Cancer Genome Sequencing Research Articles

Base editing screens define the genetic landscape of cancer drug resistance mechanisms.

Drug resistance is a principal limitation to the long-term efficacy of cancer therapies. Cancer genome sequencing can retrospectively delineate the genetic basis of drug resistance, but this requires large numbers of post-treatment samples to nominate causal variants. Here we prospectively identify genetic mechanisms of resistance to ten oncology drugs from CRISPR base editing mutagenesis screens in four cancer cell lines using a guide RNA library predicted to install 32,476 variants in 11 cancer genes. We identify four functional classes of protein variants modulating drug sensitivity and use single-cell transcriptomics to reveal how these variants operate through distinct mechanisms, including eliciting a drug-addicted cell state. We identify variants that can be targeted with alternative inhibitors to overcome resistance and functionally validate an epidermal growth factor receptor (EGFR) variant that sensitizes lung cancer cells to EGFR inhibitors. Our variant-to-function map has implications for patient stratification, therapy combinations and drug scheduling in cancer treatment.

Shedding light on the DICER1 mutational spectrum of uncertain significance in malignant neoplasms.

The Dicer protein is an indispensable player in such fundamental cell pathways as miRNA biogenesis and regulation of protein expression in a cell. Most recently, both germline and somatic mutations in DICER1 have been identified in diverse types of cancers, which suggests Dicer mutations can lead to cancer progression. In addition to well-known hotspot mutations in RNAase III domains, DICER1 is characterized by a wide spectrum of variants in all the functional domains; most are of uncertain significance and unstated clinical effects. Moreover, various new somatic DICER1 mutations continuously appear in cancer genome sequencing. The latest contemporary methods of variant effect prediction utilize machine learning algorithms on bulk data, yielding suboptimal correlation with biological data. Consequently, such analysis should be conducted based on the functional and structural characteristics of each protein, using a well-grounded targeted dataset rather than relying on large amounts of unsupervised data. Domains are the functional and evolutionary units of a protein; the analysis of the whole protein should be based on separate and independent examinations of each domain by their evolutionary reconstruction. Dicer represents a hallmark example of a multidomain protein, and we confirmed the phylogenetic multidomain approach being beneficial for the clinical effect prediction of Dicer variants. Because Dicer was suggested to have a putative role in hematological malignancies, we examined variants of DICER1 occurring outside the well-known hotspots of the RNase III domain in this type of cancer using phylogenetic reconstruction of individual domain history. Examined substitutions might disrupt the Dicer function, which was demonstrated by molecular dynamic simulation, where distinct structural alterations were observed for each mutation. Our approach can be utilized to study other multidomain proteins and to improve clinical effect evaluation.

Performance of somatic structural variant calling in lung cancer using Oxford Nanopore sequencing technology

BackgroundLung cancer is a heterogeneous disease and the primary cause of cancer-related mortality worldwide. Somatic mutations, including large structural variants, are important biomarkers in lung cancer for selecting targeted therapy. Genomic studies in lung cancer have been conducted using short-read sequencing. Emerging long-read sequencing technologies are a promising alternative to study somatic structural variants, however there is no current consensus on how to process data and call somatic events. In this study, we preformed whole genome sequencing of lung cancer and matched non-tumour samples using long and short read sequencing to comprehensively benchmark three sequence aligners and seven structural variant callers comprised of generic callers (SVIM, Sniffles2, DELLY in generic mode and cuteSV) and somatic callers (Severus, SAVANA, nanomonsv and DELLY in somatic modes).ResultsDifferent combinations of aligners and variant callers influenced somatic structural variant detection. The choice of caller had a significant influence on somatic structural variant detection in terms of variant type, size, sensitivity, and accuracy. The performance of each variant caller was assessed by comparing to somatic structural variants identified by short-read sequencing. When compared to somatic structural variants detected with short-read sequencing, more events were detected with long-read sequencing. The mean recall of somatic variant events identified by long-read sequencing was higher for the somatic callers (72%) than generic callers (53%). Among the somatic callers when using the minimap2 aligner, SAVANA and Severus achieved the highest recall at 79.5% and 79.25% respectively, followed by nanomonsv with a recall of 72.5%.ConclusionLong-read sequencing can identify somatic structural variants in clincal samples. The longer reads have the potential to improve our understanding of cancer development and inform personalized cancer treatment.

