Patterns Of Genomic Instability Research Articles

OR20-04 Genetic Profiling Using A Gene Panel Identifies Pathogenic Variants And Copy Number Variations In Pituitary Adenomas

Abstract Disclosure: T. Silva: None. D. Mohan: None. J. Mebarak: None. D. Meredith: None. W. Bi: Speaker; Self; Stryker. R. Carroll: None. U.B. Kaiser: None. A. Marcondes Lerario: None. A. Abreu: None. Introduction: Pituitary adenomas (PA) are the second most common central nervous system neoplasm and arise from different cell lineages. PA are benign tumors, although a subset of them may present with aggressive behavior, resulting in significant morbidity. Several genetic alterations have been implicated in PA tumorigenesis, such as USP8 mutations in Cushing disease (CD) and GNAS mutations in somatotroph adenomas (SA). However, the genetic profiles of PA are not completely understood. The aim of this study was to identify genetic alterations associated with PA using a gene panel. Methods: Targeted next generation exome sequencing (OncoPanel) was performed to identify single nucleotide variants (SNV) and copy number variations (CNV) associated with PA. OncoPanel version 1 included 275 genes, version 2 had 300 genes, and the third, current version 447 genes. USP8 was included only in the most recent version. Patients undergoing surgery at our institution between 2013-2020, who agreed to participate in the OncoPanel initiative and who had samples with good quality DNA were enrolled. Classification of PA into subtypes was performed by combining clinical and histopathological data. A strict algorithm was used to assess SNV pathogenicity. Chi-square, Mann Whitney and Kruskal-Wallis tests were used for statistical analyses. Results: Our cohort included 171 patients with following PA subtypes: TPIT lineage - 18 CD, 14 silent ACTH; PIT-1 lineage - 29 SA, 21 lactotroph and 4 silent poorly differentiated PIT-1 adenomas; SF-1 lineage - 77 gonadotroph and 8 null cell adenomas (negative immunostaining for hormones and TF). GNAS mutations were found in 28% of SA and USP8 mutations in 50% of CD screened for USP8. Pathogenic variants in MECOM were associated with silent ACTH, in FGFR4 and NF2 with CD, and in NOTCH2 with lactotroph adenomas, whereas pathogenic variants in TP53, EGFR, MEN1, BRCA1, FH, ATR, BLM, and XPC were identified in multiple PA subtypes. The number of CNV varied significantly between different subtypes of adenomas and were higher in lactotroph (median, 46), poorly differentiated PIT-1 (41), SA (26) and silent ACTH (17) compared to CD (3), gonadotroph (1) and null cell (0.5) (p< 0.001). MIB-1 proliferation index >3% was associated with higher CNV burden (p=0.022). Interestingly, copy number loss of chromosome 16 and 1q combined were exclusively identified in SA without GNAS mutations. Conclusion: OncoPanel analyses identified pathogenic variants in PA, some well recognized variants such as USP8 and GNAS, and some rare events such as NF2 in CD. In addition, our findings highlight subtype-specific patterns of genomic instability. The association between high CNV burden and the MIB-1 proliferation index may help to predict adenoma behavior. Genomic profile using a gene panel has the potential to better classify PA and contribute to clinical management. Presentation: Saturday, June 17, 2023

Mammalian Sex Hormones as Steroid-Structured Compounds in Wheat Seedling: Template of the Cytosine Methylation Alteration and Retrotransposon Polymorphisms with iPBS and CRED-iBPS Techniques

Phytohormones are chemical compounds found naturally in plants that have a significant effect on their growth and development. The increase in research on the occurrence of mammalian sex hormones (MSHs) in plants has prompted the need to investigate the functions performed by these hormones in plant biology. In the present study, we investigated the effects of MSHs on DNA damage and DNA methylation of wheat (Triticum aestivum L.) during the seedling growth stage, using the CRED-iPBS (coupled restriction enzyme digestion/inter primer binding site) assay and iPBS analysis to determine DNA methylation status. Exogenous treatment with four MSHs (17-β-estradiol, estrogen, progesterone, and testosterone) was carried out at four different concentrations (0, 0.05, 0.5, and 5 µM). The highest genomic template stability (GTS) value (80%) was observed for 5 µM 17-β-estradiol, 0.5 µM testosterone, and 0.05 µM estrogen, while the lowest value (70.7%) was observed for 5 µM progesterone and 0.5 µM estrogen. The results of the CRED-iPBS analysis conducted on MspI indicate that the 0.05 µM estrogen-treated group had the highest polymorphism value of 40%, while the 5 µM progesterone-treated group had the lowest value of 20%. For HpaII, treatment with 0.5 µM 17-β-estradiol had the highest polymorphism value of 33.3%, while the group treated with 0.05 µM 17-β-estradiol and 0.05 µM progesterone had the lowest value of 19.4%. In conclusion, MSH treatments altered the stability of the genomic template of wheat plants and affected the cytosine methylation status at the seedling growth stage. Upon comprehensive examination of the results, it was seen that the employed methodology successfully detected alterations in cytosine methylation of genomic DNA (gDNA), as well as changes in the pattern of genomic instability.

Nonlethal Furfural Exposure Causes Genomic Alterations and Adaptability Evolution in Saccharomyces cerevisiae.

