A de novo frameshift variant in MED13 gene in a patient with autism spectrum disorder and magnetic resonance imaging abnormalities mimicking tuberous sclerosis.

The mediator complex subunit 13 (MED13) gene is implicated in neurodevelopmental disorders including autism spectrum disorder (ASD), intellectual disability, and speech delay with varying severity and course. Additional, extra central nervous system, features include eye or vision problems, hypotonia, congenital heart abnormalities, and dysmorphisms. We describe a 7-year- and 4-month-old girl evaluated for ASD whose brain magnetic resonance imaging was suggestive of multiple cortical tubers. The exome sequencing (ES - trio analysis) uncovered a unique, de novo, frameshift variant in the MED13 gene (c.4880del, D1627Vfs*17), with a truncating effect on the protein. This case report thus expands the phenotypic spectrum of MED13-related disorders to include brain abnormalities.

A Novel Heterozygous Mutation c.1627G>T (p.Gly543Cys) in the SLC34A1 Gene in a Male Patient with Recurrent Nephrolithiasis and Early Onset Osteopenia: A Case Report.

Serum phosphate concentration is regulated by renal phosphate reabsorption and mediated by sodium-phosphate cotransporters. Germline mutations in genes encoding these cotransporters have been associated with clinical phenotypes, variably characterized by hyperphosphaturia, hypophosphatemia, recurrent kidney stones, skeletal demineralization, and early onset osteoporosis. We reported a 33-year-old male patient presenting a history of recurrent nephrolithiasis and early onset osteopenia in the lumbar spine and femur. He was tested, through next generation sequencing (NGS), by using a customized multigenic panel containing 33 genes, whose mutations are known to be responsible for the development of congenital parathyroid diseases. Two further genes, SLC34A1 and SLC34A3, encoding two sodium-phosphate cotransporters, were additionally tested. A novel germline heterozygous mutation was identified in the SLC34A1 gene, c.1627G>T (p.Gly543Cys), currently not reported in databases of human gene mutations and scientific literature. SLC34A1 germline heterozygous mutations have been associated with the autosomal dominant hypophosphatemic nephrolithiasis/osteoporosis type 1 (NPHLOP1). Consistently, alongside the clinical features of NPHLOP1, our patient experienced recurrent nephrolithiasis and lumbar and femoral osteopenia at a young age. Genetic screening for the p.Gly453Cys variant and the clinical characterization of his first-degree relatives associated the presence of the variant in one younger brother, presenting renal colic and microlithiasis, suggesting p.Gly453Cys is possibly associated with renal altered function in the NPHLOP1 phenotype.

Genomic history of the Italian population recapitulates key evolutionary dynamics of both Continental and Southern Europeans.

BACKGROUND: The cline of human genetic diversity observable across Europe is recapitulated at a micro-geographic scale by variation within the Italian population. Besides resulting from extensive gene flow, this might be ascribable also to local adaptations to diverse ecological contexts evolved by people who anciently spread along the Italian Peninsula. Dissecting the evolutionary history of the ancestors of present-day Italians may thus improve the understanding of demographic and biological processes that contributed to shape the gene pool of European populations. However, previous SNP array-based studies failed to investigate the full spectrum of Italian variation, generally neglecting low-frequency genetic variants and examining a limited set of small effect size alleles, which may represent important determinants of population structure and complex adaptive traits. To overcome these issues, we analyzed 38 high-coverage whole-genome sequences representative of population clusters at the opposite ends of the cline of Italian variation, along with a large panel of modern and ancient Euro-Mediterranean genomes. RESULTS: We provided evidence for the early divergence of Italian groups dating back to the Late Glacial and for Neolithic and distinct Bronze Age migrations having further differentiated their gene pools. We inferred adaptive evolution at insulin-related loci in people from Italian regions with a temperate climate, while possible adaptations to pathogens and ultraviolet radiation were observed in Mediterranean Italians. Some of these adaptive events may also have secondarily modulated population disease or longevity predisposition. CONCLUSIONS: We disentangled the contribution of multiple migratory and adaptive events in shaping the heterogeneous Italian genomic background, which exemplify population dynamics and gene-environment interactions that played significant roles also in the formation of the Continental and Southern European genomic landscapes.

Distinct Viral and Mutational Spectrum of Endemic Burkitt Lymphoma.

