ABSTRACT
PURPOSE: To investigate the xenobiotic profiles of patients with neovascular age-related macular degeneration (nAMD) undergoing anti-vascular endothelial growth factor (anti-VEGF) intravitreal therapy (IVT) to identify biomarkers indicative of clinical phenotypes through advanced AI methodologies. METHODS: In this cross-sectional observational study, we analyzed 156 peripheral blood xenobiotic features in a cohort of 46 nAMD patients stratified by choroidal neovascularization (CNV) control under anti-VEGF IVT. We employed Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS) for measurement and leveraged an AI-driven iterative Random Forests (iRF) approach for robust pattern recognition and feature selection, aligning molecular profiles with clinical phenotypes. RESULTS: AI-augmented iRF models effectively refined the metabolite spectrum by discarding non-predictive elements. Perfluorooctanesulfonate (PFOS) and Ethyl ß-glucopyranoside were identified as significant biomarkers through this process, associated with various clinically relevant phenotypes. Unlike single metabolite classes, drug metabolites were distinctly correlated with subretinal fluid presence. CONCLUSIONS: This study underscores the enhanced capability of AI, particularly iRF, in dissecting complex metabolomic data to elucidate the xenobiotic landscape of nAMD and environmental impact on the disease. The preliminary biomarkers discovered offer promising directions for personalized treatment strategies, although further validation in broader cohorts is essential for clinical application.
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Purpose: We aimed to determine the impact of artificial sweeteners (AS), especially saccharin, on the progression and treatment efficacy of patients with neovascular age-related macular degeneration (nAMD) under anti-vascular endothelial growth factor (anti-VEGF-A) treatment. Methods: In a cross-sectional study involving 46 patients with nAMD undergoing intravitreal anti-VEGF therapy, 6 AS metabolites were detected in peripheral blood using liquid chromatography - tandem mass spectrometry (LC-MS/MS). Disease features were statistically tested against these metabolite levels. Additionally, a murine choroidal neovascularization (CNV) model, induced by laser, was used to evaluate the effects of orally administered saccharin, assessing both imaging outcomes and gene expression patterns. Polymerase chain reaction (PCR) methods were used to evaluate functional expression of sweet taste receptors in a retinal pigment epithelium (RPE) cell line. Results: Saccharin levels in blood were significantly higher in patients with well-controlled CNV activity (P = 0.004) and those without subretinal hyper-reflective material (P = 0.015). In the murine model, saccharin-treated mice exhibited fewer leaking laser scars, lesser occurrence of bleeding, smaller fibrotic areas (P < 0.05), and a 40% decrease in mononuclear phagocyte accumulation (P = 0.06). Gene analysis indicated downregulation of inflammatory and VEGFR-1 response genes in the treated animals. Human RPE cells expressed taste receptor type 1 member 3 (TAS1R3) mRNA and reacted to saccharin stimulation with changes in mRNA expression. Conclusions: Saccharin appears to play a protective role in patients with nAMD undergoing intravitreal anti-VEGF treatment, aiding in better pathological lesion control and scar reduction. The murine study supports this observation, proposing saccharin's potential in mitigating pathological VEGFR-1-induced immune responses potentially via the RPE sensing saccharin in the blood stream.
Subject(s)
Choroidal Neovascularization , Macular Degeneration , Humans , Mice , Animals , Vascular Endothelial Growth Factor Receptor-1 , Saccharin/therapeutic use , Vascular Endothelial Growth Factor A/genetics , Vascular Endothelial Growth Factor A/metabolism , Sweetening Agents , Cross-Sectional Studies , Chromatography, Liquid , Tandem Mass Spectrometry , Choroidal Neovascularization/metabolism , Macular Degeneration/metabolism , RNA, Messenger/genetics , Intravitreal Injections , Angiogenesis Inhibitors/therapeutic useABSTRACT
The noncoding genome presents a largely untapped source of new biological insights, including thousands of long noncoding RNA (lncRNA) loci. While lncRNA dysregulation has been reported in myeloid malignancies, their functional relevance remains to be systematically interrogated. We performed CRISPRi screens of lncRNA signatures from normal and malignant hematopoietic cells and identified MYNRL15 as a myeloid leukemia dependency. Functional dissection suggests an RNA-independent mechanism mediated by two regulatory elements embedded in the locus. Genetic perturbation of these elements triggered a long-range chromatin interaction and downregulation of leukemia dependency genes near the gained interaction sites, as well as overall suppression of cancer dependency pathways. Thus, this study describes a new noncoding myeloid leukemia vulnerability and mechanistic concept for myeloid leukemia. Importantly, MYNRL15 perturbation caused strong and selective impairment of leukemia cells of various genetic backgrounds over normal hematopoietic stem and progenitor cells in vitro, and depletion of patient-derived xenografts in vivo.