Furfural is a major inhibitor found in lignocellulosic hydrolysate, a promising feedstock for the biofermentation industry. In this study, we aimed to investigate the potential impact of this furan-derived chemical on yeast genome integrity and phenotypic evolution by using genetic screening systems and high-throughput analyses. Our results showed that the rates of aneuploidy, chromosomal rearrangements (including large deletions and duplications), and loss of heterozygosity (LOH) increased by 50-fold, 23-fold, and 4-fold, respectively, when yeast cells were cultured in medium containing a nonlethal dose of furfural (0.6 g/L). We observed significantly different ratios of genetic events between untreated and furfural-exposed cells, indicating that furfural exposure induced a unique pattern of genomic instability. Furfural exposure also increased the proportion of CG-to-TA and CG-to-AT base substitutions among point mutations, which was correlated with DNA oxidative damage. Interestingly, although monosomy of chromosomes often results in the slower growth of yeast under spontaneous conditions, we found that monosomic chromosome IX contributed to the enhanced furfural tolerance. Additionally, terminal LOH events on the right arm of chromosome IV, which led to homozygosity of the SSD1 allele, were associated with furfural resistance. This study sheds light on the mechanisms underlying the influence of furfural on yeast genome integrity and adaptability evolution. IMPORTANCE Industrial microorganisms are often exposed to multiple environmental stressors and inhibitors during their application. This study demonstrates that nonlethal concentrations of furfural in the culture medium can significantly induce genome instability in the yeast Saccharomyces cerevisiae. Notably, furfural-exposed yeast cells displayed frequent chromosome aberrations, indicating the potent teratogenicity of this inhibitor. We identified specific genomic alterations, including monosomic chromosome IX and loss of heterozygosity of the right arm of chromosome IV, that confer furfural tolerance to a diploid S. cerevisiae strain. These findings enhance our understanding of how microorganisms evolve and adapt to stressful environments and offer insights for developing strategies to improve their performance in industrial applications.

Abstract 3556: Assessing patterns of genomic instability in recurrent osteosarcoma

Abstract Objective: The osteosarcoma genome is characterized by high levels of genomic instability, however whether there is pervasive ongoing genomic instability, or instability introduced by an early catastrophic event, is still unsettled. Methods: We performed 30-80x whole genome sequencing (WGS) of 37 tumor samples from 8 patients with relapsed or refractory osteosarcoma. Each patient had at least one sample from a primary site and one sample from a metastatic or relapse site. A set of high confidence single nucleotide variants (SNV), copy number alterations (CNA), structural variations (SV) were called for each sample using our pediatric expanded genomics pipeline and an analysis focused on markers of genomic instability was performed using a custom pipeline of computational tools. Results: Of the 8 patients in our cohort, 4 had localized disease at diagnosis (OSCE4, OSCE5, OSCE6, OSCE9) and 4 had metastatic disease at diagnosis (OSCE1, OSCE2, OSCE3, OSCE10). There were 17 samples from primary sites, 7 pretreatment biopsies, 10 on therapy primary resections, 20 metastatic sites, 15 of which were from lung metastases. We have previously reported on the clonal evolution regarding SNVs and CNAs within this cohort. TP53 structural variants involving intron 1/2 were seen in 6/8 patients (OSCE2, OSCE3, OSCE4, OSCE6, OSCE9, OSCE10). No new structural variants in consensus driver genes emerged in metastatic or relapse samples. Comparing the earliest sample to most recent sample in each patient, only OSCE1, OSCE9, and OSCE10 demonstrated an increase of ≥100 structural variants at the final timepoint. In OSCE2, OSCE5, and OSCE6, there were less structural variants identified at the final timepoint when compared to the earliest timepoint. Homologous recombination deficiency (HRD) scores were calculated for each sample. Only 4/8 patients (OSCE3, OSCE4, OSCE9, OSCE10) had an HRD score in the most recently acquired sample higher than the score from the primary site. Regions of hypermutation consistent with kataegis were identified for each sample, with the number of kataegis events remaining unchanged or decreasing in some instances. Kataegis events co-localized with rearrangement events a majority of the time (range 50-100%, Avg= 72.6%). Complex rearrangements such as chromothripsis and chromoplexy were assesed. In 6 patients there were > 9 chromosomal arms involved in complex rearrangements that were shared between primary sites and metastatic/recurrent sites, while ≤ 4 chromosomal arms were involved with new complex rearrangements unique to the metastatic sites. Conclusion: The patterns observed in our cohort reveal that osteosarcoma is relatively stable from diagnosis through subsequent relapses, supporting the model that an early catastrophic event accounts for the genomic instability observed in osteosarcoma. Citation Format: Michael David Kinnaman, Simone Zaccaria, Alvin Makohon-Moore, Gunes Gundem, Juan E. Arango Ossa, Filemon S. Dela Cruz, Paul A. Meyers, Meera Hameed, William D. Tap, Julia Lynne Glade Bender, Elli Papaemmanuil, Andrew Kung, Christine Iacobuzio Donahue. Assessing patterns of genomic instability in recurrent osteosarcoma. [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 3556.

