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Multiple myeloma is a treatable, but currently incurable, hematological malignancy of plasma cells characterized by diverse and complex tumor genetics for which precision medicine approaches to treatment are lacking. The Multiple Myeloma Research Foundation's Relating Clinical Outcomes in Multiple Myeloma to Personal Assessment of Genetic Profile study ( NCT01454297 ) is a longitudinal, observational clinical study of newly diagnosed patients with multiple myeloma (n = 1,143) where tumor samples are characterized using whole-genome sequencing, whole-exome sequencing and RNA sequencing at diagnosis and progression, and clinical data are collected every 3 months. Analyses of the baseline cohort identified genes that are the target of recurrent gain-of-function and loss-of-function events. Consensus clustering identified 8 and 12 unique copy number and expression subtypes of myeloma, respectively, identifying high-risk genetic subtypes and elucidating many of the molecular underpinnings of these unique biological groups. Analysis of serial samples showed that 25.5% of patients transition to a high-risk expression subtype at progression. We observed robust expression of immunotherapy targets in this subtype, suggesting a potential therapeutic option.
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The emergence of single nucleus RNA sequencing (snRNA-seq) offers to revolutionize the study of Alzheimer's disease (AD). Integration with complementary multiomics data such as genetics, proteomics and clinical data provides powerful opportunities to link cell subpopulations and molecular networks with a broader disease-relevant context. We report snRNA-seq profiles from superior frontal gyrus samples from 101 well characterized subjects from the Banner Brain and Body Donation Program in combination with whole genome sequences. We report findings that link common AD risk variants with CR1 expression in oligodendrocytes as well as alterations in hematological parameters. We observed an AD-associated CD83(+) microglial subtype with unique molecular networks and which is associated with immunoglobulin IgG4 production in the transverse colon. Our major observations were replicated in two additional, independent snRNA-seq data sets. These findings illustrate the power of multi-tissue molecular profiling to contextualize snRNA-seq brain transcriptomics and reveal disease biology.
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Enfermedad de Alzheimer , Análisis de la Célula Individual , Transcriptoma , Humanos , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Masculino , Femenino , Anciano , Microglía/metabolismo , Anciano de 80 o más Años , Oligodendroglía/metabolismo , Persona de Mediana Edad , Inmunoglobulina G/metabolismo , Redes Reguladoras de Genes , Análisis de Secuencia de ARN , Encéfalo/metabolismo , Encéfalo/patología , Perfilación de la Expresión GénicaRESUMEN
In recent years, the development of sensor and wearable technologies have led to their increased adoption in clinical and health monitoring settings. One area that is in early, but promising, stages of development is the use of biosensors for therapeutic drug monitoring (TDM). Traditionally, TDM could only be performed in certified laboratories and was used in specific scenarios to optimize drug dosage based on measurement of plasma/blood drug concentrations. Although TDM has been typically pursued in settings involving medications that are challenging to manage, the basic approach is useful for characterizing drug activity. TDM is based on the idea that there is likely a clear relationship between plasma/blood drug concentration (or concentration in other matrices) and clinical efficacy. However, these relationships may vary across individuals and may be affected by genetic factors, comorbidities, lifestyle, and diet. TDM technologies will be valuable for enabling precision medicine strategies to determine the clinical efficacy of drugs in individuals, as well as optimizing personalized dosing, especially since therapeutic windows may vary inter-individually. In this mini-review, we discuss emerging TDM technologies and their applications, and factors that influence TDM including drug interactions, polypharmacy, and supplement use. We also discuss how using TDM within single subject (N-of-1) and aggregated N-of-1 clinical trial designs provides opportunities to better capture drug response and activity at the individual level. Individualized TDM solutions have the potential to help optimize treatment selection and dosing regimens so that the right drug and right dose may be matched to the right person and in the right context.
