Tumor Aneuploidy Research Articles

KaryoTap Enables Aneuploidy Detection in Thousands of Single Human Cells.

Investigating chromosomal instability and aneuploidy within tumors is essential for understanding tumorigenesis and developing diagnostic and therapeutic strategies. Single-cell DNA sequencing technologies have enabled such analyses, revealing aneuploidies specific to individual cells within the same tumor. However, it has been difficult to scale the throughput of these methods to detect rare aneuploidies while maintaining high sensitivity. To overcome this deficit, we developed KaryoTap, a method combining custom targeted DNA sequencing panels for the Tapestri platform with a computational framework to enable detection of chromosome- and chromosome arm-scale aneuploidy (gains or losses) and copy number neutral loss of heterozygosity in all human chromosomes across thousands of single cells simultaneously. KaryoTap allows detecting gains and losses with an average accuracy of 83% for arm events and 91% for chromosome events. Importantly, together with chromosomal copy number, our system allows us to detect barcodes and gRNAs integrated into the cells' genome, thus enabling pooled CRISPR- or ORF-based functional screens in single cells. As a proof of principle, we performed a small screen to expand the chromosomes that can be targeted by our recently described CRISPR-based KaryoCreate system for engineering aneuploidy in human cells. KaryoTap will prove a powerful and flexible approach for the study of aneuploidy and chromosomal instability in both tumors and normal tissues.

Clinical and molecular correlates of tumor aneuploidy in metastatic non-small cell lung cancer

Recent studies have linked elevated tumor aneuploidy to anti-tumor immune suppression and adverse survival following immunotherapy. Herein, we provide supportive evidence for tumor aneuploidy as a biomarker of response to immunotherapy in patients with non-small cell lung cancer (NSCLC). We identify a dose–response relationship between aneuploidy score and patient outcomes. In two independent NSCLC cohorts (n = 659 patients), we demonstrate a novel association between elevated aneuploidy and non-smoking-associated oncogenic driver mutations. Lastly, we report enrichment of TERT amplification and immune-suppressive phenotypes of highly aneuploid NSCLC. Taken together, our findings emphasize a potentially critical role for tumor aneuploidy in guiding immunotherapy treatment strategies.

Chromosome 7 Gain Compensates for Chromosome 10 Loss in Glioma.

The co-occurrence of chromosome 10 loss and chromosome 7 gain in gliomas is the most frequent loss-gain co-aneuploidy pair in human cancers. This phenomenon has been investigated since the late 1980s without resolution. Expanding beyond previous gene-centric studies, we investigated the co-occurrence in a genome-wide manner, taking an evolutionary perspective. Mining of large-scale tumor aneuploidy data confirmed the previous finding of a small-scale longitudinal study that the most likely order is chromosome 10 loss, followed by chromosome 7 gain. Extensive analysis of genomic and transcriptomic data from both patients and cell lines revealed that this co-occurrence can be explained by functional rescue interactions that are highly enriched on chromosome 7, which could potentially compensate for any detrimental consequences arising from the loss of chromosome 10. Transcriptomic data from various normal, noncancerous human brain tissues were analyzed to assess which tissues may be most predisposed to tolerate compensation of chromosome 10 loss by chromosome 7 gain. The analysis indicated that the preexisting transcriptomic states in the cortex and frontal cortex, where gliomas arise, are more favorable than other brain regions for compensation by rescuer genes that are active on chromosome 7. Collectively, these findings suggest that the phenomenon of chromosome 10 loss and chromosome 7 gain in gliomas is orchestrated by a complex interaction of many genes residing within these two chromosomes and provide a plausible reason why this co-occurrence happens preferentially in cancers originating in certain regions of the brain. Significance: Increased expression of multiple rescuer genes on the gained chromosome 7 could compensate for the downregulation of several vulnerable genes on the lost chromosome 10, resolving the long-standing mystery of this frequent co-occurrence in gliomas.

The TP53-activated E3 ligase RNF144B is a tumour suppressor that prevents genomic instability.

