A machine-learning method for biobank-scale genetic prediction of blood group antigens.

A key element for successful blood transfusion is compatibility of the patient and donor red blood cell (RBC) antigens. Precise antigen matching reduces the risk for immunization and other adverse transfusion outcomes. RBC antigens are encoded by specific genes, which allows developing computational methods for determining antigens from genomic data. We describe here a classification method for determining RBC antigens from genotyping array data. Random forest models for 39 RBC antigens in 14 blood group systems and for human platelet antigen (HPA)-1 were trained and tested using genotype and RBC antigen and HPA-1 typing data available for 1,192 blood donors in the Finnish Blood Service Biobank. The algorithm and models were further evaluated using a validation cohort of 111,667 Danish blood donors. In the Finnish test data set, the median (interquartile range [IQR]) balanced accuracy for 39 models was 99.9 (98.9-100)%. We were able to replicate 34 out of 39 Finnish models in the Danish cohort and the median (IQR) balanced accuracy for classifications was 97.1 (90.1-99.4)%. When applying models trained with the Danish cohort, the median (IQR) balanced accuracy for the 40 Danish models in the Danish test data set was 99.3 (95.1-99.8)%. The RBC antigen and HPA-1 prediction models demonstrated high overall accuracies suitable for probabilistic determination of blood groups and HPA-1 at biobank-scale. Furthermore, population-specific training cohort increased the accuracies of the models. This stand-alone and freely available method is applicable for research and screening for antigen-negative blood donors.

A novel nonsense variant in RHAG underlies a Nordic Rhnull phenotype.

BACKGROUND AND OBJECTIVES: The extremely rare Rhnull phenotype is characterized by the absence of all Rh antigens on erythrocytes. It is divided into the regulator and amorph types based on the underlying genetic background. The more common regulator type depends on critical variants silencing RHAG, which encodes RhAG glycoprotein, necessary for RhD/RhCE expression. Rhnull cells have altered expression of glycophorin B and LW glycoprotein. MATERIALS AND METHODS: Four unrelated Rhnull individuals were investigated. Serological testing was performed according to standard blood bank practice. RHD/RHCE and S/s allele-specific Polymerase chain reaction (PCR) genotyping was done on genomic DNA using in-house PCR assays. RHAG, and in some cases also RHD/RHCE, were sequenced. Initial s phenotyping results triggered additional serological investigation. RESULTS: Anti-Rh29 was identified in all four individuals. Extended typing with anti-S and anti-s showed that the three samples predicted to type as s+ failed to react with 2 of 5 anti-s. Sequence analysis of all 10 RHAG exons and the immediate intron/exon boundaries revealed a single nucleotide variant in the 3'-end of intron 6, c.946 -2a>g in all samples. RHD/RHCE showed no alterations. CONCLUSION: A novel Nordic Rhnull allele was identified. In addition, it was shown that s+ Rhnull red blood cells are not only U- but also have qualitative changes in their s antigen expression.

Rh Blood Group D Antigen Genotyping Using a Portable Nanopore-based Sequencing Device: Proof of Principle.

BACKGROUND: Nanopore sequencing is direct sequencing of a single-stranded DNA molecule using biological pores. A portable nanopore-based sequencing device from Oxford Nanopore Technologies (MinION) depends on driving a DNA molecule through nanopores embedded in a membrane using a voltage. Changes in current are then measured by a sensor, thousands of times per second and translated to nucleobases. METHODS: Genomic DNA (gDNA) samples (n = 13) were tested for Rh blood group D antigen (RHD) gene zygosity using droplet digital PCR. The RHD gene was amplified in 6 overlapping amplicons using long-range PCR. Amplicons were purified, and the sequencing library was prepared following the 1D Native barcoding gDNA protocol. Sequencing was carried out with 1D flow cells R9 version. Data analysis included basecalling, aligning to the RHD reference sequence, and calling variants. Variants detected were compared to the results acquired previously by the Ion Personal Genome Machine (Ion PGM). RESULTS: Up to 500× sequence coverage across the RHD gene allowed accurate variant calling. Exonic changes in the RHD gene allowed RHD allele determination for all samples sequenced except 1 RHD homozygous sample, where 2 heterozygous RHD variant alleles are suspected. There were 3 known variant RHD alleles (RHD*01W.02, RHD*11, and RHD*15) and 6 novel RHD variant alleles, as previously seen in Ion PGM sequencing data for these samples. CONCLUSIONS: MinION was effective in blood group genotyping, provided enough sequencing data to achieve high coverage of the RHD gene, and enabled confident calling of variants and RHD allele determination.

