bluuebunny/crossref_metadata_2025_split huggingface dataset:
Proteogenomic detection of circulating biomarkers for clinical oncology.
Authors: Quan Qimin , Ritchey Joshua , Wilkinson Joe , Kaiser Alaina , Geanacopoulos John , Boyce John 
Published: June 2022

e15010 Background: Circulating biomarkers have the potential to detect cancer in its earliest stages and monitor patients in remission. The integration of proteogenomics in circulating biomarkers may transform the molecular diagnostics of cancer and accelerate basic and clinical oncology research. Proteomics bridges the gaps of functional information lost due to post-transcriptional and post-translational modifications in a genomic approach. A recent study showed that adding just 8 protein biomarkers to a panel of circulating DNA biomarkers increased the diagnostic accuracy up to 98% sensitivity and 99% specificity. However, the proteogenomic approach normally requires the use of multiple different assay technologies and laboratory workflows, including mass spectrometry. Methods: MosaicNeedles are densely integrated nanoneedle sensors fabricated on a planar substrate that integrates proteogenomic analysis in one platform. 94,000 sensors with more than 2 billion total nanoneedles can be integrated on to a standard SBS plate, which can be configured into 96, 384 or 1536 well format. Each sensor contains an array of nanoneedles, dedicated to detecting one analyte of interest. All nanoneedles comprising the same sensor are functionalized with the same capture probes. The capture probe can be either an antibody for protein detection or an oligonucleotide with a specific target sequence to a DNA fragment, mRNA, or miRNA of interest. Results: At low analyte concentration, the binding of proteins to the nanoneedles follows a Poisson distribution. Therefore, statistically, no more than one molecule is bound per nanoneedle. A further addition of aptamers or antibodies will form a sandwich complex with the target analyte. Since each of the nanoneedles has an intrinsic optical resonance spectrum and will red-shift as the sandwich complex forms on its surface, the number of analytes can be quantitated by simply counting the number of nanoneedles that display a color change. At high analyte concentration, each nanoneedle has more than one analyte, so the number of analytes can be calculated by averaging the spectrum shifts of all nanoneedles. This combined single molecule counting (digital) and spectrum shift (analog) analysis allows the platform to detect both high abundance and low abundance protein analytes in one reaction. A 10,000-plex study can be achieved with a total of 2.5 billion nanoneedles on a 50mm by 50mm consumable. In this consumable, a 2,000-plex proteome and 8,000 cell-free DNA fragments can be detected. Conclusions: A full proteogenomic quantification can be performed on the NanoMosaic platform in one reaction with high sensitivity and large dynamic range. It simplifies the workflow and allows users to integrate proteomic and genomic information to discover new circulating biomarkers.
How circulating cancer cells disguise: The role of platelets.
Authors: Schott Dorothea , Pizon Monika , Pachmann Ulrich A. , Schill Erika , Pachmann Katharina 
Published: June 2022

e15011 Background: Solid tumors are notorious for their ability to form lethal metastases, sometimes several decades following initial cancer diagnosis. Development of distant metastases is a result of the primary tumor shedding cells that travel via lymphatics and the blood to distant sites where they can form metastases. Platelets are known to specifically enhance tumor cells’ survival in the bloodstream by as yet poorly understood mechanisms. To study the interplay of platelets with circulating tumor cells, we implemented our published approach to label both circulating epithelial tumor cells and platelets. Methods: Blood samples were collected from 86 non-metastatic breast cancer patients avoiding fixation and processed at 4 time points following blood collection using the maintrac approach. Samples were additionally stained with anti-CD36 antibody to visualize platelets. Results: Typically, we observed at 0h post-blood draw platelet aggregates strictly attached to single cells that did not stain with anti-EpCAM antibody. These clogged unstained cells were, by far, outnumbered by surrounding white blood cells with no platelets attached. After keeping blood samples at room temperature for 24h a completely different picture became apparent: there were still cells detectable with platelets attached to them but the underlying cell now became accessible to the anti-EpCAM antibody leading to green fluorescence staining. CETCs were detected in 94% patients on day 1 post-blood draw. The increase in single platelets in the background could be a sign that after 24h, platelets detach from CETCs. At 72 h following initial blood drawing, platelets had almost completely disappeared from the cell surface. Numbers of EpCAM-positive cells remained stable between days 1 and 3. No CETCs were observed in 2 individuals with no cancer diagnosis, neither on day zero, nor on day 1. Conclusions: Our results suggest that platelets play a key role in masking circulating tumor cells. Masking may explain the difficulties in detection of these cells and prevention of their elimination by the immune system. Our unmasking approach detects sufficient numbers of circulating tumor cells to monitor the effect on blood tumor cells of different therapeutic measures, thus contributing to improved systemic therapies for cancer.