ABSTRACT
There is early evidence of extraocular systemic signals effecting function and morphology in neovascular age-related macular degeneration (nAMD). The prospective, cross-sectional BIOMAC study is an explorative investigation of peripheral blood proteome profiles and matched clinical features to uncover systemic determinacy in nAMD under anti-vascular endothelial growth factor intravitreal therapy (anti-VEGF IVT). It includes 46 nAMD patients stratified by the level of disease control under ongoing anti-VEGF treatment. Proteomic profiles in peripheral blood samples of every patient were detected with LC-MS/MS mass spectrometry. The patients underwent extensive clinical examination with a focus on macular function and morphology. In silico analysis includes unbiased dimensionality reduction and clustering, a subsequent annotation of clinical features, and non-linear models for recognition of underlying patterns. The model assessment was performed using leave-one-out cross validation. The findings provide an exploratory demonstration of the link between systemic proteomic signals and macular disease pattern using and validating non-linear classification models. Three main results were obtained: (1) Proteome-based clustering identifies two distinct patient subclusters with the smaller one (n = 10) exhibiting a strong signature for oxidative stress response. Matching the relevant meta-features on the individual patient's level identifies pulmonary dysfunction as an underlying health condition in these patients. (2) We identify biomarkers for nAMD disease features with Aldolase C as a putative factor associated with superior disease control under ongoing anti-VEGF treatment. (3) Apart from this, isolated protein markers are only weakly correlated with nAMD disease expression. In contrast, applying a non-linear classification model identifies complex molecular patterns hidden in a high number of proteomic dimensions determining macular disease expression. In conclusion, so far unconsidered systemic signals in the peripheral blood proteome contribute to the clinically observed phenotype of nAMD, which should be examined in future translational research on AMD.
Subject(s)
Angiogenesis Inhibitors , Macular Degeneration , Humans , Angiogenesis Inhibitors/therapeutic use , Ranibizumab/therapeutic use , Vascular Endothelial Growth Factor A/metabolism , Proteome , Prospective Studies , Chromatography, Liquid , Cross-Sectional Studies , Proteomics , Tandem Mass Spectrometry , Macular Degeneration/drug therapy , PhenotypeABSTRACT
AIM: Chemoresistance is a major cause of treatment failure in colorectal cancer (CRC) therapy. In this study, the impact of the IGF2BP family of RNA-binding proteins on CRC chemoresistance was investigated using in silico, in vitro, and in vivo approaches. METHODS: Gene expression data from a well-characterized cohort and publicly available cross-linking immunoprecipitation sequencing (CLIP-Seq) data were collected. Resistance to chemotherapeutics was assessed in patient-derived xenografts (PDXs) and patient-derived organoids (PDOs). Functional studies were performed in 2D and 3D cell culture models, including proliferation, spheroid growth, and mitochondrial respiration analyses. RESULTS: We identified IGF2BP2 as the most abundant IGF2BP in primary and metastastatic CRC, correlating with tumor stage in patient samples and tumor growth in PDXs. IGF2BP2 expression in primary tumor tissue was significantly associated with resistance to selumetinib, gefitinib, and regorafenib in PDOs and to 5-fluorouracil and oxaliplatin in PDX in vivo. IGF2BP2 knockout (KO) HCT116 cells were more susceptible to regorafenib in 2D and to oxaliplatin, selumitinib, and nintedanib in 3D cell culture. Further, a bioinformatic analysis using CLIP data suggested stabilization of target transcripts in primary and metastatic tumors. Measurement of oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) revealed a decreased basal OCR and an increase in glycolytic ATP production rate in IGF2BP2 KO. In addition, real-time reverse transcriptase polymerase chain reaction (qPCR) analysis confirmed decreased expression of genes of the respiratory chain complex I, complex IV, and the outer mitochondrial membrane in IGF2BP2 KO cells. CONCLUSIONS: IGF2BP2 correlates with CRC tumor growth in vivo and promotes chemoresistance by altering mitochondrial respiratory chain metabolism. As a druggable target, IGF2BP2 could be used in future CRC therapy to overcome CRC chemoresistance.
