Multiple Tumor Suppressor Genes Research Articles

Abstract B018: miRNA-130a promotes Breast cancer progression by inhibiting multiple tumour suppressor genes as a Gene-TF-miRNA network

Abstract Cancer is a disease that has implications worldwide, including in developing countries. It is known for its notorious ways of adapting and gaining resistance to the therapies available. Breast cancer (BC) majorly affects women and leading cause of death in women worldwide. Despite studying BC for many decades, we are not yet certain about a definitive therapy that can cure it with confidence. One of the challenges associated with treating BC is its metastasis and resistance to available therapies. Non-cancerous cells exhibit a tightly regulated transcription profile. An altered transcription profile is considered one of the main causes of the origin of cancers and their progression, including resistance and metastasis. Tumor suppressor genes are associated with mitigating any oncogenic signals induced by external factors or internal dysregulation in oncogenic genes. Changes in the expression profiles of tumour suppressor and oncogenic genes are associated with BC progression and metastasis. Transcription factors (TFs) play a significant role in marinating the tight regulation of transcription. In this study, we have analyzed Genes(N=1057), TFs(N =233) and miRNAs (N = 246) that are promoting BC from literature and publicly available databases; we have analyzed the regulation of Gene-Gene, miRNA-miRNA, TF-TF, miRNA-Gene, miRNA-TF, TF-Gene, TF-miRNA as a network of three node Feed Forward Loops (FFLs). Upon constructing 3, 4,5 and 6 node FFLs, we have identified that the miRNA130a is promoting BC by inhibiting crucial Tumor suppressor genes like TGFBR1, TGFBR2, SOD2, PTEN, SMAD4, RUNX3 etc. Citation Format: Prasanna Kumar Reddy Gayam, Jesil Mathew A, Fayaz SM. miRNA-130a promotes Breast cancer progression by inhibiting multiple tumour suppressor genes as a Gene-TF-miRNA network [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: RNAs as Drivers, Targets, and Therapeutics in Cancer; 2024 Nov 14-17; Bellevue, Washington. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(11_Suppl):Abstract nr B018.

Suppressive cancer nonstop extension mutations increase C-terminal hydrophobicity and disrupt evolutionarily conserved amino acid patterns

Nonstop extension mutations, a.k.a. stop-lost or stop-loss mutations, convert a stop codon into a sense codon resulting in translation into the 3’ untranslated region until the next in-frame stop codon, thereby extending the C-terminus of a protein. In cancer, only nonstop mutations in SMAD4 have been functionally characterized, while the impact of other nonstop mutations remain unknown. Here, we exploit our pan-cancer NonStopDB dataset and test all 2335 C-terminal extensions arising from somatic nonstop mutations in cancer for their impact on protein expression. In a high-throughput screen, 56.1% of the extensions effectively reduce protein abundance. Extensions of multiple tumor suppressor genes like PTEN, APC, B2M, CASP8, CDKN1B and MLH1 are effective and validated for their suppressive impact. Importantly, the effective extensions possess a higher hydrophobicity than the neutral extensions linking C-terminal hydrophobicity with protein destabilization. Analyzing the proteomes of eleven different species reveals conserved patterns of amino acid distribution in the C-terminal regions of all proteins compared to the proteomes like an enrichment of lysine and arginine and a depletion of glycine, leucine, valine and isoleucine across species and kingdoms. These evolutionary selection patterns are disrupted in the cancer-derived effective nonstop extensions.

Functional mapping of epigenetic regulators uncovers coordinated tumor suppression by the HBO1 and MLL1 complexes.

Epigenetic dysregulation is widespread in cancer. However, the specific epigenetic regulators and the processes they control to drive cancer phenotypes are poorly understood. Here, we employed a novel, scalable and high-throughput in vivo method to perform iterative functional screens of over 250 epigenetic regulatory genes within autochthonous oncogenic KRAS-driven lung tumors. We identified multiple novel epigenetic tumor suppressor and tumor dependency genes. We show that a specific HBO1 complex and the MLL1 complex are among the most impactful tumor suppressive epigenetic regulators in lung. The histone modifications generated by the HBO1 complex are frequently absent or reduced in human lung adenocarcinomas. The HBO1 and MLL1 complexes regulate chromatin accessibility of shared genomic regions, lineage fidelity and the expression of canonical tumor suppressor genes. The HBO1 and MLL1 complexes are epistatic during lung tumorigenesis, and their functional correlation is conserved in human cancer cell lines. Together, these results demonstrate the value of quantitative methods to generate a phenotypic roadmap of epigenetic regulatory genes in tumorigenesis in vivo .