Using nucleosome levels, copy number alterations, and DNA methylation to profile disease state through liquid biopsy.
Authors: Harrison Sarah , Poplawski Shane G. , Orlanski Shari , Wei Melvin , Atteberry Brandi , Berman Benjamin P. , Kelly Terry 
Published: June 2022

e15014 Background: Nucleosomes are the repeating unit of chromatin that contain important signals for proper genomic function and transcriptional activity. Upon cell death, chromatin is fragmented, and nucleosomes are released into circulation and thus, detectable in liquid biopsy. Cell death naturally occurs over time and increases in a variety of diseases resulting in elevated levels of circulating nucleosomes and the genetic and epigenetic signatures they carry. These signatures contain important information about the cells from which they were derived, as well as the reason for them being in circulation. Methods: We measured circulating nucleosome levels, using Volition’s Nu.Q H3.1 sandwich ELISA across a healthy cohort spanning 40-85 years of age as well as patients whom had been diagnosed with Non-Hodgkin's Lymphoma (NHL), some of which were untreated and others that had undergone treatment. Additionally, to demonstrate that the correlation between nucleosome (Nu.Q) levels and copy number alterations is specific to cancer, we assessed the genomic integrity of an NHL sample with a Nu.Q level of 744 ng/ml and a sepsis sample with Nu.Q level of 3,281 ng/ml using low coverage Oxford Nanopore whole genome sequencing to identify genomic amplifications and deletions. We further used cell type specific methylation patterns to identify the cell of origin of cfDNA in each sample. Results: Importantly, we found no significant correlation between nucleosome level and age in our healthy cohort (R2= 0.0004; 40-49: N = 10, mean = 39 ng/ml; 50-59: N = 10, mean = 18 ng/ml; 60-69: N = 10, mean = 35 ng/ml; 70-79: N = 15, mean = 34 ng/ml; 80-85: N = 5, mean = 32 ng/ml). We did however find elevated nucleosome levels in untreated NHL patients (N = 4, mean 646 ng/ml) and that nucleosome levels in NHL patients that had undergone treatment were reduced (N = 29, mean = 180 ng/ml). Using copy number changes, we found that the fraction of cfDNA estimated to be derived from the tumor in the NHL sample was 54% and were able to detect amplifications in chromosomes 10, 11 and 18 and deletions in chromosomes 1, 8, 17, 18 and 23, whereas we found no copy number alterations present in the cfDNA sample derived from a sepsis patient and a tumor fraction of 0. Furthermore, using cell type specific methylation patterns to deconvolute the data we found that a large fraction of the cfDNA in the NHL sample was derived from B-cells consistent with the NHL sample being of B-cell origin whereas we determined that the cell of origin of the cfDNA in the sepsis sample was Neutrophils. Conclusions: This data shows that nucleosome levels, as measured by Nu.Q, can be used in conjunction with shallow sequencing and DNA methylation profiles to define tumor fraction and cell of origin in liquid biopsy.
High-multiplexing quantitative CodePlex proteomic profiling of platelets in triple-negative breast cancer (TNBC) patients and healthy subjects.