Endemic Burkitt lymphoma (eBL) is primarily found in children in equatorial regions and represents the first historical example of a virus-associated human malignancy. Although Epstein-Barr virus (EBV) infection and MYC translocations are hallmarks of the disease, it is unclear whether other factors may contribute to its development. We performed RNA-Seq on 20 eBL cases from Uganda and showed that the mutational and viral landscape of eBL is more complex than previously reported. First, we found the presence of other herpesviridae family members in 8 cases (40%), in particular human herpesvirus 5 and human herpesvirus 8 and confirmed their presence by immunohistochemistry in the adjacent non-neoplastic tissue. Second, we identified a distinct latency program in EBV involving lytic genes in association with TCF3 activity. Third, by comparing the eBL mutational landscape with published data on sporadic Burkitt lymphoma (sBL), we detected lower frequencies of mutations in MYC, ID3, TCF3 and TP53, and a higher frequency of mutation in ARID1A in eBL samples. Recurrent mutations in two genes not previously associated with eBL were identified in 20% of tumors: RHOA and cyclin F (CCNF). We also observed that polyviral samples showed lower numbers of somatic mutations in common altered genes in comparison to sBL specimens, suggesting dual mechanisms of transformation, mutation versus virus driven in sBL and eBL respectively.

Epigenetically induced ectopic expression of UNCX impairs the proliferation and differentiation of myeloid cells.

We here describe a leukemogenic role of the homeobox gene UNCX, activated by epigenetic modifications in acute myeloid leukemia (AML). We found the ectopic activation of UNCX in a leukemia patient harboring a t(7;10)(p22;p14) translocation, in 22 of 61 of additional cases [a total of 23 positive patients out of 62 (37.1%)], and in 6 of 75 (8%) of AML cell lines. UNCX is embedded within a low-methylation region (canyon) and encodes for a transcription factor involved in somitogenesis and neurogenesis, with specific expression in the eye, brain, and kidney. UNCX expression turned out to be associated, and significantly correlated, with DNA methylation increase at its canyon borders based on data in our patients and in archived data of patients from The Cancer Genome Atlas. UNCX-positive and -negative patients displayed significant differences in their gene expression profiles. An enrichment of genes involved in cell proliferation and differentiation, such as MAP2K1 and CCNA1, was revealed. Similar results were obtained in UNCX-transduced CD34+ cells, associated with low proliferation and differentiation arrest. Accordingly, we showed that UNCX expression characterizes leukemia cells at their early stage of differentiation, mainly M2 and M3 subtypes carrying wild-type NPM1 We also observed that UNCX expression significantly associates with an increased frequency of acute promyelocytic leukemia with PML-RARA and AML with t(8;21)(q22;q22.1); RUNX1-RUNX1T1 classes, according to the World Health Organization disease classification. In summary, our findings suggest a novel leukemogenic role of UNCX, associated with epigenetic modifications and with impaired cell proliferation and differentiation in AML.

The epigenetic landscape of age-related diseases: the geroscience perspective.

In this review, we summarize current knowledge regarding the epigenetics of age-related diseases, focusing on those studies that have described DNA methylation landscape in cardio-vascular diseases, musculoskeletal function and frailty. We stress the importance of adopting the conceptual framework of "geroscience", which starts from the observation that advanced age is the major risk factor for several of these pathologies and aims at identifying the mechanistic links between aging and age-related diseases. DNA methylation undergoes a profound remodeling during aging, which includes global hypomethylation of the genome, hypermethylation at specific loci and an increase in inter-individual variation and in stochastic changes of DNA methylation values. These epigenetic modifications can be an important contributor to the development of age-related diseases, but our understanding on the complex relationship between the epigenetic signatures of aging and age-related disease is still poor. The most relevant results in this field come from the use of the so called "epigenetics clocks" in cohorts of subjects affected by age-related diseases. We report these studies in final section of this review.

Bile salt export pump deficiency disease: two novel, late onset, ABCB11 mutations identified by next generation sequencing.

 Progressive familial intrahepatic cholestasis (PFIC) is a heterogeneous group of autosomal recessive cholestatic diseases of childhood and represents the main indication for liver transplantation at this age; PFIC2 involves ABCB11 gene, that encodes the ATPdependent canalicular bile salt export pump (BSEP). Benign intrahepatic cholestasis (BRIC) identifies a group of diseases involving the same genes and characterized by intermittent attacks of cholestasis with no progression to liver cirrhosis. Diagnosis with standard sequencing techniques is expensive and available only at a few tertiary centers. We report the application of next generation sequencing (NGS) in the diagnosis of the familial intrahepatic cholestasis with a parallel sequencing of three causative genes. We identified the molecular defects in ABCB11 gene in two different probands who developed a severe cholestatic disease of unknown origin. In the first patient a compound heterozygosity for the novel frameshift mutation p.Ser1100GlnfsX38 and the missense variant p.Glu135Lys was detected. In the second patient, triggered by contraceptive therapy, we identified homozygosity for a novel missense variant p.Ala523Gly. In conclusion, these mutations seem to have a late onset and a less aggressive clinical impact, acting as an intermediate form between BRIC and PFIC.