Adenosine-to-Inosine RNA editing in cancer: molecular mechanisms and downstream targets.

Adenosine-to-Inosine (A-to-I), one of the most prevalent RNA modifications, has recently garnered significant attention. The A-to-I modification actively contributes to biological and pathological processes by affecting the structure and function of various RNA molecules, including double stranded RNA, transfer RNA, microRNA, and viral RNA. Increasing evidence suggests that A-to-I plays a crucial role in the development of human disease, particularly in cancer, and aberrant A-to-I levels are closely associated with tumorigenesis and progression through regulation of the expression of multiple oncogenes and tumor suppressor genes. Currently, the underlying molecular mechanisms of A-to-I modification in cancer are not comprehensively understood. Here, we review the latest advances regarding the A-to-I editing pathways implicated in cancer, describing their biological functions and their connections to the disease.

Activation of AMPK inhibits cervical cancer growth by hyperacetylation of H3K9 through PCAF

BackgroundDysregulation in histone acetylation, a significant epigenetic alteration closely associated with major pathologies including cancer, promotes tumorigenesis, inactivating tumor-suppressor genes and activating oncogenic pathways. AMP-activated protein kinase (AMPK) is a cellular energy sensor that regulates a multitude of biological processes. Although a number of studies have identified the mechanisms by which AMPK regulates cancer growth, the underlying epigenetic mechanisms remain unknown.MethodsThe impact of metformin, an AMPK activator, on cervical cancer was evaluated through assessments of cell viability, tumor xenograft model, pan-acetylation analysis, and the role of the AMPK-PCAF-H3K9ac signaling pathway. Using label-free quantitative acetylproteomics and chromatin immunoprecipitation-sequencing (ChIP) technology, the activation of AMPK-induced H3K9 acetylation was further investigated.ResultsIn this study, we found that metformin, acting as an AMPK agonist, activates AMPK, thereby inhibiting the proliferation of cervical cancer both in vitro and in vivo. Mechanistically, AMPK activation induces H3K9 acetylation at epigenetic level, leading to chromatin remodeling in cervical cancer. This also enhances the binding of H3K9ac to the promoter regions of multiple tumor suppressor genes, thereby promoting their transcriptional activation. Furthermore, the absence of PCAF renders AMPK activation incapable of inducing H3K9 acetylation.ConclusionsIn conclusion, our findings demonstrate that AMPK mediates the inhibition of cervical cancer growth through PCAF-dependent H3K9 acetylation. This discovery not only facilitates the clinical application of metformin but also underscores the essential role of PCAF in AMPK activation-induced H3K9 hyperacetylation.

EZH2 G553C significantly increases the risk of brain metastasis from lung cancer due to salt bridge instability

BackgroundThe incidence and mortality of lung cancer is the highest in China and the world. Brain is the most common distant metastasis site of lung cancer. Its transfer mechanism and predictive biomarkers are still unclear. EZH2 participates in the catalysis of transcriptional inhibition complex, mediates chromatin compactness, leads to the silencing of its downstream target genes, participates in the silencing of multiple tumor suppressor genes, and is related to cell proliferation, apoptosis and cycle regulation. In physiology, EZH2 has high activity in stem cells or progenitor cells, inhibits genes related to cell cycle arrest and promotes self-renewal. To detect the expression and mutation of EZH2 gene in patients with brain metastasis of lung cancer, and provide further theoretical basis for exploring the pathogenesis of brain metastasis of lung cancer and finding reliable biomarkers to predict brain metastasis of lung cancer.MethodsThis study investigated susceptible genes for brain metastasis of lung cancer. The second-generation sequencing technology was applied to screen the differential genes of paired samples (brain metastasis tissues, lung cancer tissues and adjacent tissues) of lung cancer patients with brain metastasi.ResultsIt revealed that there was a significant difference in the G553C genotype of EZH2 between lung cancer brain metastasis tissues and lung cancer tissues (p = 0.045). The risk of lung cancer brain metastasis in G allele carriers was 2.124 times higher than that in C allele carriers. Immunohistochemistry showed that compared with lung cancer patients and lung cancer patients with brain metastasis, the expression level of EZH2 in lung cancer tissues of lung cancer patients was significantly higher than that in adjacent lung tissues (p < 0.0001), and higher than that in brain metastasis tissues (p = 0.0309). RNA in situ immunohybridization showed that EZH2 mRNA expression was gradually high in lung cancer adjacent tissues, lung cancer tissues and lung cancer brain metastasis tissues.ConclusionsEZH2 G553C polymorphism contributes to the prediction of brain metastasis of lung cancer, in which G allele carriers are more prone to brain metastasis.