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The emergence of technologies that can support high-throughput profiling of single cell transcriptomes offers to revolutionize the study of brain tissue from persons with and without Alzheimer's disease (AD). Integration of these data with additional complementary multiomics data such as genetics, proteomics and clinical data provides powerful opportunities to link observed cell subpopulations and molecular network features within a broader disease-relevant context. We report here single nucleus RNA sequencing (snRNA-seq) profiles generated from superior frontal gyrus cortical tissue samples from 101 exceptionally well characterized, aged subjects from the Banner Brain and Body Donation Program in combination with whole genome sequences. We report findings that link common AD risk variants with CR1 expression in oligodendrocytes as well as alterations in peripheral hematological lab parameters, with these observations replicated in an independent, prospective cohort study of ageing and dementia. We also observed an AD-associated CD83(+) microglial subtype with unique molecular networks that encompass many known regulators of AD-relevant microglial biology, and which are associated with immunoglobulin IgG4 production in the transverse colon. These findings illustrate the power of multi-tissue molecular profiling to contextualize snRNA-seq brain transcriptomics and reveal novel disease biology. The transcriptomic, genetic, phenotypic, and network data resources described within this study are available for access and utilization by the scientific community.
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Background: Small bowel carcinoids are insidious tumors that are often metastatic when diagnosed. Limited mutation landscape studies of carcinoids indicate that these tumors have a relatively low mutational burden. The development of targeted therapies will depend upon the identification of mutations that drive the pathogenesis and metastasis of carcinoid tumors. Methods: Whole exome and RNA sequencing of 5 matched sets of normal tissue, primary small intestine carcinoid tumors, and liver metastases were investigated. Germline and somatic variants included: single nucleotide variants (SNVs), insertions/deletions (indels), structural variants, and copy number alterations (CNAs). The functional impact of mutations was predicted using Ensembl Variant Effect Predictor. Results: Large-scale CNAs were observed including the loss of chromosome 18 in all 5 metastases and 3/5 primary tumors. Certain somatic SNVs were metastasis-specific; including mutations in ATRX, CDKN1B, MXRA5 (leading to the activation of a cryptic splice site and loss of mRNA), SMARCA2, and the loss of UBE4B. Additional mutations in ATRX, and splice site loss of PYGL, leading to intron retention observed in primary and metastatic tumors. Conclusions: We observed novel mutations in primary/metastatic carcinoid tumor pairs, and some have been observed in other types of neuroendocrine tumors. We confirmed a previously observed loss of chromosome 18 and CDKN1B. Transcriptome sequencing added relevant information that would not have been appreciated with DNA sequencing alone. The detection of several splicing mutations on the DNA level and their consequences at the RNA level suggests that RNA splicing aberrations may be an important mechanism underlying carcinoid tumors.
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Tumor Carcinoide , Neoplasias Intestinales , Tumores Neuroendocrinos , Humanos , Multiómica , Tumor Carcinoide/genética , Tumor Carcinoide/patología , Tumor Carcinoide/secundario , Tumores Neuroendocrinos/genética , Tumores Neuroendocrinos/patología , Neoplasias Intestinales/genética , Neoplasias Intestinales/patología , Ubiquitina-Proteína LigasasRESUMEN
PURPOSE: Hyperthermic intraperitoneal chemotherapy (HIPEC) with cisplatin confers a survival benefit in epithelial ovarian cancer (EOC) but is associated with renal toxicity. Sodium thiosulfate (ST) is used for nephroprotection for HIPEC with cisplatin, but standard HIPEC practices vary. METHODS: A prospective, nonrandomized, clinical trial evaluated safety outcomes of HIPEC with cisplatin 75 mg/m2 during cytoreductive surgery (CRS) in patients with EOC (n = 34) and endometrial cancer (n = 6). Twenty-one patients received no ST (nST), and 19 received ST. Adverse events (AEs) were reported according to CTCAE v.5.0. Serum creatinine (Cr) was collected preoperatively and postoperatively (Days 5-8). Progression-free survival (PFS) was followed. Normal peritoneum was biopsied before and after HIPEC for whole transcriptomic sequencing to identify RNAseq signatures correlating with AEs. RESULTS: Forty patients had HIPEC at the time of interval or secondary CRS. Renal toxicities in the nST group were 33% any grade AE and 9% grade 3 AEs. The ST group demonstrated no renal AEs. Median postoperative Cr in the nST group was 1.1 mg/dL and 0.5 mg/dL in the ST group (p = 0.0001). Median change in Cr from preoperative to postoperative levels were + 53% (nST) compared with - 9.6% (ST) (p = 0.003). PFS did not differ between the ST and nST groups in primary or recurrent EOC patients. Renal AEs were associated with downregulation of metabolic pathways and upregulation of immune pathways. CONCLUSIONS: ST significantly reduces acute renal toxicity associated with HIPEC with cisplatin in ovarian cancer patients. As nephrotoxicity is high in HIPEC with cisplatin, nephroprotective agents should be considered.