Abstract A084: Journey to understanding indigenous peoples engagement in cancer genomic research

Abstract Introduction: Tribal Nations across the Southwest have striking disparities in cancer incidence and mortality. Compounded by disproportionately low rates of cancer screening and limited access to healthcare, Native Americans are more often diagnosed at later stages of disease and have the poorest outcomes in all types of cancer when compared to any other racial and ethnic group in the United States. At the University of New Mexico Comprehensive Cancer Center, the Participant Engagement Cancer Genomic Sequencing (PE-CGS) Center is developing culturally appropriate materials and strategies to enhance the engagement and recruitment of Native Americans into genomic research. This effort aims to determine participant knowledges, attitudes, cultural beliefs, and communication preferences regarding clinical genomics testing. Ultimately, targeting improvement of cancer outcomes in Southwestern Tribal communities. Cultural adaptations are crucial with regards to biospecimen collection, storage, and usage, due to historical mistrust of research among Native communities. (i.e., Havasupai Tribe v. the Arizona Board of Regents). A Tribal Advisory Committee, consisting of Tribal leaders, Native physicians, and patient advocates oversee all aspects of the research study. The goal is to identify appropriate optimized health literacy strategies for engaging Native peoples in cancer genomic research. Methods: The team conducted semi-structured discussions with participants to understand their perspectives on motivations to join, cultural safety components, and effectiveness of engagement strategies. With participant consent, the discussions were recorded and transcribed. Analysis included the transcripts and notes taken by the interviewer and followed Grounded Theory principles. All transcripts and notes followed line-by-line analysis by three of the authors (JS, JR and TG) who met regularly to identify and discuss emergent themes. Results: As of May 31, 2024, the PE-CGS study has enrolled 112 participants and completed 39 discussions. Participants emphasized altruistic motives tied to family and community, echoing traditional values, in their decision to participate in the study. This notion emerged during interactions with Native team members, where comfort stemmed from shared experiences (healthcare system, barriers), use of Native language, and adherence to Native social norms. Additional considerations included present autonomy throughout various stages of the study (sample use, data sharing, discussions). Return of genetic testing results improved understanding of treatment options, hereditary risk, and health literacy. Conclusion: Feedback highlights the importance of aligning cultural beliefs with Western medicine through a strengths-based approach. The findings will inform improvements in respectful engagement, communication, information exchange, and recruitment processes with Native American cancer patients and cancer survivors to genomic testing studies. Citation Format: Jessica Sanchez, Koby Blackwater, Joseph Rodman, Tamar Ginossar, Andrew Sussman, Shiraz Mishra, Cindy Blair, Mallery Quetawki, Debra MacKenzie, V. Shane Pankratz, Ursa Brown-Glaberman, Jeffery Trent, Cheryl Willman. Journey to understanding indigenous peoples engagement in cancer genomic research [abstract]. In: Proceedings of the 17th AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2024 Sep 21-24; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2024;33(9 Suppl):Abstract nr A084.

Reconstructing extrachromosomal DNA structural heterogeneity from long-read sequencing data using Decoil.

Circular extrachromosomal DNA (ecDNA) is a form of oncogene amplification found across cancer types and associated with poor outcome in patients. ecDNA can be structurally complex and can contain rearranged DNA sequences derived from multiple chromosome locations. As the structure of ecDNA can impact oncogene regulation and may indicate mechanisms of its formation, disentangling it at high resolution from sequencing data is essential. Even though methods have been developed to identify and reconstruct ecDNA in cancer genome sequencing, it remains challenging to resolve complex ecDNA structures, in particular amplicons with shared genomic footprints. We here introduce Decoil, a computational method that combines a breakpoint-graph approach with LASSO regression to reconstruct complex ecDNA and deconvolve co-occurring ecDNA elements with overlapping genomic footprints from long-read nanopore sequencing. Decoil outperforms de novo assembly and alignment-based methods in simulated long-read sequencing data for both simple and complex ecDNAs. Applying Decoil on whole-genome sequencing data uncovered different ecDNA topologies and explored ecDNA structure heterogeneity in neuroblastoma tumors and cell lines, indicating that this method may improve ecDNA structural analyses in cancer.

Chromatin remodellers as therapeutic targets.