High clonal diversity and spatial genetic admixture in early prostate cancer and surrounding normal tissue

Somatic copy number alterations (SCNAs) are pervasive in advanced human cancers, but their prevalence and spatial distribution in early-stage, localized tumors and their surrounding normal tissues are poorly characterized. Here, we perform multi-region, single-cell DNA sequencing to characterize the SCNA landscape across tumor-rich and normal tissue in two male patients with localized prostate cancer. We identify two distinct karyotypes: ‘pseudo-diploid’ cells harboring few SCNAs and highly aneuploid cells. Pseudo-diploid cells form numerous small-sized subclones ranging from highly spatially localized to broadly spread subclones. In contrast, aneuploid cells do not form subclones and are detected throughout the prostate, including normal tissue regions. Highly localized pseudo-diploid subclones are confined within tumor-rich regions and carry deletions in multiple tumor-suppressor genes. Our study reveals that SCNAs are widespread in normal and tumor regions across the prostate in localized prostate cancer patients and suggests that a subset of pseudo-diploid cells drive tumorigenesis in the aging prostate.

Severe induction of aberrant DNA methylation by nodular gastritis in adults.

Nodular gastritis (NG) is characterized by marked antral lymphoid follicle formation, and is a strong risk factor for diffuse-type gastric cancer in adults. However, it is unknown whether aberrant DNA methylation, which is induced by atrophic gastritis (AG) and is a risk for gastric cancer, is induced by NG. Here, we analyzed methylation induction by NG. Gastric mucosal samples were obtained from non-cancerous antral tissues of 16 NG and 20 AG patients with gastric cancer and 5 NG and 6 AG patients without, all age- and gender-matched. Genome-wide methylation analysis and expression analysis were conducted by a BeadChip array and RNA-sequencing, respectively. Clustering analysis of non-cancerous antral tissues of NG and AG patients with gastric cancer was conducted using methylation levels of 585 promoter CpG islands (CGIs) of methylation-resistant genes, and a large fraction of NG samples formed a cluster with strong methylation induction. Promoter CGIs of CDH1 and DAPK1 tumor-suppressor genes were more methylated in NG than in AG. Notably, methylation levels of these genes were also higher in the antrum of NG patients without cancer. Genes related to lymphoid follicle formation, such as CXCL13/CXCR5 and CXCL12/CXCR4, had higher expression in NG, and genes involved in DNA demethylation TET2 and IDH1, had only half the expression in NG. Severe aberrant methylation, involving multiple tumor-suppressor genes, was induced in the gastric antrum and body of patients with NG, in accordance with their high gastric cancer risk.

Spheroid architecture strongly enhances miR-221/222 expression and promotes oxidative phosphorylation in an ovarian cancer cell line through a mechanism that includes restriction of miR-9 expression.

Tumor cell spheroids are organized multicellular structures that form during the expansive growth of carcinoma cells. Spheroids formation is thought to contribute to metastasis by supporting growth and survival of mobile tumor cell populations. We investigated how spheroid architecture affects OXPHOS activity, microRNA expression, and intraperitoneal survival of an ovarian carcinoma cell line using high resolution respirometry, quantitative RT-PCR, and a rodent intraperitoneal growth model. Rates of oxidative phosphorylation/respiration per cell of cells growing as spheroids were nearly double those of a variant of the same cell type growing in suspension as loosely aggregated cells. Further, inhibition of spheroid formation by treatment with CDH2 (N-cadherin) siRNA reduced the rate of OXPHOS to that of the non-spheroid forming variant. Cells growing as spheroids showed greatly enhanced expression of miR-221/222, an oncomiR that targets multiple tumor suppressor genes and promotes invasion, and reduced expression of miR-9, which targets mitochondrial tRNA-modification enzymes and inhibits OXPHOS. Consistent with greater efficiency of ATP generation, tumor cells growing as spheroids injected into the nutrient-poor murine peritoneum survived longer than cells growing in suspension as loosely associated aggregates. The data indicate that growth in spheroid form enhances the OXPHOS activity of constituent tumor cells. In addition, spheroid architecture affects expression of microRNA genes involved in growth control and mitochondrial function. During the mobile phase of metastasis, when ovarian tumor cells disperse through nutrient-poor environments such as the peritoneum, enhanced OXPHOS activity afforded by spheroid architecture would enhance survival and metastatic potential.

