Impact of Sodium Thiosulfate on Prevention of Nephrotoxicities in HIPEC: An Ancillary Evaluation of Cisplatin-Induced Toxicities in Ovarian Cancer.

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.

A virome-wide clonal integration analysis platform for discovering cancer viral etiology.

Oncoviral infection is responsible for 12%-15% of cancer in humans. Convergent evidence from epidemiology, pathology, and oncology suggests that new viral etiologies for cancers remain to be discovered. Oncoviral profiles can be obtained from cancer genome sequencing data; however, widespread viral sequence contamination and noncausal viruses complicate the process of identifying genuine oncoviruses. Here, we propose a novel strategy to address these challenges by performing virome-wide screening of early-stage clonal viral integrations. To implement this strategy, we developed VIcaller, a novel platform for identifying viral integrations that are derived from any characterized viruses and shared by a large proportion of tumor cells using whole-genome sequencing (WGS) data. The sensitivity and precision were confirmed with simulated and benchmark cancer data sets. By applying this platform to cancer WGS data sets with proven or speculated viral etiology, we newly identified or confirmed clonal integrations of hepatitis B virus (HBV), human papillomavirus (HPV), Epstein-Barr virus (EBV), and BK Virus (BKV), suggesting the involvement of these viruses in early stages of tumorigenesis in affected tumors, such as HBV in TERT and KMT2B (also known as MLL4) gene loci in liver cancer, HPV and BKV in bladder cancer, and EBV in non-Hodgkin's lymphoma. We also showed the capacity of VIcaller to identify integrations from some uncharacterized viruses. This is the first study to systematically investigate the strategy and method of virome-wide screening of clonal integrations to identify oncoviruses. Searching clonal viral integrations with our platform has the capacity to identify virus-caused cancers and discover cancer viral etiologies.

A phenotypically diverse family with an atypical 22q11.2 deletion due to an unbalanced 18q23;22q11.2 translocation.

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.

An Analysis of Patients with DNA Repair Pathway Mutations Treated with a PARP Inhibitor.

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.

Integrated genomic analyses reveal frequent TERT aberrations in acral melanoma.

Genomic analyses of cutaneous melanoma (CM) have yielded biological and therapeutic insights, but understanding of non-ultraviolet (UV)-derived CMs remains limited. Deeper analysis of acral lentiginous melanoma (ALM), a rare sun-shielded melanoma subtype associated with worse survival than CM, is needed to delineate non-UV oncogenic mechanisms. We thus performed comprehensive genomic and transcriptomic analysis of 34 ALM patients. Unlike CM, somatic alterations were dominated by structural variation and absence of UV-derived mutation signatures. Only 38% of patients demonstrated driver BRAF/NRAS/NF1 mutations. In contrast with CM, we observed PAK1 copy gains in 15% of patients, and somatic TERT translocations, copy gains, and missense and promoter mutations, or germline events, in 41% of patients. We further show that in vitro TERT inhibition has cytotoxic effects on primary ALM cells. These findings provide insight into the role of TERT in ALM tumorigenesis and reveal preliminary evidence that TERT inhibition represents a potential therapeutic strategy in ALM.

Comprehensive molecular profiling of 718 Multiple Myelomas reveals significant differences in mutation frequencies between African and European descent cases.

Multiple Myeloma (MM) is a plasma cell malignancy with significantly greater incidence and mortality rates among African Americans (AA) compared to Caucasians (CA). The overall goal of this study is to elucidate differences in molecular alterations in MM as a function of self-reported race and genetic ancestry. Our study utilized somatic whole exome, RNA-sequencing, and correlated clinical data from 718 MM patients from the Multiple Myeloma Research Foundation CoMMpass study Interim Analysis 9. Somatic mutational analyses based upon self-reported race corrected for ancestry revealed significant differences in mutation frequency between groups. Of interest, BCL7A, BRWD3, and AUTS2 demonstrate significantly higher mutation frequencies among AA cases. These genes are all involved in translocations in B-cell malignancies. Moreover, we detected a significant difference in mutation frequency of TP53 and IRF4 with frequencies higher among CA cases. Our study provides rationale for interrogating diverse tumor cohorts to best understand tumor genomics across populations.

A pilot precision medicine trial for children with diffuse intrinsic pontine glioma-PNOC003: A report from the Pacific Pediatric Neuro-Oncology Consortium.