TP53, the most frequently mutated gene in human cancers, orchestrates a complex transcriptional program crucial for cancer prevention. While certain TP53-dependent genes have been extensively studied, others, like the recently identified RNF144B, remained poorly understood. This E3 ubiquitin ligase has shown potent tumor suppressor activity in murine Eμ Myc-driven lymphoma, emphasizing its significance in the TP53 network. However, little is known about its targets and its role in cancer development, requiring further exploration. In this work, we investigate RNF144B's impact on tumor suppression beyond the hematopoietic compartment in human cancers. Employing TP53 wild-type cells, we generated models lacking RNF144B in both non-transformed and cancerous cells of human and mouse origin. By using proteomics, transcriptomics, and functional analysis, we assessed RNF144B's impact in cellular proliferation and transformation. Throughin vitroand in vivo experiments, we explored proliferation, DNA repair, cell cycle control, mitotic progression, and treatment resistance. Findings were contrasted with clinical datasets and bioinformatics analysis. Our research underscores RNF144B's pivotal role as a tumor suppressor, particularly in lung adenocarcinoma. In both human and mouse oncogene-expressing cells, RNF144B deficiency heightened cellular proliferation and transformation. Proteomic and transcriptomic analysis revealed RNF144B's novel function in mediating protein degradation associated with cell cycle progression, DNA damage response and genomic stability. RNF144B deficiency induced chromosomal instability, mitotic defects, and correlated with elevated aneuploidy and worse prognosis in human tumors. Furthermore, RNF144B-deficient lung adenocarcinoma cells exhibited resistance to cell cycle inhibitors that induce chromosomal instability. Supported by clinical data, our study suggests that RNF144B plays a pivotal role in maintaining genomic stability during tumor suppression.

Abstract 962: Monitoring on-treatment tumor molecular dynamics using plasma whole genome sequencing (pWGS) and whole exome sequencing (pWES) of circulating tumor DNA (ctDNA) in advanced esophageal cancer: An exploratory analysis of KEYNOTE-590

Abstract Background: We evaluated on-treatment tumor molecular dynamics using ctDNA-based pWGS and pWES in patients (pts) with locally advanced or metastatic esophageal cancer that responded to pembro + chemo or chemo alone and later experienced PD in KEYNOTE-590 (NCT03189719). Methods: Pts with locally advanced, unresectable, or metastatic esophageal or Siewert type 1 gastroesophageal junction cancer were randomized 1:1 to first-line pembro 200 mg or placebo + chemo (5-fluorouracil + cisplatin) Q3W for ≤35 cycles. Cell-free DNA (cfDNA) was isolated and sequenced from plasma samples collected at baseline (BL), during on-treatment response (cycle 5 [C5]), and at PD (discontinuation [D]) from pts with response and subsequent PD. ctDNA burden (tumor fraction [TF]) was estimated using low-pass pWGS by quantifying tumor aneuploidy in cfDNA. All samples with TF >1.5% underwent high-depth pWES. ctDNA burden from pWES was estimated as maximum somatic allele frequency (MSAF). Associations of TF with MSAF were assessed using Spearman correlation coefficient r. For each subject, Jaccard index (JI) was calculated to evaluate the overlap of variants between BL and D. Variant allele frequency (VAF) and normalized VAF (nVAF) were used to determine variant clonality. Results: 108 pts (pembro + chemo, 59; chemo, 49) with initial response and subsequent PD had plasma samples available from BL, C5, and D. Samples from 94 pts had TF >1.5% by pWGS and underwent pWES. MSAF correlated with TF at BL (r = 0.84), C5 (0.56), and D (0.86). ctDNA TF was lower at C5 than at BL and D, with a mean of 4.4% (C5) vs 20.9% (BL) and 14.0% (D) with pembro + chemo and 4.0% (C5) vs 15.8% (BL) and 15.5% (D) with chemo. In variant-level analyses of pWES data from samples with high TF (≥2.5%) at BL and D (n = 65), 39.2% and 44.2% of variants were shared between BL and D with pembro + chemo and chemo, respectively. Higher overlap was observed for variants between BL and D for the chemo arm (JI: median, 0.54; range, 0.04-0.91) vs pembro + chemo (0.37; 0.01-0.92). Shared variants between BL and D had higher clonality compared with variants at BL-only or D-only with pembro + chemo (BL and D mean nVAF = 0.50 vs BL-only = 0.39, and D-only = 0.38) and chemo (0.49 vs 0.38 and 0.38). In pooled variant analyses, TP53 was the most commonly observed mutation and was rarely lost or acquired at D. Conclusions: Comprehensive pWES was feasible with sufficient ctDNA burden (TF ≥2.5%). ctDNA burden dynamics followed high disease burden at BL and D, consistent with radiographic determination of response/PD. Tumor variants at BL and D generally had a common clonal trunk and a higher overlap of variants was observed at D in the chemo arm. These results provide preliminary molecular evidence of immune editing in pts receiving immunotherapy that warrants further study. Citation Format: Minita Shah, E.J. Dettman, Andrew Albright, Cai Chen, Julie Kobie, Sukrut Shah, Carol Peña, Razvan Cristescu, Z Alexander Cao. Monitoring on-treatment tumor molecular dynamics using plasma whole genome sequencing (pWGS) and whole exome sequencing (pWES) of circulating tumor DNA (ctDNA) in advanced esophageal cancer: An exploratory analysis of KEYNOTE-590 [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 962.