Molecular docking, 3D-QASR and molecular dynamics simulations of benzimidazole Pin1 inhibitors.

Pin1 (protein interacting with never-in-mitosis akinase-1) is a member of the family of peptidylprolyl cis-trans isomerases (PPIases) that specifically recognize and isomerize substrates containing phosphorylated Ser/Thr-Pro sequences. Pin1 is involved in many cellular processes and plays a key role in the cell cycle, transcriptional regulation, cell metabolism, proliferation and differentiation, and its abnormalities lead to degenerative and neoplastic diseases. Pin1 is highly expressed in human cancers and promotes the development of tumors by activating multiple oncogenes and inactivating multiple tumor suppressor genes, making it an attractive target for cancer therapy. In this study, we investigated the binding mechanism and conformational relationship between benzimidazole Pin1 inhibitors and Pin1 proteins by molecular docking, three-dimensional quantitative structure-activity relationship (3D-QSAR) modeling, binding free energy calculations and decomposition, and molecular dynamics simulations. Molecular docking and molecular dynamics simulations disclosed the most likely binding pose of benzimidazoles with the Pin1 protein. The results of 3D-QSAR modeling indicated that electrostatic fields, hydrophobic fields and hydrogen bonding play important roles in the binding process of inhibitors to proteins. The binding free energy calculations and energy decomposition indicated that Lys63, Arg69, Cys113, Leu122, Met130, and Ser154 may be key residues in the binding of benzimidazole-based inhibitors to the Pin1 protein. This study provides an important theoretical basis for the design and optimization of benzimidazole compounds.

Co-Occurring Alterations in Multiple Tumor Suppressor Genes Are Associated With Worse Outcomes in Patients With EGFR-Mutant Lung Cancer

IntroductionPatients with metastatic EGFR-mutant NSCLC inevitably have disease progression while on tyrosine kinase inhibitor (TKI) therapy. Co-occurring tumor suppressor gene (TSG) alterations have been associated with poor outcomes, however, detailed analyses of their impact on patient outcomes are limited. MethodsPatients with EGFR-mutant NSCLC treated with EGFR TKIs who had tumor genomic profiling were included. Alterations in TP53 and five additional TSGs (RB1, NF1, ARID1A, BRCA1, and PTEN) were used to stratify the cohort into the following three subgroups: patients with tumors harboring a TP53 mutation plus a mutation in at least one additional TSG (TP53mut/TSGmut), those having a TP53 mutation without additional TSG mutations (TP53mut/TSGwt), and those with TP53wt. Patient characteristics and clinical outcomes were assessed in two independent cohorts. ResultsA total of 101 patients from the Yale Cancer Center and 182 patients from the American Association for Cancer Research Project GENIE database were included. In the Yale cohort, TP53 mutations were identified in 65 cases (64%), of which 23 were TP53mut/TSGmut and 42 were TP53mut/TSGwt. Although the presence of a TP53 mutation was associated with worse outcomes, the additional TSG alteration in TP53mut tumors identified a subset of patients associated with particularly aggressive disease and inferior clinical outcome in both the Yale and the GENIE cohorts. Specifically, in the Yale cohort for patients receiving first-line TKIs, those with TP53mut/TSGmut tumors had shorter progression-free survival (PFS) and overall survival (OS) than TP53mut/TSGwt (PFS: hazard ratio [HR] = 2.03, confidence interval [CI]: 1.12–3.69, p < 0.01, OS: HR = 1.58, CI: 0.82–3.04, p = 0.12) or TP53wt cases (PFS: HR 2.4, CI: 1.28–4.47, p < 0.001, OS: HR = 2.54, CI: 1.21–5.34, p < 0.005). Inferior outcomes in patients with TP53mut/TSGmut tumors were also found in those receiving osimertinib as second-line therapy. Similar findings were seen in patients in the GENIE cohort. ConclusionsPatients with TP53mut/TSGmut tumors represent a patient subgroup characterized by an aggressive disease phenotype and inferior outcomes on EGFR TKIs. This information is important for understanding the biological underpinnings of differential outcomes with TKI treatment and has implications for identifying patients who may benefit from additional therapeutic interventions beyond osimertinib monotherapy.