210O Mutational signature analysis reveals patterns of genomic instability linked to resistance to endocrine therapy (ET) +/- CDK 4/6 inhibition (CDK4/6i) in estrogen receptor-positive/HER2-negative (ER+/HER2-) metastatic breast cancer (MBC)

APOBEC mutagenesis is enriched in MBCs. The predictive role of APOBEC and other mutational processes on standard regimens in the MBC setting is largely unknown. We retrieved clinical and genomic data of 4,595 BCs that underwent targeted sequencing by an FDA-cleared panel comprising >400 genes. Mutational signatures were computed by the SigMA algorithm for samples with at least 5 single nucleotide variants and manually reviewed. BCs were classified as APOBEC+, HRD+ or APOBEC-/HRD- based on the dominant mutational process as defined by SigMA. Progression-free survival (PFS) to any line (L) of therapy was assessed using univariate and multivariate Cox models. APOBEC+ and HRD+ were found at higher proportion in MBCs than in primary tumors (24% and 24% vs 15% and 13%; p<0.001). APOBEC+ were highly represented in ER+/HER2- (19%) and HER2+ (47%) subtypes, while HRD+ in triple-negative (36%). Lobular BCs had a higher proportion of APOBEC+ than HRD+ and APOBEC-/HRD- (34% vs 10% vs 21%, p<0.001). ER+/HER2- MBCs with APOBEC displayed lower ESR1 hotspot mutations (p<0.001). Survival analyses were restricted to patients with available pre-treatment samples. In ER+/HER2- MBCs treated with 1L or 2L ET monotherapy (n=395), APOBEC+ tumors were independently associated to shorter mPFS (HR 1.6, 95%CI 1.2-2.2, p=0.001), regardless of the ET agent. APOBEC+ and HRD+ MBCs were associated with lower mPFS on 1L CDK4/6i+ET (n=419) than APOBEC-/HRD- (HR 1.5, 95%CI 1.1-2, p=0.01 and HR 1.7, 95%CI 1.2-2.5, p=0.006, respectively), regardless of ET partner. HRD+ BCs had a reduced mPFS on 2L (n=207) and 3L (n=372) of CDK4/6i+ET (HR 2.1, 95%CI 1-4.1, p=0.03 and HR 1.8, 95%CI 1.3-2.5, p=0.001, respectively). Patients with APOBEC+ and HRD+ ER+/HER2- MBCs experience shorter survival on ET +/- CDK4/6i, suggesting that the genomic instability conferred by APOBEC and HRD results in early resistance. Novel treatments tailored on these BC subgroups are needed. Additional genomic analyses to unravel mutational patterns related to APOBEC and HRD in this setting are underway.

Copy number alterations in stage I epithelial ovarian cancer highlight three genomic patterns associated with prognosis

BackgroundStage I epithelial ovarian cancer (EOC) encompasses five histologically different subtypes of tumors confined to the ovaries with a generally favorable prognosis. Despite the intrinsic heterogeneity, all stage I EOCs are treated with complete resection and adjuvant therapy in most of the cases. Owing to the lack of robust prognostic markers, this often leads to overtreatment. Therefore, a better molecular characterization of stage I EOCs could improve the assessment of the risk of relapse and the refinement of optimal treatment options. Materials and methods205 stage I EOCs tumor biopsies with a median follow-up of eight years were gathered from two independent Italian tumor tissue collections, and the genome distribution of somatic copy number alterations (SCNAs) was investigated by shallow whole genome sequencing (sWGS) approach. ResultsDespite the variability in SCNAs distribution both across and within the histotypes, we were able to define three common genomic instability patterns, namely stable, unstable, and highly unstable. These patterns were based on the percentage of the genome affected by SCNAs and on their length. The genomic instability pattern was strongly predictive of patients’ prognosis also with multivariate models including currently used clinico-pathological variables. ConclusionsThe results obtained in this study support the idea that novel molecular markers, in this case genomic instability patterns, can anticipate the behavior of stage I EOC regardless of tumor subtype and provide valuable prognostic information. Thus, it might be propitious to extend the study of these genomic instability patterns to improve rational management of this disease.

2. Analytical tools to support detection of homologous recombination deficiency (HRD) using cytogenomic scar markers

Homologous Recombination Deficiency (HRD) has been shown to be an effective pharmacogenetic biomarker for determining the efficacy of PARP inhibitor therapy across various tumor types. There is continued research on the best approach to measure HRD status including assessments of sequence variants of homologous recombination repair pathway genes and consequential patterns of genomic instability. Recent technological advancements have enabled for more robust measurement of the cytogenomic aberrations associated with genomic instability emphasizing the importance of its analysis for HRD.

Abstract GS4-08: Comprehensive genomic profiling of patients with breast cancer identifies germline-somatic interactions mediating therapy resistance