Chromosome evolution screens recapitulate tissue-specific tumor aneuploidy patterns.

Whole chromosome and arm-level copy number alterations occur at high frequencies in tumors, but their selective advantages, if any, are poorly understood. Here, utilizing unbiased whole chromosome genetic screens combined with in vitro evolution to generate arm- and subarm-level events, we iteratively selected the fittest karyotypes from aneuploidized human renal and mammary epithelial cells. Proliferation-based karyotype selection in these epithelial lines modeled tissue-specific tumor aneuploidy patterns in patient cohorts in the absence of driver mutations. Hi-C-based translocation mapping revealed that arm-level events usually emerged in multiples of two via centromeric translocations and occurred more frequently in tetraploids than diploids, contributing to the increased diversity in evolving tetraploid populations. Isogenic clonal lineages enabled elucidation of pro-tumorigenic mechanisms associated with common copy number alterations, revealing Notch signaling potentiation as a driver of 1q gain in breast cancer. We propose that intrinsic, tissue-specific proliferative effects underlie tumor copy number patterns in cancer.

Abstract A018: Optimizing cancer immunotherapy response prediction by tumor aneuploidy score and fraction of copy number alterations

Abstract Identifying patients that are likely to respond to cancer immunotherapy is an important, yet highly challenging clinical need. Using 3139 patients across 17 different cancer types, we comprehensively studied the ability of two common copy-number alteration (CNA) scores—the tumor aneuploidy score (AS) and the fraction of genome single nucleotide polymorphism encompassed by copy-number alterations (FGA)—to predict survival following immunotherapy in both pan-cancer and individual cancer types. First, we show that choice of cutoff during CNA calling significantly influences the predictive power of AS and FGA for patient survival following immunotherapy. Remarkably, by using proper cutoff during CNA calling, AS and FGA can predict pan-cancer survival following immunotherapy for both high-TMB and low-TMB patients. However, at the individual cancer level, our data suggest that the use of AS and FGA for predicting immunotherapy response is currently limited to only a few cancer types. Therefore, larger sample sizes are needed to evaluate the clinical utility of these measures for patient stratification in other cancer types. Finally, we propose a simple, non-parameterized, elbow-point-based method to help determine the cutoff used for calling CNAs. Citation Format: Tian-Gen Chang, Yingying Cao, Eldad D Shulman, Uri Ben-David, Alejandro A. Schäffer, Eytan Ruppin. Optimizing cancer immunotherapy response prediction by tumor aneuploidy score and fraction of copy number alterations [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr A018.

Concurrent Radiation and Immunotherapy Augments Local Immunity and Improves Survival in Aneuploid NSCLC

Over 500 clinical trials combining radiation (RT) and immune checkpoint blockade (ICB) have been initiated based on preclinical evidence that RT can augment local immunity and improve the efficacy of ICB. However, many recent clinical trials have not found a benefit of combining RT and ICB, raising questions about whether a synergy exists. We examined whether RT and ICB interact to beneficially stimulate the immune response in patients and identified biomarkers of response to RT and ICB. We performed a molecular analysis of 1,740 patients from 3 cohorts. The COSINR dataset is a randomized clinical trial of 22 non-small cell lung cancer (NSCLC) patients treated with concurrent or sequential SBRT and ipilimumab/nivolumab. Paired pre- and on-treatment biopsies of an irradiated metastasis underwent whole exome sequencing and RNA-seq. On-treatment biopsies were obtained after SBRT and prior to ICB (sequential) or after SBRT and one cycle of ICB (concurrent). The UC cohort consisted of targeted DNA sequencing of 58 NSCLC patients treated with ICB alone, sequential RT+ICB, or concurrent RT+ICB. The MSKCC dataset is a pan-cancer cohort of targeted DNA sequencing of 1,660 patients treated with ICB. Aneuploidy score (AS) was defined as the fraction of chromosome arms with arm-level copy number alterations. Survival analyses utilized the Kaplan-Meier method and multivariable Cox proportional hazards models. In the COSINR trial, SBRT+ICB increased, whereas SBRT alone decreased, expression of effector T cell IFN-gamma and adaptive immune signatures (P<0.05). Established biomarkers of ICB response, including IFN-gamma signature, tumor mutational burden (TMB), PD-L1 expression, and neoantigen burden were not associated with survival (P>0.05). However, patients whose tumors exhibited high (≥median) but not low, AS had improved survival when treated with concurrent vs. sequential SBRT+ICB (1-year overall survival [OS] 100% vs. 17%, P = 0.025). Our findings were corroborated in the UC cohort: high AS tumors treated with RT + ICB had superior 1-year OS compared to those treated with ICB alone (59% vs. 31%, P = 0.021). Among those who received RT + ICB, concurrent treatment improved OS relative to sequential (1-year OS 76% vs. 38%). RT did not improve OS in patients with low (<median) AS. In the MSKCC cohort, high AS was associated with poor OS following ICB after controlling for TMB and ICB paradigm (hazard ratio [95% CI]: 1.44 [1.07-1.94], P = 0.014). Among tumors with low TMB (<80th percentile), but not high TMB, high AS (≥median) was associated with poor OS (P = 0.003). Our findings distinguish the genomic and transcriptomic effects of RT versus RT+ICB and challenge the prevailing paradigm that local ablative RT positively stimulates the immune response. We propose the use of tumor aneuploidy as a biomarker in personalizing treatment approaches for patients with various cancers.

