Wilms Tumor Cell Research Articles

ZSTK474 targeting PIK3R3 inhibits the Wilms' tumor through G0 / G1 phase arrest.

Wilms' tumor (WT), also known as nephroblastoma, is the predominant form of primary malignant renal cancer. The unfavorable prognoses linked to anaplastic nephroblastoma and recurrent nephroblastoma emphasize the crucial requirement for the exploration of innovative treatment modalities for WT. Our study conducted one-way Cox regression and Kaplan-Meier analyses using TARGET-WT nephroblastoma data to identify differentially expressed genes in nephroblastoma and evaluate their prognostic relevance. Utilizing the Connectivity Map database, ZSTK474 emerged as a viable therapeutic option for WT. The effect of ZSTK474 on WT and related underlying mechanisms were further investigated through in vitro and in vivo investigations. The in vivo experiment results indicated that ZSTK474 effectively inhibited subcutaneous tumor growth in WT mice. CCK-8 assays revealed two nephroblastoma cell lines exhibited half-inhibitory concentrations of 2μM and 2.51μM for ZSTK474, respectively. ZSTK474 was shown to inhibit the migration and invasion capabilities of WT cells in both Transwell and wound healing assays. Flow cytometry apoptosis and TUNEL assays demonstrated that ZSTK474 induced apoptosis in WT cells. Cell cycle analysis revealed that ZSTK474 led to the induction of G0/G1 phase arrest. Sequencing of ZSTK474-treated WiT49 cells suggested that the impact of ZSTK474 on WT might be mediated by the PI3K/Akt pathway, specifically by inhibiting PIK3R3. Knock-down of PIK3R3 confirmed that ZSTK474 downregulated PIK3R3, reducing Akt phosphorylation, cyclin D and CDK4 levels and elevating P21 expression in nephroblastoma cells. However, current research has limitations, including a lack of understanding of the long-term effects and potential resistance mechanisms of new therapies. This research provides insight into the potential of ZSTK474 and other PI3K inhibitors for treating nephroblastoma.

Retraction Note: Wilms tumor-suppressing peptide inhibits proliferation and induces apoptosis of Wilms tumor cells in vitro and in vivo

Retraction Note: Wilms tumor-suppressing peptide inhibits proliferation and induces apoptosis of Wilms tumor cells in vitro and in vivo

Spectrum of Renal Tumors Other Than Renal Cell Carcinoma with Emphasis on Cytomorphological, Immunohistochemical, and Cytogenetic Study

Abstract Background: The role of fine needle aspiration cytology (FNAC) in the diagnosis of renal malignancies is established and has been getting more precise and important over a period of time. Knowledge of the pathology of uncommon renal neoplasms along with radiological and clinical correlations often aids in correct diagnosis. Aims: The present study aims to describe the cytomorphological and immunohistochemical findings in the varied spectrum of renal tumors, other than renal cell carcinomas (RCC). Materials and Methods: Data of 238 cases of ultrasound-guided renal FNAC performed in our tertiary cancer institute over 4 years were collected from the department registry. All nondiagnostic cases and cases diagnosed as RCC were excluded from the study, so 57 cases of renal tumors were reviewed along with the cell blocks and ancillary studies. Results: Out of the 57 cases, 35 cases were primary renal neoplasms, which included renal oncocytoma (3.5%), angiomyolipoma (3.5%), rhabdoid tumor (1.8%), Wilms tumor (28.0%), Ewing sarcoma (3.5%), urothelial carcinoma (8.8%), and small round cell tumor unclassified (12.3%). Twenty two cases were metastatic tumors, which included hematolymphoid neoplasm (14%) and metastatic carcinomas (24.6%) from various other primary carcinomas. Conclusion: Our study shows that renal FNAC is safe and fairly accurate in diagnosing a wide spectrum of renal tumors and has high diagnostic accuracy, when performed along with cell block and immunohistochemistry. Awareness of the pathology of uncommon renal tumors along with relevant clinical history and radiological findings may aid in identifying the type of tumor for further appropriate management.

