Breast Cancer.

Objective (s): In this mini-review, we aimed to discuss the Wnt/β-catenin signaling pathway modulation in triple-negative breast cancer, particularly the contribution of lncRNAs and miRNAs in its regulation and their possible entwining role in breast cancer pathogenesis, proliferation, migration, or malignancy. Background: Malignant tumor formation is very high for breast cancer in women and is a leading cause of death all over the globe. Among breast cancer subtypes, triple-negative breast cancer is rife in premenopausal women, most invasive, and prone to metastasis. Complex pathways are involved in this cancer’s pathogenesis, advancement, and malignancy, including the Wnt/β-catenin signaling pathway. This pathway is conserved among vertebrates and is necessary for sustaining cell homeostasis. It is regulated by several elements such as transcription factors, enhancers, non-coding RNAs (lncRNAs and miRNAs), etc. Methods: We evaluated lncRNAs and miRNAs differentially expressed in triple-negative breast cancer (TNBC) from the cDNA microarray data set literature survey. Using in silico analyses combined with a review of the current literature, we anticipated identifying lncRNAs and miRNAs that might modulate the Wnt/β-catenin signaling pathway. Result: The miRNAs and lncRNAs specific to triple-negative breast cancer have been identified based on literature and database searches. Tumorigenesis, metastasis, and EMT were all given special attention. Apart from cross-talk being essential for TNBC tumorigenesis and treatment outcomes, our results indicated eight upregulated and seven downregulated miRNAs and 19 upregulated and three downregulated lncRNAs that can be used as predictive or diagnostic markers. This consolidated information could be useful in the clinic and provide a combined literature resource for TNBC researchers working on the Wnt/β-catenin miRNA/lncRNA axis. Conclusion: In conclusion, because the Wnt pathway and miRNAs/lncRNAs can modulate TNBC, their intertwinement results in a cascade of complex reactions that affect TNBC and related processes. Their function in TNBC pathogenesis has been highlighted in molecular processes underlying the disease progression.

Exosomal CTCF Confers Cisplatin Resistance in Osteosarcoma by Promoting Autophagy via the IGF2-AS/miR-579-3p/MSH6 Axis.

Cancer-derived exosomes participate in carcinogenesis and progression of cancers, including metastasis and drug-resistance. Of note, CTCF has been suggested to induce drug resistance in various cancers. Herein, we aim to investigate the role of cisplatin- (CDDP-) resistant osteosarcoma- (OS-) derived exosomal CTCF in OS cell resistance to CDDP and its mechanistic basis. Differentially expressed transcription factors, long noncoding RNAs (lncRNAs), miRNAs, and genes in OS were retrieved using bioinformatics approaches. Exosomes were extracted from CDDP-resistant OS cells and then cocultured with parental OS cells, followed by lentiviral transduction to manipulate the expression of CTCF, IGF2-AS, miR-579-3p, and MSH6. We assessed the in vitro and in vivo effects on malignant phenotypes, autophagy, CDDP sensitivity, and tumor formation of OS cells. It was established that CTCF and IGF2-AS were highly expressed in CDDP-resistant OS cells, and the CDDP-resistant OS cell-derived exosomal CTCF enhanced IGF2-AS transcription. CDDP-resistant OS-derived exosomes transmitted CTCF to OS cells and increased CDDP resistance in OS cells by activating an autophagy-dependent pathway. Mechanistically, CTCF activated IGF2-AS transcription and IGF2-AS competitively bound to miR-579-3p to upregulate MSH6 expression. Additionally, the promoting function of exosomal CTCF-mediated IGF2-AS/miR-579-3p/MSH6 in OS cell resistance to CDDP was confirmed in vivo. Taken together, CDDP-resistant OS-derived exosomal CTCF enhanced resistance of OS cells to CDDP via activating the autophagy-dependent pathway, providing a potential therapeutic consideration for OS treatment.

Assessing Nordihydroguaiaretic Acid Therapeutic Effect for Glioblastoma Multiforme.

In this study, we demonstrate that Raman microscopy combined with computational analysis is a useful approach to discriminating accurately between brain tumor bio-specimens and to identifying structural changes in glioblastoma (GBM) bio-signatures after nordihydroguaiaretic acid (NDGA) administration. NDGA phenolic lignan was selected as a potential therapeutic agent because of its reported beneficial effects in alleviating and inhibiting the formation of multi-organ malignant tumors. The current analysis of NDGA’s impact on GBM human cells demonstrates a reduction in the quantity of altered protein content and of reactive oxygen species (ROS)-damaged phenylalanine; results that correlate with the ROS scavenger and anti-oxidant properties of NDGA. A novel outcome presented here is the use of phenylalanine as a biomarker for differentiating between samples and assessing drug efficacy. Treatment with a low NDGA dose shows a decline in abnormal lipid-protein metabolism, which is inferred by the formation of lipid droplets and a decrease in altered protein content. A very high dose results in cell structural and membrane damage that favors transformed protein overexpression. The information gained through this work is of substantial value for understanding NDGA’s beneficial as well as detrimental bio-effects as a potential therapeutic drug for brain cancer.

Systemic Deficiency of PTEN Accelerates Breast Cancer Growth and Metastasis.

Mutation or loss of the tumor suppressor gene PTEN or its functional status in tumor stromal cells may affect tumor occurrence, development, invasion, and metastasis, in which, however, the role of overall low PTEN expression, mutation, or deletion in the tumor-bearing host has rarely been reported. Breast cancer is a common highly invasive metastatic tumor. We therefore treated mouse breast cancer 4T1 cells with the specific PTEN inhibitor VO-OHpic to study the effects of PTEN suppression or deletion on malignant behavior in vivo and in vitro. VO-OHpic effectively inhibited PTEN gene/protein expression in 4T1 cells, accelerated cell proliferation, and enhanced cell migration and invasion. We also transplanted 4T1 cells with VO-OHpic-inhibited PTEN into mice to create orthotopic and metastatic breast cancer models. The proliferation of 4T1 cells in mouse mammary gland was increased and distant metastasis was enhanced, with metastatic foci in the lung, liver, and intestinal tract. In addition, injection of mice with VO-OHpic to inhibit PTEN in the overall microenvironment accelerated the proliferation of transplanted 4T1 cells and enhanced distant metastasis and the formation of metastatic tumors. Metastatic foci formed in the lung, liver, intestine, thymus, and brain, and PTEN levels in the organ/tissues were negatively associated with the formation of metastatic foci. Similarly, inoculation of PTEN-deficient 4T1 cells into systemic PTEN-inhibited mice further enhanced the orthotopic growth and distant metastasis of 4T1 breast cancer. VO-OHpic inhibition of PTEN in 4T1 cells was also associated with significantly increased phosphorylation of Akt and phosphoinositide 3-kinase (PI3K), suggesting that inhibition of PTEN could activate the PI3K-Akt pathway, as a key signaling pathway regulating cell proliferation and death. These results confirmed that functional loss or deletion of the tumor suppressor gene PTEN significantly enhanced the proliferation, invasion, and metastasis of 4T1 cells. Systemic decrease or deletion of PTEN in the organism or organ/tissue microenvironment was conducive to the proliferation of breast cancer cells in situ and distant metastasis. These results suggest that, as well the PTEN in cancer cells the systemic microenvironment PTEN intensely mediates the proliferation, invasion and metastasis of mouse breast cancer cells via regulating the PI3K-Akt signaling pathway.