Abstract Background: Germline genetic alterations are established mediators of breast carcinogenesis, often giving rise to specific forms of genomic instability. BRCA1/2 pathogenic variants (PVs) are emblematic of this phenomenon through their induction of homologous recombination deficiency. While specific patterns of genomic instability may sensitize cancers to therapies such as PARP inhibitors (PARPi) or platinum chemotherapy, their implications for lineage-directed therapies such as endocrine therapy (ET) or CDK4/6 inhibitors (CDK4/6i) are unknown. Herein, we systematically investigated the patterns of association of germline alterations with specific somatic alterations and explored the resulting effect on clinical outcomes. Methods: Patients who underwent germline and matched tumor tissue sequencing utilizing MSK-IMPACT from April 2014 to May 2021 and had available germline analysis results were included. The final analysis presented at SABCS will include 6000 tumors from 5,150 patients, anonymized according to established institutional IRB guidelines to allow for germline analysis on the full cohort. We analyzed genomic data to inform the full spectrum of somatic and germline mutations, ploidy, and allele-specific copy number to determine loss of heterozygosity (LOH). We performed gene- and pathway-level enrichment analyses between somatic variants and germline PVs. Univariable and multivariable Cox proportional hazards models were constructed to assess the association of therapy-specific progression-free survival (PFS) with select germline PVs and germline-somatic interactions. Results: The preliminary analysis includes 2,798 tumors from 2,242 patients with germline and somatic sequencing results. The most frequent germline PVs were: BRCA2 (n = 81), BRCA1 (n = 67), CHEK2 (n = 57), ATM (n = 32), PALB2 (n = 19). The cohort robustly confirmed previously established relationships such as mutual exclusivity of gATM and TP53 variants (OR 0.10, 95% CI 0.032 - 0.33, q = 0.005). Alterations of TP53 were seen in 83% (56/67) of gBRCA1 patients; however, this did not achieve significance when adjusted for receptor subtype (OR 3.90, 95% CI 1.34-11.38, q = 0.15). The size of the cohort allowed discovery of several novel relationships. For instance, gBRCA2 loss was associated with alterations in TGF-B pathway components (OR 3.58, 95% CI 1.70 - 7.56, q = 0.002), potentially relevant to metastatic disease progression. PIK3CA mutations were significantly less prevalent in both gBRCA2 (OR 0.52, 95% CI 0.31 - 0.87, q = 0.063) and gBRCA1 PVs (OR 0.21, 95% CI 0.085 - 0.51, q = 0.014). Our analysis uncovered a strong association between gBRCA2 and somatic RB1 pathogenic alterations (OR 3.58, 95% CI 1.70 - 7.56, q = 0.011), with most variants (80%) encountered in metastatic gBRCA2 tumors. Given the essential role of RB1 in CDK4/6i response, we investigated the effect of BRCA2 status on clinical efficacy of CDK4/6i-ET. Strikingly, gBRCA2 PVs were significantly associated with inferior PFS (HR 2.17, 95% CI 1.46-3.22, p &amp;lt; 0.001) on first line treatment with CDK4/6i-ET. We posited the enrichment of somatic RB1 loss as a potential mechanism of resistance to CDK4/6i. Given the proximity of RB1 to BRCA2 on chromosome 13, we hypothesized that co-LOH of BRCA2 and RB1 predisposes the cancer cells to bi-allelic loss under therapeutic pressure of CDK4/6i. Indeed, 18/26 gBRCA2 (69.2%) tumors evaluable for allele-specific copy number had evidence of RB1 LOH. Discussion: Analysis of germline-somatic interactions yielded novel associations relevant to breast cancer progression and treatment resistance. Among these, we demonstrated BRCA2 carriers to have inferior outcomes to first line CDK4/6i-ET with potential implications for optimal first line therapy and sequencing of CDK4/6i vs PARPi in this patient population. Citation Format: Anton Safonov, Chai Bandlamudi, Paulino Tallón de Lara, Emanuela Ferraro, Fatemeh Derakhshan, Marie Will, Mark Donoghue, Pier Selenica, Joshua Drago, Ezra Rosen, Carlos dos Anjos, Elaine Walsh, Elizabeth A Comen, Mehnaj Ahmed, Barbara Acevedo, Ahmet Zehir, Michael F Berger, David Solit, Larry Norton, Ronglai Shen, Zsofia Stadler, Simon Powell, Jorge S Reis-Filho, Sarat Chandarlapaty, Mark Robson, Pedram Razavi. Comprehensive genomic profiling of patients with breast cancer identifies germline-somatic interactions mediating therapy resistance [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr GS4-08.

Patterns of Genomic Instability in Interspecific Yeast Hybrids With Diverse Ancestries.

The genomes of hybrids often show substantial deviations from the features of the parent genomes, including genomic instabilities characterized by chromosomal rearrangements, gains, and losses. This plastic genomic architecture generates phenotypic diversity, potentially giving hybrids access to new ecological niches. It is however unclear if there are any generalizable patterns and predictability in the type and prevalence of genomic variation and instability across hybrids with different genetic and ecological backgrounds. Here, we analyzed the genomic architecture of 204 interspecific Saccharomyces yeast hybrids isolated from natural, industrial fermentation, clinical, and laboratory environments. Synchronous mapping to all eight putative parental species showed significant variation in read depth indicating frequent aneuploidy, affecting 44% of all hybrid genomes and particularly smaller chromosomes. Early generation hybrids with largely equal genomic content from both parent species were more likely to contain aneuploidies than introgressed genomes with an older hybridization history, which presumably stabilized the genome. Shared k-mer analysis showed that the degree of genomic diversity and variability varied among hybrids with different parent species. Interestingly, more genetically distant crosses produced more similar hybrid genomes, which may be a result of stronger negative epistasis at larger genomic divergence, putting constraints on hybridization outcomes. Mitochondrial genomes were typically inherited from the species also contributing the majority nuclear genome, but there were clear exceptions to this rule. Together, we find reliable genomic predictors of instability in hybrids, but also report interesting cross- and environment-specific idiosyncrasies. Our results are an important step in understanding the factors shaping divergent hybrid genomes and their role in adaptive evolution.