Large-scale cancer genome sequencing studies have revealed that chromatin regulators are frequently mutated in cancer. In particular, more than 20% of cancers harbour mutations in genes that encode subunits of SWI/SNF (BAF) chromatin remodelling complexes. Additional links of SWI/SNF complexes to disease have emerged with the findings that some oncogenes drive transformation by co-opting SWI/SNF function and that germline mutations in select SWI/SNF subunits are the basis of several neurodevelopmental disorders. Other chromatin remodellers, including members of the ISWI, CHD and INO80/SWR complexes, have also been linked to cancer and developmental disorders. Consequently, therapeutic manipulation of SWI/SNF and other remodelling complexes has become of great interest, and drugs that target SWI/SNF subunits have entered clinical trials. Genome-wide perturbation screens in cancer cell lines with SWI/SNF mutations have identified additional synthetic lethal targets and led to further compounds in clinical trials, including one that has progressed to FDA approval. Here, we review the progress in understanding the structure and function of SWI/SNF and other chromatin remodelling complexes, mechanisms by which SWI/SNF mutations cause cancer and neurological diseases, vulnerabilities that arise because of these mutations and efforts to target SWI/SNF complexes and synthetic lethal targets for therapeutic benefit.

Uncovering of NRAGE/JNK and PI3K/AKT pathways through serial WGS of patients with breast cancer.

e12577 Background: Cancer genomic sequencing has fundamentally advanced our understanding of the basic biology of this disease, and more recently, has offered method to guide and evaluate treatment in clinic. We performed whole-genome sequencing (WGS) on three consecutive tissue and blood samples from breast cancer patients: diagnosis, post-neoadjuvant chemotherapy, recurrence after curative resection. Methods: We performed WGS to compare the genetic profiles of the primary, surgical and recurrent samples (tissue and blood) to determine: (1) how closely related the genetics are among the primary, post-neoadjuvant chemotherapy, and the recurrent tumors, (2) whether there are variations in mutational processes among the primary, post- neoadjuvant chemotherapy, and recurrent tumor. WGS was performed on three patients for whom both tissue and blood were available at diagnosis, at surgery after neoadjuvant chemotherapy and at a recurrence. Results: As a result of somatic protein coding variant analysis, PIK3CA mutation was confirmed at the time of diagnosis tissue in all cases. Mutations were found at p.E545K in patient 14, p.R916C in patient 18, and p.R916C in patient 3. Among these, patients 14 and 18 had PIK3CA mutations even at the time of recurrence. In patient 14, the same PIK3CA mutation was found in 3 consecutive samples, but in patient 18, it changed from p.R916C at diagnosis to p.G480E at relapse. Furthermore, functional enrichment analysis identified a common NRAGE signaling system in the cfDNA. The NRAG pathway was found in the blood at diagnosis, surgery, and recurrence in all three patients. Conclusions: This study was a proof-of-concept study, in which we hypothesized that NRAGE/JNK and PI3K/AKT signaling pathways are involved in breast cancer recurrence and treatment response through WGS of consecutive samples. Further research is ongoing with larger numbers of patients.

Approaches to engaging participants in a national cancer genomics research network.

11022 Background: Although patient engagement is thought to improve the relevance and acceptability of research across diverse patient populations, a paucity of studies actively evaluate engagement activities to assess their impact. The NCI’s Participant Engagement and Cancer Genome Sequencing (PE-CGS) Network was established to advance cancer genomics for understudied cancers and among underserved populations by engaging patients and survivors as participants in research. We sought to characterize strategies for engaging and evaluating engagement among research centers within PE-CGS, and to describe factors impacting each center’s unique engagement strategies and evaluation methods. Methods: We used a multi-method approach to document centers’ engagement strategies and methods to evaluate engagement.We first completed a document review of centers’ study materials including protocols, specific aims, and publicly available information. We then conducted semi-structured key informant interviews with research center members to validate findings from the document review and to identify factors that influenced engagement strategies. The semi-structured interview guide was developed using the Health Equity Implementation Framework to ensure that we accounted for multi-level factors (societal, contextual, and clinical factors) relevant to engagement. Engagement strategies and evaluation methods were collated and analyzed descriptively. Information about factors that influenced their engagement strategies were analyzed using deductive thematic analysis. Results: Across the five PE-CGS research centers, we identified 9 engagement strategies including: convening at network meetings (n=5), use of advisory boards (n=5), brochures (n=3), newsletters (n=2), community outreach through advocacy organizations (n=1) and in-person events (n=2), and use of branded websites (n=4), social media (n=3), and health technology apps (n=1). Methods used for evaluating engagement included: surveys (n=4), interviews/focus groups (n=4), informal feedback from participants (n=2), randomized experiments (n=1), and N-of-1 studies (n=1). Multi-level factors that influenced engagement included those related to the language and culture of the patient population, availability of validated measures, timing of interactions with participants within their health journey, and patient burden imposed by the experienced cancer. Conclusions: Across a national research network with a priority to advance participant engagement in cancer genomics research, we observed a range of unique engagement strategies and evaluation methods. Notably evaluation approaches were not typically tied to specific engagement strategies, but rather sought to evaluate broader aspects of engagement in the respective research studies, such as their ability to engender inclusivity, collaboration, and trust among participants.

