Structural variants shape the genomic landscape and clinical outcome of multiple myeloma

Deciphering genomic architecture is key to identifying novel disease drivers and understanding the mechanisms underlying myeloma initiation and progression. In this work, using the CoMMpass dataset, we show that structural variants (SV) occur in a nonrandom fashion throughout the genome with an increased frequency in the t(4;14), RB1, or TP53 mutated cases and reduced frequency in t(11;14) cases. By mapping sites of chromosomal rearrangements to topologically associated domains and identifying significantly upregulated genes by RNAseq we identify both predicted and novel putative driver genes. These data highlight the heterogeneity of transcriptional dysregulation occurring as a consequence of both the canonical and novel structural variants. Further, it shows that the complex rearrangements chromoplexy, chromothripsis and templated insertions are common in MM with each variant having its own distinct frequency and impact on clinical outcome. Chromothripsis is associated with a significant independent negative impact on clinical outcome in newly diagnosed cases consistent with its use alongside other clinical and genetic risk factors to identify prognosis

BRAF and DIS3 Mutations Associate with Adverse Outcome in a Long-term Follow-up of Patients with Multiple Myeloma

Purpose: Copy number changes and translocations have been studied extensively in many datasets with long term follow-up. The impact of mutations remains debated given the short time to follow-up of most datasets. Methods: we performed targeted panel sequencing covering 125 myeloma-specific genes and the loci involved in translocations in 223 newly diagnosed myeloma samples recruited into one of the Total Therapy Trials (TT). Results: As expected, the most commonly mutated genes were NRAS, KRAS, and BRAF making up 44% of patients. Double-Hit, BRAF and DIS3 mutations had an impact on outcome alongside classical risk factors in the context of an intensive treatment approach. We were able to identify both V600E and non-V600E BRAF mutations, 58% of which were predicted to be hypoactive or kinase dead. Interestingly, 44% of the hypoactive/kinase dead BRAF mutated patients showed co-occurring alterations in KRAS, NRAS or activating BRAF mutations suggesting they play a role in the oncogenesis of multiple myeloma (MM) by facilitating MAPK activation and may lead to chemo resistance. Conclusion: Overall, these data highlight the importance of mutational screening to better understand newly diagnosed MM (NDMM) and may lead to patient specific mutation-driven treatment approaches

Kryptoracemates

Racemic crystals normally crystallise in centrosymmetric spacegroups containing equal numbers of enantiomers. More rarely, racemates may crystallise in non-centrosymmetric space-groups having glide symmetry or, even more rarely, in space-groups devoid of a centre of inversion, having no rotary-inversion axes nor glide plane. The latter class of crystals form the subject of the present bibliographic review – a survey of kryptoracemic behaviour. The term kryptoracemic alludes to the presence of a hidden or non-crystallographic centre of inversion between two molecules that might otherwise be expected to crystallise in an achiral space-group, often about a centre of inversion. Herein, examples of molecules with stereogenic centres crystallising in one of the 65 Sohncke space-groups are described. Genuine kryptoracemates, i.e. crystals comprising only enantiomorphous pairs are described followed by an overview of non-genuine kryptoracemates whereby the crystal also contains other species such as solvent and/or counterions. A full range, i.e. one to six, stereogenic centres are noted in genuine kryptoracemates. Examples will also be described whereby there are more that one enantiomeric pair of molecules in the crystallographic asymmetric unit. A more diverse range of examples are available for non-genuine kryptoracemates. There are unbalanced species where in addition to the enantiomeric pair of molecules, there is another enantiomeric molecule present. There are examples of genuine co-crystals, solvated species and of salts. Finally, special examples will be highlighted where the counterions are chiral and where they are disparate, both circumstances promoting kryptoracemic behaviour

Single-dose administration and the influence of the timing of the booster dose on immunogenicity and efficacy of ChAdOx1 nCoV-19 (AZD1222) vaccine: a pooled analysis of four randomised trials.