Recommendation for validation and quality assurance of non-invasive prenatal testing for foetal blood groups and implications for IVD risk classification according to EU regulations.

BACKGROUND AND OBJECTIVES: Non-invasive assays for predicting foetal blood group status in pregnancy serve as valuable clinical tools in the management of pregnancies at risk of detrimental consequences due to blood group antigen incompatibility. To secure clinical applicability, assays for non-invasive prenatal testing of foetal blood groups need to follow strict rules for validation and quality assurance. Here, we present a multi-national position paper with specific recommendations for validation and quality assurance for such assays and discuss their risk classification according to EU regulations. MATERIALS AND METHODS: We reviewed the literature covering validation for in-vitro diagnostic (IVD) assays in general and for non-invasive foetal RHD genotyping in particular. Recommendations were based on the result of discussions between co-authors. RESULTS: In relation to Annex VIII of the In-Vitro-Diagnostic Medical Device Regulation 2017/746 of the European Parliament and the Council, assays for non-invasive prenatal testing of foetal blood groups are risk class D devices. In our opinion, screening for targeted anti-D prophylaxis for non-immunized RhD negative women should be placed under risk class C. To ensure high quality of non-invasive foetal blood group assays within and beyond the European Union, we present specific recommendations for validation and quality assurance in terms of analytical detection limit, range and linearity, precision, robustness, pre-analytics and use of controls in routine testing. With respect to immunized women, different requirements for validation and IVD risk classification are discussed. CONCLUSION: These recommendations should be followed to ensure appropriate assay performance and applicability for clinical use of both commercial and in-house assays.

Time points and risk factors for RhD immunizations after the implementation of targeted routine antenatal anti-D prophylaxis: A retrospective nationwide cohort study.

INTRODUCTION: Targeted routine antenatal anti-D prophylaxis was introduced to the national prophylaxis program in Finland in late 2013. The aim of this study was to assess the incidence, time-points, and risk factors for Rhesus D immunization after the implementation of routine antenatal anti-D prophylaxis, in all women in Finland with antenatal anti-D antibodies detected in 2014-2017. MATERIAL AND METHODS: In a nationwide population-based retrospective cohort study, the incidence, time-points, and risk factors of anti-D immunizations were analyzed. Information on antenatal screening was obtained from the Finnish Red Cross Blood Service database, and obstetric data from hospital records and the Finnish Medical Birth Register. RESULTS: The study included a total of 228 women (197 with complete data for all pregnancies). After the implementation of routine antenatal anti-D prophylaxis, the prevalence of pregnancies with anti-D antibodies decreased from 1.52% in 2014 to 0.88% in 2017, and the corresponding incidence of new immunizations decreased from 0.33% to 0.10%. Time-points for detection of new anti-D antibodies before and after 2014 were the first screening sample at 8-12 weeks of gestation in 52% vs 19%, the second sample at 24-26 weeks in 20% vs 50%, and the third screening at 36 weeks in 28% vs 32%. CONCLUSIONS: The incidence of new anti-D immunizations decreased as expected after the implementation of routine antenatal anti-D prophylaxis. True failures are rare and they mainly occur when the prophylaxis is not given appropriately, suggesting a need for constant education of healthcare professionals on the subject.

Maternal HLA genotyping is not useful for predicting severity of fetal and neonatal alloimmune thrombocytopenia.

Lack of reliable laboratory parameters is the main challenge in the management of fetal and neonatal alloimmune thrombocytopenia (FNAIT). Despite the long-known association between the HLA-DRB3*01:01 allele and human platelet antigen 1a (HPA-1a) alloimmunisation, maternal human leucocyte antigen (HLA) typing has been of little clinical value. Recently, other DRB3 allele variants have been suggested to predict the severity of FNAIT. In this nationwide population-based retrospective cohort study, we performed extensive HLA typing of 96 women, accounting for 87% of our cohort of 110 families with confirmed or possible HPA-1a-immunisation. The HLA type was compared with anti-HPA-1a levels, severity of neonatal disease and responsiveness to maternally administrated intravenous gammaglobulin (IVIG). HLA haplotypes were constructed to investigate further HLA associations. Despite significantly lower anti-HPA-1a levels in DRB3*01:01-negative women, the carrier status of this particular allele could not be used to confirm or rule out FNAIT in the absence of detectable antibodies. In the haplotype analysis, the DRB3*01:01 allele was the actual factor associated with FNAIT. No other HLA allele was shown to be of additional value as a predictor of severe FNAIT or non-responsiveness to IVIG treatment. Thus, HLA genotyping was not found useful in differentiating high- and low-risk pregnancies or in guiding antenatal treatment in affected families.