Authors: Han Edward Xu , Gu Sean , Steinle Matthew , Gu Vivian , Tyagi Tarun , Mackay Sean , Hwa John , Zhou Jing 
Published: June 2022

e15017 Background: Platelets play significant roles in inflammation, immune regulation, and intracellular communication processes involved in tumor pathogenesis. Aberrant platelet activation has been associated with tumor progression and metastasis, however, cancer patient platelet deep profiling remains underutilized. New tools are needed for rapid high throughput quantification of potential altered platelet secretome in cancer patients. Such technology may provide novel insights into impactful cancer-associated biomarkers. Methods: We have developed a high throughput multiplexed proteomic technology to profile platelet cytokine content and releasates and performed a pilot study in healthy patients versus triple negative breast cancer (TNBC). Macrochambers of the CodePlex chip were pre-patterned with a 23-plex antibody IsoPlexis Human Adaptive Immune Panel: GM-CSF, Granzyme B, IFN-g, IL-10, IL-13, IL-15, IL-17A, IL-2, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IP-10, MCP-1, MIP-1a, MIP-1b, perforin, sCD137, TGF-b1, TNF-a, TNF-b. 2% BSA/PBS was used as background control. The chips are loaded into an IsoLight automation system, releasates measured by fluorescence ELISA, and analysis performed by IsoSpeak software. In a pilot study to test feasibility, platelets were meticulously harvested from triple negative breast cancer patients (n=2) and healthy controls (n=2) and stimulated for 30 minutes (ADP or TRAP-6) to promote cytokine release. Both the supernatants and platelet lysates were then loaded on CodePlex chips and analyzed. Results: The methodology of bulk proteomic measurements has previously been validated in a variety of biofluids such as serum/plasma, cerebrospinal fluid, tracheal washes, and cell culture supernatants. Freshly isolated platelets activated by TRAP6 generally released more cytokines than those activated by ADP stimulation in both healthy controls and disease samples. Interestingly, platelet releasates in supernatants from TNBC patients demonstrated increased pro-tumor IL-8 and IL-9, and reduced antitumor IFN-g compared to healthy controls. Higher levels of IL-8 and IL-9 and minimal IFN-g were detected at similar activation concentrations, in platelet lysates from both healthy and TNBC platelets. This suggests that TNBC may alter platelet content and/or activation. Increased numbers are being performed to validate these initial results. Conclusions: From this small pilot study, multiplexing quantitative proteomic analysis is potentially useful in detecting differential cytokine profile in platelet releasates and lysate contents between TNBC patients and healthy controls. Quantifying the platelet proteomes in cancer patients using high throughput proteomics may provide valuable insights to dissect cancer-platelet crosstalk and potential pathogenesis mechanisms.
Comprehensive genomic profiling: Insights into diagnostic value and treatment strategy.
Authors: Shiller Michelle , Zhang Haiying , Yin Lihui , Mukherjee Sandeep , Coldren Christopher , Mennel Robert Gary , Konduri Kartik , Wood David , Chandra Pranil 
Published: June 2022

e15028 Background: The clinical application of next-generation sequencing (NGS) has continued to advance precision oncology by enabling the identification of biomarkers that may predict potential response to targeted therapy, yet the benefit of tissue-based NGS to patient care is not solely confined to this narrow application. Not uncommonly, NGS can reveal genomic characteristics of the tumor that may be inconsistent with initial pathology review and clinical presentation. When this occurs, these genomic insights along with other clinical and diagnostic findings may be utilized to reconsider the initial diagnosis and seek to explore alternative diagnostic explanations that may be more consistent with the genomic findings. Methods: Patient diagnoses and clinical data were reviewed and compared against NGS results to determine if the genomic profile was consistent with and supports the initial diagnosis. If the genomic profile of the tumor was found to be inconsistent with the initial diagnosis, a secondary review of the case was initiated. Out of a cohort of >2,500 patient cases, notable examples of cases were identified that highlight the impact of NGS findings on the diagnostic classification of the tumor. Genomic profiling was conducted utilizing PathGroup’s molecular pathology-directed tumor profiling solution, Endeavor, which is powered by the FDA-cleared PGDx elio tissue complete test. Results: Among the identified patient cases where initial pathological diagnosis was altered following review of NGS results, several examples include 1) a sarcoma case being re-classified as melanoma, 2) a lung lesion initially diagnosed as NSCLC that was metastatic basal cell carcinoma and 3) a cancer of unknown primary that was determined to be a cholangiocarcinoma. Treating oncologists were kept apprised of all diagnostic findings and modifications to either the initial diagnosis or to the tumor classification which led to an altered treatment strategy in each case. Conclusions: NGS can be broadly applied throughout the cancer care continuum, with a new perspective on its potential value diagnostically. In these cases, re-evaluation and subsequent re-classification led to treatment strategy shifts that are more specific to the patient’s tumor type. These findings provide critical real-world evidence that supports the value of broad genomic profiling in complex cancer cases, especially when the genomic findings are inconsistent with the initial diagnosis, or the diagnosis is unclear.