Abstract 3932: Multi-omics characterization of molecular features and global-local genomic ancestry analysis of colorectal cancer in Hispanic-Latinos

Abstract Introduction: Clinical factors and molecular characteristics play pivotal roles in shaping therapeutic strategies and prognoses in colorectal cancer (CRC), the second leading cause of cancer-related mortality in the United States. Despite an overall decline in mortality, the Hispanic/Latino (H/L) population in the Los Angeles area exhibits mortality rates up to 20% higher than their Caucasian American counterparts, often with diagnoses at a younger age and advanced disease stages. Few studies have utilized substantial H/L sample sizes and integrated comprehensive clinical and multi-omics data to conduct integrative translational analysis of reported genomic alterations and their implications for colorectal tumorigenesis. Methods: We collected clinical, DNA, and RNA sequencing data from 36 primary CRC Tumor/Normal (T/N) samples and 82 primary CRC T/N samples within the H/L population in the Los Angeles area through the Participant Engagement and Cancer Genome Sequencing (PE-CGS) Network and the Oncology Research Information Exchange Network (ORIEN), respectively. Additionally, we retrieved two CRC samples with spatial transcriptomic (ST) data from the 10xGenomics database. This data underwent Whole Exome and RNA sequencing analysis. Furthermore, we conducted global and local genomic ancestry analyses to establish correlations between self-ancestral identification and the five main superpopulations. ST data was analyzed using recently released bioinformatics tools. Clinical and genomic data were integrated. Results: In these studies, distinct age-of-onset, Amerindian (AMR), and survival patterns were observed, with significant mutations identified in key genes, including APC, TP53, KRAS, and PIK3CA. Cancer heterogeneity was evident, influenced by mutation type, microsatellite instability, subsite, and ethnicity. Ancestry analysis indicated genetic diversity, with a predominant AMR heritage and cases showcasing substantial European genetic proportions alongside AMR ancestry. Additionally, RNA sequencing has been conducted, with ongoing transcriptomic analysis in both studies, and the results are set for presentation at the annual meeting. The ST analysis generated initial maps, delineating diverse cellular populations and revealing the samples' distinct immunological and inflammatory signals. Conclusion: Our study has delved into the molecular profiling of CRC tumors among H/L patients, contributing essential insights into the DNA and RNA level characterization of CRC tumors and the multifaceted clinical and genomic heterogeneity within our aimed population. It also provides crucial insights into CRC tumor heterogeneity and the tumor microenvironment. These findings serve as the cornerstone for forthcoming sample analyses within our PE-CGS project to potentially lead to the development of personalized treatments. Citation Format: Enrique I. Velazquez Villarreal, Seeta Rajpara, Yuxin Jin, Mackenzie Postel, Brigette Waldrup, Flemming Wu, Donna Loza, Heinz-Josef Lenz, David W. Craig, John D. Carpten. Multi-omics characterization of molecular features and global-local genomic ancestry analysis of colorectal cancer in Hispanic-Latinos [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3932.

Abstract A057: Inter-tumor heterogeneity revealed by multi-site whole genome sequencing of high grade serous ovarian cancer

Abstract A057: Inter-tumor heterogeneity revealed by multi-site whole genome sequencing of high grade serous ovarian cancer

The New Era of Cancer Cytogenetics and Cytogenomics.

The promises of the cancer genome sequencing project, combined with various -omics technologies, have raised questions about the importance of cancer cytogenetic analyses. It is suggested that DNA sequencing provides high resolution, speed, and automation, potentially replacing cytogenetic testing. We disagree with this reductionist prediction. On the contrary, various sequencing projects have unexpectedly challenged gene theory and highlighted the importance of the genome or karyotype in organizing gene network interactions. Consequently, profiling the karyotype can be more meaningful than solely profiling gene mutations, especially in cancer where karyotype alterations mediate cellular macroevolution dominance. In this chapter, recent studies that illustrate the ultimate importance of karyotype in cancer genomics and evolution are briefly reviewed. In particular, the long-ignored non-clonal chromosome aberrations or NCCAs are linked to genome or chromosome instability, genome chaos is linked to genome reorganization under cellular crisis, and the two-phased cancer evolution reconciles the relationship between genome alteration-mediated punctuated macroevolution and gene mutation-mediated stepwise microevolution. By further synthesizing, the concept of karyotype coding is discussed in the context of information management. Altogether, we call for a new era of cancer cytogenetics and cytogenomics, where an array of technical frontiers can be explored further, which is crucial for both basic research and clinical implications in the cancer field.