Targeted antenatal anti-D prophylaxis program for RhD-negative pregnant women - outcome of the first two years of a national program in Finland.

INTRODUCTION: The aim of this study was to assess the accuracy of the non-invasive fetal RHD test at 24-26 weeks of gestation as part of the national antenatal screening program to target routine antenatal anti-D prophylaxis (RAADP) at 28-30 weeks at women carrying an RhD-positive fetus. MATERIAL AND METHODS: A prospective cohort study involving all maternity care centers and delivery hospitals in Finland between February 2014 and January 2016. Fetal RHD genotyping using cell-free fetal DNA in maternal plasma was performed with real-time polymerase chain reaction in a centralized setting. The results were systematically compared with the serological newborn RhD typing. The main outcome measure was the accuracy of the fetal RHD assay; the secondary variable was compliance with the newly introduced RAADP program. RESULTS: Fetal RHD was screened from 10 814 women. For the detection of fetal RHD, sensitivity was 99.99% [95% confidence interval (CI) 99.92-99.99] and specificity 99.81% (95% CI 99.60-99.92). One false-negative and seven false-positive results were reported by the delivery hospitals in two years. The negative predictive value of the test was 99.97% (95% CI 99.81-99.99). At the end of the study period, over 98% of the RhD-negative women participated in the new screening program. CONCLUSIONS: The targeted RAAPD program was implemented effectively in the national maternity care program in Finland. An accurate fetal RHD screening test allows discontinuation of newborn testing without risking the postnatal prophylaxis program. In the future, the main area to investigate will be the clinical effect of RAADP on subsequent pregnancies.

High-density SNP mapping of the HLA region identifies multiple independent susceptibility loci associated with selective IgA deficiency.

Selective IgA deficiency (IgAD; serum IgA<0.07 g/l) is the most common form of human primary immune deficiency, affecting approximately 1∶600 individuals in populations of Northern European ancestry. The polygenic nature of IgAD is underscored by the recent identification of several new risk genes in a genome-wide association study. Among the characterized susceptibility loci, the association with specific HLA haplotypes represents the major genetic risk factor for IgAD. Despite the robust association, the nature and location of the causal variants in the HLA region remains unknown. To better characterize the association signal in this region, we performed a high-density SNP mapping of the HLA locus and imputed the genotypes of common HLA-B, -DRB1, and -DQB1 alleles in a combined sample of 772 IgAD patients and 1,976 matched controls from 3 independent European populations. We confirmed the complex nature of the association with the HLA locus, which is the result of multiple effects spanning the entire HLA region. The primary association signal mapped to the HLA-DQB1*02 allele in the HLA Class II region (combined P = 7.69×10(-57); OR = 2.80) resulting from the combined independent effects of the HLA-B*0801-DRB1*0301-DQB1*02 and -DRB1*0701-DQB1*02 haplotypes, while additional secondary signals were associated with the DRB1*0102 (combined P = 5.86×10(-17); OR = 4.28) and the DRB1*1501 (combined P = 2.24×10(-35); OR = 0.13) alleles. Despite the strong population-specific frequencies of HLA alleles, we found a remarkable conservation of these effects regardless of the ethnic background, which supports the use of large multi-ethnic populations to characterize shared genetic association signals in the HLA region. We also provide evidence for the location of association signals within the specific extended haplotypes, which will guide future sequencing studies aimed at characterizing the precise functional variants contributing to disease pathogenesis.

Case report: Severe hemolytic disease of the fetus and newborn due to anti-C+G.

Anti-G is commonly present with anti-D and/or anti-C and can confuse serological investigations. in general, anti-G is not considered a likely cause of severe hemolytic disease of the fetus and newborn (HDFN), but it is important to differentiate it from anti-D in women who should be administered anti-D immunoglobulin prophylaxis. We report one woman with three pregnancies severely affected by anti-C+G requiring intrauterine treatment and a review of the literature. In our case, the identification of the correct antibody was delayed because the differentiation of anti-C+G and anti-D+C was not considered important during pregnancy since the father was D-. In addition, anti-C+G and anti-G titer levels were not found to be reliable as is generally considered in Rh immunization. Severe HDFN occurred at a maternal anti-C+G antibody titer of S and anti-G titer of 1 in comparison with the critical titer level of 16 or more in our laboratory. close collaboration between the immunohematology laboratory and the obstetric unit is essential. In previously affected families, early assessment for fetal anemia is required even when titers are low.