BACKGROUND: The ChAdOx1 nCoV-19 (AZD1222) vaccine has been approved for emergency use by the UK regulatory authority, Medicines and Healthcare products Regulatory Agency, with a regimen of two standard doses given with an interval of 4-12 weeks. The planned roll-out in the UK will involve vaccinating people in high-risk categories with their first dose immediately, and delivering the second dose 12 weeks later. Here, we provide both a further prespecified pooled analysis of trials of ChAdOx1 nCoV-19 and exploratory analyses of the impact on immunogenicity and efficacy of extending the interval between priming and booster doses. In addition, we show the immunogenicity and protection afforded by the first dose, before a booster dose has been offered. METHODS: We present data from three single-blind randomised controlled trials-one phase 1/2 study in the UK (COV001), one phase 2/3 study in the UK (COV002), and a phase 3 study in Brazil (COV003)-and one double-blind phase 1/2 study in South Africa (COV005). As previously described, individuals 18 years and older were randomly assigned 1:1 to receive two standard doses of ChAdOx1 nCoV-19 (5 × 1010 viral particles) or a control vaccine or saline placebo. In the UK trial, a subset of participants received a lower dose (2·2 × 1010 viral particles) of the ChAdOx1 nCoV-19 for the first dose. The primary outcome was virologically confirmed symptomatic COVID-19 disease, defined as a nucleic acid amplification test (NAAT)-positive swab combined with at least one qualifying symptom (fever ≥37·8°C, cough, shortness of breath, or anosmia or ageusia) more than 14 days after the second dose. Secondary efficacy analyses included cases occuring at least 22 days after the first dose. Antibody responses measured by immunoassay and by pseudovirus neutralisation were exploratory outcomes. All cases of COVID-19 with a NAAT-positive swab were adjudicated for inclusion in the analysis by a masked independent endpoint review committee. The primary analysis included all participants who were SARS-CoV-2 N protein seronegative at baseline, had had at least 14 days of follow-up after the second dose, and had no evidence of previous SARS-CoV-2 infection from NAAT swabs. Safety was assessed in all participants who received at least one dose. The four trials are registered at ISRCTN89951424 (COV003) and ClinicalTrials.gov, NCT04324606 (COV001), NCT04400838 (COV002), and NCT04444674 (COV005). FINDINGS: Between April 23 and Dec 6, 2020, 24 422 participants were recruited and vaccinated across the four studies, of whom 17 178 were included in the primary analysis (8597 receiving ChAdOx1 nCoV-19 and 8581 receiving control vaccine). The data cutoff for these analyses was Dec 7, 2020. 332 NAAT-positive infections met the primary endpoint of symptomatic infection more than 14 days after the second dose. Overall vaccine efficacy more than 14 days after the second dose was 66·7% (95% CI 57·4-74·0), with 84 (1·0%) cases in the 8597 participants in the ChAdOx1 nCoV-19 group and 248 (2·9%) in the 8581 participants in the control group. There were no hospital admissions for COVID-19 in the ChAdOx1 nCoV-19 group after the initial 21-day exclusion period, and 15 in the control group. 108 (0·9%) of 12 282 participants in the ChAdOx1 nCoV-19 group and 127 (1·1%) of 11 962 participants in the control group had serious adverse events. There were seven deaths considered unrelated to vaccination (two in the ChAdOx1 nCov-19 group and five in the control group), including one COVID-19-related death in one participant in the control group. Exploratory analyses showed that vaccine efficacy after a single standard dose of vaccine from day 22 to day 90 after vaccination was 76·0% (59·3-85·9). Our modelling analysis indicated that protection did not wane during this initial 3-month period. Similarly, antibody levels were maintained during this period with minimal waning by day 90 (geometric mean ratio [GMR] 0·66 [95% CI 0·59-0·74]). In the participants who received two standard doses, after the second dose, efficacy was higher in those with a longer prime-boost interval (vaccine efficacy 81·3% [95% CI 60·3-91·2] at ≥12 weeks) than in those with a short interval (vaccine efficacy 55·1% [33·0-69·9] at <6 weeks). These observations are supported by immunogenicity data that showed binding antibody responses more than two-fold higher after an interval of 12 or more weeks compared with an interval of less than 6 weeks in those who were aged 18-55 years (GMR 2·32 [2·01-2·68]). INTERPRETATION: The results of this primary analysis of two doses of ChAdOx1 nCoV-19 were consistent with those seen in the interim analysis of the trials and confirm that the vaccine is efficacious, with results varying by dose interval in exploratory analyses. A 3-month dose interval might have advantages over a programme with a short dose interval for roll-out of a pandemic vaccine to protect the largest number of individuals in the population as early as possible when supplies are scarce, while also improving protection after receiving a second dose. FUNDING: UK Research and Innovation, National Institutes of Health Research (NIHR), The Coalition for Epidemic Preparedness Innovations, the Bill & Melinda Gates Foundation, the Lemann Foundation, Rede D'Or, the Brava and Telles Foundation, NIHR Oxford Biomedical Research Centre, Thames Valley and South Midland's NIHR Clinical Research Network, and AstraZeneca