Impact of somatic mutations and transcriptomic alterations on cancer aneuploidy.

Aneuploidy has been recognized as one of hallmark of tumorigenesis since the early 20th century. Recent developments in structural variation analysis in the human genome have revealed the diversity of aneuploidy in cancer. However, the effects of gene mutation and expression in tumors on aneuploidy remain poorly understood. Here, we performed whole exome analysis of over 5,000 Japanese cancer cases and investigated the impact of somatic mutations and gene expression alterations on aneuploidy. First, we evaluated tumor content and genomic alterations that could influence aneuploidy. Next, we compared the aneuploidy frequency in 18 cancer types and observed that TP53 mutations were associated with the aneuploidy on specific chromosomes in colorectal and gastric cancers. Finally, we used expression analysis to isolate pathways involved in aneuploidy accumulation from tumors without TP53 mutations. Chromosomal instability and cell cycle aberration were associated with aneuploidy in TP53 wild-type tumors, and 26 commonly upregulated genes were identified in aneuploidy-high solid tumors without TP53 mutations. Among them, two cancer-related genes (CENPA and PBK) were involved in aneuploidy. Our integrated analysis revealed that both TP53 mutations and transcriptomic alterations independent of somatic mutations affect aneuploidy accumulation. Our findings will facilitate further understanding of diverse aneuploidies in the tumorigenesis.

Gene set enrichment analysis identifies immune subtypes of kidney renal clear cell carcinoma with significantly different molecular and clinical properties.

Kidney renal clear cell carcinoma (KIRC) is the most prevalent renal malignancy, marked by a high abundance of tumor-infiltrating lymphocytes (TILs) and an unfavorable prognosis upon metastasis. Numerous studies have demonstrated that KIRC possesses a tumor microenvironment that is highly heterogeneous, and this is associated with significant variations in the effectiveness of most first-line drugs administered to KIRC patients. Therefore, it is crucial to classify KIRC based on the tumor microenvironment, although these subtyping techniques are still inadequate. By applying gene set enrichment scores of 28 immune signatures, we conducted a hierarchical clustering of KIRC and determined its immune subtypes. In addition, we conducted a comprehensive exploration of the molecular and clinical features of these subtypes, including survival prognosis, proliferation, stemness, angiogenesis, tumor microenvironment, genome instability, intratumor heterogeneity, and pathway enrichment. Through cluster analysis, two immune subtypes of KIRC were identified and termed Immunity-High (Immunity-H) and Immunity-Low (Immunity-L). This clustering outcome was consistent in four independent KIRC cohorts. The subtype Immunity-H exhibited elevated levels of TILs, tumor aneuploidy, homologous recombination deficiency, stemness, and proliferation potential, along with a poorer prognosis for survival. Despite this, the Immunity-L subtype demonstrated elevated intratumor heterogeneity and a stronger angiogenesis signature in contrast to Immunity-H. According to the results of pathway enrichment analysis, the Immunity-H subtype was found to be highly enriched in immunological, oncogenic, and metabolic pathways, whereas the Immunity-L subtype was highly enriched in angiogenic, neuroactive ligand-receptor interaction, and PPAR pathways. Based on the enrichment of immune signatures in the tumor microenvironment, KIRC can be categorized into two immune subtypes. The two subtypes demonstrate considerably distinct molecular and clinical features. In KIRC, an increase in immune infiltration is linked to a poor prognosis. Patients with Immunity-H KIRC may exhibit active responses to PPAR and immune checkpoint inhibitors, whereas patients with Immunity-L may manifest favorable responses to anti-angiogenic agents and immune checkpoint inhibitors. The immunological classification provides molecular insights into KIRC immunity, as well as clinical implications for the management of this disease.