[Where to find a suitable red cell product, if the patient has a rare blood group?]. / Mistä sopiva punasoluvalmiste, jos potilaalla on harvinainen veriryhmä?

A rare blood group and the associated anti-red cell antibody against a very common blood group factor make it significantly more difficult to find suitable red cell products, possibly endangering the additional treatment of the patient. The need of rare blood should be anticipated early enough. The availability of red cells is secured by extensive blood group studies among blood donors, by donor registers, freezing of rare red cells and through international collaboration. The determination of rare blood groups by genotyping provides a longed-for addition to the investigational repertoire of both patients and blood donors. Systematic freezing of red cells was started in Finland in 2010.

Hippocampal sclerosis is associated with celiac disease type immunity in patients with drug-resistant temporal lobe epilepsy.

BACKGROUND: A prior small-scale single center study suggested an association between celiac disease (CD)-type immunity and refractory temporal lobe epilepsy (TLE) with hippocampal sclerosis (HS). The present study addresses this putative association in a large, well-characterized group of drug-resistant epilepsy (DRE) patients. These patients were grouped based on the spectrum of CD and gluten sensitivity-associated antibodies. METHODS: In this cross-sectional study, 253 consecutive adult epilepsy patients (135 females, 118 males; age 16-76 years) were categorized into three groups: (i) CD-positive group with either prior diagnosis of CD or CD-specific TG2/EmA antibodies, (ii) AGA-positive group with antigliadin antibodies (AGA) but without CD, and (iii) CD/AGA-negative group without any gluten sensitivity-associated antibodies or CD. Clinical and immunological findings were then compared among the groups. RESULTS: TLE with HS was more common in the CD-positive group compared to CD/AGA-negative group (31.8% versus 11.9%, P = 0.019). Autoimmune disorders were more common in the AGA-positive group than in the CD/AGA-negative group (P = 0.025). Considering HS lateralization; left lateralization was more common in CD-positive group compared to CD/AGA-negative group (71.4% versus 25%, P = 0.030). TG6 seropositivity did not differ among the groups (P > 0.05). CONCLUSIONS: This study provides further evidence linking TLE with HS and CD-type autoimmunity suggesting that CD-type immune response to gluten can be one potential mechanism as a disease modifier leading to DRE and HS. Understanding these immunological factors is imperative for developing immunomodulatory or dietary treatments for DRE potentially preventing HS progression.

Degree of damage to the small bowel and serum antibody titers correlate with clinical presentation of patients with celiac disease.

BACKGROUND & AIMS: In patients with celiac disease, gluten-induced lesions of the small-bowel mucosa develop gradually. However, it is not clear whether clinical presentation correlates with the degree of mucosal damage based on histology analysis. We investigated whether the degree of mucosal damage to the small bowel correlates with clinical presentation and serum markers of celiac disease. METHODS: We collected results from serology tests and mucosal biopsy samples from 638 consecutive patients with celiac disease and compared them with reported gastrointestinal symptoms, health-related quality-of-life scores, results from laboratory tests, and bone mineral densities of patients. We assessed mucosal injury based on the ratio of villous height to crypt depth, numbers of intraepithelial CD3(+) cells, and semiquantitative Marsh classification criteria. Correlations were established based on the Pearson or Spearman coefficients. RESULTS: The ratio of the villous height to crypt depth correlated with the severity of gastrointestinal symptoms, quality-of-life scores, laboratory test results, numbers of intraepithelial CD3(+) cells, and serum levels of antibodies associated with celiac disease. There was no correlation between the ratio of villous height to crypt depth and bone mineral density. The number of intraepithelial CD3(+) cells was not associated with symptoms, whereas the Marsh classification and serum levels of antibodies associated with celiac disease correlated with gastrointestinal symptoms, laboratory test results, and numbers of intraepithelial CD3(+) cells. CONCLUSIONS: The ratio of small-bowel villous height to crypt depth and results from serology tests correlate with reported symptoms and quality of life of patients with celiac disease. Patient-reported outcomes are therefore of value, in addition to histology findings, in assessing patients with celiac disease.

