No abstract

The main problems in drawing causal inferences from epidemiological case-control studies are confounding by unmeasured extraneous factors, selection bias and differential misclassification of exposure. In genetics the first of these, in the form of population structure, has dominated recent debate. Population structure explained part of the significant +11.2% inflation of test statistics we observed in an analysis of 6,322 nonsynonymous SNPs in 816 cases of type 1 diabetes and 877 population-based controls from Great Britain. The remainder of the inflation resulted from differential bias in genotype scoring between case and control DNA samples, which originated from two laboratories, causing false-positive associations. To avoid excluding SNPs and losing valuable information, we extended the genomic control method by applying a variable downweighting to each SNP.

SUMMARY Background Diffuse large-B-cell lymphoma (DLBCL) is curable but when treatment fails, outcome is poor. Imaging scans help identify patients at risk of treatment failure but are often imprecise, and the radiation exposure is a potential health risk. Specific, sensitive and readily available biomarkers of treatment failure are needed. Methods We retrospectively analyzed cell-free circulating tumor DNA (ctDNA) in patients treated on one of 3 treatment protocols using quantitative next-generation DNA sequencing. Eligible patients had DLBCL, no evidence of indolent lymphoma and were previously untreated. Serial serum samples and concurrent computed tomography scans were obtained at specified times during most treatment cycles and 5-years of follow-up. VDJ gene segments of the rearranged immunoglobulin receptor genes were amplified and sequenced from pre-treatment specimens and serum ctDNA encoding the VDJ rearrangements was quantitated. Findings Tumor clonotype(s) were identified in pretreatment specimens from 126 patients who were followed for a median (interquartile range) of 11 (6.8 to 14.2) years. Interim ctDNA monitoring at the end of 2 treatment cycles in 108 patients showed a time to progression (TTP) of 41.7% (95% Confidence Interval (CI): 22.2% to 60.1%) and 80.2% (95% CI: 69.6% to 87.3%), at 5-years (p<0.0001) in patients with and without detectable ctDNA, respectively, and a positive and negative predicative value (PPV and NPV) of 63% and 80%, respectively. Surveillance ctDNA monitoring was performed in 107 patients who achieved complete remission. A Cox proportional hazards model showed patients who developed detectable ctDNA during surveillance had a hazard ratio 228 times that of patients with undetectable ctDNA for clinical disease progression (95% CI: 51 to 1022) (p<0.0001). Surveillance ctDNA had a PPV and NPV of 88% and 98%, respectively, and identified recurrence a median (range) of 3.5 months (0 to 200) before evidence of clinical disease. Interpretation Surveillance ctDNA identifies patients at risk of recurrence before clinical evidence of disease in most patients and results in lower disease burden at relapse. Interim ctDNA is a promising biomarker to identify patients at high risk of treatment failure.

Key Points• MRD assessment by sequencing is prognostic of TTP and OS in multiple myeloma patients.• Among patients in complete response, MRD assessment by sequencing enables identification of 2 distinct subgroups with different TTP.We assessed the prognostic value of minimal residual disease (MRD) detection in multiple myeloma (MM) patients using a sequencing-based platform in bone marrow samples from 133 MM patients in at least very good partial response (VGPR) after front-line therapy.Deep sequencing was carried out in patients in whom a high-frequency myeloma clone was identified and MRD was assessed using the IGH-VDJ H , IGH-DJ H , and IGK assays. The results were contrasted with those of multiparametric flow cytometry (MFC) and allelespecific oligonucleotide polymerase chain reaction (ASO-PCR).

Cytotoxic T lymphocyte–associated antigen-4 (CTLA-4) blockade can promote antitumor T cell immunity and clinical responses. The mechanism by which anti–CTLA-4 antibodies induces antitumor responses is controversial. To determine the effects of CTLA-4 blockade on the T cell repertoire, we used next-generation deep sequencing to measure the frequency of individual rearranged T cell receptor β (TCRβ) genes, thereby characterizing the diversity of rearrangements, known as T cell clonotypes. CTLA-4 blockade in patients with metastatic castration-resistant prostate cancer and metastatic melanoma resulted in both expansion and loss of T cell clonotypes, consistent with a global turnover of the T cell repertoire. Overall, this treatment increased TCR diversity as reflected in the number of unique TCR clonotypes. The repertoire of clonotypes continued to evolve over subsequent months of treatment. Whereas the number of clonotypes that increased with treatment was not associated with clinical outcome, improved overall survival was associated with maintenance of high-frequency clones at baseline. In contrast, the highest-frequency clonotypes fell with treatment in patients with short overall survival. Stably maintained clonotypes included T cells having high-avidity TCR such as virus-reactive T cells. Together, these results suggest that CTLA-4 blockade induces T cell repertoire evolution and diversification. Moreover, improved clinical outcomes are associated with less clonotype loss, consistent with the maintenance of high-frequency TCR clonotypes during treatment. These clones may represent the presence of preexisting high-avidity T cells that may be relevant in the antitumor response.

Key Points MRD using NGS-identified patients with an excellent outcome in multiple myeloma. MRD should be assessed in every prospective trial, and is a candidate to become a primary end point.

Progressive myoclonus epilepsy of the Unverricht-Lundborg type (EPM1) is an autosomal recessive inherited form of epilepsy, previously linked to human chromosome 21q22.3. The gene encoding cystatin B was shown to be localized to this region, and levels of messenger RNA encoded by this gene were found to be decreased in cells from affected individuals. Two mutations, a 3' splice site mutation and a stop codon mutation, were identified in the gene encoding cystatin B in EPM1 patients but were not present in unaffected individuals. These results provide evidence that mutations in the gene encoding cystatin B are responsible for the primary defect in patients with EPM1.

Early events in neuronal differentiation are generally considered to be regulated by factors independent of alterations in membrane permeability. Weaver mice harbour a mutation that blocks neuronal differentiation just after cessation of cell division, prior to cell migration and synaptogenesis. Cerebellar granule cells in homozygous weaver mice fail to differentiate, either because intrinsic cues are absent or because the granule cells are unable to respond to those cues. We now report that weaver mice have a missense mutation in a gene encoding a G-protein coupled inward rectifier potassium channel. The mutation alters the putative ion-permeable, pore-forming domain of the protein, suggesting that granule cell differentiation is regulated by changes in membrane permeability.