Modeling high-risk Wilms tumors enables the discovery of therapeutic vulnerability

Wilms tumor (WT) is the most common pediatric kidney cancer treated with standard chemotherapy. However, less-differentiated blastemal type of WT often relapses. To model the high-risk WT for therapeutic intervention, we introduce pluripotency factors into WiT49, a mixed-type WT cell line, to generate partially reprogrammed cells, namely WiT49-PRCs. When implanted into the kidney capsule in mice, WiT49-PRCs form kidney tumors and develop both liver and lung metastases, whereas WiT49 tumors do not metastasize. Histological characterization and gene expression signatures demonstrate that WiT49-PRCs recapitulate blastemal-predominant WTs. Moreover, drug screening in isogeneic WiT49 and WiT49-PRCs leads to the identification of epithelial- or blastemal-predominant WT-sensitive drugs, whose selectivity is validated in patient-derived xenografts (PDXs). Histone deacetylase (HDAC) inhibitors (e.g., panobinostat and romidepsin) are found universally effective across different WT and more potent than doxorubicin in PDXs. Taken together, WiT49-PRCs serve as a blastemal-predominant WT model for therapeutic intervention to treat patients with high-risk WT.

N6-methyladenosine modification of circMARK2 enhances cytoplasmic export and stabilizes LIN28B, contributing to the progression of Wilms tumor

BackgroundThe potential involvement of circular RNAs (circRNAs) and N6-methyladenosine (m6A) modification in the progression of Wilms tumor (WT) has not been fully elucidated. This study investigates the regulatory mechanisms and clinical significance of m6A-modified circMARK2 and its role in WT progression.MethodsWe identified dysregulated circRNAs through deep sequencing and validated their expression by qRT-PCR in WT tissues. The biological functions of circMARK2 were assessed using clone formation, transwell migration, and orthotopic animal models. To dissect the underlying mechanisms, we employed RNA immunoprecipitation, RNA pull-down, dual-luciferase reporter assays, Western blotting, and immunofluorescence and immunohistochemical staining.ResultsCircMARK2, upregulated in WT tissues, was found to be m6A-modified and promoted cytoplasmic export. It facilitated WT progression by stabilizing LIN28B mRNA through the circMARK2/IGF2BP2 interaction. In vitro and in vivo studies demonstrated that circMARK2 enhances the malignant behavior of WT cells. Clinically, higher circMARK2 levels in tumor tissues of WT patients were linked to increased tumor aggressiveness and reduced survival rates.ConclusionsOur study provides the first comprehensive evidence that m6A-modified circMARK2 contributes to WT progression by enhancing LIN28B mRNA stability, promoting cellular aggressiveness. CircMARK2 emerges as a potential biomarker for prognosis and a promising target for therapeutic intervention in WT, underscoring the clinical relevance of m6A modification in pediatric renal cancer.

Actinomycin D and bortezomib disrupt protein homeostasis in Wilms tumor.

Wilms tumor is the most common kidney cancer in children, and diffusely anaplastic Wilms tumor is the most chemoresistant histological subtype. Here we explore how Wilms tumor cells evade the common chemotherapeutic drug actinomycin D, which inhibits ribosomal RNA biogenesis. Using ribosome profiling, protein arrays, and a genome-wide knockout screen, we describe how actinomycin D disrupts protein homeostasis and blocks cell cycle progression. We found that, when ribosomal capacity is limited by actinomycin D treatment, anaplastic Wilms tumor cells preferentially translate proteasome components and upregulate proteasome activity. Furthermore, the proteasome inhibitor bortezomib sensitizes cells to actinomycin D treatment by inducing apoptosis both in vitro and in vivo. Lastly, we show that increased levels of proteasome components are associated with anaplastic histology and with worse prognosis in non-anaplastic Wilms tumor. In sum, maintaining protein homeostasis is critical for Wilms tumor proliferation, and it can be therapeutically disrupted by blocking protein synthesis or turnover.

DROSHA Regulates Mesenchymal Gene Expression in Wilms Tumor.