This clinical trial evaluated whether whole exome sequencing (WES) and RNA sequencing (RNAseq) of paired normal and tumor tissues could be incorporated into a personalized treatment plan for newly diagnosed patients (<25 years of age) with diffuse intrinsic pontine glioma (DIPG). Additionally, whole genome sequencing (WGS) was compared to WES to determine if WGS would further inform treatment decisions, and whether circulating tumor DNA (ctDNA) could detect the H3K27M mutation to allow assessment of therapy response. Patients were selected across three Pacific Pediatric Neuro-Oncology Consortium member institutions between September 2014 and January 2016. WES and RNAseq were performed at diagnosis and recurrence when possible in a CLIA-certified laboratory. Patient-derived cell line development was attempted for each subject. Collection of blood for ctDNA was done prior to treatment and with each MRI. A specialized tumor board generated a treatment recommendation including up to four FDA-approved agents based upon the genomic alterations detected. A treatment plan was successfully issued within 21 business days from tissue collection for all 15 subjects, with 14 of the 15 subjects fulfilling the feasibility criteria. WGS results did not significantly deviate from WES-based therapy recommendations; however, WGS data provided further insight into tumor evolution and fidelity of patient-derived cell models. Detection of the H3F3A or HIST1H3B K27M (H3K27M) mutation using ctDNA was successful in 92% of H3K27M mutant cases. A personalized treatment recommendation for DIPG can be rendered within a multicenter setting using comprehensive next-generation sequencing technology in a clinically relevant timeframe.

Comprehensive Genomic Profiling of Hodgkin Lymphoma Reveals Recurrently Mutated Genes and Increased Mutation Burden.

BACKGROUND: The genomic landscape of Hodgkin lymphoma (HL) has been difficult to characterize due to the paucity of neoplastic cells and an abundant microenvironment. Such characterization is needed in order to improve treatment strategies. MATERIALS AND METHODS: We performed comprehensive genomic profiling (CGP) using targeted next-generation sequencing on archival formalin-fixed paraffin embedded tumor samples from 63 patients to analyze the landscape of HL. RESULTS: CGP was successful for 49/63 archival specimens (78%), and revealed aberrations impacting genes including B2M, TP53, and XPO1 (E571). Of the 34 patients for whom total mutation burden (TMB; mutations/megabase [Mb]) was assessed, 5 (15%) had high TMB (≥20 mutations/Mb), 18 (53%) had intermediate TMB (6-19 mutations/Mb), and 11 (32%) had low TMB (≤5 mutations/Mb). We next tested 13 patients' plasma cell-free DNA with droplet digital polymerase chain reaction for the presence of XPO1 E571 mutation, which was confirmed in the plasma of 31% of patients. In three patients with serially collected plasma samples, XPO1 E571K allelic frequency changes corresponded with changes in tumor size on conventional radiographic imaging. CONCLUSION: The study demonstrates that comprehensive genomic profiling of archival Hodgkin lymphoma tumor samples is feasible and leads to the identification of genes that are recurrently mutated and that Hodgkin lymphoma has increased mutation burden in the majority of samples analyzed. Furthermore, tracking of XPO1 E571 mutant allele frequency in a subset of patients may also represent a potential disease-monitoring strategy and warrants further investigation. IMPLICATIONS FOR PRACTICE: This study provides the first evidence that comprehensive genomic profiling can be performed to map the genomic landscape of Hodgkin lymphoma and that a subpopulation of patients has mutations in TP53, B2M, XPO1, and other genes. It was found that 15% of patients have high mutation burden, which, in cancers such as melanoma, may indicate sensitivity to immune checkpoint inhibitors, and may thus be explored for Hodgkin lymphoma. Lastly, this work demonstrates that changes in the mutant allele frequency of XPO1 in serially collected plasma cell-free DNA samples correspond with treatment outcomes measured with conventional radiographic imaging.

E6201, an intravenous MEK1 inhibitor, achieves an exceptional response in BRAF V600E-mutated metastatic malignant melanoma with brain metastases.

Malignant melanoma (MM) exhibits a high propensity for central nervous system dissemination with ~50% of metastatic MM patients developing brain metastases (BM). Targeted therapies and immune checkpoint inhibitors have improved overall survival for MM patients with BM. However, responses are usually of short duration and new agents that effectively penetrate the blood brain barrier (BBB) are needed. Here, we report a MM patient with BM who experienced an exceptional response to E6201, an ATP-competitive MEK1 inhibitor, on a Phase 1 study, with ongoing near-complete response and overall survival extending beyond 8 years. Whole exome and transcriptome sequencing revealed a high mutational burden tumor (22 mutations/Megabase) with homozygous BRAF V600E mutation. Correlative preclinical studies demonstrated broad activity for E6201 across BRAF V600E mutant melanoma cell lines and effective BBB penetration in vivo. Together, these results suggest that E6201 may represent a potential new treatment option for BRAF-mutant MM patients with BM.

Circular RNA expression and regulatory network prediction in posterior cingulate astrocytes in elderly subjects.

