Transplantation of hematopoietic stem cells from partially matched family donors is a promising therapy for patients who have a hematologic cancer and are at high risk for relapse. The donor T-cell infusions associated with such transplantation can promote post-transplantation immune reconstitution and control residual disease. We identified 43 patients who underwent haploidentical transplantation and infusion of donor T cells for acute myeloid leukemia or myelodysplastic syndrome and conducted post-transplantation studies that included morphologic examination of bone marrow, assessment of hematopoietic chimerism with the use of short-tandem-repeat amplification, and HLA typing. The genomic rearrangements in mutant variants of leukemia were studied with the use of genomic HLA typing, microsatellite mapping, and single-nucleotide–polymorphism arrays. The post-transplantation immune responses against the original cells and the mutated leukemic cells were analyzed with the use of mixed lymphocyte cultures. In 5 of 17 patients with leukemia relapse after haploidentical transplantation and infusion of donor T cells, we identified mutant variants of the original leukemic cells. In the mutant leukemic cells, the HLA haplotype that differed from the donor's haplotype had been lost because of acquired uniparental disomy of chromosome 6p. T cells from the donor and the patient after transplantation did not recognize the mutant leukemic cells, whereas the original leukemic cells taken at the time of diagnosis were efficiently recognized and killed. After transplantation of haploidentical hematopoietic stem cells and infusion of donor T cells, leukemic cells can escape from the donor's antileukemic T cells through the loss of the mismatched HLA haplotype. This event leads to relapse.

Gonçalo Abecasis, Francesco Cucca, David Schlessinger, Serena Sanna and colleagues report ~17.6 million genetic variants from whole-genome sequencing of 2,120 Sardinians. They assess the impact of these variants on circulating lipid levels and five inflammatory biomarkers. We report ∼17.6 million genetic variants from whole-genome sequencing of 2,120 Sardinians; 22% are absent from previous sequencing-based compilations and are enriched for predicted functional consequences. Furthermore, ∼76,000 variants common in our sample (frequency >5%) are rare elsewhere (<0.5% in the 1000 Genomes Project). We assessed the impact of these variants on circulating lipid levels and five inflammatory biomarkers. We observe 14 signals, including 2 major new loci, for lipid levels and 19 signals, including 2 new loci, for inflammatory markers. The new associations would have been missed in analyses based on 1000 Genomes Project data, underlining the advantages of large-scale sequencing in this founder population.

Genomewide association studies of autoimmune diseases have mapped hundreds of susceptibility regions in the genome. However, only for a few association signals has the causal gene been identified, and for even fewer have the causal variant and underlying mechanism been defined. Coincident associations of DNA variants affecting both the risk of autoimmune disease and quantitative immune variables provide an informative route to explore disease mechanisms and drug-targetable pathways. Using case–control samples from Sardinia, Italy, we performed a genomewide association study in multiple sclerosis followed by TNFSF13B locus–specific association testing in systemic lupus erythematosus (SLE). Extensive phenotyping of quantitative immune variables, sequence-based fine mapping, cross-population and cross-phenotype analyses, and gene-expression studies were used to identify the causal variant and elucidate its mechanism of action. Signatures of positive selection were also investigated. A variant in TNFSF13B, encoding the cytokine and drug target B-cell activating factor (BAFF), was associated with multiple sclerosis as well as SLE. The disease-risk allele was also associated with up-regulated humoral immunity through increased levels of soluble BAFF, B lymphocytes, and immunoglobulins. The causal variant was identified: an insertion–deletion variant, GCTGT→A (in which A is the risk allele), yielded a shorter transcript that escaped microRNA inhibition and increased production of soluble BAFF, which in turn up-regulated humoral immunity. Population genetic signatures indicated that this autoimmunity variant has been evolutionarily advantageous, most likely by augmenting resistance to malaria. A TNFSF13B variant was associated with multiple sclerosis and SLE, and its effects were clarified at the population, cellular, and molecular levels. (Funded by the Italian Foundation for Multiple Sclerosis and others.)

Methylation at 5-cytosine (5-mC) is a fundamental epigenetic DNA modification associated recently with cardiac disease. In contrast, the role of 5-hydroxymethylcytosine (5-hmC)—5-mC’s oxidation product—in cardiac biology and disease is unknown. Here we assess the hydroxymethylome in embryonic, neonatal, adult and hypertrophic mouse cardiomyocytes, showing that dynamic modulation of hydroxymethylated DNA is associated with specific transcriptional networks during heart development and failure. DNA hydroxymethylation marks the body of highly expressed genes as well as distal regulatory regions with enhanced activity. Moreover, pathological hypertrophy is characterized by a shift towards a neonatal 5-hmC distribution pattern. We also show that the ten-eleven translocation 2 (TET2) enzyme regulates the expression of key cardiac genes, such as Myh7, through 5-hmC deposition on the gene body and at enhancers. Thus, we provide a genome-wide analysis of 5-hmC in the cardiomyocyte and suggest a role for this epigenetic modification in heart development and disease.

DNA sequencing identifies common and rare genetic variants for association studies, but studies typically focus on variants in nuclear DNA and ignore the mitochondrial genome. In fact, analyzing variants in mitochondrial DNA (mtDNA) sequences presents special problems, which we resolve here with a general solution for the analysis of mtDNA in next-generation sequencing studies. The new program package comprises 1) an algorithm designed to identify mtDNA variants (i.e., homoplasmies and heteroplasmies), incorporating sequencing error rates at each base in a likelihood calculation and allowing allele fractions at a variant site to differ across individuals; and 2) an estimation of mtDNA copy number in a cell directly from whole-genome sequencing data. We also apply the methods to DNA sequence from lymphocytes of ~2,000 SardiNIA Project participants. As expected, mothers and offspring share all homoplasmies but a lesser proportion of heteroplasmies. Both homoplasmies and heteroplasmies show 5-fold higher transition/transversion ratios than variants in nuclear DNA. Also, heteroplasmy increases with age, though on average only ~1 heteroplasmy reaches the 4% level between ages 20 and 90. In addition, we find that mtDNA copy number averages ~110 copies/lymphocyte and is ~54% heritable, implying substantial genetic regulation of the level of mtDNA. Copy numbers also decrease modestly but significantly with age, and females on average have significantly more copies than males. The mtDNA copy numbers are significantly associated with waist circumference (p-value = 0.0031) and waist-hip ratio (p-value = 2.4×10^-5), but not with body mass index, indicating an association with central fat distribution. To our knowledge, this is the largest population analysis to date of mtDNA dynamics, revealing the age-imposed increase in heteroplasmy, the relatively high heritability of copy number, and the association of copy number with metabolic traits. We present a new program that provides a general solution for the analysis of variation of mtDNA (the small circular genome in mitochondria, separate from the DNA in the nucleus). This is needed because many large-scale genetic studies are using new DNA sequencing technologies to help assess genetic variation and its effects on disease, but the mitochondrial genome is often ignored because it exists in many copies in a cell, complicating analyses. Our approach both identifies variants on mitochondrial genome and estimates mtDNA copy number. Applying the programs to DNA sequence from ~2,000 SardiNIA project participants, we show that heteroplasmies (mtDNA variants with more than one allele at a DNA site) increase with age, and that copy number is relatively highly heritable and is correlated with metabolic traits, particularly central fat levels. The program package can facilitate comprehensive mtDNA analysis from any whole-genome sequencing data, with an increase in the understanding of mtDNA dynamics and its potential role in aging and metabolism.