Wilms tumor, the most common pediatric kidney cancer, resembles embryonic renal progenitors. Currently, there are no ways to therapeutically target Wilms tumor driver mutations, such as in the microRNA processing gene DROSHA. In this study, we used a "multiomics" approach to define the effects of DROSHA mutation in Wilms tumor. We categorized Wilms tumor mutations into four mutational subclasses with unique transcriptional effects: microRNA processing, MYCN activation, chromatin remodeling, and kidney developmental factors. In particular, we find that DROSHA mutations are correlated with de-repressing microRNA target genes that regulate differentiation and proliferation and a self-renewing, mesenchymal state. We model these findings by inhibiting DROSHA expression in a Wilms tumor cell line, which led to upregulation of the cell cycle regulator cyclin D2 (CCND2). Furthermore, we observed that DROSHA mutations in Wilms tumor and DROSHA silencing in vitro were associated with a mesenchymal state with aberrations in redox metabolism. Accordingly, we demonstrate that Wilms tumor cells lacking microRNAs are sensitized to ferroptotic cell death through inhibition of glutathione peroxidase 4, the enzyme that detoxifies lipid peroxides. Implications: This study reveals genotype-transcriptome relationships in Wilms tumor and points to ferroptosis as a potentially therapeutic vulnerability in one subset of Wilms tumor.

A Multidisciplinary Hyper-Modeling Scheme in Personalized In Silico Oncology: Coupling Cell Kinetics with Metabolism, Signaling Networks, and Biomechanics as Plug-In Component Models of a Cancer Digital Twin.

The massive amount of human biological, imaging, and clinical data produced by multiple and diverse sources necessitates integrative modeling approaches able to summarize all this information into answers to specific clinical questions. In this paper, we present a hypermodeling scheme able to combine models of diverse cancer aspects regardless of their underlying method or scale. Describing tissue-scale cancer cell proliferation, biomechanical tumor growth, nutrient transport, genomic-scale aberrant cancer cell metabolism, and cell-signaling pathways that regulate the cellular response to therapy, the hypermodel integrates mutation, miRNA expression, imaging, and clinical data. The constituting hypomodels, as well as their orchestration and links, are described. Two specific cancer types, Wilms tumor (nephroblastoma) and non-small cell lung cancer, are addressed as proof-of-concept study cases. Personalized simulations of the actual anatomy of a patient have been conducted. The hypermodel has also been applied to predict tumor control after radiotherapy and the relationship between tumor proliferative activity and response to neoadjuvant chemotherapy. Our innovative hypermodel holds promise as a digital twin-based clinical decision support system and as the core of future in silico trial platforms, although additional retrospective adaptation and validation are necessary.

Identification of key genes in Wilms tumor based on high-throughput RNA sequencing and their impacts on prognosis and immune responses

To identify the key genes differentially expressed in Wilms tumor and analyze their potential impacts on prognosis and immune responses of the patients. High-throughput RNA sequencing was used to identify the differentially expressed mRNAs in clinical samples of Wilms tumor and paired normal tissues, and their biological functions were analyzed using GO, KEGG and GSEA enrichment analyses. The hub genes were identified using STRING database, based on which a prognostic model was constructed using LASSO regression. The mutations of the key hub genes were analyzed and their impacts on immunotherapy efficacy was predicted using the cBioPortal platform. RT-qPCR was used to verify the differential expressions of the key hub genes in Wilms tumor. Of the 1612 differentially expressed genes identified in Wilms tumor, 1030 were up-regulated and 582 were down-regulated, involving mainly cell cycle processes and immune responses. Ten hub genes were identified, among which 4 genes (TP53, MED1, CCNB1 and EGF) were closely related to the survival of children with Wilms tumor. A 3-gene prognostic signature was constructed through LASSO regression analysis, and the patients stratified into with high- and low-risk groups based on this signature had significantly different survival outcomes (HR=1.814, log-rank P=0.002). The AUCs of the 3-, 5- and 7-year survival ROC curves of this model were all greater than 0.7. The overall mutations in the key hub genes or the individual mutations in TP53/CCNB1 were strongly correlated with a lower survival rates, and a high TP53 expression was correlated with a poor immunotherapy efficacy. RT-qPCR confirmed that the key hub genes had significant differential expressions in Wilms tumor tissues and cells. TP53 gene plays an important role in the Wilms tumor and may potentially serve as a new immunotherapeutic biomarker as well as a therapeutic target.