Somatic mutations of MLL4/COMPASS induce cytoplasmic localization providing molecular insight into cancer prognosis and treatment.

Cancer genome sequencing consortiums have recently catalogued an abundance of somatic mutations, across a wide range of human cancers, in the chromatin-modifying enzymes that regulate gene expression. Defining the molecular mechanisms underlying the potentially oncogenic functions of these epigenetic mutations could serve as the basis for precision medicine approaches to cancer therapy. MLL4 encoded by the KMT2D gene highly mutated in a large number of human cancers, is a key histone lysine monomethyltransferase within the Complex of Proteins Associated with Set1 (COMPASS) family that regulates gene expression through enhancer function, potentially functioning as a tumor suppressor. We report that the KMT2D mutations which cause MLL4 protein truncation also alter MLL4's subcellular localization, resulting in loss-of-function in the nucleus and gain-of-function in the cytoplasm. We demonstrate that isogenic correction of KMT2D truncation mutation rescues the aberrant localization phenotype and restores multiple regulatory functions of MLL4, including COMPASS integrity/stabilization, histone H3K4 mono-methylation, enhancer activation, and therefore transcriptional regulation. Moreover, isogenic correction diminishes the sensitivity of KMT2D-mutated cancer cells to targeted metabolic inhibition. Using immunohistochemistry, we identified that cytoplasmic MLL4 is unique to the tissue of bladder cancer patients with KMT2D truncation mutations. Using a preclinical carcinogen model of bladder cancer in mouse, we demonstrate that truncated cytoplasmic MLL4 predicts response to targeted metabolic inhibition therapy for bladder cancer and could be developed as a biomarker for KMT2D-mutated cancers. We also highlight the broader potential for prognosis, patient stratification and treatment decision-making based on KMT2D mutation status in MLL4 truncation-relevant diseases, including human cancers and Kabuki Syndrome.