Subject(s)
Colorectal Neoplasms , Humans , Oxaliplatin/pharmacology , Colorectal Neoplasms/drug therapy , Colorectal Neoplasms/genetics , Colorectal Neoplasms/pathology , Drug Resistance, Neoplasm/genetics , RNA-Binding Proteins/genetics , RNA-Binding Proteins/metabolism , Cell Line, Tumor , Cell Proliferation/genetics , Gene Expression Regulation, NeoplasticABSTRACT
Gain of chromosome 21 (Hsa21) is among the most frequent aneuploidies in leukemia. However, it remains unclear how partial or complete amplifications of Hsa21 promote leukemogenesis and why children with Down syndrome (DS) (ie, trisomy 21) are particularly at risk of leukemia development. Here, we propose that RUNX1 isoform disequilibrium with RUNX1A bias is key to DS-associated myeloid leukemia (ML-DS). Starting with Hsa21-focused CRISPR-CRISPR-associated protein 9 screens, we uncovered a strong and specific RUNX1 dependency in ML-DS cells. Expression of the RUNX1A isoform is elevated in patients with ML-DS, and mechanistic studies using murine ML-DS models and patient-derived xenografts revealed that excess RUNX1A synergizes with the pathognomonic Gata1s mutation during leukemogenesis by displacing RUNX1C from its endogenous binding sites and inducing oncogenic programs in complex with the MYC cofactor MAX. These effects were reversed by restoring the RUNX1A:RUNX1C equilibrium in patient-derived xenografts in vitro and in vivo. Moreover, pharmacological interference with MYC:MAX dimerization using MYCi361 exerted strong antileukemic effects. Thus, our study highlights the importance of alternative splicing in leukemogenesis, even on a background of aneuploidy, and paves the way for the development of specific and targeted therapies for ML-DS, as well as for other leukemias with Hsa21 aneuploidy or RUNX1 isoform disequilibrium.
Subject(s)
Core Binding Factor Alpha 2 Subunit , Down Syndrome , Leukemia, Myeloid , Animals , Child , Humans , Mice , Aneuploidy , Core Binding Factor Alpha 2 Subunit/genetics , Down Syndrome/complications , Down Syndrome/genetics , Leukemia, Myeloid/genetics , Protein Isoforms/genetics , Trisomy/geneticsABSTRACT
BACKGROUND: Targeted therapies for metastatic uveal melanoma have shown limited benefit in biomarker-unselected populations. The Treat20 Plus study prospectively evaluated the feasibility of a precision oncology strategy in routine clinical practice. PATIENTS AND METHODS: Fresh biopsies were analyzed by high-throughput genomics (whole-genome, whole-exome, and RNA sequencing). A multidisciplinary molecular and immunologic tumor board (MiTB) made individualized treatment recommendations based on identified molecular aberrations, patient situation, drug, and clinical trial availability. Therapy selection was at the discretion of the treating physician. The primary endpoint was the feasibility of the precision oncology clinical program. RESULTS: Molecular analyses were available for 39/45 patients (87%). The MiTB provided treatment recommendations for 40/45 patients (89%), of whom 27/45 (60%) received ≥1 matched therapy. First-line matched therapies included MEK inhibitors (n = 15), MET inhibitors (n = 10), sorafenib (n = 1), and nivolumab (n = 1). The best response to first-line matched therapy was partial response in one patient (nivolumab based on tumor mutational burden), mixed response in two patients, and stable disease in 12 patients for a clinical benefit of 56%. The matched therapy population had a median progression-free survival and overall survival of 3.3 and 13.9 months, respectively. The growth modulation index with matched therapy was >1.33 in 6/17 patients (35%) with prior systemic therapy, suggesting clinical benefit. CONCLUSIONS: A precision oncology approach was feasible for patients with metastatic uveal melanoma, with 60% receiving a therapy matched to identify molecular aberrations. The clinical benefit after checkpoint inhibitors highlights the value of tumor mutational burden testing.