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Antineoplásicos , Hipertermia Inducida , Neoplasias Ováricas , Humanos , Femenino , Cisplatino/uso terapéutico , Quimioterapia Intraperitoneal Hipertérmica , Antineoplásicos/uso terapéutico , Estudios Prospectivos , Hipertermia Inducida/efectos adversos , Recurrencia Local de Neoplasia , Neoplasias Ováricas/patología , Carcinoma Epitelial de Ovario , Procedimientos Quirúrgicos de Citorreducción/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Terapia CombinadaRESUMEN
The development of effective treatments to prevent and slow Alzheimer's disease (AD) pathogenesis is needed in order to tackle the steady increase in the global prevalence of AD. This challenge is complicated by the need to identify key health shifts that precede the onset of AD and cognitive decline as these represent windows of opportunity for intervening and preventing disease. Such shifts may be captured through the measurement of biomarkers that reflect the health of the individual, in particular those that reflect brain age and biological age. Brain age biomarkers provide a composite view of the health of the brain based on neuroanatomical analyses, while biological age biomarkers, which encompass the epigenetic clock, provide a measurement of the overall health state of an individual based on DNA methylation analysis. Acceleration of brain and biological ages is associated with changes in cognitive function, as well as neuropathological markers of AD. In this mini-review, we discuss brain age and biological age research in the context of cognitive decline and AD. While more research is needed, studies show that brain and biological aging trajectories are variable across individuals and that such trajectories are non-linear at older ages. Longitudinal monitoring of these biomarkers may be valuable for enabling earlier identification of divergent pathological trajectories toward AD and providing insight into points for intervention.
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The posterior cingulate cortex, a key hub of the default mode network, underlies autobiographical memory retrieval and displays hypometabolic changes early in Alzheimer disease. To obtain an unbiased understanding of the molecular pathobiology of the aged posterior cingulate cortex, we performed RNA sequencing (RNA-seq) on tissue obtained from 26 participants of the Rush Religious Orders Study (11 males/15 females; aged 76-96 years) with a pre-mortem clinical diagnosis of no cognitive impairment and post-mortem neurofibrillary tangle Braak Stages I/II, III, and IV. Transcriptomic data were gathered using next-generation sequencing of RNA extracted from posterior cingulate cortex generating an average of 60 million paired reads per subject. Normalized expression of RNA-seq data was calculated using a global gene annotation and a microRNA profile. Differential expression (DESeq2, edgeR) using Braak staging as the comparison structure isolated genes for dimensional scaling, associative network building and functional clustering. Curated genes were correlated with the Mini-Mental State Examination and semantic, working and episodic memory, visuospatial ability, and a composite Global Cognitive Score. Regulatory mechanisms were determined by co-expression networks with microRNAs and an overlap of transcription factor binding sites. Analysis revealed 750 genes and 12 microRNAs significantly differentially expressed between Braak Stages I/II and III/IV and an associated six groups of transcription factor binding sites. Inputting significantly different gene/network data into a functional annotation clustering model revealed elevated presynaptic, postsynaptic and ATP-related expression in Braak Stages III and IV compared with Stages I/II, suggesting these pathways are integral for cognitive resilience seen in unimpaired elderly subjects. Principal component analysis and Kruskal-Wallis testing did not associate Braak stage with cognitive function. However, Spearman correlations between genes and cognitive test scores followed by network analysis revealed upregulation of classes of synaptic genes positively associated with performance on the visuospatial perceptual orientation domain. Upregulation of key synaptic genes suggests a role for these transcripts and associated synaptic pathways in cognitive resilience seen in elders despite Alzheimer disease pathology and dementia.