Comments: Unconventional Shale Reservoirs Meet Cancer Genome Sequencing

The cross-industry development and application of technologies is the epitome of innovation and ingenuity. The more divergent the industries are, the more the unique ways of thinking strike us as remarkable. Technology transfer between the oil and gas industry and other sectors isn’t new. Many examples can be found in medical science, space exploration, and renewable and sustainable technologies. In this JPT issue, the technologies of Oceaneering Space Systems and Impossible Sensing Energy are featured. A case study presented at the 2023 SPE Annual Technical Conference and Exhibition took medical science to a new low—into the depths of the Permian Basin for downhole reservoir drainage diagnostics (RDD). The coauthors, a subsurface and wells manager and a chapter manager, surveillance, analysis, and optimization and pilots at Chevron Technology Ventures (CTV), described using genome sequencing for cancer detection and treatment in a successful proof-of-concept (POC) test conducted from 2017 to 2019. Nampetch Yamali and Daniel Emery explained in SPE 215052 how in 2017 CTV was given the task of identifying technology to assess vertical drainage dimensions to estimate drainage volumes from fractured wells. The goal was to minimize interference from offset wells and co-developments to optimize reservoir recovery in unconventional fields. At that time, the technologies and their capabilities were limited. Although subsurface microbial DNA sequencing for upstream assets has found applications in enhanced biocide and corrosion inhibition, reservoir sweet spot indicators, and tracer technology, use of the technique was thought also to have potential in horizontal well development planning via RDD. Learning of a genomic sequencing technique for cancer cell identification in humans at the biology and biochemistry department at the University of Houston, the wheels started turning at CTV with thoughts it could be developed and adopted for RDD. Similarities were seen because RDD uses in-situ subsurface microbial DNA to infer the depths from which fluids drain after fracturing a horizontal well and optimizes well spacing on a pad, especially in stacked reservoirs. The predictive analytics platform mapped the hydrocarbon footprints of geologic subzones by using noninvasive DNA testing tools. The unique DNA markers for RDD were extracted from mud and cuttings from the horizontal and vertical drilling sections. DNA from produced fluids was also collected. The authors wrote, “Produced fluids collected at two-to-four-week intervals at the wellhead are mapped to the RDD framework. They are ‘lineage traced’ back to shale and tight rock. Drainage heights and percentage of contribution from these locations is computed adapting DNA sequencing and data analytics pipelines which were developed for lineage tracing cancer cells, breaking off from the primary tumor and metastasizing to distant sites back to the tissue of origin.” After validating the DNA extractions, CTV conducted a blind test using only the DNA data to determine the landing zones for each of the four wells in the pad. Using clustering methods, the results of the blind test accurately identified all landing zones within an acceptable margin, according to Yamali and Emery. The next step in the trial analyzed the DNA of the produced fluids. “Preliminary results showed drainage height estimates for each horizontal well.” In 2021, the study continued with an additional four new field trials in the Permian Basin for application of the technique to zonal production identification. The authors wrote, “Although the results from the POC were positive, subsurface DNA RDD technology is a novel technique within the oil and gas industry, and advanced applications are still evolving. This combination naturally gives rise to more general skepticism and a need to thoroughly evaluate the technology from operational, economic, and accuracy perspectives.” The paper describes the ongoing cross-disciplinary work being done by petroleum engineers, geologists, microbiologists, and stratigraphers to fully evaluate this method of RDD. It further details CTV’s wider approach to “accelerate the innovation life cycle, from pilot stages to widespread adoption at scale,” emphasizing the need for leaders’ support for nonconventional thinking and approaches. As the close of 2023 approaches, I’d like to take this opportunity to wish you the best in 2024 on behalf of the JPT Editorial Review Board and the JPT staff. We’ll be kicking off the new year with the commemoration of JPT’s 75th anniversary. Each issue will include an article dedicated to the evolution of technology and industry practices over the seven and a half decades JPT has covered the upstream industry. In January, Trent Jacobs and Stephen Rassenfoss will revisit technological advancements since JPT’s 50th anniversary in 1999. Since then, over 25 million B/D have been added to the global supply thanks in very large part to advancements in drilling, completions, and reservoir technologies. Join us as we explore these along with some predictions made 25 years ago and the surprising realities of where the industry stands today.

Short-Read Sequencing Detects Large Structural Variants in Cancer Genomes.

Most large structural variants (SV) can be detected by short-read sequencing (SRS) of cancer genomes.

DISP-04. EXPLORING ENGAGEMENT IN LOW GRADE GLIOMA GENOMIC RESEARCH: A COMPARATIVE ANALYSIS OF THREE APPROACHES

Abstract BACKGROUND Engaging patients and care partners in research planning and conduct improves equity and patient-centeredness of research. Determining the most effective engagement methods remains challenging. This study evaluated participation and experience using three engagement methods with the Low Grade Glioma (LGG) Registry’s OPTIMUM project, part of the Participant Engagement in Cancer Genome Sequencing (PE-CGS) Network. METHODS We evaluated participation and experience among participants in LGG Registry Research Advisory Council (RAC) meetings, Twitter (“tweet chats”), and Facebook discussions across four engagement activities. Participants completed validated engagement experience surveys, including the Public & Patient Engagement Evaluation Tool (PPEET), Research Engagement Survey Tool (REST), Patient Engagement in Research Scale (PEIRS-22), and the Trust in Medical Researchers scale. Researchers recorded discussions and performed qualitative analyses across groups. RESULTS RAC engagements involved 25 unique participants representing diverse backgrounds, while tweet chats and Facebook discussions had 197 and 133 participants, respectively. Post-engagement surveys (n = 52 post-engagement 1; n = 40 post-engagement 4) showed no statistically significant differences between engagement approaches in the overall meaningfulness of engagement on PEIRS-22 or REST scales, nor were there differences in post-engagement 1 trust. RAC members rated PEIRS-22 procedural requirements more positively and reported recalling a wide range of expressed views more than social media participants. Conversely, RAC members gave less positive ratings on PPEET items related to feeling heard. Qualitative findings highlighted differences in the nature of interactions across groups, but agreement around trust and similar recommendations for promoting participation in genomic research across engagement methods.