Optimizing cancer immunotherapy response prediction by tumor aneuploidy score and fraction of copy number alterations

Identifying patients that are likely to respond to cancer immunotherapy is an important, yet highly challenging clinical need. Using 3139 patients across 17 different cancer types, we comprehensively studied the ability of two common copy-number alteration (CNA) scores—the tumor aneuploidy score (AS) and the fraction of genome single nucleotide polymorphism encompassed by copy-number alterations (FGA)—to predict survival following immunotherapy in both pan-cancer and individual cancer types. First, we show that choice of cutoff during CNA calling significantly influences the predictive power of AS and FGA for patient survival following immunotherapy. Remarkably, by using proper cutoff during CNA calling, AS and FGA can predict pan-cancer survival following immunotherapy for both high-TMB and low-TMB patients. However, at the individual cancer level, our data suggest that the use of AS and FGA for predicting immunotherapy response is currently limited to only a few cancer types. Therefore, larger sample sizes are needed to evaluate the clinical utility of these measures for patient stratification in other cancer types. Finally, we propose a simple, non-parameterized, elbow-point-based method to help determine the cutoff used for calling CNAs.

Impact of pathogenic germline BRCA1/2 and PALB2 mutations and tumor aneuploidy in patients with HR+/HER2- metastatic breast cancer treated with CDK4/6 inhibitors.

1075 Background: CDK4/6 inhibitors lack established predictors of efficacy in HR+/HER2- mBC. Pathogenic germline BRCA2 variants, somatic homologous recombination deficiency (HRD) and APOBEC signatures conferred worse survival in small series. Broader tumor DNA panels enable the detection of these and other parameters such as aneuploidy and tumor mutational burden. Methods: Clinical and molecular data from a single institution cohort of CDK4/6i-treated HR+/HER2- mBC pts were analyzed. ECOG PS was 0-1. Clinical variables were collected retrospectively. Primary endpoints were progression-free survival (PFS) and overall survival (OS). Tumor-only sequencing was performed on clinical samples with 161-gene OCAv3, 523-gene TSO500 or 555-gene lab-developed NGS panels. HRD was assessed with the Signature Multivariate Analysis (SigMA) tool using a likelihood-based measure to estimate single base substitution signature 3. Aneuploidy scores were defined as the fraction of covered arms harboring arm-level somatic copy number alterations as determined by ASCETS v.1.1. Results: 153 pts were available for analysis. Median age was 54 y (29–88). 74% were postmenopausal, 28% de novo metastatic and 67% had visceral disease. Palbociclib was used in 80%, ribociclib in 17% and abemaciclib in 3%, mostly as early lines (1L 65%, 2L 22%). Median follow up was 46 mo (7–84) and median time from sample collection to CDK4/6i was 0.6 y (-18.5–+2.6). 76% of samples have been sequenced. The median number of mutations (muts) was 3 (1–32), most commonly in PIK3CA (37%), GATA3 (26%) and TP53 (20%). ESR1, KRAS and ARID1A, but not RB1 muts were numerically enriched in post–CDK4/6i mets (n=8). TP53 (HR 1.85 CI 1.02–3.33) and GATA3 (HR 0.48, CI 0.28–0.83) muts were associated with PFS. Germline ( g) testing was available in 68 (44%) pts, with 21 pts carrying pathogenic variants including 11 BRCA2, 2 BRCA1 and 2 PALB2. Cox regression showed a strong negative impact of gBRCA1/2-PALB2 on PFS (HR 4.0 CI 2.0–7.9) and OS (HR 3.5 CI 1.6–7.7) adjusted for age, visceral disease and prior chemo. In 1L, mPFS/OS was 9.9/28.1 mo for gBRCA1/2-PALB2 carriers, 26.8/NR mo for non-carriers and 23.7/49.2 mo for untested (Cox HR PFS 5.1 CI 2.1-14.0; HR OS 4.0 CI 1.4-11.2). Pts with gATM/ CHEK2/ BRIP1 had a 1L PFS/OS&gt;15/45 mo. Among 36 pts assessed for HRD, 22 were HRDlow and 14 HRDhigh with no significant survival association. 3/4 known gBRCA1/2 carriers were HRDhigh and 7/8 gWT HRDlow. In these pts, higher aneuploidy was associated with shorter PFS and OS (Cox HR PFS 2.0 CI 0.9-4.3, p=0.07; HR OS 2.6 CI 1.0-7.1; p=0.05). Conclusions: Pathogenic gBRCA1/2- PALB2 and higher aneuploidy were associated with shorter survival. Profiling of aneuploidy, HRD, TMB and APOBEC in all pts is ongoing including 55 baseline ctDNA samples and 361 samples from the GENIE BPC cohort. Results will account for FDR, left truncation and optimal cutoffs.