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PURPOSE: Hyperthermic intraperitoneal chemotherapy (HIPEC) confers a survival benefit in epithelial ovarian cancer (EOC) and in preclinical models. However, the molecular changes induced by HIPEC have not been corroborated in humans. PATIENTS AND METHODS: A feasibility trial evaluated clinical and safety outcomes of HIPEC with cisplatin during optimal cytoreductive surgery (CRS) in patients with EOC diagnosed with stage III, IV, or recurrent EOC. Pre- and post-HIPEC biopsies were comprehensively profiled with genomic and transcriptomic sequencing to identify mutational and RNAseq signatures correlating with response; the tumor microenvironment was profiled to identify potential immune biomarkers; and transcriptional signatures of tumors and normal samples before and after HIPEC were compared to investigate HIPEC-induced acute transcriptional changes. RESULTS: Thirty-five patients had HIPEC at the time of optimal CRS; all patients had optimal CRS. The median progression-free survival (PFS) was 24.7 months for primary patients and 22.4 for recurrent patients. There were no grade 4 or 5 adverse events. Anemia was the most common grade 3 adverse event (43%). Hierarchical cluster analyses identified distinct transcriptomic signatures of good versus poor responders to HIPEC correlating with a PFS of 29.9 versus 7.3 months, respectively. Among good responders, significant HIPEC-induced molecular changes included immune pathway upregulation and DNA repair pathway downregulation. Within cancer islands, % programmed cell death protein 1 expression in CD8+ T cells significantly increased after HIPEC. An exceptional responder (PFS 58 months) demonstrated the highest programmed cell death protein 1 increase. Heat shock proteins comprised the top differentially upregulated genes in HIPEC-treated tumors. CONCLUSION: Distinct transcriptomic signatures identify responders to HIPEC, and preclinical model findings are confirmed for the first time in a human cohort.
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Carcinoma Epitelial de Ovario , Quimioterapia Intraperitoneal Hipertérmica , Neoplasias Ováricas , Carcinoma Epitelial de Ovario/tratamiento farmacológico , Estudios de Factibilidad , Femenino , Humanos , Quimioterapia Intraperitoneal Hipertérmica/efectos adversos , Recurrencia Local de Neoplasia/tratamiento farmacológico , Neoplasias Ováricas/tratamiento farmacológico , Microambiente TumoralRESUMEN
The RNA-binding protein HuD (a.k.a., ELAVL4) is involved in neuronal development and synaptic plasticity mechanisms, including addiction-related processes such as cocaine conditioned-place preference (CPP) and food reward. The most studied function of this protein is mRNA stabilization; however, we have recently shown that HuD also regulates the levels of circular RNAs (circRNAs) in neurons. To examine the role of HuD in the control of coding and non-coding RNA networks associated with substance use, we identified sets of differentially expressed mRNAs, circRNAs and miRNAs in the striatum of HuD knockout (KO) mice. Our findings indicate that significantly downregulated mRNAs are enriched in biological pathways related to cell morphology and behavior. Furthermore, deletion of HuD altered the levels of 15 miRNAs associated with drug seeking. Using these sets of data, we predicted that a large number of upregulated miRNAs form competing endogenous RNA (ceRNA) networks with circRNAs and mRNAs associated with the neuronal development and synaptic plasticity proteins LSAMP and MARK3. Additionally, several downregulated miRNAs form ceRNA networks with mRNAs and circRNAs from MEF2D, PIK3R3, PTRPM and other neuronal proteins. Together, our results indicate that HuD regulates ceRNA networks controlling the levels of mRNAs associated with neuronal differentiation and synaptic physiology.
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The robust detection of disease-associated splice events from RNAseq data is challenging due to the potential confounding effect of gene expression levels and the often limited number of patients with relevant RNAseq data. Here we present a novel statistical approach to splicing outlier detection and differential splicing analysis. Our approach tests for differences in the percentages of sequence reads representing local splice events. We describe a software package called Bisbee which can predict the protein-level effect of splice alterations, a key feature lacking in many other splicing analysis resources. We leverage Bisbee's prediction of protein level effects as a benchmark of its capabilities using matched sets of RNAseq and mass spectrometry data from normal tissues. Bisbee exhibits improved sensitivity and specificity over existing approaches and can be used to identify tissue-specific splice variants whose protein-level expression can be confirmed by mass spectrometry. We also applied Bisbee to assess evidence for a pathogenic splicing variant contributing to a rare disease and to identify tumor-specific splice isoforms associated with an oncogenic mutation. Bisbee was able to rediscover previously validated results in both of these cases and also identify common tumor-associated splice isoforms replicated in two independent melanoma datasets.