TP53 mutations in Romanian patients with colorectal cancer

BackgroundColorectal cancer (CRC) has been ranked as the second most deadly cancer and the third most diagnosed cancer cases for the year 2020. Specifically for Romania, the number of CRC-related deaths in 2019 was estimated at 6307 people, with a standardized mortality rate of 33.8 per 100,000 inhabitants. Although the tumor protein 53 (TP53) gene is intensively studied, there are few data on TP53 mutations in Romanian CRC. Furthermore, since genetic alterations may show geographical differences, our study aimed to analyze the clinical status and TP53 somatic variation in Romanian CRC patients.Subjects and methodsDNA from 40 randomly selected cases of CRC was extracted from formalin-fixed paraffin-embedded tissues and sequenced using direct Sanger sequencing techniques, and variants were annotated according to the recommendations of the Human Genome Variation Society. Novel variants were analyzed using MutationTaster2021 to predict their effects.ResultsThe mean age was 63.6 years (range 33–85 years) with a male to female ratio of 2.3. More than 45% (18/40) had an advanced cancer stage (≥ stage III). Mutations were found in 21/40 cases (52.5%), with one case having two mutations, giving a total of twenty-two mutations in the TP53 coding DNA. These mutations include 3 (13.6%) insertion-deletion mutations, two of which are novel frameshift mutations: c.165delT (in exon 4) and c.928_935dup (in exon 9), both of which are predicted to lead to nonsense-mediated mRNA decay and are classified as deleterious. The remaining 19 (86.36%) were substitution mutations: 1 nonsense and 18 (81.8%) missense mutations, with G > A (n = 7/19; 36.8%) and C > T (n = 6/19; 31.5%) transitions being the most common. The G > T transversion was found in 21.05% (4/19) of the substitution mutations.ConclusionWe have described two novel frameshift mutations in TP53. The discovery of novel mutations following the efforts of The Cancer Genome Atlas and other large-scale cancer genome sequencing projects may be further evidence of the heterogeneous nature of mutations in cancer and may indicate that the identification of carcinogenic mutations is not yet saturated. Further sequencing is therefore needed, especially in less studied populations. Importantly, consideration of their geographical environment will shed light on population-specific carcinogenesis.

Unique Genomic Alterations and Microbial Profiles Identified in Patients With Gastric Cancer of African, European, and Asian Ancestry: A Novel Path for Precision Oncology.

Here, we characterize differences in the genetic and microbial profiles of GC in patients of African (AFR), European, and Asian ancestry. Gastric cancer (GC) is a heterogeneous disease with clinicopathologic variations due to a complex interplay of environmental and biological factors, which may affect disparities in oncologic outcomes.. We identified 1042 patients with GC with next-generation sequencing data from an institutional Integrated Mutation Profiling of Actionable Cancer Targets assay and the Cancer Genomic Atlas group. Genetic ancestry was inferred from markers captured by the Integrated Mutation Profiling of Actionable Cancer Targets and the Cancer Genomic Atlas whole exome sequencing panels. Tumor microbial profiles were inferred from sequencing data using a validated microbiome bioinformatics pipeline. Genomic alterations and microbial profiles were compared among patients with GC of different ancestries. We assessed 8023 genomic alterations. The most frequently altered genes were TP53 , ARID1A , KRAS , ERBB2 , and CDH1 . Patients of AFR ancestry had a significantly higher rate of CCNE1 alterations and a lower rate of KRAS alterations ( P < 0.05), and patients of East Asian ancestry had a significantly lower rate of PI3K pathway alterations ( P < 0.05) compared with other ancestries. Microbial diversity and enrichment did not differ significantly across ancestry groups ( P > 0.05). Distinct patterns of genomic alterations and variations in microbial profiles were identified in patients with GC of AFR, European, and Asian ancestry. Our findings of variation in the prevalence of clinically actionable tumor alterations among ancestry groups suggest that precision medicine can mitigate oncologic disparities.

Replication DNA polymerases, genome instability and cancer therapies.

It has been over a decade since the initial identification of exonuclease domain mutations in the genes encoding the catalytic subunits of replication DNA polymerases ϵ and δ (POLE and POLD1) in tumors from highly mutated endometrial and colorectal cancers. Interest in studying POLE and POLD1 has increased significantly since then. Prior to those landmark cancer genome sequencing studies, it was well documented that mutations in replication DNApolymerases that reduced their DNA synthesis accuracy, their exonuclease activity or their interactions with other factors could lead to increased mutagenesis, DNA damage and even tumorigenesis in mice. There are several recent, well-written reviews of replication DNA polymerases. The aim of this review is to gather and review in some detail recent studies of DNA polymerases ϵ and δ as they pertain to genome instability, cancer and potential therapeutic treatments. The focus here is primarily on recent informative studies on the significance of mutations in genes encoding their catalytic subunits (POLE and POLD1), mutational signatures, mutations in associated genes, model organisms, and the utility of chemotherapy and immune checkpoint inhibition in polymerase mutant tumors.