Abstract 1393: Persistent mutation burden drives sustained anti-tumor immune responses in human cancers

Abstract INTRODUCTION: Tumor mutation burden (TMB) is a commonly used biomarker for cancer immunotherapy however TMB only partially captures tumor foreignness. We hypothesized that mutations in single-copy regions of the genome or mutations present in multiple copies (hereafter referred to as persistent mutations) are retained during cancer evolution and immunoediting, may render the tumor continuously visible to the immune system and promote sustained tumor control during immune checkpoint blockade (ICB). METHODS: We performed pan-cancer analyses of whole exome sequencing data across 31 tumor types in TCGA to quantify the landscape of persistent mutations (n=9,242). We then evaluated the association between persistent tumor mutation burden (pTMB) and ICB response compared to TMB in eight ICB-treated cohorts of patients with NSCLC, melanoma, mesothelioma, and head and neck cancer (n=524). To investigate the clonal evolution of persistent mutations we serially analyzed whole exome sequence data from NSCLCs prior to and at emergence of acquired resistance to ICB. Finally, we evaluated the composition of the tumor microenvironment (TME) in baseline and on-ICB melanomas by RNA sequencing differential enrichment analyses and deconvolution. RESULTS: Integration of sequence alterations in only-copy and multi-copy states for 9,242 tumors across 31 tumor types revealed a cancer lineage-dependent distribution of persistent mutations that was largely independent of the overall TMB. In evaluating differential classification based on pTMB- vs TMB-high, we found re-classification rates as high as 53% in individual tumor types, with a median reclassification rate of 33% across all tumor types (range 15% - 53%). We then evaluated the clonal composition of persistent mutations and found a wide range of correlations between pTMB and fraction of clonal mutations (Spearman ρ range: -0.11 - 0.59). In ICB-treated cohorts, pTMB better distinguished responding tumors compared to TMB, and a number of mutation and copy-number related features including tumor aneuploidy (melanoma: Mann-Whitney p=2.3e-06, NSLC: p&amp;lt;2.0e-03, mesothelioma p=0.03, HNSCC p=0.05). Using in silico simulations, we found a similar advantage for pTMB when estimated from gene-panel targeted next generation sequencing. To support the biological plausibility of pTMB in the context of tumor evolution, we evaluated the rate loss of persistent mutations in longitudinal analyses of pre- and post-ICB NSCLC and found that a rate of loss significantly lower for persistent compared to loss-prone mutations (odds ratio 61.43, p&amp;lt;2.2e-16). Consistent with our hypothesis, pTMB-high tumors had a more inflamed TME (p&amp;lt;1e-11). CONCLUSIONS: Persistent mutations represent a biologically distinct subset within the overall TMB that is unlikely to be lost under selective pressure of ICB and may function as an intrinsic driver of sustained immunologic tumor control. Citation Format: Noushin Niknafs, Archana Balan, Christopher Cherry, Karlijn Hummelink, Kim Monkhorst, Xiaoshan M. Shao, Zineb Belcaid, Kristen A. Marrone, Joseph Murray, Kellie N. Smith, Benjamin Levy, Josephine Feliciano, Christine L. Hann, Vincent Lam, Drew M. Pardoll, Rachel Karchin, Tanguy Y. Seiwert, Julie R. Brahmer, Patrick M. Forde, Victor E. Velculescu, Valsamo K. Anagnostou. Persistent mutation burden drives sustained anti-tumor immune responses in human cancers [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 1393.

One toolkit to bring them all, and in silico analyze them.

One toolkit to bring them all, and in silico analyze them.

Highlights from Recent Cancer Literature.