BACKGROUND: Circular RNAs (circRNAs) are a novel class of endogenous, non-coding RNAs that form covalently closed continuous loops and that are both highly conserved and abundant in the mammalian brain. A role for circRNAs in sponging microRNAs (miRNAs) has been proposed, but the circRNA-miRNA-mRNA interaction networks in human brain cells have not been defined. Therefore, we identified circRNAs in RNA sequencing data previously generated from astrocytes microdissected from the posterior cingulate (PC) of Alzheimer's disease (AD) patients (N = 10) and healthy elderly controls (N = 10) using four circRNA prediction algorithms - CIRI, CIRCexplorer, find_circ and KNIFE. RESULTS: Overall, utilizing these four tools, we identified a union of 4438 unique circRNAs across all samples, of which 70.3% were derived from exonic regions. Notably, the widely reported CDR1as circRNA was detected in all samples across both groups by find_circ. Given the putative miRNA regulatory function of circRNAs, we identified potential miRNA targets of circRNAs, and further, delineated circRNA-miRNA-mRNA networks using in silico methods. Pathway analysis of the genes regulated by these miRNAs identified significantly enriched immune response pathways, which is consistent with the known function of astrocytes as immune sensors in the brain. CONCLUSIONS: In this study, we performed circRNA detection on cell-specific transcriptomic data and identified potential circRNA-miRNA-mRNA regulatory networks in PC astrocytes. Given the known function of astrocytes in cerebral innate immunity and our identification of significantly enriched immune response pathways, the circRNAs we identified may be associated with such key functions. While we did not detect recurrent differentially expressed circRNAs in the context of healthy controls or AD, we report for the first time circRNAs and their potential regulatory impact in a cell-specific and region-specific manner in aged subjects. These predicted regulatory network and pathway analyses may help provide new insights into transcriptional regulation in the brain.

Oligomeric amyloid ß preferentially targets neuronal and not glial mitochondrial-encoded mRNAs.

INTRODUCTION: Our laboratories have demonstrated that accumulation of oligomeric amyloid ß (OAß) in neurons is an essential step leading to OAß-mediated mitochondrial dysfunction. METHODS: Alzheimer's disease (AD) and matching control hippocampal neurons, astrocytes, and microglia were isolated by laser-captured microdissection from the same subjects, followed by whole-transcriptome sequencing. Complementary in vitro work was performed in OAß-treated differentiated SH-SY5Y, followed by the use of a novel CoQ10 analogue for protection. This compound is believed to be effective both in suppressing reactive oxygen species and also functioning in mitochondrial electron transport. RESULTS: We report decreases in the same mitochondrial-encoded mRNAs in Alzheimer's disease laser-captured CA1 neurons and in OAß-treated SH-SY5Y cells, but not in laser-captured microglia and astrocytes. Pretreatment with a novel CoQ10 analogue, protects neuronal mitochondria from OAß-induced mitochondrial changes. DISCUSSION: Similarity of expression changes in neurons from Alzheimer's disease brain and neuronal cells treated with OAß, and the effect of a CoQ10 analogue on the latter, suggests a pretreatment option to prevent OAß toxicity, long before the damage is apparent.

Integrated genomic characterization reveals novel, therapeutically relevant drug targets in FGFR and EGFR pathways in sporadic intrahepatic cholangiocarcinoma.

Advanced cholangiocarcinoma continues to harbor a difficult prognosis and therapeutic options have been limited. During the course of a clinical trial of whole genomic sequencing seeking druggable targets, we examined six patients with advanced cholangiocarcinoma. Integrated genome-wide and whole transcriptome sequence analyses were performed on tumors from six patients with advanced, sporadic intrahepatic cholangiocarcinoma (SIC) to identify potential therapeutically actionable events. Among the somatic events captured in our analysis, we uncovered two novel therapeutically relevant genomic contexts that when acted upon, resulted in preliminary evidence of anti-tumor activity. Genome-wide structural analysis of sequence data revealed recurrent translocation events involving the FGFR2 locus in three of six assessed patients. These observations and supporting evidence triggered the use of FGFR inhibitors in these patients. In one example, preliminary anti-tumor activity of pazopanib (in vitro FGFR2 IC50≈350 nM) was noted in a patient with an FGFR2-TACC3 fusion. After progression on pazopanib, the same patient also had stable disease on ponatinib, a pan-FGFR inhibitor (in vitro, FGFR2 IC50≈8 nM). In an independent non-FGFR2 translocation patient, exome and transcriptome analysis revealed an allele specific somatic nonsense mutation (E384X) in ERRFI1, a direct negative regulator of EGFR activation. Rapid and robust disease regression was noted in this ERRFI1 inactivated tumor when treated with erlotinib, an EGFR kinase inhibitor. FGFR2 fusions and ERRFI mutations may represent novel targets in sporadic intrahepatic cholangiocarcinoma and trials should be characterized in larger cohorts of patients with these aberrations.

Long insert whole genome sequencing for copy number variant and translocation detection.