Targeting TRIP13 in favorable histology Wilms tumor with nuclear export inhibitors synergizes with doxorubicin

Wilms tumor (WT) is the most common renal malignancy of childhood. Despite improvements in the overall survival, relapse occurs in ~15% of patients with favorable histology WT (FHWT). Half of these patients will succumb to their disease. Identifying novel targeted therapies remains challenging in part due to the lack of faithful preclinical in vitro models. Here we establish twelve patient-derived WT cell lines and demonstrate that these models faithfully recapitulate WT biology using genomic and transcriptomic techniques. We then perform loss-of-function screens to identify the nuclear export gene, XPO1, as a vulnerability. We find that the FDA approved XPO1 inhibitor, KPT-330, suppresses TRIP13 expression, which is required for survival. We further identify synergy between KPT-330 and doxorubicin, a chemotherapy used in high-risk FHWT. Taken together, we identify XPO1 inhibition with KPT-330 as a potential therapeutic option to treat FHWTs and in combination with doxorubicin, leads to durable remissions in vivo.

Abstract 1084: Preclinical testing of a cell adhesion molecule 1 (CADM1) targeting antibody-drug conjugate in osteosarcoma and other pediatric solid tumor models

Abstract Introduction: Survival outcomes of osteosarcoma have remained stagnant for decades, emphasizing the urgent need to identify new cell surface targets and develop novel therapies. Antibody-drug conjugates (ADCs), known for their targeted delivery of cytotoxic agents to specific targets, present an attractive therapeutic class for potential use in osteosarcoma. Methods: We previously identified Cell Adhesion Molecule 1 (CADM1) is highly expressed on the cell surface in osteosarcoma using an integrated proteomic and transcriptomic surfaceome profiling approach. As a proof of concept, a humanized CADM1 antibody (clone PTA021-A1) was conjugated with Tesirine (SG3249), a pyrrolobenzodiazepine (PBD) dimer payload. The antitumor activity of this CADM1 ADC was validated in vivo in 8 osteosarcoma patient-derived xenografts (PDXs). CADM1 is also highly expressed in other pediatric solid malignancies such as neuroblastoma and Wilms tumor. The CADM1 ADC was tested in vitro in neuroblastoma and Wilms tumor models. Negative controls included a CD19 targeting ADC with the same payloads and CADM1 negative Ewing sarcoma models. Results: The expression of the CADM1 was validated by flow cytometry in osteosarcoma, neuroblastoma, and Wilms tumor cell lines, and by immunohistochemistry (IHC) in osteosarcoma patient samples and PDX models. In vivo testing of CADM1 ADC in 8 osteosarcoma PDX models showed that the ADC was well-tolerated and significantly prolonged progressive-free survival in all osteosarcoma models. Complete response (CR) and maintained CR was observed in 3 models. In vitro testing for neuroblastoma and Wilms tumor is ongoing, and preliminary data suggest promising antitumor activities. Conclusions: CADM1 is highly expressed in osteosarcoma and other pediatric solid tumors. CADM1 ADC demonstrated promising antitumor in vitro and in vivo activity in osteosarcoma, and initial in vitro studies in neuroblastoma and Wilms tumor show similarly promising responses. These data support the need for further studies assessing the potential of a CADM1 ADC as a novel agent for the treatment of pediatric solid cancers. Citation Format: Zhongting Zhang, Yifei Wang, Wendong Zhang, Xiangjun Tian, Qi Wang, Rossana N Lazcano Segura, Michael Roth, Jonathan Gill, Douglas Harrison, Zhaohui Xu, Yianhua Yi, Sylvester Jusu, Giuseppe Longo, Xin Zhou, Jing Wang, Richard Gorlick. Preclinical testing of a cell adhesion molecule 1 (CADM1) targeting antibody-drug conjugate in osteosarcoma and other pediatric solid tumor models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1084.