Breaking Insights| March 02 2023 Highlights from Recent Cancer Literature Author & Article Information Online ISSN: 1538-7445 Print ISSN: 0008-5472 ©2023 American Association for Cancer Research2023American Association for Cancer Research Cancer Res (2023) 83 (5): 653–654. https://doi.org/10.1158/0008-5472.CAN-83-5-BI Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Tools Icon Tools Get Permissions Cite Icon Cite Search Site Article Versions Icon Versions Version of Record March 2 2023 Citation Highlights from Recent Cancer Literature. Cancer Res 1 March 2023; 83 (5): 653–654. https://doi.org/10.1158/0008-5472.CAN-83-5-BI Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest Search Advanced Search Despite hundreds of trials examining the combination of radiotherapy with immunotherapy, evidence for synergy as well as predictive biomarkers are lacking. Spurr and colleagues conducted a trial to examine the efficacy of the combination of immunotherapy plus multisite ablative radiotherapy in non-small cell lung cancer patients. Genomic and transcriptomic analyses of baseline and on-treatment biopsies demonstrated that concurrent radiation and immunotherapy increased the adaptative immune response while radiation alone decreased this response. Furthermore, high baseline aneuploidy predicted improved outcomes in patients treated with concurrent therapy. Expert Commentary: This study suggests that high tumor aneuploidy may be a biomarker of response to concurrent radiotherapy and immunotherapy and should be validated in future trials. Spurr LF, Martinez CA, Kang W, Chen M, Zha Y, Hseu R, et al. Highly aneuploid non-small cell lung cancer shows enhanced responsiveness to concurrent radiation and immune checkpoint blockade. Nat Cancer 2022;3:1498–512. The potential impact of... You do not currently have access to this content.

Abstract P2-11-24: A Novel, Rapid and Economical Prognostic Tool For Adjuvant therapy Decisions in Hormone Positive Breast Cancer

Abstract Several multigene-based, risk scoring methods are available and their use is recommended to support treatment decision making in breast cancer. Although the value of these tests is established, critical challenges remain that limit their use, especially regarding turnaround time for fast decision making and the associated costs. We have developed DigiStain - a novel technology that uses mid-infrared imaging to precisely measure a surrogate of tumour aneuploidy, across unstained biopsy sections. Considering that genomic context shapes the pattern and consequences of aneuploidy during cancer development and progression, aneuploidy may have valuable prognostic or predictive value. Using bespoke software, DigiStain allows a quantitative score ‘DigiStain index’ (DI) to be reproducibly extracted from an objective physical measurement of a cancer i.e., aneuploidy. This information is generated within minutes and is available at the same time as routine H&amp;E staining. We previously showed that DI significantly correlates with tumour grade and survival. To further validate the technology, we have investigated the ability of the DI to predict 5-year survival in a retrospective analysis of patients with oestrogen receptor (ER) positive breast cancer. Methods: Clinical samples were obtained from patients (N=944) treated at the University Hospital of Nottingham UK. Using the DigiStain imager, H&amp;E images were registered before acquiring the DI score for the section. Statistical methodology followed the recommended approach of Royston et al. 2009, The BMJ. A multivariate logistic model modelling the ‘event’ of death by 5 years was generated. The predictive performance of the logistic regression model was assessed using Area Under the receiver operating characteristic Curve (AUC) plots. The predicted vs observed event probability was determined at various cut-offs and the accuracy of the classification was measured by its sensitivity and specificity. Results: Of 944 ER+ patients with follow up information, 77 (8.2%) died in the first 5 years. The age distribution was approximately normal, ranging from 26 to 70 (mean = 54.8 years). Tumour size ranged from 0.2 to 7.5 mm (mean = 1.90 mm and median = 1.70 mm) and had normal distribution when the natural log of size was considered. DI contributed to the prognostic model with (p=0.0018). Odds ratios showed increased odds of death by 5 years with higher DI. The AUC obtained for the model was 0.7722. Due to the number of events the sample was not split into training and validation sets. Instead bootstrapping (n=500) was used to assess performance. When adjusted by bootstrapping the AUC was reduced only slightly to 0.76. Classification tables for sensitivity and specificity across different risk cut-offs and distribution of 5-year predicted probabilities showed the best distinction between groups at low levels of predicted probabilities. As expected, there was a trade-off between sensitivity and specificity when choosing an appropriate cut-off point to define high risk of death by 5 years. Conclusions: The risk score obtained using the DI is a significant prognostic factor for 5-year survival in ER+ breast cancer (p=0.0018) with good diagnostic accuracy. Analysis of 10-year survival data is ongoing and plans to expand the sample collection to include patients from other sites are underway. Citation Format: Hemmel Amrania. A Novel, Rapid and Economical Prognostic Tool For Adjuvant therapy Decisions in Hormone Positive Breast Cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P2-11-24.

Author Correction: Tumor aneuploidy predicts survival following immunotherapy across multiple cancers

Author Correction: Tumor aneuploidy predicts survival following immunotherapy across multiple cancers

NamiRNA-mediated high expression of KNSTRN correlates with poor prognosis and immune infiltration in hepatocellular carcinoma.