Subject(s)
Neoplasms, Second Primary , Uveal Neoplasms , Biomarkers, Tumor/genetics , Feasibility Studies , Humans , Melanoma , Neoplasms, Second Primary/drug therapy , Nivolumab/therapeutic use , Precision Medicine , Prospective Studies , Uveal Neoplasms/drug therapy , Uveal Neoplasms/geneticsABSTRACT
Metastasis-Associated in Colon Cancer 1 (MACC1) is a strong prognostic biomarker inducing proliferation, migration, invasiveness, and metastasis of cancer cells. The context of MACC1 dysregulation in cancers is, however, still poorly understood. Here, we investigated whether chromosomal instability and somatic copy number alterations (SCNA) frequently occurring in CRC contribute to MACC1 dysregulation, with prognostic and predictive impacts. Using the Oncotrack and Charité CRC cohorts of CRC patients, we showed that elevated MACC1 mRNA expression was tightly dependent on increased MACC1 gene SCNA and was associated with metastasis and shorter metastasis free survival. Deep analysis of the COAD-READ TCGA cohort revealed elevated MACC1 expression due to SCNA for advanced tumors exhibiting high chromosomal instability (CIN), and predominantly classified as CMS2 and CMS4 transcriptomic subtypes. For that cohort, we validated that elevated MACC1 mRNA expression correlated with reduced disease-free and overall survival. In conclusion, this study gives insights into the context of MACC1 expression in CRC. Increased MACC1 expression is largely driven by CIN, SCNA gains, and molecular subtypes, potentially determining the molecular risk for metastasis that might serve as a basis for patient-tailored treatment decisions.
ABSTRACT
Given the plasticity of hematopoietic stem and progenitor cells, multiple routes of differentiation must be blocked in the the pathogenesis of acute myeloid leukemia, the molecular basis of which is incompletely understood. We report that posttranscriptional repression of the transcription factor ARID3A by miR-125b is a key event in the pathogenesis of acute megakaryoblastic leukemia (AMKL). AMKL is frequently associated with trisomy 21 and GATA1 mutations (GATA1s), and children with Down syndrome are at a high risk of developing the disease. The results of our study showed that chromosome 21-encoded miR-125b synergizes with Gata1s to drive leukemogenesis in this context. Leveraging forward and reverse genetics, we uncovered Arid3a as the main miR-125b target behind this synergy. We demonstrated that, during normal hematopoiesis, this transcription factor promotes megakaryocytic differentiation in concert with GATA1 and mediates TGFß-induced apoptosis and cell cycle arrest in complex with SMAD2/3. Although Gata1s mutations perturb erythroid differentiation and induce hyperproliferation of megakaryocytic progenitors, intact ARID3A expression assures their megakaryocytic differentiation and growth restriction. Upon knockdown, these tumor suppressive functions are revoked, causing a blockade of dual megakaryocytic/erythroid differentiation and subsequently of AMKL. Inversely, restoring ARID3A expression relieves the arrest of megakaryocytic differentiation in AMKL patient-derived xenografts. This work illustrates how mutations in lineage-determining transcription factors and perturbation of posttranscriptional gene regulation can interact to block multiple routes of hematopoietic differentiation and cause leukemia. In AMKL, surmounting this differentiation blockade through restoration of the tumor suppressor ARID3A represents a promising strategy for treating this lethal pediatric disease.