Identification of clinically actionable biomarkers via routine comprehensive genomic profiling across a large community health system.
Authors: Weerasinghe Roshanthi K. , Meng Ryan , Dowdell Alexa K. , Bapat Bela , Vita Ann , Schroeder Brock , Stein Alisha , Harold Lauren , Schmidt Mark , Chang Shu-Ching , Ward Thomas , Wagner Josiah , Piotrowski Stanley , Febbo Phillip G. , Bifulco Carlo Bruno , Piening Brian 
Published: June 2022

e15035 Background: Precision therapies and immunotherapies have revolutionized cancer care, with novel genomic biomarker-associated therapies being introduced into clinical practice rapidly. We assessed the utility of comprehensive genomic profiling (CGP)-based testing for identifying biomarkers associated with approved therapies, and therapies in precision medicine basket clinical trials (CT) across a large cohort of advanced cancer patients in the Providence health system. Methods: Advanced cancer patients were tested utilizing the Providence CGP workflow between 2019-2021. Clinical actionability was assessed for CGP and compared with results from an in silico 50-gene panel based on a previously utilized lab-developed test at Providence. Clinical actionability was assessed based on OncoKB and alterations were assigned as: FDA recognized (Level 1), standard of care (Level 2), FDA approved/investigational drug in another indication or having compelling clinical evidence (Level 3). CT matching was assessed based on enrollment criteria for ASCO-TAPUR, NCI-MATCH and My Pathway CTs at time of testing. Pooled electronic medical record and genomic data were curated and standardized. Results: Of the 3,218 advanced cancer patients tested with CGP, 52% were female, 80% were white, and median age was 67 years. Across 31 tumor types, the most commonly tested were lung (26%), bowel/colon (16%), and breast (9%). Overall, 48% of patients tested with CGP harbored at least one actionable biomarker (OncoKB Levels1/2/3). Clinical actionability was significantly higher in the CGP cohort compared to the in silico cohort based on presence of at least one Level 1 biomarker (45% vs. 19%, p < 0.001). CGP cohort had higher proportion of patients with multiple/co-occurring Level 1 biomarkers compared to in silico cohort (20% vs. 9%, p < 0.001). Of the most prevalent tumor types, 57% lung, 94% bowel/colon, and 37% breast had Level 1 alterations with CGP testing. Notably, 49% of CGP cohort vs. 23% in silico cohort (p < 0.001) harbored a biomarker matching to one or more arms of the three basket CTs. Conclusions: CGP and small panel testing can both identify patients eligible for approved therapies and/or basket CTs with CGP having significantly higher clinical actionability and CT eligibility. We expect value of CGP to increase as biomarker actionability transforms clinical practice.
Utility and timeliness of somatic genetic testing for patients enrolled in the molecular screening and therapeutics (MoST) study at Princess Alexandra Hospital (PAH).