Serodiagnostic assays for celiac disease based on the open or closed conformation of the autoantigen, transglutaminase 2.

The autoantigen of celiac disease, transglutaminase 2 (TG2), adopts an open conformation during enzymatic activation. We studied diagnostic accuracy of serodiagnostic assays using TG2 in its open and closed conformation as antigens in patients with diagnostic difficulties. The open TG2 antibody (TG2ab) test identified 93% of untreated celiac patients in contrast to 44%, 27%, and 68% detected by closed and conventional TG2ab and endomysial antibody (EmA) tests, respectively. The assay was able to detect 60% of non-responding celiac patients seronegative for conventional TG2ab and EmA. The titers of the open TG2abs were higher than those of the closed TG2abs. The serological test utilizing TG2 in an open conformation was more accurate than the other assays in finding active celiac disease even in patients having negative or borderline conventional celiac autoantibodies and in revealing poor dietary response non-invasively. It thus offers a promising tool in the diagnostics and follow-up of celiac disease.

Gluten-sensitive hypertransaminasemia in celiac disease: an infrequent and often subclinical finding.

OBJECTIVES: Earlier studies suggest that 40-50% of untreated celiac disease patients have elevated transaminase levels. Celiac disease can also be the underlying reason for unexplained hypertransaminasemia or even liver failure. We investigated the prevalence and gluten dependency of hypertransaminasemia in celiac patients also diagnosed with minor or atypical symptoms. METHODS: In the cross-sectional study, serum aspartate (AST) and alanine (ALT) transaminase levels were measured in 313 untreated and 339 treated adult celiac patients and 237 non-celiac controls. In the prospective part, transaminase levels were investigated in 130 celiac patients at diagnosis and after 1 year on a gluten-free diet and in 25 treated celiac patients in clinical remission before and after gluten challenge. RESULTS: The proportion of subjects with elevated serum AST values in the untreated celiac disease group (11%) did not differ significantly from that in the treated celiac disease (8%) or non-celiac control groups (9%) (P=0.587). Although the serum transaminase values were within normal range in the majority of untreated patients, initially normal liver enzyme levels decreased significantly on a gluten-free diet. In treated celiac disease, gluten challenge led to mild and transient hypertransaminasemia. CONCLUSIONS: In our area, where the prevalence of celiac disease is high, hypertransaminasemia is less frequent in untreated celiac disease patients than previously reported. The regular screening of transaminases in celiac disease needs to be re-evaluated. Although the liver enzyme levels were within the reference values in the majority of celiac patients, an obvious gluten dependence of transaminase levels was observed even in these subjects.

Antibodies against deamidated gliadin peptides in early-stage celiac disease.

BACKGROUND AND GOALS: The widely used serum endomysial (EmA) and transglutaminase 2 (TG2-ab) antibodies predict forthcoming villous damage and celiac disease when the small-bowel mucosa structure is still normal. However, these autoantibodies may remain negative in this early stage of the disease. We hypothesized that the antibodies against deamidated gliadin peptides (DGP-AGA) might appear before the other antibodies and would thus be useful in the diagnosis and follow-up of patients with early-stage celiac disease. STUDY: Serum DGP-AGA, TG2-ab, and EmA were measured at baseline and after 1 year on a gluten-free diet in 42 adults proven to have early-stage celiac disease despite normal small-bowel mucosal morphology (Marsh I-II), and in 20 celiac subjects evincing villous atrophy (Marsh III). Thirty-nine subjects with no signs of celiac disease served as nonceliac controls. RESULTS: Sensitivity to detect early-stage celiac disease was 79% for DGP-AGA, 64% for TG2-ab, and 81% for EmA. Specificities were 95%, 100%, and 100%, respectively. The corresponding efficiencies of the tests were 89% for DGP-AGA, 81% for TG2-ab, and 91% for EmA. All 3 antibodies were significantly decreased on a gluten-free diet. CONCLUSIONS: This study showed that the sensitivity of DGP-AGA was superior to TG2-ab and comparable to EmA in celiac patients having early-stage celiac disease with normal villous morphology. On the basis of these results, DGP-AGA would seem to offer a promising new method for case-finding and follow-up in this entity.

Diagnosing mild enteropathy celiac disease: a randomized, controlled clinical study.