The tumor immune microenvironmental analysis of 2,033 transcriptomes across 7 cancer types

Understanding the tumor microenvironment is important to efficiently identify appropriate patients for immunotherapies in a variety of cancers. Here, we presented the tumor microenvironmental analysis of 2,033 cancer samples across 7 cancer types: colon adenocarcinoma, skin cutaneous melanoma, kidney renal papillary cell carcinoma, sarcoma, pancreatic adenocarcinoma, glioblastoma multiforme, and pheochromocytoma / paraganglioma from The Cancer Genome Atlas cohort. Unsupervised hierarchical clustering based on the gene expression profiles separated the cancer samples into two distinct clusters, and characterized those into immune-competent and immune-deficient subtypes using the estimated abundances of infiltrated immune and stromal cells. We demonstrated differential tumor microenvironmental characteristics of immune-competent subtypes across 7 cancer types, particularly immunosuppressive tumor microenvironment features in kidney renal papillary cell carcinoma with significant poorer survival rates and immune-supportive features in sarcoma and skin cutaneous melanoma. Additionally, differential genomic instability patterns between the subtypes were found across the cancer types, and discovered that immune-competent subtypes in most of cancer types had significantly higher immune checkpoint gene expressions. Overall, this study suggests that our subtyping approach based on transcriptomic data could contribute to precise prediction of immune checkpoint inhibitor responses in a wide range of cancer types.

The development of a homologous recombination deficiency (HRD) score to identify HR-deficient tumors.

e18085 Background: BRCA1/2 mutated cancers are homologous recombination repair (HRR) deficient and often sensitive to drugs targeting DNA repair pathway. Emerging evidences suggest tumors with mutations in other HRR genes might also respond favorably to DNA repair pathway targeted drugs; however, the results were inconclusive. Genomic instability is characterized by telomeric allelic imbalance (TAI), large-scale transition (LST) and loss of heterozygosity (LOH). Here, we developed an HRD score based on assessing the status of these patterns in tumors. Methods: 73 cancer patients, including 36 breast cancer (BC), 35 ovarian cancer (OC) and 2 prostate cancer (PC) patients were recruited. 28 were BRCA-deficient tumors (10 BCs, 17OCs and 1 PC) harboring pathogenic/likely pathogenic (P/LP) germline mutation in BRCA1/2 with LOH. 13 BRCA-mutated tumors (10 BCs, 2 OCs and 1 PC) carried P/LP BRCA1/2 mutation without LOH. The rest 33 were HRR wild type controls (16 BCs and 16 OCs). All samples were profiled using a panel consisting of 24 classic HRR genes and their adjacent SNPs. The scores for TAI, LST and LOH were developed separately and an incorporated HRD score based on the 3 patterns was established. Clinical data of 42 OC pts were collected to validate the algorithm. Results: All pairwise correlations of TAI, LST and LOH scores showed high correlations (TAI vs. LOH: r = 0.89; TAI vs. LST: r = 0.92; LST vs. LOH: r = 0.83). The univariate logistic regression was used to assess the correlation of the score with BRCA deficiency. In the cohort of BRCA-deficient vs. HRR-negative, the scores of TAI (P = 9.72 x 10−4), LST (P = 8.75 x 10−5), LOH (P = 0.01) and HRD (P = 3.35 x 10−4) all showed significant associations with BRCA-deficiency. In the cohort of BRCA-deficient vs. BRCA-mutated +HRR-negative, all the scores also highly correlated with BRCA-deficiency (TAI: P = 2.35 x 10−4, LST: P = 1.48 x 10−5; LOH: P = 2.36 x 10−4, HRD: P = 6.29 x 10−5). Using 42 as a cutoff, it achieved sensitivities of 94.1%, 60% and 82% for identifying BRCA-deficient tumors in the 35 OCs, 36 BCs and in all 73 tumors, respectively. We further validated the predictive value of HRD score in an additional 42 OC pts and found significantly higher HRD sore in pts sensitive to platinum therapy (P = 0.036). Patients having an HRD score &gt; 42 had a significantly longer PFS than pts with a score &lt; 42 (p = 0.04). Conclusions: Our study developed a HRD score base on assessing 3 patterns of genomic instability (TAI, LST and LOH). The HRD score revealed a promising value in detecting HR-deficient tumors (especially BRCA-deficient).

Distinct genomic instability landscape of lung adenocarcinoma associated with treatment and metastasis.

9534 Background: Lung adenocarcinoma (LUAD) is the most common subtype of non-small cell lung cancer (NSCLC). Genomic instability, defined as genome-wide copy number alterations, is a key pathogenic signature which occurs at the early stage of most cancers and is associated with an increased risk of recurrence or death. We examined the pattern of genomic instability in primary and metastatic LUAD. Methods: We performed deep targeted sequencing (425 genes) of 3395 tissue samples and whole exome sequencing (WES) of 60 tissue samples from LUAD patients. Whole-genome doubling (WGD) and arm level aneuploidy were analyzed to uncover correlation with clinical phenotypes and other genomic alterations including driver mutations, tumor mutation burden (TMB), and microsatellite instability (MSI). Results: Overall, targeted sequencing revealed that WGD occurred in 64.33% LUAD samples, which was comparable with WES results. Compared to primary site, metastasis exhibited higher proportion of WGD (1.14 fold). Specifically, liver metastasis has the highest WGD percentage among metastasis sites (~87.5%; 1.40 fold increase compared to primary). Interestingly, patients who received tyrosine kinase inhibitors (TKI) had higher frequency of WGD than patients without TKI treatment. In addition, TMB was higher in WGD+ patients but MSI status was not significantly different between groups. Arm-level aneuploidy was prevalent in this cohort. The most common amplification events were 7p gain (62%), 5p gain (54%), and 8q gain (53%); top deletion events were 19p loss (47%), 15q loss (42%), and 10 q loss (41%). Patients with EGFR or TP53 mutation were more likely to have aneuploidy compared to wildtypes. Subgroup analysis showed distinct patterns of aneuploidy among metastasis sites, suggesting organ-specific alterations. Evolution analysis showed 7p gain was an early event common in primary tumor whereas metastatic tumor had multiple distinct evolutionary trajectories following 7p gain. Several copy number signatures were associated with specific TKI and chemotherapies. For example, TKI-naïve tumors lacked 7p gain but had 19p loss as the most common alteration. Conclusions: The genomic landscape of LUAD was characterized by widespread large-scale copy number alterations including WGD and chromosomal aneuploidy. Metastasis had elevated level of aneuploidy compared to primary tumors which were specific to metastatic site. Copy number signature associated with different treatments may contribute to distinct long-term survival and side effects among patients.