Authors: Watson Benjamin , Arneil Melissa , Leo Paul , O'Byrne Kenneth John , Atkinson Victoria 
Published: June 2022

e15039 Background: Next generation sequencing (NGS) technologies are increasingly employed to deliver optimal precision therapy for patients with cancer. The Molecular Screening and Therapeutics (MoST) study led by Prof David Thomas, Garvan Institute, Sydney, Australia, was established to identify patients for enrolment into clinical trials and for targeted or immune therapies not routinely indicated for their illness, specifically focusing on those with rare or neglected malignancies or with more common malignancies that had completed standard therapy. Methods: 111 patients were enrolled at the PAH between 1/5/2020 and 31/5/2021. Of these the results of the first 100 patients with complete medical records and sufficient tissue for NGS analysis are reported. Sources included electronic medical record, cancer state-wide database and the Molecular Tumor Board reports. The NGS Illumina TSO500 platform was employed for genetic testing. Determining utility of the results/reports included analyzing the proportion of patients with clinically relevant mutations detected, treatment recommendations, clinical trial recommendations and clinical trial enrolments. The timeframe from enrolment to results/recommendations conveyed to the patient was recorded. Demographic data was collected at enrolment. The final date of data collection was 31/12/2021, ensuring a minimum of six months follow-up. Local ethical regulations were complied with. Results: The study included 51 men and 49 women, ECOG 0-1, median age 57 years (range 21 – 84 years). Gastrointestinal malignancies were the most common, 31 patients, followed by sarcoma, 16, and breast cancer, 14. At enrolment, 64 patients were receiving chemotherapy, with the lines of treatments received ranging from 0 – 7, mean of 2.6. Mutations were detected in the following number of specimens: Tier 1A - 21, Tier 2C - 27, Tier 2D - 69 and Tier 3 - 69. High tumor mutational burden, defined as >10Muts/Mb, was present in 12 specimens and potential germline mutations were detected in 22 patients. There were 34 treatment recommendations and 27 clinical trial recommendations with 14 patients enrolled in clinical trials. The mean timeframe from enrolment to the result of testing being conveyed to the patient was 85.1 days. The overall median survival was 9.72 months. Conclusions: Results demonstrate that somatic genetic testing generates clinical trial, treatment and genetics health recommendations for a significant number of patients with 1:7 being recruited into clinical trials. Median length of time between recruitment and delivery of results to the patient was almost 3 months, which will improve through streamlining of clinical and logistical processes and advances in testing technologies, ensuring timely results for clinicians and their patients.
Comprehensive genomic and clinical analysis of cancer of unknown primary site in Chinese patients.
Authors: Xue Hongsheng , Yuan Shaohua , Chen Hui , Zhang Yuman , Wang Aodi , Mei Lei , Wang Liwei , Dong Xiaowei , Wang Kai 
Published: June 2022

e15071 Background: Cancer of unknown primary (CUP) comprises 2-5% of all diagnosed cancers worldwide and is one of most common cause of death. Deep understanding of CUP plays an imported role in improving the disease diagnoses and finding the primary sites. In this study, our data revealed the genomic and clinical features of CUP in Chinese patients. Methods: FFPE samples and matched peripheral blood were collected for NGS based 450 cancer genes panel assay. Somatic variants were analyzed following CAP/CLIA-certificated workflows, including Single Nucleotide Variant (SNV), Copy Number Variation (CNV), large insertion and deletions (Lindel) and gene rearrangements. MSI status and TMB were also acquired by an NGS algorithm in house. PD-L1 status was analyzed by IHC. Results: Of the 264 CUP patients, 54.2% were male and median age was 61 years old (range, 2-88 years old). The median value of TMB was 4.6 muts/Mb (range, 0-61.2). Only 1 patient was found MSI-H status with the TMB value of 61.2 muts/Mb. In addition, 23.9% and 28.4% of the CUP patients presented PDL1 positive (> 1%) and PDL1 negative respectively, with others unavailable. In the patients of available stage status, they were all advanced stage (stage ≥ III). Top metastatic sites were lymphaden (17.8%), liver(15.9%) and bone (15.5%). TP53 was the most common mutated gene (62.1%), followed by KRAS (22.3%), CDKN2A (15.9%), TERT (10.2%), EGFR (9.8%) and PIK3CA (9.8%). In the alterations of KRAS, G12D, G12V and G12C were occupied 32.2%, 28.8% and 8.4% respectively. Among the alterations of EGFR, 38.5% were L858R and 19.2% were 19del. Besides, amplification of MYC (4.9%) and 11q13 (5.3%) obviously appeared in our data. Fusion/Rearrangements of ALK and ROS1 accounted for 0.8% and 1.1% respectively . The gene alterations of tyrosine kinase were found in 48.4% of CUP patients such as ALK, BRAF, EGFR, FGFR1/2/3/4, RET, ROS1 and so on. PI3K−Akt signaling pathway and MAPK signaling pathway were the most significantly pathways (fdr < 0.001) enriched in our data. A CUP patient with positive PDL1 and high TMB achieved a 9-month PFS after treatment of TP chemotherapy and immunotherapy. Conclusions: In this study, we revealed comprehensive genomic and clinical features of cancer of unknown primary site in Chinese patients. It shows promise to identify the origin anatomic site of tumor while targeted therapeutic approaches to improve outcomes.