Inferring the genetic history of lactase persistence along the Italian peninsula from a large genomic interval surrounding the LCT gene.

OBJECTIVE: Although genetic variants related to lactase persistence in European populations were supposed to have firstly undergone positive selection in farmers from the Balkans and Central Europe, demographic and evolutionary dynamics that subsequently shaped the distribution of this adaptive trait across the continent have still to be elucidated. To deepen the knowledge about potential routes of diffusion of lactase persistence to Western Europe we investigated variation at a large genomic region surrounding the LCT gene along the Italian peninsula, a geographical area that played a key role in population movements responsible for Neolithic diffusion across Europe. METHODS: By genotyping 40 highly selected SNPs in more than 400 Italian individuals we described gradients of nucleotide and haplotype variation potentially related to lactase persistence and compared them with those observed in several European and Mediterranean human groups. RESULTS: Multiple migratory events responsible for earlier introduction of the examined alleles in Italy than in Northern European regions could be invoked. Different demic processes occurred along the western and eastern sides of the peninsula were also inferred via linkage disequilibrium and population structure analyses. CONCLUSION: The appreciable genetic continuum observed between people from Northern or Central-Western Italy and Central European populations suggested a local arrival of lactase persistence-related variants mainly via overland routes. On the contrary, diversity of Central-Eastern and Southern Italian groups entailed also gene flow from South-Eastern Mediterranean regions, in accordance to the earlier entrance of the Neolithic in Southern Italy via maritime population movements along the Mediterranean coastlines.

High-throughput genotyping of high-risk Human Papillomavirus by MALDI-TOF Mass Spectrometry-based method.

A high-throughput matrix-assisted laser desorption ionisation-time of flight (MALDI-TOF) mass spectrometry (MS)-based method was here developed to genotype 16 high-risk human papillomavirus (HPV) types in cervical cytology specimens. This method was compared to a commercial kit, the Inno-LiPA HPV genotyping assay, which detects a broad spectrum of HPV types. HPV DNA was assessed by the two methods in a total of 325 cervical cytology specimens collected in PreservCyt® solution. The overall agreement was almost perfect (Cohen's k=0.86) in term of positive and negative cases. Indeed, HPV types 16, 35, 56 and 66 showed the highest agreement values (>0.80). The highest agreement values (K >0.80) were found for all 16 HPV types in single infections, but only for HPV 16, 35, 45 and 56 in multiple infections. In conclusion, the high-throughput MS-based method developed here is well-suited for broad spectrum HPV genotyping in large-scale epidemiological studies.

Glutathione transferase-A2 S112T polymorphism predicts survival, transplant-related mortality, busulfan and bilirubin blood levels after allogeneic stem cell transplantation.

Busulfan liver metabolism depends on glutathione, a crucial mediator of cellular and systemic stress. Here we investigated 40 polymorphisms at 27 loci involved in hepatic glutathione homeostasis, with the aim of testing their impact on the clinical outcome of 185 busulfan-conditioned allogeneic transplants. GSTA2 S112T serine allele homozygosity is an independent prognostic factor for poorer survival (RR=2.388), for increased any time- and 100-day transplant-related mortality (RR=4.912 and RR=5.185, respectively). The genotype also predicts a wider busulfan area under the concentration-time curve (1214.36 ± 570.06 vs. 838.10 ± 282.40 mMol*min) and higher post-transplant bilirubin serum levels (3.280 ± 0.422 vs. 1.874+0.197 mg/dL). In vitro, busulfan elicits pro-inflammatory activation (increased NF-KappaB activity and interleukin-8 expression) in human hepatoma cells. At the same time, the drug down-regulates a variety of genes involved in bilirubin liver clearance: constitutive androstane receptor, multidrug resistance-associated protein, solute carrier organic anion transporters, and even GSTA2. It is worthy of note that GSTA2 also acts as an intra-hepatic bilirubin binding protein. These data underline the prognostic value of GSTA2 genetic variability in busulfan-conditioned allotransplants and suggest a patho-physiological model in which busulfan-induced inflammation leads to the impairment of post-transplant bilirubin metabolism.