Abstract 243: Long-read sequencing of pediatric cancer genomes identifies multiple clinically relevant variants

Abstract Genetic characterization of pediatric cancer samples clinically for diagnostic, prognostic, or therapeutic information currently relies on an assortment of methods that complement and partially overlap one another, such as fluorescence in situ hybridization (FISH), karyotyping, microarray, and sequencing. This need for multiple tests results in longer analysis time, increased total costs, and higher consumption of sometimes limited tumor material. Long-read sequencing (lrSeq) using HiFi technology, i.e., circular consensus sequencing of reads 12-16kb in length, is able to provide sequence level changes with highly accurate base quality (&amp;gt;99%), as well as information on copy number variants, structural variants, and methylation status, all fully phased. Here, we assess the ability of lrSeq to detect clinically relevant genetic alterations (CRGAs) in pediatric cancer samples. We selected 10 patient cases of solid and liquid tumors with known CRGAs from prior clinical testing. These included 3 cases of T-cell acute lymphoblastic leukemia (T-ALL): one with t(9;17) involving NOTCH1, one with t(7;7) involving TRB, and one with biallelic CDKN2A deletion; 2 high-grade glioma (HGG) cases: one with t(2;9) causing a WDCP-NTRK2 fusion, and one with a TP53 mutation; 2 cases of acute myeloid leukemia (AML): one with biallelic mutations in CEPBA and one with a KMT2A-rearrangement (KMT2A-r; t(v;11q23.3)); 1 case of B-cell ALL with KMT2A-r, t(v;11q23.3); 1 neuroblastoma (NBL) case with regional amplifications of chromosome 12 including MDM2; and 1 case of anaplastic large cell lymphoma (ALCL) with an NPM1-ALK fusion (t(2;5)). DNA from these samples was sequenced on the PacBio Sequel II System to a target depth of 25x. HiFi reads were processed using the PacBio Human WGS Workflow. Methylation prediction was made from Primrose and 5mC modification probability of CpG sites were computed by pb-CpG-tools. We then assessed phased CpG methylation in known imprinted genes.We found that lrSeq detected translocations in 6 out of 6 cases, providing precise break ends and often clarifying unclear cytogenetic findings (such as partner genes). HiFi sequencing also detected 3 of 3 sequence alterations in CEBPA and TP53; biallelic deletion of CDKN2A; and known regions with loss of heterozygosity (LOH) in 2 out of 2 cases. However, it had difficulty detecting all amplified regions of chromosome 12 in the case of NBL, such as the one containing MDM2. Finally, we were able to detect methylation differences in a phased manner by parental haplotype in known imprinting regions across the genome. The difference in 5mC modification probability between the two alleles of the known imprinted genes was 81.5. Our results demonstrate that lrSeq can detect multiple CRGAs across several different types of genetic variants and pediatric cancers, bringing us closer to the promise of an all-in-one genetic test for cancer characterization. Citation Format: Byunggil Yoo, Warren Cheung, Irina Pushel, Lisa Lansdon, Chengpeng Bi, Erin Guest, Tomi Pastinen, Midhat Farooqi. Long-read sequencing of pediatric cancer genomes identifies multiple clinically relevant variants [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 243.

Advances in the role of SWI/SNF complexes in tumours.

Cancer development is a complex process involving both genetic and epigenetic changes. The SWI/SNF (switch/sucrose non-fermentable) chromatin remodelling complex, one of the most studied ATP-dependent complexes, plays an important role in coordinating chromatin structural stability, gene expression and post-translational modifications. The SWI/SNF complex can be classified into BAF, PBAF and GBAF according to their constituent subunits. Cancer genome sequencing studies have shown a high incidence of mutations in genes encoding subunits of the SWI/SNF chromatin remodelling complex, with abnormalities in one or more of these genes present in nearly 25% of all cancers, which indicating that stabilizing normal expression of genes encoding subunits in the SWI/SNF complex may prevent tumorigenesis. In this paper, we will review the relationship between the SWI/SNF complex and some clinical tumours and its mechanism of action. The aim is to provide a theoretical basis to guide the diagnosis and treatment of tumours caused by mutations or inactivation of one or more genes encoding subunits of the SWI/SNF complex in the clinical setting.