Fully accessible fitness landscape of oncogene-negative lung adenocarcinoma

Cancer genomes are almost invariably complex with genomic alterations cooperating during each step of carcinogenesis. In cancers that lack a single dominant oncogene mutation, cooperation between the inactivation of multiple tumor suppressor genes can drive tumor initiation and growth. Here, we shed light on how the sequential acquisition of genomic alterations generates oncogene-negative lung tumors. We couple tumor barcoding with combinatorial and multiplexed somatic genome editing to characterize the fitness landscapes of three tumor suppressor genes NF1, RASA1, and PTEN, the inactivation of which jointly drives oncogene-negative lung adenocarcinoma initiation and growth. The fitness landscape was surprisingly accessible, with each additional mutation leading to growth advantage. Furthermore, the fitness landscapes remained fully accessible across backgrounds with the inactivation of additional tumor suppressor genes. These results suggest that while predicting cancer evolution will be challenging, acquiring the multiple alterations that drive the growth of oncogene-negative tumors can be facilitated by the lack of constraints on mutational order.

Genetic analysis of low-grade adenosquamous carcinoma of the breast progressing to high-grade metaplastic carcinoma

PurposeLow-grade adenosquamous carcinoma (LGASC) is a rare type of metaplastic carcinoma of the breast (MBC) with an indolent clinical course. A few LGASC cases with high-grade transformation have been reported; however, the genetics underlying malignant progression of LGASC remain unclear.MethodsWe performed whole-genome sequencing analysis on five MBCs from four patients, including one case with matching primary LGASC and a lymph node metastatic tumor consisting of high-grade MBC with a predominant metaplastic squamous cell carcinoma component (MSC) that progressed from LGASC and three cases of independent de novo MSC.ResultsUnlike de novo MSC, LGASC and its associated MSC showed no TP53 mutation and tended to contain fewer structural variants than de novo MSC. Both LGASC and its associated MSC harbored the common GNAS c.C2530T:p.Arg844Cys mutation, which was more frequently detected in the cancer cell fraction of MSC. MSC associated with LGASC showed additional pathogenic deletions of multiple tumor-suppressor genes, such as KMT2D and BTG1. Copy number analysis revealed potential 18q loss of heterozygosity in both LGASC and associated MSC. The frequency of SMAD4::DCC fusion due to deletions increased with progression to MSC; however, chimeric proteins were not detected. SMAD4 protein expression was already decreased at the LGASC stage due to unknown mechanisms.ConclusionNot only LGASC but also its associated high-grade MBC may be genetically different from de novo high-grade MBC. Progression from LGASC to high-grade MBC may involve the concentration of driver mutations caused by clonal selection and inactivation of tumor-suppressor genes.

MDB-34. “INDUCING IRON IMBALANCE BY TARGETING THE ABCB7/GPX4 AXIS ACCELERATES FERROPTOSIS IN MEDULLOBLASTOMA”