The NSL complex maintains nuclear architecture stability via lamin A/C acetylation.

While nuclear lamina abnormalities are hallmarks of human diseases, their interplay with epigenetic regulators and precise epigenetic landscape remain poorly understood. Here, we show that loss of the lysine acetyltransferase MOF or its associated NSL-complex members KANSL2 or KANSL3 leads to a stochastic accumulation of nuclear abnormalities with genomic instability patterns including chromothripsis. SILAC-based MOF and KANSL2 acetylomes identified lamin A/C as an acetylation target of MOF. HDAC inhibition or acetylation-mimicking lamin A derivatives rescue nuclear abnormalities observed in MOF-deficient cells. Mechanistically, loss of lamin A/C acetylation resulted in its increased solubility, defective phosphorylation dynamics and impaired nuclear mechanostability. We found that nuclear abnormalities include EZH2-dependent histone H3 Lys 27 trimethylation and loss of nascent transcription. We term this altered epigenetic landscape "heterochromatin enrichment in nuclear abnormalities" (HENA). Collectively, the NSL-complex-dependent lamin A/C acetylation provides a mechanism that maintains nuclear architecture and genome integrity.

Genomic Instability Changes of Circulating Tumor DNA Reflect the Responses to Chemotherapy or Targeted Therapy in Advanced Gastric Cancer

Background: Gastric cancer is characterized by frequent somatic copy number alterations (SCNV) and a high level of genomic instability. In this study, we investigated the genomic instability changes of circulating tumor DNA (ctDNA). Methods: A total of 101 plasmas and paired blood cells from 26 patients with AGC before (n = 26) and after (n = 75) drug treatment were collected. Plasma ctDNA was separated and assessed for SCNV by whole-genome sequencing (WGS), and the whole genome abnormality score (WGAS) was calculated. We next assessed the correlations between WGAS and therapeutic response. Findings: Genomic instability was observed in 20 of 26 patients (77%) prior to drug treatment. For patients with genomic instability prior to treatment, the response rate (55%, 11/20) was higher than that (17%, 1/6) of patients with stable genomes. Notably, genomic instability fluctuated during therapy, with 93% (14/15) of post-therapeutic ctDNAs showing low-genomic instability when sensitive to drug therapy and 52% (13/25) of ctDNAs with developed resistance to therapy showing high genomic instability. For ctDNAs with developed resistance, the genomic instability patterns were identical to those before treatment, but the instability level was lower than the pretherapeutic level. Interpretation: We found that genomic instability based on ctDNA could be used to predict and monitor therapeutic response in gastric cancer, although validation in a larger cohort will be necessary. Funding Statement: This work was supported by the National Key Research and Development Program of China (No. 2017YFC1308900, 2017YFC0908400) and the Beijing Municipal Science & Technology Commission Program (No. Z161100002616036). Declaration of Interests: The authors declare no conflict of competing interest. Ethics Approval Statement: This study was approved by the Medical Ethics Committee of Peking University Cancer Hospital, and written informed consent was obtained from all of the patients for their samples to be used in the future.

Integration of genomic copy number variations and chemotherapy-response biomarkers in pediatric sarcoma