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Empalme Alternativo , Melanoma/genética , Proteínas Proto-Oncogénicas/metabolismo , Análisis de Secuencia de ARN/métodos , Biología Computacional/métodos , Perfilación de la Expresión Génica , Humanos , Espectrometría de Masas , Melanoma/metabolismo , Mutación , Especificidad de Órganos , Proteínas Proto-Oncogénicas/genética , Programas InformáticosRESUMEN
Although combination BRAF and MEK inhibitors are highly effective for the 40-50% of cutaneous metastatic melanomas harboring BRAFV600 mutations, targeted agents have been ineffective for BRAFV600wild-type (wt) metastatic melanomas. The SU2C Genomics-Enabled Medicine for Melanoma Trial utilized a Simon two-stage optimal design to assess whether comprehensive genomic profiling improves selection of molecular-based therapies for BRAFV600wt metastatic melanoma patients who had progressed on standard-of-care therapy, which may include immunotherapy. Of the response-evaluable patients, binimetinib was selected for 20 patients randomized to the genomics-enabled arm, and nine were treated on the alternate treatment arm. Response rates for 27 patients treated with targeted recommendations included one (4%) partial response, 18 (67%) with stable disease, and eight (30%) with progressive disease. Post-trial genomic and protein pathway activation mapping identified additional drug classes that may be considered for future studies. Our results highlight the complexity and heterogeneity of metastatic melanomas, as well as how the lack of response in this trial may be associated with limitations including monotherapy drug selection and the dearth of available single and combination molecularly-driven therapies to treat BRAFV600wt metastatic melanomas.
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Bencimidazoles/administración & dosificación , Genómica , Melanoma , Proteínas Proto-Oncogénicas B-raf , Neoplasias Cutáneas , Adulto , Anciano , Femenino , Humanos , Masculino , Melanoma/tratamiento farmacológico , Melanoma/genética , Melanoma/metabolismo , Persona de Mediana Edad , Metástasis de la Neoplasia , Proyectos Piloto , Estudios Prospectivos , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas B-raf/metabolismo , Neoplasias Cutáneas/tratamiento farmacológico , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo , Melanoma Cutáneo MalignoRESUMEN
The 22q11.2 deletion syndrome (22q11.2 DS) is the most common deletion syndrome in humans. In most cases, it occurs de novo. A rare family of three with 22q11.2 deletion syndrome (22q11.2 DS) resulting from an unbalanced 18q;22q translocation is reported here. Their deletion region is atypical in that it includes only 26 of the 36 genes in the minimal critical 22q11.2 DS region but it involves the loss of the centromeric 22q region and the entire p arm. The deletion region overlaps with seven other rare atypical cases; common to all cases was the loss of a region including SEPT5-GP1BB proximally and most of ARVCF distally. Interrogation of the deleted 22q region proximal to the canonical 22q11.2 deletion region in the DECIPHER database showed seven cases with isolated or combined traits of 22q11.2 DS, including three with clefts. The phenotypes in the present family thus may result from the loss of a subset of genes in the critical region, or alternatively the loss of other genes or sequences in the proximal 22q deletion region, or interactive effects among these. Despite the identical deletion locus in the three affected family members, expression of the 22q11.2 DS traits differed substantially among them. These three related cases thus contribute to knowledge of 22q11.2 DS in that their unusual deletion locus co-occurred with the cardinal features of the syndrome while their identical deletions are associated with variable phenotypic expression.
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Proteínas de Ciclo Celular/genética , Síndrome de DiGeorge/genética , Complejo GPIb-IX de Glicoproteína Plaquetaria/genética , Septinas/genética , Translocación Genética/genética , Adolescente , Adulto , Niño , Deleción Cromosómica , Cromosomas Humanos Par 18/genética , Cromosomas Humanos Par 22/genética , Síndrome de DiGeorge/patología , Femenino , Predisposición Genética a la Enfermedad , Humanos , Masculino , Fenotipo , Adulto JovenRESUMEN
BACKGROUND: Tumor heterogeneity underlies resistance and disease progression in glioblastoma (GBM), and tumors most commonly recur adjacent to the surgical resection margins in contrast non-enhancing (NE) regions. To date, no targeted therapies have meaningfully altered overall patient survival in the up-front setting. The aim of this study was to characterize intratumoral heterogeneity in recurrent GBM using bulk samples from primary resection and recurrent samples taken from contrast-enhancing (EN) and contrast NE regions. METHODS: Whole exome and RNA sequencing were performed on matched bulk primary and multiple recurrent EN and NE tumor samples from 16 GBM patients who received standard of care treatment alone or in combination with investigational clinical trial regimens. RESULTS: Private mutations emerge across multi-region sampling in recurrent tumors. Genomic clonal analysis revealed increased enrichment in gene alterations regulating the G2M checkpoint, Kras signaling, Wnt signaling, and DNA repair in recurrent disease. Subsequent functional studies identified augmented PI3K/AKT transcriptional and protein activity throughout progression, validated by phospho-protein levels. Moreover, a mesenchymal transcriptional signature was observed in recurrent EN regions, which differed from the proneural signature in recurrent NE regions. CONCLUSIONS: Subclonal populations observed within bulk resected primary GBMs transcriptionally evolve across tumor recurrence (EN and NE regions) and exhibit aberrant gene expression of common signaling pathways that persist despite standard or targeted therapy. Our findings provide evidence that there are both adaptive and clonally mediated dependencies of GBM on key pathways, such as the PI3K/AKT axis, for survival across recurrences.