Abstract Medulloblastomas (MB), the most common malignant pediatric brain tumor and a leading cause of childhood mortality, are stratified into four primary subgroups. Deletions within chromosomal locus 17p13.3, which houses multiple tumor suppressor genes including miR-1253, characterize high-risk group 3 tumors. These aggressive tumors also enrich iron transport genes to satisfy their high proliferative need. MiR-1253 targets iron transport by inhibiting the mitochondrial Fe-S transporter, ABCB7. This study elucidated the impact of repressing ABCB7 on cisplatin cytotoxicity in group 3 MB and whether these effects were mediated by ferroptosis. In silico and in vitro analyses revealed specific enrichment of ABCB7 and GPX4, a critical regulator of ferroptosis, in group 3 MB cell lines and tumors. MiR-1253 overexpression (miR-1253OE) resulted in downregulation of both ABCB7 and GPX4, concurrently increasing mitochondrial iron overload, mitochondrial oxidative stress, and lipid peroxidation, leading to cell death and abrogation of medullosphere formation; ABCB7 knockdown (ABCB7KD) recapitulated these effects and abrogated GPX4 expression. Fractionation studies confirmed the inhibitory impact of miR-1253OE and ABCB7KD on GPX4 expression in the cytosol and mitochondria. Seahorse studies revealed that the bulk of ATP generation was occurring in the cytoplasm through glycolysis and not via oxidative phosphorylation, suggesting mitochondrial dysfunction was triggered when ABCB7 was repressed. Cisplatin, a chemotherapeutic agent used in group 3 MB treatment, induces cell death by DNA crosslinking; it also inhibits GPX4 expression. In miR-1253OE and ABCB7KD group 3 cancer cells, cisplatin IC50 was reduced 2-fold. Resultantly, cisplatin treatment augmented oxidative stress and lipid peroxidation, depleted glutathione stores, and culminated in a higher index of ferroptosis. In a mouse model of group 3 tumors, ABCB7KD potentiated cisplatin effects and dramatically prolonged survival. The current study illustrates how targeting iron transport can augment ferroptosis to potentiate cisplatin cytotoxicity in group 3 MB tumors.

Association of alterations in multiple tumor suppressor genes with poor outcomes in patients with EGFR-mutant lung cancer.

9104 Background: Co-occurring alterations in tumor suppressor genes (TSG) have been described as determinants of disease heterogeneity in EGFR-mutant non-small cell lung cancer (NSCLC), however, detailed analyses of their impact on patient outcomes are limited. Methods: Patients with EGFR-mutant NSCLC treated with EGFR tyrosine kinase inhibitors (TKIs) at the Yale Cancer Center who had tumor genomic profiling either before EGFR-TKI therapy (pre-TKI) or at disease progression (post-TKI) were included. Whole exome sequencing (WES) of paired pre- and post-TKI tumor specimens were performed for a subset of patients. The cohort was divided into three subgroups based on alterations in TP53 and five additional TSGs ( RB1, NF1, ARID1A, BRCA1 and PTEN): patients with tumors harboring a TP53 mutation plus a mutation in at least 1 additional TSG ( TP53mut/TSGmut), patients with tumors harboring a TP53 mutation without an additional TSG mutation ( TP53mut/TSGwt), and patients with TP53wt tumors. Clinical characteristics including progression-free (PFS) and overall survival (OS) were assessed. Results: One-hundred-one patients were included in this retrospective study. TP53 mutations were identified in 65 (64%) tumors, of which 23 (35%) and 42 (65%) were classified as TP53mut/TSGmut and TP53mut/TSGwt, respectively. Among those cases with paired WES available (n = 34), frequencies of alterations in the six included TSGs did not significantly differ between pre- and post-TKI tumor specimens. In the full study cohort, the presence of a TP53 mutation was associated with numerically worse PFS (HR 1.46, CI 0.97 – 2.21, p = 0.09) and OS (HR 1.68, CI 1.05 – 2.70, p = 0.04) on first-line EGFR-TKI. Strikingly, after dividing the TP53mut cohort into TP53mut /TSGmut and TP53mut /TSGwt cases, alterations in additional TSG were found to drive the poor outcomes: TP53mut /TSGmut cases had significantly worse PFS and OS on first-line EGFR TKI than TP53mut /TSGwt(mPFS 8.0 vs 10.6 months, p = 0.006; mOS 30.0 vs 33.3 months, p = 0.12) or TP53wt cases (mPFS 8.0 vs 12.6 months, p &lt; 0.0001; mOS 30.0 vs 48.8 months, p = 0.001). There was no significant difference in PFS or OS between patients with TP53mut /TSGwt and TP53wt tumors. Similar outcome differences between the three groups were found in patients who received osimertinib in the second-line setting. Conclusions: The inferior outcomes associated with EGFR/TP53-mutant NSCLC tumors may be due to additional TSG alterations rather than TP53 mutational status alone. Alterations in the investigated TSGs did not appear to emerge under EGFR-TKI therapy, which suggests that they are truncal. Our findings may have implications for understanding the biologic underpinnings of differential outcomes to EGFR TKIs.