Overexpression of aquaporin-1 plays a vital role in proliferation, apoptosis, and pyroptosis of Wilms’ tumor cells

BackgroundNephroblastoma, also known as Wilms’ tumor (WT), is an embryonic malignant tumor and one of the most common malignant tumors in the abdominal region of children. The exact role and underlying mechanisms of aquaporin-1 (AQP1) in the occurrence and development of nephroblastoma remain unclear.MethodsAfter overexpression of AQP1, cell proliferation was assessed using the CCK-8 proliferation assay and EdU staining. Flow cytometry was employed to assess cell apoptosis, and Western blotting (WB) analysis was conducted to validate the expression of relevant protein markers. mRNA sequencing (mRNA-Seq) was performed on WT cells overexpressing AQP1 to predict and characterize the associated mechanisms. Transmission electron microscopy was utilized to observe changes in the ultrastructure of WT cells undergoing apoptosis and pyroptosis following AQP1 overexpression. Functional in vivo validation was conducted through animal experiments.ResultsWe validated that overexpression of AQP1 inhibited cell proliferation and promoted cell apoptosis and pyroptosis both in vitro and in vivo. mRNA-Seq analysis of WT cells with AQP1 overexpression suggested that these effects might be mediated through the inhibition of the JAK-STAT signaling pathway. Additionally, we discovered that overexpression of AQP1 activated the classical pyroptosis signaling pathway dependent on caspase-1, thereby promoting pyroptosis in WT.ConclusionThese findings highlight the important functional role of AQP1 in the pathobiology of nephroblastoma, providing novel insights into the development of this disease. Moreover, these results offer new perspectives on the potential therapeutic targeting of AQP1 as a treatment strategy for nephroblastoma.

Abstract B001: Elucidating metastatic pathways in pediatric cancers

Abstract The most common cause of death in pediatric cancer patients is a treatment resistant tumor, often further complicated by metastatic spread to multiple sites, making both surgery and radiation therapy unfeasible as curative treatment options. How metastatic spread occurs in pediatric tumors is to a large extent completely unexplored. We aimed to elucidate the evolutionary trajectories of metastatic disease in the pediatric tumors, neuroblastoma, Wilms tumor and germ cell tumors, which are the most common tumors in children besides tumors of the central nervous system and leukemias. More specifically we wanted to decipher when in the evolutionary history of the tumor, metastases appear, whether there were signs of monoclonal or polyclonal spread (one or several subclones from the primary tumor colonizing the same metastatic site), mono- or polyphyletic spread (one subclone or several subclones colonizing several metastatic sites) and intermetastatic spread (subclones from one metastatic site spreading to another). Due to the spatiotemporal genetic heterogeneity of tumors, DNA was extracted from multiple sites from the primary tumor and several metastatic sites for each patient. A total of 174 samples from 18 patients were included. All samples underwent whole genome copy number profiling and a subset also targeted deep sequencing. Phylogenetic trees were reconstructed, allowing spatiotemporal tracking of subclones for each patient. Having a median of 8 (range 4 to 22) samples available per patient we could confirm an extensive genetic heterogeneity within and between the primary tumor and metastases in each patient, stressing the importance of multiregional sampling to fully characterize the disease. In 13 of 18 patients, early metastatic spread was identified and a total of 6 of 18 patients displayed both polyphyletic and polyclonal seeding. Hence metastatic spread can occur several times during tumor evolution, several subclones possess metastatic capability and several different subclones may colonize the same site. We also found that a higher number of subclones from the primary tumor metastasize to the lymph nodes compared to distant sites, suggesting a weaker evolutionary bottleneck for locoregional lymphatic spread. Finally, intermetastatic spread could be identified in 7 out of 11 patients having metastases in at least 2 different distant sites. Our results provide further knowledge of the evolutionary dynamics of metastatic spread in pediatric cancer patients. Metastases often have a different genomic profile than the primary tumor, stressing the need to sample metastases for assessment of therapeutic targets, since metastases may completely lack targets present in the primary, resulting in progression at distant sites despite the primary tumor responding to treatment. We also found that metastatic spread can occur early and several times during tumor evolution. Strikingly, intermetastatic spread is identified as a common feature in pediatric tumors, indicating that metastases alone may act as a hub for further metastatic spread. Citation Format: Natalie Andersson, Michele Ferro, Caroline Jansson, Jenny Karlsson, Linda Holmquist Mengelbier, Subhayan Chattopadhyay, David Gisselsson. Elucidating metastatic pathways in pediatric cancers [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Translating Cancer Evolution and Data Science: The Next Frontier; 2023 Dec 3-6; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(3 Suppl_2):Abstract nr B001.