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Circular RNAs (circRNAs) are evolutionarily conserved RNA species that are formed when exons "back-splice" to each other. Current computational algorithms to detect these back-splicing junctions produce divergent results, and hence there is a need for a method to distinguish true-positive circRNAs. To this end, we developed Assembly based CircRNA Validator (ACValidator) for in silico verification of circRNAs. ACValidator extracts reads from a user-defined window on either side of a circRNA junction and assembles them to generate contigs. These contigs are aligned against the circRNA sequence to find contigs spanning the back-spliced junction. When evaluated on simulated datasets, ACValidator achieved over â¼80% sensitivity on datasets with an average of 10 circRNA-supporting reads and with read lengths of at least 100 bp. In experimental datasets, ACValidator produced higher verification percentages for samples treated with ribonuclease R compared to nontreated samples. Our workflow is applicable to non-polyA-selected RNAseq datasets and can also be used as a candidate selection strategy for prioritizing experimental validations. All workflow scripts are freely accessible on our GitHub page https://github.com/tgen/ACValidator along with detailed instructions to set up and run ACValidator.
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BACKGROUND: Molecular analysis has revealed four subtypes of pancreatic ductal adenocarcinoma (PDAC). One subtype identified for the presence of DNA damage repair deficiency can be targeted therapeutically with the poly (ADP-ribose) polymerase (PARP) inhibitor olaparib. We performed a single institution retrospective analysis of treatment response in patients with PDAC treated with olaparib who have DNA damage repair deficiency mutations. SUBJECTS, MATERIALS, AND METHODS: Patients with germline or somatic mutations involving the DNA repair pathway were identified and treated with olaparib. The primary objective was to examine the objective response rate (ORR). The secondary objectives were assessing tolerability, overall survival, and change in cancer antigen 19-9. Quantitative texture analysis (QTA) was evaluated from CT scans to explore imaging biomarkers. RESULTS: Thirteen individuals with metastatic PDAC were treated with Olaparib. The ORR to Olaparib was 23%. Median overall survival (OS) was 16.47 months. Four of seven patients with BRCA mutations had an effect on RAD51 binding, with a median OS of 24.60 months. Exploratory analysis of index lesions using QTA revealed correlations between lesion texture and OS (hepatic lesion tumor texture correlation coefficient [CC], 0.683, p = .042) and time on olaparib (primary pancreatic lesion tumor texture CC, 0.778, p = .023). CONCLUSION: In individuals with metastatic PDAC who have mutations involved in DNA repair, Olaparib may provide clinical benefit. BRCA mutations affecting RAD51 binding domains translated to improved median OS. QTA of individual tumors may allow for additional information that predicts outcomes to treatment with PARP inhibitors. IMPLICATIONS FOR PRACTICE: Pursuing germline and somatic DNA sequencing in individuals with pancreatic ductal adenocarcinoma may yield abnormalities in DNA repair pathways. These individuals may receive benefit with poly (ADP-ribose) polymerase (PARP) inhibition. Radiomics and deep sequencing analysis may yet uncover additional information that may predict outcome to treatment with PARP inhibitors.