Global methylation analysis identifies prognostically important epigenetically inactivated tumor suppressor genes in multiple myeloma

Outcome in multiple myeloma is highly variable and a better understanding of the factors that influence disease biology is essential to understand and predict behavior in individual patients. In the present study, we analyzed combined genomewide DNA methylation and gene expression data of patients treated in the Medical Research Council Myeloma IX trial. We used these data to identify epigenetically repressed tumor suppressor genes with prognostic relevance in myeloma. We identified 195 genes with changes in methylation status that were significantly associated with prognosis. Combining DNA methylation and gene expression data led to the identification of the epigenetically regulated tumor modulating genes GPX3, RBP1, SPARC, and TGFBI. Hypermethylation of these genes was associated with significantly shorter overall survival, independent of age, International Staging System score, and adverse cytogenetics. The 4 differentially methylated and expressed genes are known to mediate important tumor suppressive functions including response to chemotherapy (TGFBI), interaction with the microenvironment (SPARC), retinoic acid signaling (RBP1), and the response to oxidative stress (GPX3), which could explain the prognostic impact of their differential methylation. Assessment of the DNA methylation status of the identified genes could contribute to the molecular characterization of myeloma, which is prerequisite for an individualized treatment approac

Translocations at 8q24 juxtapose MYC with genes that harbor superenhancers resulting in overexpression and poor prognosis in myeloma patients.

Secondary MYC translocations in myeloma have been shown to be important in the pathogenesis and progression of disease. Here, we have used a DNA capture and massively parallel sequencing approach to identify the partner chromosomes in 104 presentation myeloma samples. 8q24 breakpoints were identified in 21 (20%) samples with partner loci including IGH, IGK and IGL, which juxtapose the immunoglobulin (Ig) enhancers next to MYC in 8/23 samples. The remaining samples had partner loci including XBP1, FAM46C, CCND1 and KRAS, which are important in B-cell maturation or myeloma pathogenesis. Analysis of the region surrounding the breakpoints indicated the presence of superenhancers on the partner chromosomes and gene expression analysis showed increased expression of MYC in these samples. Patients with MYC translocations had a decreased progression-free and overall survival. We postulate that translocation breakpoints near MYC result in colocalization of the gene with superenhancers from loci, which are important in the development of the cell type in which they occur. In the case of myeloma these are the Ig loci and those important for plasma cell development and myeloma pathogenesis, resulting in increased expression of MYC and an aggressive disease phenotype

Search for rare protein altering variants influencing susceptibility to multiple myeloma

The genetic basis underlying the inherited risk of developing multiple myeloma (MM) is largely unknown. To examine the impact of rare protein altering variants on the risk of developing MM we analyzed high-coverage exome sequencing data on 513 MM cases and 1,569 healthy controls, performing both single variant and gene burden tests. We did not identify any recurrent coding low-frequency alleles (1–5%) with moderate effect that were statistically associated with MM. In a gene burden analysis we did however identify a promising relationship between variation in the marrow kinetochore microtubule stromal gene KIF18A, which plays a role in control mitotic chromosome positioning dynamics, and risk of MM (P =3.6x10−6). Further analysis showed KIF18A displays a distinct pattern of expression across molecular subgroups of MM as well as being associated with patient survival. Our results inform future study design and provide a resource for contextualizing the impact of candidate MM susceptibility genes