CT radiomics to differentiate between Wilms tumor and clear cell sarcoma of the kidney in children

BackgroundTo investigate the role of CT radiomics in distinguishing Wilms tumor (WT) from clear cell sarcoma of the kidney (CCSK) in pediatric patients.MethodsWe retrospectively enrolled 83 cases of WT and 33 cases of CCSK. These cases were randomly stratified into a training set (n = 81) and a test set (n = 35). Several imaging features from the nephrographic phase were analyzed, including the maximum tumor diameter, the ratio of the maximum CT value of the tumor solid portion to the mean CT value of the contralateral renal vein (CTmax/CT renal vein), and the presence of dilated peritumoral cysts. Radiomics features from corticomedullary phase were extracted, selected, and subsequently integrated into a logistic regression model. We evaluated the model's performance using the area under the curve (AUC), 95% confidence interval (CI), and accuracy.ResultsIn the training set, there were statistically significant differences in the maximum tumor diameter (P = 0.021) and the presence of dilated peritumoral cysts (P = 0.005) between WT and CCSK, whereas in the test set, no statistically significant differences were observed (P > 0.05). The radiomics model, constructed using four radiomics features, demonstrated strong performance in the training set with an AUC of 0.889 (95% CI: 0.811–0.967) and an accuracy of 0.864. Upon evaluation using fivefold cross-validation in the training set, the AUC remained high at 0.863 (95% CI: 0.774–0.952), with an accuracy of 0.852. In the test set, the radiomics model achieved an AUC of 0.792 (95% CI: 0.616–0.968) and an accuracy of 0.857.ConclusionCT radiomics proves to be diagnostically valuable for distinguishing between WT and CCSK in pediatric cases.

Delineating the interplay between oncogenic pathways and immunity in anaplastic Wilms tumors

Wilms tumors are highly curable in up to 90% of cases with a combination of surgery and radio-chemotherapy, but treatment-resistant types such as diffuse anaplastic Wilms tumors pose significant therapeutic challenges. Our multi-omics profiling unveils a distinct desert-like diffuse anaplastic Wilms tumor subtype marked by immune/stromal cell depletion, TP53 alterations, and cGAS-STING pathway downregulation, accounting for one-third of all diffuse anaplastic cases. This subtype, also characterized by reduced CD8 and CD3 infiltration and active oncogenic pathways involving histone deacetylase and DNA repair, correlates with poor clinical outcomes. These oncogenic pathways are found to be conserved in anaplastic Wilms tumor cell models. We identify histone deacetylase and/or WEE1 inhibitors as potential therapeutic vulnerabilities in these tumors, which might also restore tumor immunogenicity and potentially enhance the effects of immunotherapy. These insights offer a foundation for predicting outcomes and personalizing treatment strategies for aggressive pediatric Wilms tumors, tailored to individual immunological landscapes.

Oncogenic Cells of Renal Embryonic Lineage Sensitive to the Small-Molecule Inhibitor QC6352 Display Depletion of KDM4 Levels and Disruption of Ribosome Biogenesis.

The histone lysine demethylases KDM4A-C are involved in physiologic processes including stem cell identity and self-renewal during development, DNA damage repair, and cell-cycle progression. KDM4A-C are overexpressed and associated with malignant cell behavior in multiple human cancers and are therefore potential therapeutic targets. Given the role of KDM4A-C in development and cancer, we aimed to test the potent, selective KDM4A-C inhibitor QC6352 on oncogenic cells of renal embryonic lineage. The anaplastic Wilms tumor cell line WiT49 and the tumor-forming human embryonic kidney cell line HEK293 demonstrated low nanomolar QC6352 sensitivity. The cytostatic response to QC6352 in WiT49 and HEK293 cells was marked by induction of DNA damage, a DNA repair-associated protein checkpoint response, S-phase cell-cycle arrest, profound reduction of ribosomal protein gene and rRNA transcription, and blockade of newly synthesized proteins. QC6352 caused reduction of KDM4A-C levels by a proteasome-associated mechanism. The cellular phenotype caused by QC6352 treatment of reduced migration, proliferation, tumor spheroid growth, DNA damage, and S-phase cell-cycle arrest was most closely mirrored by knockdown of KDM4A as determined by siRNA knockdown of KDM4A-C. QC6352 sensitivity correlated with high basal levels of ribosomal gene transcription in more than 900 human cancer cell lines. Targeting KDM4A may be of future therapeutic interest in oncogenic cells of embryonic renal lineage or cells with high basal expression of ribosomal protein genes.