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Reparación del ADN/genética , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Adulto , Anciano , Humanos , Persona de Mediana Edad , Mutación , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Estudios RetrospectivosRESUMEN
Glioma is recognized to be a highly heterogeneous CNS malignancy, whose diverse cellular composition and cellular interactions have not been well characterized. To gain new clinical- and biological-insights into the genetically-bifurcated IDH1 mutant (mt) vs wildtype (wt) forms of glioma, we integrated data from protein, genomic and MR imaging from 20 treatment-naïve glioma cases and 16 recurrent GBM cases. Multiplexed immunofluorescence (MxIF) was used to generate single cell data for 43 protein markers representing all cancer hallmarks, Genomic sequencing (exome and RNA (normal and tumor) and magnetic resonance imaging (MRI) quantitative features (protocols were T1-post, FLAIR and ADC) from whole tumor, peritumoral edema and enhancing core vs equivalent normal region were also collected from patients. Based on MxIF analysis, 85,767 cells (glioma cases) and 56,304 cells (GBM cases) were used to generate cell-level data for 24 biomarkers. K-means clustering was used to generate 7 distinct groups of cells with divergent biomarker profiles and deconvolution was used to assign RNA data into three classes. Spatial and molecular heterogeneity metrics were generated for the cell data. All features were compared between IDH mt and IDHwt patients and were finally combined to provide a holistic/integrated comparison. Protein expression by hallmark was generally lower in the IDHmt vs wt patients. Molecular and spatial heterogeneity scores for angiogenesis and cell invasion also differed between IDHmt and wt gliomas irrespective of prior treatment and tumor grade; these differences also persisted in the MR imaging features of peritumoral edema and contrast enhancement volumes. A coherent picture of enhanced angiogenesis in IDHwt tumors was derived from multiple platforms (genomic, proteomic and imaging) and scales from individual proteins to cell clusters and heterogeneity, as well as bulk tumor RNA and imaging features. Longer overall survival for IDH1mt glioma patients may reflect mutation-driven alterations in cellular, molecular, and spatial heterogeneity which manifest in discernable radiological manifestations.
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Glioma/genética , Isocitrato Deshidrogenasa/genética , Adulto , Anciano , Biomarcadores de Tumor/genética , Neoplasias Encefálicas/patología , Estudios de Casos y Controles , Femenino , Técnica del Anticuerpo Fluorescente/métodos , Heterogeneidad Genética , Humanos , Isocitrato Deshidrogenasa/metabolismo , Imagen por Resonancia Magnética/métodos , Masculino , Persona de Mediana Edad , Mutación , Clasificación del Tumor , Proteómica , Análisis de Secuencia de ARN/métodos , Análisis de la Célula Individual , Secuenciación del Exoma/métodosRESUMEN
Circular RNAs (circRNAs) are a class of non-coding RNAs involved in functions including micro-RNA (miRNA) regulation, mediation of protein-protein interactions, and regulation of parental gene transcription. In classical next generation RNA sequencing (RNA-seq), circRNAs are typically overlooked as a result of poly-A selection during construction of mRNA libraries, or are found at very low abundance, and are therefore difficult to isolate and detect. Here, a circRNA library construction protocol was optimized by comparing library preparation kits, pre-treatment options and various total RNA input amounts. Two commercially available whole transcriptome library preparation kits, with and without RNase R pre-treatment, and using variable amounts of total RNA input (1 to 4 µg), were tested. Lastly, multiple tissue types; including liver, lung, lymph node, and pancreas; as well as multiple brain regions; including the cerebellum, inferior parietal lobe, middle temporal gyrus, occipital cortex, and superior frontal gyrus; were compared to evaluate circRNA abundance across tissue types. Analysis of the generated RNA-seq data using six different circRNA detection tools (find_circ, CIRI, Mapsplice, KNIFE, DCC, and CIRCexplorer) revealed that a stranded total RNA library preparation kit with RNase R pre-treatment and 4 µg RNA input is the optimal method for identifying the highest relative number of circRNAs. Consistent with previous findings, the highest enrichment of circRNAs was observed in brain tissues compared to other tissue types.