Subject(s)
DNA-Binding Proteins/genetics , Leukemia, Megakaryoblastic, Acute/genetics , Transcription Factors/genetics , Animals , Child , GATA1 Transcription Factor/genetics , Gene Expression Regulation, Leukemic , Genes, Tumor Suppressor , Humans , Leukemia, Megakaryoblastic, Acute/pathology , Mice, Inbred C57BL , MicroRNAs/genetics , MutationABSTRACT
The current standard therapies for advanced, recurrent or metastatic colon cancer are the 5-fluorouracil and oxaliplatin or irinotecan schedules (FOxFI) +/- targeted drugs cetuximab or bevacizumab. Treatment with the FOxFI cytotoxic chemotherapy regimens causes significant toxicity and might induce secondary cancers. The overall low efficacy of the targeted drugs seen in colon cancer patients still is hindering the substitution of the chemotherapy. The ONCOTRACK project developed a strategy to identify predictive biomarkers based on a systems biology approach, using omics technologies to identify signatures for personalized treatment based on single drug response data. Here, we describe a follow-up project focusing on target-specific drug combinations. Background for this experimental preclinical study was that, by analyzing the tumor growth inhibition in the PDX models by cetuximab treatment, a broad heterogenic response from complete regression to tumor growth stimulation was observed. To provide confirmation of the hypothesis that drug combinations blocking alternatively activated oncogenic pathways may improve therapy outcomes, 25 models out of the well-characterized ONCOTRACK PDX panel were subjected to treatment with a drug combination scheme using four approved, targeted cancer drugs.
ABSTRACT
Genetic investigation of tumor heterogeneity and clonal evolution in solid cancers could be assisted by the analysis of liquid biopsies. However, tumors of various entities might release different quantities of circulating tumor cells (CTCs) and cell-free DNA (cfDNA) into the bloodstream, potentially limiting the diagnostic potential of liquid biopsy in distinct tumor histologies. Patients with advanced colorectal cancer (CRC), head and neck squamous cell carcinoma (HNSCC), and melanoma (MEL) were enrolled in the study, representing tumors with different metastatic patterns. Mutation profiles of cfDNA, CTCs, and tumor tissue were assessed by panel sequencing, targeting 327 cancer-related genes. In total, 30 tissue, 18 cfDNA, and 7 CTC samples from 18 patients were sequenced. Best concordance between the mutation profile of tissue and cfDNA was achieved in CRC and MEL, possibly due to the remarkable heterogeneity of HNSCC (63%, 55% and 11%, respectively). Concordance especially depended on the amount of cfDNA used for library preparation. While 21 of 27 (78%) tissue mutations were retrieved in high-input cfDNA samples (30-100 ng, N = 8), only 4 of 65 (6%) could be detected in low-input samples (<30 ng, N = 10). CTCs were detected in 13 of 18 patients (72%). However, downstream analysis was limited by poor DNA quality, allowing targeted sequencing of only seven CTC samples isolated from four patients. Only one CTC sample reflected the mutation profile of the respective tumor. Private mutations, which were detected in CTCs but not in tissue, suggested the presence of rare subclones. Our pilot study demonstrated superiority of cfDNA- compared to CTC-based mutation profiling. It was further shown that CTCs may serve as additional means to detect rare subclones possibly involved in treatment resistance. Both findings require validation in a larger patient cohort.
Subject(s)
Colorectal Neoplasms , Cell-Free Nucleic Acids , Female , Humans , Liquid Biopsy , Male , Mutation , Neoplastic Cells, Circulating , Oncogenes , Pilot ProjectsABSTRACT
Up to 40% of advance lung, melanoma and breast cancer patients suffer from brain metastases (BM) with increasing incidence. Here, we assessed whether circulating tumor cells (CTCs) in peripheral blood can serve as a disease surrogate, focusing on CD44 and CD74 expression as prognostic markers for BM. We show that a size-based microfluidic approach in combination with a semi-automated cell recognition system are well suited for CTC detection in BM patients and allow further characterization of tumor cells potentially derived from BM. CTCs were found in 50% (7/14) of breast cancer, 50% (9/18) of non-small cell lung cancer (NSCLC) and 36% (4/11) of melanoma patients. The next-generation sequencing (NGS) analysis of nine single CTCs from one breast cancer patient revealed three different CNV profile groups as well as a resistance causing ERS1 mutation. CD44 and CD74 were expressed on most CTCs and their expression was strongly correlated, whereas matched breast cancer BM tissues were much less frequently expressing CD44 and CD74 (negative in 46% and 54%, respectively). Thus, plasticity of CD44 and CD74 expression during trafficking of CTCs in the circulation might be the result of adaptation strategies.