TRPV1 channels regulate cortical excitability in humans.

Studies in rodents show that transient receptor potential vanilloid 1 (TRPV1) channels regulate glutamate release at central and peripheral synapses. In humans, a number of nonsynonymous single-nucleotide polymorphisms (SNPs) have been described in the TRPV1 gene, and some of them significantly alter the functionality of the channel. To address the possible role of TRPV1 channels in the regulation of synaptic transmission in humans, we studied how TRPV1 genetic polymorphisms affect cortical excitability measured with transcranial magnetic stimulation (TMS). Two SNPs of the TRPV1 gene were selected and genotyped (rs222747 and rs222749) in a sample of 77 healthy subjects. In previous cell expression studies, the "G" allele of rs222747 was found to enhance the activity of the channel, whereas rs222749 had no functional effect. Allelic variants in the rs222749 region were not associated with altered cortical response to single, paired, and repetitive TMS. In contrast, subjects homozygous for the G allele in rs222747 exhibited larger short-interval intracortical facilitation (a measure of glutamate transmission) explored through paired-pulse TMS of the primary motor cortex. Recruitment curves, short-interval intracortical inhibition, intracortical facilitation, and long-interval intracortical inhibition were unchanged. LTP- and LTD-like plasticity explored through intermittent or continuous theta-burst stimulation was also similar in the "G" and "non-G" subjects. To our knowledge, our results provide the first evidence that TRPV1 channels regulate cortical excitability to paired-pulse stimulation in humans.

In hepatocellular carcinoma miR-519d is up-regulated by p53 and DNA hypomethylation and targets CDKN1A/p21, PTEN, AKT3 and TIMP2.

MiR-519d belongs to the chromosome 19 miRNA cluster (C19MC), the largest human miRNA cluster. One of its members, miR-519d, is over-expressed in hepatocellular carcinoma (HCC) and we characterized its contribution to hepatocarcinogenesis. In HCC cells, the over-expression of miR-519d promotes cell proliferation, invasion and impairs apoptosis following anticancer treatments. These functions are, at least in part, exerted through the direct targeting of CDKN1A/p21, PTEN, AKT3 and TIMP2. The mechanisms underlying miR-519d aberrant expression in HCC were assayed by genomic DNA amplification, methylation analysis and ChIP assay. The aberrant hypomethylation of C19MC and TP53 were respectively identified as an epigenetic change allowing the aberrant expression of miR-519d and one of the factors able to activate its transcription. In conclusion, we assessed the oncogenic role of miR-519d in HCC by characterizing its biological functions, including the modulation of response to anticancer treatments and by identifying CDKN1A/p21, PTEN, AKT3 and TIMP2 among its targets.

High frequency of heterozygous rare variants of the SLC34A1 and SLC9A3R1 genes in patients with atypical femur fracture.

OBJECTIVE: Atypical femur fractures (AFFs) are rare fragility fractures originating at the lateral cortex of the femur, affecting the subtrochanteric or diaphyseal area of thebone with a transverse morphology. Occurrence of AFF is specifically associated with a small number of rare monogenic congenital metabolic bone disorders, such as hypophosphatasia, and with long-term treatment with antiresorptiondrugs. The exact pathogenesis of these fractures remains poorly understood and, except for cases of diagnosed HPP or other AFF-causing bone diseases, it is not possible to assess which patients are at higher riskof developing AFFs as a consequence of anti-resorption therapy. DESIGN: We genetically screened 25 unrelated patients who had developed at least one AFF. INTERVENTION: Genetic screening was performed through a nextgeneration sequencing analysis with a customized panel containing 76 human genes involved in the regulation of the mineralization processWe genetically screened 25 unrelated patients who had developed at least one AFF. RESULTS: We found a relatively high frequency (32.0%) of heterozygous rare variants inthe SLC34A1 and SLC9A3R1 genes, two genes whose heterozygous inactivating mutations have been respectively associated with autosomal dominant hypophosphatemic nephrolithiasis/osteoporosis types 1 and 2 (NPHLOP1and NPHLOP2). Other heterozygous rare variants were found in the BMPR1B, CYP27B1, FBN1, MEPE, PIGO, and PHOSPHO1 genes, each in a single AFF case (4.0%). CONCLUSIONS AND RELEVANCE: Our findings suggest that rarevariants of SLC34A1 and SLC9A3R1 could represent a possible genetic risk factor for the occurrence of AFFs. On the other hand, AFFs could represent an unsuspected clinical manifestation and/or an anti-resorption therapycorrelatedadverse event in patients with NPHLOP disorders.