Mutations of kinetochore-localized astrin/sperm-associated antigen 5 (KNSTRN) can interfere with chromatid cohesion, increase aneuploidy in tumours, and enhance tumourigenesis. However, the role of the KNSTRN-binding protein in hepatocellular carcinoma (HCC) remains unclear. Using The Cancer Genome Atlas databases, we investigated the potential oncogenic functions of KNSTRN in HCC along with R and various computational tools. Detailed results revealed that elevated expression of KNSTRN was considerably associated with poor overall survival (HR = 1.48, 95% CI: 1.05-2.09, p = 0.027) and progress-free interval (HR = 1.41, 95% CI: 1.05-1.89, p = 0.021) in HCC. Gene ontology/Kyoto Encyclopedia of Genes and Genomes functional enrichment analysis showed that KNSTRN is closely related to organelle fission, chromosomal region, tubulin binding, and cell cycle signalling pathway. TIMER database analysis showed the correlations between KNSTRN expression and tumour-infiltrating immune cells, biomarkers of immune cells, and immune checkpoint expression. Moreover, the KNSTRN level was significantly positively associated with immunosuppressive cells in the tumour microenvironment, including regulatory T-cells, myeloid-derived suppressor cells, and cancer-associated fibrocytes. Finally, a possible nuclear activating miRNA (NamiRNA)-enhancer network of hsa-miR-107, which activates the KNSTRN expression in liver hepatocellular carcinoma, was constructed by correlation analysis. NamiRNA-mediated upregulation of KNSTRN correlated with poor prognosis and tumour immune infiltration in HCC. KNSTRN could serve as an effective biomarker for the diagnosis and prognosis of HCC and support the development of novel therapeutic strategies.

Molecular Classification of Appendiceal Adenocarcinoma.

Appendiceal adenocarcinomas (ACs) are rare, histologically diverse malignancies treated as colorectal cancers despite having distinct biology and clinical behavior. To guide clinical decision making, we defined molecular subtypes of AC associated with patient survival, metastatic burden, and chemotherapy response. A comprehensive molecular analysis was performed in patients with AC to define molecular subtypes. Associations between molecular subtype and overall survival, intraoperative peritoneal cancer index, and first-line chemotherapy response were assessed adjusting for histopathologic and clinical variables using multivariable Cox proportional hazards, linear regression, and logistic regression models. We defined distinct molecular lineages of mucinous appendiceal adenocarcinoma (MAAP) from co-occurring mutations in GNAS, RAS, and TP53. Of 164 MAAP tumors, 24 were RAS-mutant (mut) predominant (RAS-mut/GNAS-wild-type [wt]/TP53-wt) with significantly decreased mutations and chromosomal alterations compared with tumors with GNAS mutations (GNAS-mut predominant) or TP53 mutations (TP53-mut predominant). No patient with RAS-mut predominant subtype metastatic MAAP died of cancer, and overall survival in this subgroup was significantly improved compared with patients with GNAS-mut (P = .05) and TP53-mut (P = .004) predominant subtypes. TP53-mut predominant subtypes were highly aneuploid; increased tumor aneuploidy was independently (P = .001) associated with poor prognosis. The findings retained significance in patients with any metastatic AC. RAS-mut predominant metastases exhibited reduced peritoneal tumor bulk (P = .04) and stromal invasion (P < .001) compared with GNAS-mut or TP53-mut predominant tumors, respectively. Patients with RAS-mut predominant MAAP responded more to first-line chemotherapy (50%) compared with patients with GNAS-mut predominant tumors (6%, P = .03). AC molecular patterns identify distinct molecular subtypes: a clinically indolent RAS-mut/GNAS-wt/TP53-wt subtype; a chemotherapy-resistant GNAS-mut predominant subtype; and an aggressive, highly aneuploid TP53-mut predominant subtype. Each subtype exhibits conserved clinical behavior irrespective of histopathology.

Tumor aneuploidy predicts survival following immunotherapy across multiple cancers.

Tumor mutational burden (TMB) has emerged as a promising biomarker of immunotherapy response across multiple cancer types; however, clinical outcomes among patients with low TMB tumors are heterogeneous. Herein, we demonstrate that tumor aneuploidy provides independent prognostic value among patients with lower TMB (<80th percentile) tumors treated with immunotherapy. A higher aneuploidy score is associated with poor prognosis following immunotherapy among tumors with low TMB, but not those with high TMB. Importantly, aneuploidy scores can be calculated from existing clinical targeted sequencing infrastructure, facilitating deployment of aneuploidy scores as a clinical biomarker.