A Gradual Transition Toward Anaplasia in Wilms Tumor Through Tolerance to Genetic Damage

Patients with Wilms tumor (WT) in general have excellent survival, but the prognosis of patients belonging to the subgroup of WT with diffuse anaplasia (DA) is poor due to frequent resistance to chemotherapy. We hypothesized that DA WT cells might undergo changes, such as acquiring a persistent tolerance to DNA damage and copy number aberrations (CNAs), which could eventually lead to their resistance to chemotherapy treatment. Tissue sections from chemotherapy-treated DA WTs (n = 12) were compared with chemotherapy-treated nonanaplastic WTs (n = 15) in a tissue microarray system, enabling analysis of 769 tumor regions. All regions were scored for anaplastic features and immunohistochemistry was used to quantify p53 expression, proliferation index (Ki67), and DNA double-strand breaks (γH2AX). CNAs were assessed by array-based genotyping and TP53 mutations using targeted sequencing. Proliferation index and the frequency of DNA double-strand breaks (γH2AX dot expression) increased with higher anaplasia scores. Almost all (95.6%) areas with full-scale anaplasia had TP53 mutations or loss of heterozygosity, along with an increased amount of CNAs. Interestingly, areas with wild-type TP53 with loss of heterozygosity and only one feature of anaplasia (anaplasia score 1) also had significantly higher proliferation indices, more DNA double-strand breaks, and more CNAs than regions without any anaplastic features (score 0); such areas may be preanaplastic cell populations under selective pressure for TP53 mutations. In conclusion, we suggest that chemoresistance of DA WTs may be partly explained by a high proliferative capability of anaplastic cells, which also have a high burden of double-stranded DNA breaks and CNAs, and that there is a gradual emergence of anaplasia in WT.

Identification of key genes and validation of key gene aquaporin 1 on Wilms' tumor metastasis.

Wilms' tumor (WT) is one of the most common solid tumors in children with unsatisfactory prognosis, but few molecular prognostic markers have been discovered for it. Many genes are associated with the occurrence and prognosis of WT. This study aimed to explore the key genes and potential molecular mechanisms through bioinformatics and to verify the effects of aquaporin 1 (AQP1) on WT metastasis. Differentially expressed genes (DEGs) were generated from WT gene expression data sets from the Gene Expression Omnibus (GEO) database. Gene functional enrichment analysis was carried out with the Database for Annotation, Visualization and Integrated Discovery (DAVID). A protein-protein interaction network (PPI) was constructed and visualized by the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database and Cytoscape software. Minimal Common Oncology Data Elements (MCODE) was used to detect the important modules in the PPI network, and the important nodes (genes) in the PPI module were sorted by CytoHubba. RT-qPCR was performed to validate the expression of the key genes in WT. Wound healing and Transwell assays were used to detect the cell migration and invasion abilities of AQP1-overexpressing cells. Phalloidin-iFlour 488 was used to stain the cytoskeleton to observe how AQP1 overexpression affects cytoskeletal microfilament structure. A total of 73 co-expressed DEGs were chosen for further investigation. The importance of homeostasis and transmembrane transport of ions and water were highlighted by functional analysis. Gene regulatory network and PPI network were predicted. MCODE plug identified two important modules. Finally, top five key genes were identified using CytoHubba, including Renin (REN), nephrosis 2 (NPHS2), Solute Carrier Family 12 Member 3 (SLC12A3), Solute Carrier Family 12 Member 1 (SLC12A1) and AQP1. The five key genes were mainly enriched in cell volume and ion homeostasis. RT-qPCR confirmed the expression of the five key genes in WT. AQP1 was validated to be expressed at significantly lower levels in WT than in normal tissue. AQP1 overexpression significantly reduced the migratory and invasive capacity of Wit-49 cells, as evidenced by reducing the scratch healing rate and the number of perforated control cells by Wit-49 cells. AQP1 overexpression also reduced the expression of biomarkers of epithelial-mesenchymal transformation, decreased levels of vimentin and N-cadherin and increased expression of E-cadherin, resulting in decreased formation of conspicuous lamellipodial protrusions, characteristic of diminished WT cell invasion and migration. Our study reveals the key genes of WT. These key genes may provide novel insight for the mechanism and diagnosis of WT. AQP1 overexpression inhibited invasion, migration, EMT, and cytoskeletal rearrangement of WT cells, indicating that AQP1 plays a role in the pathogenesis of WT.