Synchronous thyroid cancer and malignant struma ovarii: concordant mutations and microRNA profile, discordant loss of heterozygosity loci

BackgroundStruma ovarii is an unusual ovarian teratoma containing predominantly thyroid tissue. Less than 10% of cases undergo malignant transformation in the thyroid tissue and are considered malignant struma ovarii (MSO). MSO have been reported with concurrent thyroid lesions, but molecular data is lacking.Case presentationA 42-year-old female developed MSO and synchronous multifocal subcentimeter papillary thyroid carcinoma (PTC). The patient underwent a salpingo-oophrectomy, thyroidectomy, and low-dose radioactive iodine ablation. Both the thyroid subcentimeter PTC and MSO were positive for BRAF V600E mutation, and microRNA expression profiles were similar across all tumor deposits. However, only the malignant component demonstrated extensive loss of heterozygosity (LOH) involving multiple tumor suppressor gene (TSG) chromosomal loci.ConclusionsWe present the first reported case of MSO with synchronous multifocal subcentimeter PTC in the thyroid containing concordant BRAF V600E mutations and resulting with discordant LOH findings. This data suggests that loss of expression in tumor suppressor gene(s) may be an important contributor to phenotypic expression of malignancy.

G-quadruplex-enhanced circular single-stranded DNA (G4-CSSD) adsorption of miRNA to inhibit colon cancer progression.

Chromosomal heterogeneity leads to the abnormal expression and mutation of tumor-specific genes. Drugs targeting oncogenes have been extensively developed. However, given the random mutation of tumor suppressor genes, the development of its targeted drugs is difficult. Our early research revealed that artificial circular single-stranded DNA (CSSD) can restore multiple tumor suppressor genes to inhibit tumor malignant progression by adsorbing miRNA. Here, we improved CSSD to a fully closed single-stranded DNA with G quadruplex DNA secondary structure (G4-CSSD), which made G4-CSSD with higher acquisition rate and decreased degradation. The Cancer Genome Atlas (TCGA) and Human Protein Atlas database were used to predict tumour suppressor genes in colon cancer. Cellular and animal experiments were performed to validate the role of G4-CSSD in cancer cell progression. In colon cancer, we observed the simultaneous low expressions of chloride channel accessory 1 (CLCA1), UDP-GlcNAc:betaGal beta-1,3-N-acetylglucosaminyltransferase 6 (B3GNT6) and UDP glucuronosyltransferase family 2 member A3 (UGT2A3), which indicated an favourable prognosis. After repressing miR-590-3p with G4-CSSD590, the upregulation of CLCA1, B3GNT6 and UGT2A3 inhibited the proliferation and metastasis of colon cancer cells. This study may provide basis for new treatment methods for colon cancer by restoration of tumor suppressor genes.

Hot water extract from spent mushroom substrate of Ganoderma lucidu m improves immune function in immune deficient mice

Purpose: To investigate the effects of hot water extract of the spent mushroom substrate from Ganoderma lucidum (HWE) on the immune functions of immune-deficient mice in the presence or absence of cyclophosphamide.&#x0D; Methods: The C57BL/6 mice were treated with varying doses of HWE and cyclophosphamide, and the spleen transcriptome was evaluated by high-throughput sequencing. Pathway enrichment analysis was conducted to generate an overview of differentially-expressed genes (DEGs) functions and interactions.&#x0D; Results: Significant DEGs were observed among the control (CK), normal control (HWE), model (CY), and high-dose HWE (CY + HWEH) groups (p &lt; 0.05). Compared with CK group, HWE and CY + HWEH groups showed upregulation of genes (Alas2, CCNE1, and CCNA2), whereas genes encoding major histocompatibility complex components, costimulatory factors, proinflammatory chemokines, and inflammatory chemokines were significantly downregulated (p &lt; 0.05). Compared with CY group, multiple tumor suppressor and tumorigenesis genes, such as CDKN1A, CDKN1B, MAPK10, and TN-C, were downregulated in CK, HWE, and CY + HWEH groups.&#x0D; Conclusion: This study highlights changes in the spleen transcriptomic profiles of C57BL/6 mice treated with HWE and CY, indicating that HWE improves immunomodulation in a mouse model with immune deficiency. Hot water extract of the spent mushroom substrate from Ganoderma lucidum (HWE) has potentials as an immune enhancer in immunocompromised patients.

Identification and verification of m7G-Related genes as biomarkers for prognosis of sarcoma.