Subject(s)
Antigens, Differentiation, B-Lymphocyte/genetics , Brain Neoplasms/genetics , Brain Neoplasms/secondary , Histocompatibility Antigens Class II/genetics , Hyaluronan Receptors/genetics , Neoplastic Cells, Circulating/metabolism , Antigens, Differentiation, B-Lymphocyte/metabolism , Biomarkers, Tumor , Brain Neoplasms/diagnosis , Breast Neoplasms/pathology , Carcinoma, Non-Small-Cell Lung/pathology , Cell Line, Tumor , Female , Histocompatibility Antigens Class II/metabolism , Humans , Hyaluronan Receptors/metabolism , Immunohistochemistry , Male , Mutation , Whole Genome SequencingABSTRACT
PURPOSE: Immune checkpoint inhibitors (ICIs) are increasingly being used in non-small-cell lung cancer (NSCLC), yet biomarkers predicting their benefit are lacking. We evaluated if on-treatment changes of circulating tumor DNA (ctDNA) from ICI start (t0) to after two cycles (t1) assessed with a commercial panel could identify patients with NSCLC who would benefit from ICI. PATIENTS AND METHODS: The molecular ctDNA response was evaluated as a predictor of radiographic tumor response and long-term survival benefit of ICI. To maximize the yield of ctDNA detection, de novo mutation calling was performed. Furthermore, the impact of clonal hematopoiesis (CH)-related variants as a source of biologic noise was investigated. RESULTS: After correction for CH-related variants, which were detected in 75 patients (44.9%), ctDNA was detected in 152 of 167 (91.0%) patients. We observed only a fair agreement of the molecular and radiographic response, which was even more impaired by the inclusion of CH-related variants. After exclusion of those, a ≥ 50% molecular response improved progression-free survival (10 v 2 months; hazard ratio [HR], 0.55; 95% CI, 0.39 to 0.77; P = .0011) and overall survival (18.4 v 5.9 months; HR, 0.44; 95% CI, 0.31 to 0.62; P < .0001) compared with patients not achieving this end point. After adjusting for clinical variables, ctDNA response and STK11/KEAP1 mutations (HR, 2.08; 95% CI, 1.4 to 3.0; P < .001) remained independent predictors for overall survival, irrespective of programmed death ligand-1 expression. A landmark survival analysis at 2 months (n = 129) provided similar results. CONCLUSION: On-treatment changes of ctDNA in plasma reveal predictive information for long-term clinical benefit in ICI-treated patients with NSCLC. A broader NSCLC patient coverage through de novo mutation calling and the use of a variant call set excluding CH-related variants improved the classification of molecular responders, but had no significant impact on survival.
Subject(s)
Carcinoma, Non-Small-Cell Lung/blood , Carcinoma, Non-Small-Cell Lung/drug therapy , Circulating Tumor DNA/blood , Immune Checkpoint Inhibitors/therapeutic use , Lung Neoplasms/blood , Lung Neoplasms/drug therapy , Adult , Aged , Aged, 80 and over , Carcinoma, Non-Small-Cell Lung/pathology , Female , Humans , Lung Neoplasms/pathology , Male , Middle Aged , Neoplasm Staging , Predictive Value of Tests , Treatment OutcomeABSTRACT
Transcriptional profiling of hematopoietic cell subpopulations has helped to characterize the developmental stages of the hematopoietic system and the molecular bases of malignant and non-malignant blood diseases. Previously, only the genes targeted by expression microarrays could be profiled genome-wide. High-throughput RNA sequencing, however, encompasses a broader repertoire of RNA molecules, without restriction to previously annotated genes. We analyzed the BLUEPRINT consortium RNA-sequencing data for mature hematopoietic cell types. The data comprised 90 total RNA-sequencing samples, each composed of one of 27 cell types, and 32 small RNA-sequencing samples, each composed of one of 11 cell types. We estimated gene and isoform expression levels for each cell type using existing annotations from Ensembl. We then used guided transcriptome assembly to discover unannotated transcripts. We identified hundreds of novel non-coding RNA genes and showed that the majority have cell type-dependent expression. We also characterized the expression of circular RNA and found that these are also cell type-specific. These analyses refine the active transcriptional landscape of mature hematopoietic cells, highlight abundant genes and transcriptional isoforms for each blood cell type, and provide a valuable resource for researchers of hematologic development and diseases. Finally, we made the data accessible via a web-based interface: https://blueprint.haem.cam.ac.uk/bloodatlas/.