Nuclear medicine techniques in paediatric body oncology: Present and future

Modern nuclear medicine techniques combining anatomical and functional imaging have transformed our ability to diagnose, stage and prognosticate in paediatric oncology. Progress in the use of positron emission tomography (PET) in children with cancer has been relatively restrained compared to adults due to radiation concerns, limited research, availability, and the practical challenges associated with imaging young children. However, there have been several developments in the last decade and the role of molecular imaging in paediatric oncology will only broaden with forthcoming novel tracers and molecular radiotherapy. This review article summarises the practical aspects of nuclear imaging in paediatric oncology, discusses the main imaging methods including PET and single photon emission computed tomography (SPECT), appraises the available established and novel tracers, and summarises the evidence behind molecular imaging in paediatric lymphoma, neuroblastoma, sarcomas, thyroid cancer and other tumours. 18F-FDG PET/CT is the established method for staging and response assessment after induction chemotherapy in paediatric Hodgkin lymphoma and has high sensitivity for the detection of bone marrow disease, reducing the need for bone marrow biopsy in some cases. 123I-mIBG planar with SPECT imaging remains the mainstay for neuroblastoma imaging, however there is a well-established role for 18F-FDG PET/CT in mIBG non-avid disease, and 18F-DOPA and 68Ga-DOTA PET tracers have shown promising results as upcoming alternatives. 18F-FDG PET/CT is advocated in the staging assessment of rhabdomyosarcoma and has been shown to have excellent sensitivity in the detection of bone metastases in osteosarcoma and Ewing sarcoma, although standard chest CT is still recommended for the detection of lung metastases. The role of PET in germ cell tumours, hepatoblastoma, Wilms tumour and Langerhans cell histiocytosis is complementary to standard techniques.

Cytotoxicity of WT1-reactive T cells against Wilms tumor: An implication for antigen-specific adoptive immunotherapy.

T cells that recognize WT1 peptides have been shown to efficiently eliminate WT1-expressing tumor cells. This study was designed to investigate the feasibility of isolating WT1-reactive T cells from peripheral blood mononuclear cells (PBMCs) from healthy donors and patients with Wilms tumor, and to assess the cytotoxicity mediated by these cells against Wilms tumor cells (WiTu cells). WT1-reactive T cells were enriched and isolated by stimulating PBMCs with a WT1 peptide pool and interferon-γ capture-based immunomagnetic separation (IMS). Using the lactate dehydrogenase release assay, the in vitro cytotoxicity of the isolated cells and standard chemotherapy was evaluated on WiTu cells. Higher proportions of WT1-reactive T cells were isolated from patients with Wilms tumor compared to those isolated from HDs. WT1-reactive T cells produced > 50% specific lysis when co-cultured with WT1+ WiTu cells at the highest effector-to-target (E:T) ratio in this study (i.e., 5:1), compared to <23% when co-cultured with WT1- WiTu cells at the same ratio. WT1-reactive T cells showed anti-tumoral activity in a dose-dependent manner and mediated significantly greater cytotoxicity than the non-WT1-reactive fraction of PBMCs on WT1+ WiTu cells. The cytotoxicity of standard chemotherapy was significantly lower than that of WT1-reactive T cells when co-cultured with WT1+ WiTu cells at E:T ratios of 2:1 and 5:1. WT1-reactive T cells can be effectively enriched from the PBMCs of patients with Wilms tumor. Ex vivo generated WT1-reactive T cells might be considered an adoptive immunotherapeutic option for WT1+ Wilms tumors.