Background: Increasing evidence indicates a crucial role for N7-methylguanosine (m7G) methylation modification in human disease development, particularly cancer, and aberrant m7G levels are closely associated with tumorigenesis and progression via regulation of the expression of multiple oncogenes and tumor suppressor genes. However, the role of m7G in sarcomas (SARC) has not been adequately evaluated. Materials and methods: Transcriptome and clinical data were gathered from the TCGA database for this study. Normal and SARC groups were compared for the expression of m7G-related genes (m7GRGs). The expression of m7GRGs was verified using real-time quantitative PCR (RT-qPCR) in SARC cell lines. Then, differentially expressed genes (DEGs) were identified between high and low m7GRGs expression groups in SARC samples, and GO enrichment and KEGG pathways were evaluated. Next, prognostic values of m7GRGs were evaluated by Cox regression analysis. Subsequently, a prognostic model was constructed using m7GRGs with good prognostic values by Lasso regression analysis. Besides, the relationships between prognostic m7GRGs and immune infiltration, clinical features, cuproptosis-related genes, and antitumor drugs were investigated in patients with SARC. Finally, a ceRNA regulatory network based on m7GRGs was constructed. Results: The expression of ten m7GRGs was higher in the SARC group than in the control group. DEGs across groups with high and low m7GRGs expression were enriched for adhesion sites and cGMP-PKG. Besides, we constructed a prognostic model that consists of EIF4A1, EIF4G3, NCBP1, and WDR4 m7GRGs for predicting the survival likelihood of sarcoma patients. And the elevated expression of these four prognostic m7GRGs was substantially associated with poor prognosis and elevated expression in SARC cell lines. Moreover, we discovered that these four m7GRGs expressions were negatively correlated with CD4+ T cell levels, dendritic cell level and tumor purity, and positively correlated with tumor mutational burden, microsatellite instability, drug sensitivity and cuproptosis-related genes in patients with sarcomas. Then, a triple regulatory network of mRNA, miRNA, and lncRNA was established. Conclusion: The current study identified EIF4A1, EIF4G3, NCBP1, and WDR4 as prognostic genes for SARC that are associated with m7G.These findings extend our knowledge of m7G methylation in SARC and may guide the development of innovative treatment options.

Engraftment of Allotransplanted Tumor Cells in Adult rag2 Mutant Xenopus tropicalis.

Simple SummaryWhile the mouse is without doubt the most studied animal for experimental cancer research, aquatic vertebrates such as zebrafish have also contributed to the field. More recently, thanks to the Nobel-prize winning technology of CRISPR/Cas mediated genomic engineering, the frog Xenopus tropicalis has emerged as an additional powerful model for studying human cancer. Via CRISPR-mediated genome editing, several models for different human cancers have been obtained in this animal. However, what has been lacking in Xenopus is the possibility to transplant tumor cells between different frogs. This is important to allow better characterization of the tumor cells and exploration of therapeutic opportunities. In this paper, we describe the generation of a genetic mutant in Xenopus tropicalis that has a compromised immune system, thereby allowing the grafting and expansion of tumors obtained in this species. In addition, an optimized protocol is provided for the irradiation of wild-type Xenopus frogs that subsequently are temporarily immunocompromised and during that period allow tumor engraftment. This work will expand the toolbox for modeling human cancer in Xenopus tropicalis, thereby further establishing it as a powerful experimental cancer model.Modeling human genetic diseases and cancer in lab animals has been greatly aided by the emergence of genetic engineering tools such as TALENs and CRISPR/Cas9. We have previously demonstrated the ease with which genetically engineered Xenopus models (GEXM) can be generated via injection of early embryos with Cas9 recombinant protein loaded with sgRNAs targeting single or multiple tumor suppressor genes. What has been lacking so far is the possibility to propagate and characterize the induced cancers via transplantation. Here, we describe the generation of a rag2 knockout line in Xenopus tropicalis that is deficient in functional T and B cells. This line was validated by means of allografting experiments with primary tp53−/− and apc+/−/tp53−/− donor tumors. In addition, we optimized available protocols for the sub-lethal irradiation of wild-type X. tropicalis froglets. Irradiated animals also allowed the stable, albeit transient, engraftment of transplanted X. tropicalis tumor cells. The novel rag2−/− line and the irradiated wild-type froglets will further expand the experimental toolbox in the diploid amphibian X. tropicalis and help to establish it as a versatile and relevant model for exploring human cancer.