Comparison of actionable events detected in cancer genomes by whole-genome sequencing, in silico whole-exome and mutation panels.

BACKGROUND: Next-generation sequencing is used in cancer research to identify somatic and germline mutations, which can predict sensitivity or resistance to therapies, and may be a useful tool to reveal drug repurposing opportunities between tumour types. Multigene panels are used in clinical practice for detecting targetable mutations. However, the value of clinical whole-exome sequencing (WES) and whole-genome sequencing (WGS) for cancer care is less defined, specifically as the majority of variants found using these technologies are of uncertain significance. PATIENTS AND METHODS: We used the Cancer Genome Interpreter and WGS in 726 tumours spanning 10 cancer types to identify drug repurposing opportunities. We compare the ability of WGS to detect actionable variants, tumour mutation burden (TMB) and microsatellite instability (MSI) by using in silico down-sampled data to mimic WES, a comprehensive sequencing panel and a hotspot mutation panel. RESULTS: We reveal drug repurposing opportunities as numerous biomarkers are shared across many solid tumour types. Comprehensive panels identify the majority of approved actionable mutations, with WGS detecting more candidate actionable mutations for biomarkers currently in clinical trials. Moreover, estimated values for TMB and MSI vary when calculated from WGS, WES and panel data, and are dependent on whether all mutations or only non-synonymous mutations were used. Our results suggest that TMB and MSI thresholds should not only be tumour-dependent, but also be sequencing platform-dependent. CONCLUSIONS: There is a large opportunity to repurpose cancer drugs, and these data suggest that comprehensive sequencing is an invaluable source of information to guide clinical decisions by facilitating precision medicine and may provide a wealth of information for future studies. Furthermore, the sequencing and analysis approach used to estimate TMB may have clinical implications if a hard threshold is used to indicate which patients may respond to immunotherapy.

Whole-genome sequencing reveals clinically relevant insights into the aetiology of familial breast cancers.

BACKGROUND: Whole-genome sequencing (WGS) is a powerful method for revealing the diversity and complexity of the somatic mutation burden of tumours. Here, we investigated the utility of tumour and matched germline WGS for understanding aetiology and treatment opportunities for high-risk individuals with familial breast cancer. PATIENTS AND METHODS: We carried out WGS on 78 paired germline and tumour DNA samples from individuals carrying pathogenic variants in BRCA1 (n = 26) or BRCA2 (n = 22) or from non-carriers (non-BRCA1/2; n = 30). RESULTS: Matched germline/tumour WGS and somatic mutational signature analysis revealed patients with unreported, dual pathogenic germline variants in cancer risk genes (BRCA1/BRCA2; BRCA1/MUTYH). The strategy identified that 100% of tumours from BRCA1 carriers and 91% of tumours from BRCA2 carriers exhibited biallelic inactivation of the respective gene, together with somatic mutational signatures suggestive of a functional deficiency in homologous recombination. A set of non-BRCA1/2 tumours also had somatic signatures indicative of BRCA-deficiency, including tumours with BRCA1 promoter methylation, and tumours from carriers of a PALB2 pathogenic germline variant and a BRCA2 variant of uncertain significance. A subset of 13 non-BRCA1/2 tumours from early onset cases were BRCA-proficient, yet displayed complex clustered structural rearrangements associated with the amplification of oncogenes and pathogenic germline variants in TP53, ATM and CHEK2. CONCLUSIONS: Our study highlights the role that WGS of matched germline/tumour DNA and the somatic mutational signatures can play in the discovery of pathogenic germline variants and for providing supporting evidence for variant pathogenicity. WGS-derived signatures were more robust than germline status and other genomic predictors of homologous recombination deficiency, thus impacting the selection of platinum-based or PARP inhibitor therapy. In this first examination of non-BRCA1/2 tumours by WGS, we illustrate the considerable heterogeneity of these tumour genomes and highlight that complex genomic rearrangements may drive tumourigenesis in a subset of cases.

Poster - Thurs Eve-42: A revision of the γ-evaluation: Initial interpretation of dose disagreements on γ-vector fields.

Gamma evaluations are a common clinical tool used as a quantitative comparison between dose-distributions, combining both dose difference and distance to agreement criteria. Because gamma evaluations permit rapid analysis of agreement between complex dose distributions, they are often a preferred comparison method for assessing delivery of conformal radiotherapy distributions. Although the comparison provides a useful measure of agreement between distributions when the index is less than one, the scalar gamma value provides little information into the clinical significance or source of disagreements of failing gamma values (i.e., when γ>1). Previously, Stock et al., have presented the gamma angle as an indicator of the relative influence of the distance to agreement versus the dose difference on gamma. We present a modification to the gamma evaluation such that the complete 3D gamma vector information is considered. The predictive nature of each vector component was investigated by simulating various dose disagreements in test distributions. Misalignment tests revealed that the mean gamma vector components indicate the offset direction and relative magnitude for all test distributions. The mean dose component of the gamma vector was prognostic of double Gaussian overdoses and underdoses in a virtual conformal delivery. The response of the vector field depends on properties distinctive to each distribution, such as the local dose gradient. Understanding how these unique properties affect the vector field may permit better diagnosis of dose disagreement sources. Other vector field properties, such as curl and divergence, may yet provide more information for interpreting the cause and significance of γ>1.

Insights into the structure/function of hepatocyte growth factor/scatter factor from studies with individual domains.

Hepatocyte growth factor/scatter factor (HGF/SF), the ligand for the receptor tyrosine kinase encoded by the c-Met proto-oncogene, is a multidomain protein structurally related to the pro-enzyme plasminogen and with major roles in development, tissue regeneration and cancer. We have expressed the N-terminal (N) domain, the four kringle domains (K1 to K4) and the serine proteinase homology domain (SP) of HGF/SF individually in yeast or mammalian cells and studied their ability to: (i) bind the Met receptor as well as heparan sulphate and dermatan sulphate co-receptors, (ii) activate Met in target cells and, (iii) map their binding sites onto the beta-propeller domain of Met. The N, K1 and SP domains bound Met directly with comparable affinities (K(d)=2.4, 3.3 and 1.4 microM). The same domains also bound heparin with decreasing affinities (N>K1>>SP) but only the N domain bound dermatan sulphate. Three kringle domains (K1, K2 and K4) displayed agonistic activity on target cells. In contrast, the N and SP domains, although capable of Met binding, displayed no or little activity. Further, cross-linking experiments demonstrated that both the N domain and kringles 1-2 bind the beta-chain moiety (amino acid residues 308-514) of the Met beta-propeller. In summary, the K1, K2 and K4 domains of HGF/SF are sufficient for Met activation, whereas the N and SP domains are not, although the latter domains contribute additional binding sites necessary for receptor activation by full length HGF/SF. The results provide new insights into the structure/function of HGF/SF and a basis for engineering the N and K1 domains as receptor antagonists for cancer therapy.