Subject(s)
RNA, Long Noncoding , Transcriptome , Gene Expression Profiling , High-Throughput Nucleotide Sequencing , RNA, Circular , RNA, Long Noncoding/genetics , Sequence Analysis, RNASubject(s)
Diet, Ketogenic , Epilepsy, Generalized , Mental Disorders , Humans , Infant, Newborn , Syndrome , Vigabatrin/therapeutic useABSTRACT
Chromophobe renal cell carcinoma (chRCC) accounts for approximately 5% of all renal cancers and around 30% of chRCC cases have mutations in TP53. chRCC is poorly supported by microvessels and has markably lower glucose uptake than clear cell RCC and papillary RCC. Currently, the metabolic status and mechanisms by which this tumor adapts to nutrient-poor microenvironments remain to be investigated. In this study, we performed proteome and metabolome profiling of chRCC tumors and adjacent kidney tissues and identified major metabolic alterations in chRCC tumors, including the classical Warburg effect, the downregulation of gluconeogenesis and amino acid metabolism, and the upregulation of protein degradation and endocytosis. chRCC cells depended on extracellular macromolecules as an amino acid source by activating endocytosis to sustain cell proliferation and survival. Inhibition of the phospholipase C gamma 2 (PLCG2)/inositol 1,4,5-trisphosphate (IP3)/Ca2+/protein kinase C (PKC) pathway significantly impaired the activation of endocytosis for amino acid uptakes into chRCC cells. In chRCC, whole-exome sequencing revealed that TP53 mutations were not related to expression of PLCG2 and activation of endocytosis. Our study provides novel perspectives on metabolic rewiring in chRCC and identifies the PLCG2/IP3/Ca2+/PKC axis as a potential therapeutic target in patients with chRCC. SIGNIFICANCE: This study reveals macropinocytosis as an important process utilized by chRCC to gain extracellular nutrients in a p53-independent manner.
Subject(s)
Amino Acids/metabolism , Carcinoma, Renal Cell/metabolism , Cell Proliferation/drug effects , Cell Survival/drug effects , Endocytosis/drug effects , Kidney Neoplasms/metabolism , Amiloride/analogs & derivatives , Amiloride/pharmacology , Boron Compounds/pharmacology , Calcium/metabolism , Calcium Signaling/drug effects , Carcinoma, Renal Cell/pathology , Cell Line, Tumor , Egtazic Acid/analogs & derivatives , Egtazic Acid/pharmacology , Estrenes/pharmacology , Gluconeogenesis , Humans , Indoles/pharmacology , Inositol 1,4,5-Trisphosphate/antagonists & inhibitors , Inositol 1,4,5-Trisphosphate/metabolism , Kidney Neoplasms/pathology , Maleimides/pharmacology , Metabolome , Phospholipase C gamma/antagonists & inhibitors , Phospholipase C gamma/metabolism , Protein Kinase C/antagonists & inhibitors , Protein Kinase C/metabolism , Proteome , Pyrrolidinones/pharmacologyABSTRACT
Most relapses of acute lymphoblastic leukemia (ALL) occur in patients with a medium risk (MR) for relapse on the Associazione Italiana di Ematologia e Oncologia Pediatrica and Berlin-Frankfurt-Münster (AIEOP-BFM) ALL protocol, based on persistence of minimal residual disease (MRD). New insights into biological features that are associated with MRD are needed. Here, we identify the glycosylphosphatidylinositol-anchored cell surface protein vanin-2 (VNN2; GPI-80) by charting the cell surface proteome of MRD very high-risk (HR) B-cell precursor (BCP) ALL using a chemoproteomics strategy. The correlation between VNN2 transcript and surface protein expression enabled a retrospective analysis (ALL-BFM 2000; N = 770 cases) using quantitative polymerase chain reaction to confirm the association of VNN2 with MRD and independent prediction of worse outcome. Using flow cytometry, we detected VNN2 expression in 2 waves, in human adult bone marrow stem and progenitor cells and in the mature myeloid compartment, in line with proposed roles for fetal hematopoietic stem cells and inflammation. Prospective validation by flow cytometry in the ongoing clinical trial (AIEOP-BFM 2009) identified 10% (103/1069) of VNN2+ BCP ALL patients at first diagnosis, primarily in the MRD MR (48/103, 47%) and HR (37/103, 36%) groups, across various cytogenetic subtypes. We also detected frequent mutations in epigenetic regulators in VNN2+ ALLs, including histone H3 methyltransferases MLL2, SETD2, and EZH2 and demethylase KDM6A. Inactivation of the VNN2 gene did not impair leukemia repopulation capacity in xenografts. Taken together, VNN2 marks a cellular state of increased resistance to chemotherapy that warrants further investigations. Therefore, this marker should be included in diagnostic flow cytometry panels.