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Encéfalo/fisiología , ARN Circular/genética , Análisis de Secuencia de ARN/métodos , Secuencia de Bases , Biblioteca de Genes , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , MicroARNs/genética , ARN Mensajero/genética , Transcriptoma/genética , Secuenciación del Exoma/métodosRESUMEN
Importance: Pediatric cancers are epigenetic diseases; therefore, considering tumor gene expression information is necessary for a complete understanding of the tumorigenic processes. Objective: To evaluate the feasibility and utility of incorporating comparative gene expression information into the precision medicine framework for difficult-to-treat pediatric and young adult patients with cancer. Design, Setting, and Participants: This cohort study was conducted as a consortium between the University of California, Santa Cruz (UCSC) Treehouse Childhood Cancer Initiative and clinical genomic trials. RNA sequencing (RNA-Seq) data were obtained from the following 4 clinical sites and analyzed at UCSC: British Columbia Children's Hospital (n = 31), Lucile Packard Children's Hospital at Stanford University (n = 80), CHOC Children's Hospital and Hyundai Cancer Institute (n = 46), and the Pacific Pediatric Neuro-Oncology Consortium (n = 24). The study dates were January 1, 2016, to March 22, 2017. Exposures: Participants underwent tumor RNA-Seq profiling as part of 4 separate clinical trials at partner hospitals. The UCSC either downloaded RNA-Seq data from a partner institution for analysis in the cloud or provided a Docker pipeline that performed the same analysis at a partner institution. The UCSC then compared each participant's tumor RNA-Seq profile with more than 11â¯000 uniformly analyzed tumor profiles from pediatric and young adult patients with cancer, downloaded from public data repositories. These comparisons were used to identify genes and pathways that are significantly overexpressed in each patient's tumor. Results of the UCSC analysis were presented to clinical partners. Main Outcomes and Measures: Feasibility of a third-party institution (UCSC Treehouse Childhood Cancer Initiative) to obtain tumor RNA-Seq data from patients, conduct comparative analysis, and present analysis results to clinicians; and proportion of patients for whom comparative tumor gene expression analysis provided useful clinical and biological information. Results: Among 144 samples from children and young adults (median age at diagnosis, 9 years; range, 0-26 years; 72 of 118 [61.0%] male [26 patients sex unknown]) with a relapsed, refractory, or rare cancer treated on precision medicine protocols, RNA-Seq-derived gene expression was potentially useful for 99 of 144 samples (68.8%) compared with DNA mutation information that was potentially useful for only 34 of 74 samples (45.9%). Conclusions and Relevance: This study's findings suggest that tumor RNA-Seq comparisons may be feasible and highlight the potential clinical utility of incorporating such comparisons into the clinical genomic interpretation framework for difficult-to-treat pediatric and young adult patients with cancer. The study also highlights for the first time to date the potential clinical utility of harmonized publicly available genomic data sets.
Asunto(s)
Neoplasias/genética , ARN Neoplásico/análisis , Análisis de Secuencia de ARN , Canadá , Niño , Preescolar , Femenino , Expresión Génica , Humanos , Lactante , Recién Nacido , Masculino , Medicina de Precisión , Estados Unidos , Adulto JovenRESUMEN
PURPOSE: Naturally occurring primary canine lung cancers share clinicopathologic features with human lung cancers in never-smokers, but the genetic underpinnings of canine lung cancer are unknown. We have charted the genomic landscape of canine lung cancer and performed functional characterization of novel, recurrent HER2 (ERBB2) mutations occurring in canine pulmonary adenocarcinoma (cPAC). EXPERIMENTAL DESIGN: We performed multiplatform genomic sequencing of 88 primary canine lung tumors or cell lines. Additionally, in cPAC cell lines, we performed functional characterization of HER2 signaling and evaluated mutation-dependent HER2 inhibitor drug dose-response. RESULTS: We discovered somatic, coding HER2 point mutations in 38% of cPACs (28/74), but none in adenosquamous (cPASC, 0/11) or squamous cell (cPSCC, 0/3) carcinomas. The majority (93%) of HER2 mutations were hotspot V659E transmembrane domain (TMD) mutations comparable to activating mutations at this same site in human cancer. Other HER2 mutations were located in the extracellular domain and TMD. HER2 V659E was detected in the plasma of 33% (2/6) of dogs with localized HER2 V659E tumors. HER2 V659E cPAC cell lines displayed constitutive phosphorylation of AKT and significantly higher sensitivity to the HER2 inhibitors lapatinib and neratinib relative to HER2-wild-type cell lines (IC50 < 200 nmol/L in HER2 V659E vs. IC50 > 2,500 nmol/L in HER2 WT). CONCLUSIONS: This study creates a foundation for molecular understanding of and drug development for canine lung cancer. These data also establish molecular contexts for comparative studies in dogs and humans of low mutation burden, never-smoker lung cancer, and mutant HER2 function and inhibition.