dHPLC efficiency for semi-automated cDNA-AFLP analyses and fragment collection in the apple scab-resistance gene model.

cDNA-AFLP analysis for transcript profiling has been successfully applied to study many plant biological systems, particularly plant-microbe interactions. However, the separation of cDNA-AFLP fragments by gel electrophoresis is reported to be labor-intensive with only limited potential for automation, and the collection of differential bands for gene identification is even more cumbersome. In this work, we present the use of dHPLC (denaturing high performance liquid chromatography) and automated DNA fragment collection using the WAVE(®) System to analyze and recover cDNA-AFLP fragments. The method is successfully applied to the Malus-Venturia inaequalis interaction, making it possible to collect 66 different transcript-derived fragments for apple genes putatively involved in the defense response activated by the HcrVf2 resistance gene. The results, validated by real time quantitative RT-PCR, were consistent with the plant-pathogen interaction under investigation and this further supports the suitability of dHPLC for cDNA-AFLP transcript profiling. Features and advantages of this new approach are discussed, evincing that it is an almost fully automatable and cost-effective solution for processing large numbers of samples and collecting differential genes involved in other biological processes and non-model plants.

A Targeted Epigenetic Clock for the Prediction of Biological Age.

Epigenetic clocks were initially developed to track chronological age, but accumulating evidence indicates that they can also predict biological age. They are usually based on the analysis of DNA methylation by genome-wide methods, but targeted approaches, based on the assessment of a small number of CpG sites, are advisable in several settings. In this study, we developed a targeted epigenetic clock purposely optimized for the measurement of biological age. The clock includes six genomic regions mapping in ELOVL2, NHLRC1, AIM2, EDARADD, SIRT7 and TFAP2E genes, selected from a re-analysis of existing microarray data, whose DNA methylation is measured by EpiTYPER assay. In healthy subjects (n = 278), epigenetic age calculated using the targeted clock was highly correlated with chronological age (Spearman correlation = 0.89). Most importantly, and in agreement with previous results from genome-wide clocks, epigenetic age was significantly higher and lower than expected in models of increased (persons with Down syndrome, n = 62) and decreased (centenarians, n = 106; centenarians' offspring, n = 143; nutritional intervention in elderly, n = 233) biological age, respectively. These results support the potential of our targeted epigenetic clock as a new marker of biological age and open its evaluation in large cohorts to further promote the assessment of biological age in healthcare practice.

Genetic variants of the NMDA receptor influence cortical excitability and plasticity in humans.

N-methyl-d-aspartate (NMDA) receptors play crucial roles in glutamate-mediated synaptic transmission and plasticity and are involved in a variety of brain functions. Specific single nucleotide polymorphisms (SNPs) in the genes encoding NMDA receptor subunits have been associated with some neuropsychiatric disorders involving altered glutamate transmission, but how these polymorphisms impact on synaptic function in humans is unknown. Here, the role of NMDA receptors in the control of cortical excitability and plasticity was explored by comparing the response to single, paired, and repetitive transcranial magnetic stimulations of the motor cortex in 77 healthy subjects carrying specific allelic variants of the NR1 subunit gene (GRIN1 rs4880213 and rs6293) or of the NR2B subunit gene (GRIN2B rs7301328, rs3764028, and rs1805247). Our results showed that individuals homozygous for the T allele in the rs4880213 GRIN1 SNP had reduced intracortical inhibition, as expected for enhanced glutamatergic excitation in these subjects. Furthermore, individuals carrying the G allele in the rs1805247 GRIN2B SNP show greater intracortical facilitation and greater long-term potentiation-like cortical plasticity after intermittent -burst stimulation. Our results provide novel insights into the function of NMDA receptors in the human brain and might contribute to the clarification of the synaptic bases of severe neuropsychiatric disorders associated with defective glutamate transmission.

Clinical, Radiometabolic and Immunologic Effects of Olaparib in Locally Advanced Triple Negative Breast Cancer: The OLTRE Window of Opportunity Trial.