BackgroundWhile most pediatric sarcomas respond to front-line therapy, some bone sarcomas do not show radiographic response like soft-tissue sarcomas (rhabdomyosarccomas) but do show 90% necrosis. Though, new therapies are urgently needed to improve survival and quality of life in pediatric patients with sarcomas. Complex chromosomal aberrations such as amplifications and deletions of DNA sequences are frequently observed in pediatric sarcomas. Evaluation of copy number variations (CNVs) associated with pediatric sarcoma patients at the time of diagnosis or following therapy offers an opportunity to assess dysregulated molecular targets and signaling pathways that may drive sarcoma development, progression, or relapse. The objective of this study was to utilize publicly available data sets to identify potential predictive biomarkers of chemotherapeutic response in pediatric Osteosarcoma (OS), Rhabdomyosarcoma (RMS) and Ewing’s Sarcoma Family of Tumors (ESFTs) based on CNVs following chemotherapy (OS n = 117, RMS n = 64, ESFTs n = 25 tumor biopsies).MethodsThere were 206 CNV profiles derived from pediatric sarcoma biopsies collected from the public databases TARGET and NCBI-Gene Expression Omnibus (GEO). Through our comparative genomic analyses of OS, RMS, and ESFTs and 22,255 healthy individuals called from the Database of Genomic Variants (DGV), we identified CNVs (amplifications and deletions) pattern of genomic instability in these pediatric sarcomas. By integrating CNVs of Cancer Cell Line Encyclopedia (CCLE) identified in the pool of genes with drug-response data from sarcoma cell lines (n = 27) from Cancer Therapeutics Response Portal (CTRP) Version 2, potential predictive biomarkers of therapeutic response were identified.ResultsGenes associated with survival and/recurrence of these sarcomas with statistical significance were found on long arm of chromosome 8 and smaller aberrations were also identified at chromosomes 1q, 12q and x in OS, RMS, and ESFTs. A pool of 63 genes that harbored amplifications and/or deletions were frequently associated with recurrence across OS, RMS, and ESFTs. Correlation analysis of CNVs from CCLE with drug-response data of CTRP in 27 sarcoma cell lines, 33 CNVs out of 63 genes correlated with either sensitivity or resistance to 17 chemotherapies from which actionable CNV signatures such as IGF1R, MYC, MAPK1, ATF1, and MDM2 were identified. These CNV signatures could potentially be used to delineate patient populations that will respond versus those that will not respond to a particular chemotherapy.ConclusionsThe large-scale analyses of CNV-drug screening provides a platform to evaluate genetic alterations across aggressive pediatric sarcomas. Additionally, this study provides novel insights into the potential utilization of CNVs as not only prognostic but also as predictive biomarkers of therapeutic response. Information obtained in this study may help guide and prioritize patient-specific therapeutic options in pediatric bone and soft-tissue sarcomas.

Abstract MS1-1: Targeting DNA repair deficiency in triple negative breast cancers (TNBC) with G-quadruplex stabilisers

Abstract Triple negative breast cancers (TNBC) represent a heterogeneous group of patients where pathway targeted interventions are lacking. We have recently identified new subgroups of TNBC tumours based on patterns of genomic instability, suggesting that deficiencies in DNA repair or genome maintenance could be exploited in specific subgroups of TNBC patients. We recently discovered synthetic lethal activity for small molecules that bind and stabilise G-quadruplex (G4) structures in the human genome. Stabilized G4 structures are considered "at risk" during DNA replication, leading to single strand and double strand breaks in the absence of DNA repair. This has specific application in cancers with DNA repair deficiency. We have shown that loss of homologous recombination repair capacity in breast cancers is associated with ∼ 1 log order increase in sensitivity to G4 binders. Moreover this activity has been observed in platinum pre-treated tumours, and in PARP insensitive tumours. Recently through CRISPR screening approaches we have defined additional DNA repair lesions that confer synthetic lethality to G4 binders. This mechanism of G4 stabilisation is now being tested in a phase 1/2 clinical trial of CX5461 in breast cancer patients. Citation Format: Aparicio S. Targeting DNA repair deficiency in triple negative breast cancers (TNBC) with G-quadruplex stabilisers [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr MS1-1.

Abstract 5713: PIGN gene expression aberration weakens chromosomal stability via altering its interaction with the spindle assembly checkpoint protein complex during leukemogenesis

Abstract Previous studies have linked increased frequency of glycosylphosphatidylinositol-anchor protein (GPI-AP) deficiency with genomic instability and the risk of carcinogenesis. Recently, Phosphatidylinositol Glycan Anchor Biosynthesis; Class N (PIGN), a gene participating in GPI-AP synthesis, was suggested as a cancer chromosomal instability suppressor in a colon cancer model. We investigated the association of PIGN with genomic instability and leukemogenesis. A Random Forest analysis of the gene expression array data from 55 MDS patients (GSE4619) demonstrated a significant (p = 0.0007) correlation (Pearson r =-0.4068) between GPI-anchor biosynthesis gene expression and genomic instability, in which PIGN was ranked as the third most important in predicting risk of MDS progression. We observed that PIGN gene expression aberrations (increased transcriptional activity but diminished to no protein production) were associated with increased frequency of GPI-AP deficiency in leukemic cells during leukemic transformation/progression. The PIGN gene expression aberrations were attributed to partial intron retentions between exons 14 and 15 resulting in frameshifts and premature termination which were confirmed by examining the RNA-seq data from a group of AML patients (phs001027.v1.p1). PIGN gene expression aberration correlated with the elevation of genomic instability marker expression that was independent of the TP53 regulatory pathway. PIGN protein expression suppression/elimination caused a similar pattern of genomic instability that was rescued by PIGN restoration. Furthermore, PIGN bound to the spindle assembly checkpoint proteins and regulated their expression during the cell cycle. In conclusion, PIGN gene is crucial in the regulation of mitotic integrity to maintain chromosomal stability and prevents leukemic transformation/progression. Citation Format: Jeffrey J. Pu, Emmanuel K. Teye, Abigail Sido, Yuka I. Kawasawa, Ping Xin, Niklas K. Finnberg, Wafik S. El-Deiry, Sara Shimko. PIGN gene expression aberration weakens chromosomal stability via altering its interaction with the spindle assembly checkpoint protein complex during leukemogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5713. doi:10.1158/1538-7445.AM2017-5713

PIGN gene expression aberration is associated with genomic instability and leukemic progression in acute myeloid leukemia with myelodysplastic features.