Subject(s)
Drug Resistance, Neoplasm , Precursor Cell Lymphoblastic Leukemia-Lymphoma , Amidohydrolases/therapeutic use , Antineoplastic Combined Chemotherapy Protocols , B-Lymphocytes , Cell Adhesion Molecules , Child , Drug Resistance, Neoplasm/genetics , GPI-Linked Proteins , Hematopoietic Stem Cells , Humans , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Prospective Studies , Retrospective StudiesABSTRACT
BACKGROUND: Reliable and reproducible interpretation of molecular aberrations constitutes a bottleneck of precision medicine. Evidence-based decision management systems may improve rational therapy recommendations. To cope with an increasing amount of complex molecular data in the clinical care of patients with cancer, we established a workflow for the interpretation of molecular analyses. METHODS: A specialized physician screened results from molecular analyses for potential biomarkers, irrespective of the diagnostic modality. Best available evidence was retrieved and categorized through establishment of an in-house database and interrogation of publicly available databases. Annotated biomarkers were ranked using predefined evidence levels and subsequently discussed at a molecular tumour board (MTB), which generated treatment recommendations. Subsequent translation into patient treatment and clinical outcomes were followed up. RESULTS: One hundred patients were discussed in the MTB between January 2016 and May 2017. Molecular data were obtained for 70 of 100 patients (50 whole exome/RNA sequencing, 18 panel sequencing, 2 immunohistochemistry (IHC)/microsatellite instability analysis). The MTB generated a median of two treatment recommendations each for 63 patients. Thirty-nine patients were treated: 6 partial responses and 12 stable diseases were achieved as best responses. Genetic counselling for germline events was recommended for seven patients. CONCLUSION: The development of an evidence-based workflow allowed for the clinical interpretation of complex molecular data and facilitated the translation of personalized treatment strategies into routine clinical care. The high number of treatment recommendations in patients with comprehensive genomic data and promising responses in patients treated with combination therapy warrant larger clinical studies.
Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Decision Support Techniques , High-Throughput Nucleotide Sequencing/methods , Molecular Targeted Therapy , Neoplasms/drug therapy , Pathology, Molecular/statistics & numerical data , Precision Medicine , Adult , Aged , Aged, 80 and over , Biomarkers, Tumor/genetics , Female , Follow-Up Studies , Humans , Male , Middle Aged , Neoplasms/genetics , Neoplasms/pathology , Prognosis , Young AdultABSTRACT
The chimeric transcription factor TCF3-HLF defines an incurable acute lymphoblastic leukemia subtype. Here we decipher the regulome of endogenous TCF3-HLF and dissect its essential transcriptional components and targets by functional genomics. We demonstrate that TCF3-HLF recruits HLF binding sites at hematopoietic stem cell/myeloid lineage associated (super-) enhancers to drive lineage identity and self-renewal. Among direct targets, hijacking an HLF binding site in a MYC enhancer cluster by TCF3-HLF activates a conserved MYC-driven transformation program crucial for leukemia propagation in vivo. TCF3-HLF pioneers the cooperation with ERG and recruits histone acetyltransferase p300 (EP300), conferring susceptibility to EP300 inhibition. Our study provides a framework for targeting driving transcriptional dependencies in this fatal leukemia.