INTRODUCTION: Olaparib is effective in metastatic triple negative breast cancer (TNBC) carrying germline mutations in DNA damage repair (DDR) genes BRCA1/2 (gBRCA-mut). The OLTRE window-of-opportunity trial preliminarily investigated potential pathologic, radiometabolic and immune biomarkers of early-response to olaparib in gBRCA-wild-type (wt) TNBC and, as proof-of-concept in gBRCA-mut HER2-negative BC. METHODS: Patients received olaparib for 3 weeks (3w) before standard neoadjuvant chemotherapy and underwent multiple FDG18-PET/CT scan (basal, after olaparib), clinical assessments (basal, every 3w), tumor biopsies and blood samplings (baseline, after olaparib). Clinical and radiometabolic responses were evaluated according to RECIST1.1 and PERCIST criteria. RESULTS: 27 patients with gBRCA-wt TNBC and 8 with gBRCA-mut BC (6 TNBC, 2 HR+/HER2-negative) were enrolled. Three (11.1%) patients showed mutations in non-BRCA1/2 DDR genes and 4 (14.8%) in other genes. 3w olaparib induced 16/35 and 15/27 partial clinical and radiometabolic responses, including in 40.7% and 50.0% gBRCA-wt patients. gBRCA-mut tumors presented numerically higher tumor-infiltrating lymphocytes (TILs) levels and PD-L1 positive tumors. Clinical responders experienced a reduction in T-regs/T-eff ratio (p=0.05), B and NK lymphocytes (p=0.003 both), with an average increase in T-helpers rate (p<0.001) and CD4/CD8 ratio (p=0.02). Ki67% and TILs did not vary significantly (p=0.67 and p=0.77). A numerical increase in PD-L1 positive cases after olaparib was observed, though non-significant (p=0.134). No differences were observed according to gBRCA status and type of response. CONCLUSIONS: Early-stage TNBC might be a target population for olaparib, irrespective of gBRCA mutations. Future trials should combine TILs, PD-L1 and gBRCA status to better identify candidates for escalated/de-escalated treatment strategies including olaparib.

Whole-genome sequencing analysis of semi-supercentenarians.

Extreme longevity is the paradigm of healthy aging as individuals who reached the extreme decades of human life avoided or largely postponed all major age-related diseases. In this study, we sequenced at high coverage (90X) the whole genome of 81 semi-supercentenarians and supercentenarians [105+/110+] (mean age: 106.6 ± 1.6) and of 36 healthy unrelated geographically matched controls (mean age 68.0 ± 5.9) recruited in Italy. The results showed that 105+/110+ are characterized by a peculiar genetic background associated with efficient DNA repair mechanisms, as evidenced by both germline data (common and rare variants) and somatic mutations patterns (lower mutation load if compared to younger healthy controls). Results were replicated in a second independent cohort of 333 Italian centenarians and 358 geographically matched controls. The genetics of 105+/110+ identified DNA repair and clonal haematopoiesis as crucial players for healthy aging and for the protection from cardiovascular events.

Author Correction: Erythropoietin (EPO) haplotype associated with all-cause mortality in a cohort of Italian patients with Type-2 Diabetes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Erythropoietin (EPO) haplotype associated with all-cause mortality in a cohort of Italian patients with Type-2 Diabetes.

Type-2 Diabetes (T2D), diabetic complications, and their clinical risk factors harbor a substantial genetic component but the genetic factors contributing to overall diabetes mortality remain unknown. Here, we examined the association between genetic variants at 21 T2D-susceptibility loci and all-cause mortality in an elderly cohort of 542 Italian diabetic patients who were followed for an average of 12.08 years. Univariate Cox regression analyses detected age, waist-to-hip ratio (WHR), glycosylated haemoglobin (HbA1c), diabetes duration, retinopathy, nephropathy, chronic kidney disease (CKD), and anaemia as predictors of all-cause mortality. When Cox proportional hazards multivariate models adjusted for these factors were run, three erythropoietin (EPO) genetic variants in linkage disequilibrium (LD) with each other (rs1617640-T/G, rs507392-T/C and rs551238-A/C) were significantly (False Discovery Rate < 0.1) associated with mortality. Haplotype multivariate analysis revealed that patients carrying the G-C-C haplotype have an increased probability of survival, while an opposite effect was observed among subjects carrying the T-T-A haplotype. Our findings provide evidence that the EPO gene is an independent predictor of mortality in patients with T2D. Thus, understanding the mechanisms by which the genetic variability of EPO affects the mortality of T2D patients may provide potential targets for therapeutic interventions to improve the survival of these patients.