As next-generation sequencing continues to have an expanding presence in the clinic, the identification of the most cost-effective and robust strategy for identifying copy number changes and translocations in tumor genomes is needed. We hypothesized that performing shallow whole genome sequencing (WGS) of 900-1000-bp inserts (long insert WGS, LI-WGS) improves our ability to detect these events, compared with shallow WGS of 300-400-bp inserts. A priori analyses show that LI-WGS requires less sequencing compared with short insert WGS to achieve a target physical coverage, and that LI-WGS requires less sequence coverage to detect a heterozygous event with a power of 0.99. We thus developed an LI-WGS library preparation protocol based off of Illumina's WGS library preparation protocol and illustrate the feasibility of performing LI-WGS. We additionally applied LI-WGS to three separate tumor/normal DNA pairs collected from patients diagnosed with different cancers to demonstrate our application of LI-WGS on actual patient samples for identification of somatic copy number alterations and translocations. With the evolution of sequencing technologies and bioinformatics analyses, we show that modifications to current approaches may improve our ability to interrogate cancer genomes.

Open-access synthetic spike-in mRNA-seq data for cancer gene fusions.

BACKGROUND: Oncogenic fusion genes underlie the mechanism of several common cancers. Next-generation sequencing based RNA-seq analyses have revealed an increasing number of recurrent fusions in a variety of cancers. However, absence of a publicly available gene-fusion focused RNA-seq data impedes comparative assessment and collaborative development of novel gene fusions detection algorithms. We have generated nine synthetic poly-adenylated RNA transcripts that correspond to previously reported oncogenic gene fusions. These synthetic RNAs were spiked at known molarity over a wide range into total RNA prior to construction of next-generation sequencing mRNA libraries to generate RNA-seq data. RESULTS: Leveraging a priori knowledge about replicates and molarity of each synthetic fusion transcript, we demonstrate utility of this dataset to compare multiple gene fusion algorithms' detection ability. In general, more fusions are detected at higher molarity, indicating that our constructs performed as expected. However, systematic detection differences are observed based on molarity or algorithm-specific characteristics. Fusion-sequence specific detection differences indicate that for applications where specific sequences are being investigated, additional constructs may be added to provide quantitative data that is specific for the sequence of interest. CONCLUSIONS: To our knowledge, this is the first publicly available synthetic RNA-seq data that specifically leverages known cancer gene-fusions. The proposed method of designing multiple gene-fusion constructs over a wide range of molarity allows granular performance analyses of multiple fusion-detection algorithms. The community can leverage and augment this publicly available data to further collaborative development of analytical tools and performance assessment frameworks for gene fusions from next-generation sequencing data.

Simultaneous characterization of somatic events and HPV-18 integration in a metastatic cervical carcinoma patient using DNA and RNA sequencing.

OBJECTIVE: Integration of carcinogenic human papillomaviruses (HPVs) into the host genome is a significant tumorigenic factor in specific cancers including cervical carcinoma. Although major strides have been made with respect to HPV diagnosis and prevention, identification and development of efficacious treatments for cervical cancer patients remains a goal and thus requires additional detailed characterization of both somatic events and HPV integration. Given this need, the goal of this study was to use the next generation sequencing to simultaneously evaluate somatic alterations and expression changes in a patient's cervical squamous carcinoma lesion metastatic to the lung and to detect and analyze HPV infection in the same sample. MATERIALS AND METHODS: We performed tumor and normal exome, tumor and normal shallow whole-genome sequencing, and RNA sequencing of the patient's lung metastasis. RESULTS: We generated over 1.2 billion mapped reads and identified 130 somatic point mutations and indels, 21 genic translocations, 16 coding regions demonstrating copy number changes, and over 36 genes demonstrating altered expression in the tumor (corrected P < 0.05). Sequencing also revealed the HPV type 18 (HPV-18) integration in the metastasis. Using both DNA and RNA reads, we pinpointed 3 major events indicating HPV-18 integration into an intronic region of chromosome 6p25.1 in the patient's tumor and validated these events with Sanger sequencing. This integration site has not been reported for HPV-18. CONCLUSIONS: We demonstrate that DNA and RNA sequencing can be used to concurrently characterize somatic alterations and expression changes in a biopsy and delineate HPV integration at base resolution in cervical cancer. Further sequencing will allow us to better understand the molecular basis of cervical cancer pathogenesis.

Emerging therapeutic drug monitoring technologies: considerations and opportunities in precision medicine.

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.

Multiomic sequencing of paired primary and metastatic small bowel carcinoids.

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.

A public resource of single cell transcriptomes and multiscale networks from persons with and without Alzheimer's disease.

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.

Assessing brain and biological aging trajectories associated with Alzheimer's disease.

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.

Posterior cingulate cortex reveals an expression profile of resilience in cognitively intact elders.

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.