Previous studies have linked increased frequency of glycosylphosphatidylinositol-anchor protein (GPI-AP) deficiency with genomic instability and the risk of carcinogenesis. However, the underlying mechanism is still not clear. A randomForest analysis of the gene expression array data from 55 MDS patients (GSE4619) demonstrated a significant (p = 0.0007) correlation (Pearson r =-0.4068) between GPI-anchor biosynthesis gene expression and genomic instability, in which PIGN, a gene participating in GPI-AP biosynthesis, was ranked as the third most important in predicting risk of MDS progression. Furthermore, we observed that PIGN gene expression aberrations (increased transcriptional activity but diminished to no protein production) were associated with increased frequency of GPI-AP deficiency in leukemic cells during leukemic transformation/progression. PIGN gene expression aberrations were attributed to partial intron retentions between exons 14 and 15 resulting in frameshifts and premature termination which were confirmed by examining the RNA-seq data from a group of AML patients (phs001027.v1.p1). PIGN gene expression aberration correlated with the elevation of genomic instability marker expression that was independent of the TP53 regulatory pathway. Suppression/elimination of PIGN protein expression caused a similar pattern of genomic instability that was rescued by PIGN restoration. Finally, we found that PIGN bound to the spindle assembly checkpoint protein, MAD1, and regulated its expression during the cell cycle. In conclusion, PIGN gene is crucial in regulating mitotic integrity to maintain chromosomal stability and prevents leukemic transformation/progression.

Abstract 4574: Genomic copy number variations characterize prognosis of oropharyngeal squamous cell carcinoma, including p16 positive subset, after surgery-based treatment

Abstract Background High-risk human papillomavirus (HPV)-related (+) oropharyngeal squamous cell carcinoma (OSCC) exhibits unique clinical and molecular characteristics, compared to HPV-unrelated - OSCCs. As different patterns of genomic instability is expected in HPV-driven carcinogenesis, we questioned if patterns of genomic copy number variations (CNVs) differ depending on HPV association, and predict prognosis, after surgery-based treatment for OSCCs in Korean population. Methods Fifty-eight OSCCs patients, who underwent surgery-based treatment and followed for 42 months on median, were enrolled. HPV status was determined by p16 immunohistochemistry (IHC), and HPV DNA PCR. Genome-wide CNV analysis was performed using array-based comparative genomic hybridization of genomic DNA from paraffin-embedded specimen. Clinical association and survival analyses were followed. Results p16 predicted the overall survival (HR = 0.27, CI: 0.39-0.80, p = 0.0006), better than HPV DNA PCR (HR = 0.83, CI: 0.66-1.29, p = 0.64) or smoking history (HR = 2.18, CI: 0.92-5.18, p = 0.071). In overall, number of CNVs did not significantly differed between p16+/- groups. However, 30 locus showed different CNV pattern between these groups. PRDM2 was amplified only in p16+ group, while EGFR and 11q13.3 was more amplified in p16- patients. Loss of FGF18 lead to worse prognosis, and gain of CDK10 and RAD18 was linked to better prognosis in all patients. Amplification of HRAS and loss of KDR lead to better OS in p16+ group. Conclusions After surgery-based treatment, p16+ OSCCs exhibits different CNV patterns at some loci, compared with p16- counterpart. Specific patterns of CNVs do predict better survival, especially in p16+ OSCCs. In the future, clinical trials should stratify patients based on these patterns to apply escalated treatment to obtain better survival. Citation Format: Yuh-S. Jung, Arang Rhie, Jong-Il Kim, Jin Soo Lee, Junsun Ryu, Weon Seo Park. Genomic copy number variations characterize prognosis of oropharyngeal squamous cell carcinoma, including p16 positive subset, after surgery-based treatment. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4574. doi:10.1158/1538-7445.AM2015-4574

Genomic Complexity Profiling Reveals That HORMAD1 Overexpression Contributes to Homologous Recombination Deficiency in Triple-Negative Breast Cancers.

Triple-negative breast cancers (TNBC) are characterized by a wide spectrum of genomic alterations, some of which might be caused by defects in DNA repair processes such as homologous recombination (HR). Despite this understanding, associating particular patterns of genomic instability with response to therapy has been challenging. Here, we show that allelic-imbalanced copy-number aberrations (AiCNA) are more prevalent in TNBCs that respond to platinum-based chemotherapy, thus providing a candidate predictive biomarker for this disease. Furthermore, we show that a high level of AiCNA is linked with elevated expression of a meiosis-associated gene, HORMAD1. Elevated HORMAD1 expression suppresses RAD51-dependent HR and drives the use of alternative forms of DNA repair, the generation of AiCNAs, as well as sensitizing cancer cells to HR-targeting therapies. Our data therefore provide a mechanistic association between HORMAD1 expression, a specific pattern of genomic instability, and an association with response to platinum-based chemotherapy in TNBC. Previous studies have shown correlation between mutational "scars" and sensitivity to platinums extending beyond associations with BRCA1/2 mutation, but do not elucidate the mechanism. Here, a novel allele-specific copy-number characterization of genome instability identifies and functionally validates the inappropriate expression of the meiotic gene HORMAD1 as a driver of HR deficiency in TNBC, acting to induce allelic imbalance and moderate platinum and PARP inhibitor sensitivity with implications for the use of such "scars" and expression of meiotic genes as predictive biomarkers.