BACKGROUND & AIMS: The diagnostic criteria for celiac disease require small-bowel mucosal villous atrophy with crypt hyperplasia (Marsh III). However, mucosal damage develops gradually and patients may evince clinical symptoms before histologic changes appear. Endomysial antibodies are specific in predicting forthcoming villous atrophy. We hypothesized that patients with mild enteropathy but positive endomysial antibodies benefit from a gluten-free diet (GFD) similarly to patients with more severe enteropathy. METHODS: Small-bowel endoscopy together with clinical evaluations was performed in all together 70 consecutive adults with positive endomysial antibodies. Of these, 23 had only mild enteropathy (Marsh I-II) and they were randomized either to continue on a gluten-containing diet or start a GFD. After 1 year, clinical, serologic, and histologic evaluations were repeated. A total of 47 participants had small-bowel mucosal lesions compatible with celiac disease (Marsh III), and these served as disease controls. RESULTS: In the gluten-containing diet group (Marsh I-II) the small-bowel mucosal villous architecture deteriorated in all participants, and the symptoms and abnormal antibody titers persisted. In contrast, in the GFD group (Marsh I-II) the symptoms were alleviated, antibody titers decreased, and mucosal inflammation diminished equally to celiac controls (Marsh III). When the trial was completed, all participants chose to continue on a life-long GFD. CONCLUSIONS: Patients with endomysial antibodies benefit from a GFD regardless of the degree of enteropathy. The diagnostic criteria for celiac disease need re-evaluation: endomysial antibody positivity without atrophy belongs to the spectrum of genetic gluten intolerance, and warrants dietary treatment.

Next-generation sequencing of 35 RHD variants in 16 253 serologically D- pregnant women in the Finnish population.

Fetal RHD screening for targeted routine antenatal anti-D prophylaxis has been implemented in many countries, including Finland, since the 2010s. Comprehensive knowledge of the RHD polymorphism in the population is essential for the performance and safety of the anti-D prophylaxis program. During the first 3 years of the national screening program in Finland, over 16 000 samples from RhD- women were screened for fetal RHD; among them, 79 samples (0.5%) containing a maternal variant allele were detected. Of the detected maternal variants, 35 cases remained inconclusive using the traditional genotyping methods and required further analysis by next-generation sequencing (NGS) of the whole RHD gene to uncover the variant allele. In addition to the 13 RHD variants that have been previously reported in different populations, 8 novel variants were also detected, indicating that there is more variation of RHD in the RhD- Finnish population than has been previously known. Three of the novel alleles were identified in multiple samples; thus, they are likely specific to the original Finnish population. National screening has thus provided new information about the diversity of RHD variants in the Finnish population. The results show that NGS is a powerful method for genotyping the highly polymorphic RHD gene compared with traditional methods that rely on the detection of specific nucleotides by polymerase chain reaction amplification.

Association of HLA-DRB1, interleukin-6 and cyclin D1 polymorphisms with cervical cancer in the Swedish population--a candidate gene approach.

High-risk human papillomavirus (hrHPV) infection is the major risk factor for cervical cancer (CxCa). The role of genetic susceptibility in the disease has been suggested, but the existing data lack consistency. We conducted a nested case-control study on 973 CxCa cases and 1,763 matched controls, from two Swedish population-based cohorts to examine the association of common genetic variants with CxCa risk. Human leukocyte antigen (HLA) alleles and 24 other polymorphisms in 14 genes were selected on the basis of reported association or mechanistic plausibility with an HPV infection or cervical cancer development. Genotyping was conducted using multiplex PCR and Luminex technology. A significant association of CxCa with various polymorphisms was observed: rs1800797 in the IL-6 gene (odds ratio [OR] = 0.88, 95% confidence intervals [CI]: 0.79-0.99); rs1041981 in the LTA gene (OR = 0.87, 95% CI: 0.78-0.98), and rs9344 in the CCND1 gene (OR = 1.14, 95% CI: 1.02-1.27), for those individuals carrying the rare allele. Additionally, the alleles 0401 and 1501 of the HLA class II DRB1 locus were associated with an increased risk (OR = 1.23, 95% CI: 1.04-1.45 and OR = 1.29, 95% CI: 1.11-1.50, respectively), and allele 1301 was associated with decreased risk (OR = 0.59, 95% CI: 0.47-0.73). The effects of CCND1 and the HLA*DRB1 alleles were independent of the effect of smoking. We did not find any association of risk with polymorphisms in genes related to the innate immune system. In conclusion, our study provides evidence for genetic susceptibility to CxCa due to variations in genes involved in the immune system and in cell cycle.