Polymorphisms in the Genes Coding for TLRs, NLRs and RLRs Are Associated with Clinical Parameters of Patients with Acute Myeloid Leukemia.

Toll-like receptors (TLRs), NOD-like receptors (NLRs), and RIG-I-like receptors (RLRs) are major elements of the innate immune system that recognize pathogen-associated molecular patterns. Single-nucleotide polymorphisms (SNPs) in the TLR, NLR, and RLR genes may lead to an imbalance in the production of pro- and anti-inflammatory cytokines, changes in susceptibility to infections, the development of diseases, and carcinogenesis. Acute myeloid leukemia (AML) is a bone marrow malignancy characterized by uncontrolled proliferation of transformed myeloid precursors. We retrospectively analyzed 90 AML patients. We investigated the effect of fifteen SNPs located in the genes coding for RLR1 (rs9695310, rs10738889, rs10813831), NOD1 (rs2075820, rs6958571), NOD2 (rs2066845, rs2066847, rs2066844), TLR3 (rs5743305, rs3775296, 3775291), TLR4 (rs4986791, rs4986790), and TLR9 (rs187084, rs5743836). We observed that TLR4 rs4986791, TLR9 rs5743836, and NOD2 rs2066847 were associated with CRP levels, while RLR-1 rs10738889 was associated with LDH level. Furthermore, we found TLR3 rs5743305 AA to be more common in patients with infections. We also found TLR9 rs187084 C to be associated with more favorable risk, and RLR-1 rs9695310 GG with higher age at diagnosis. In conclusion, the current study showed that SNPs in the genes encoding TLRs, NLRs, and RLRs may be potential biomarkers in patients with AML.

Relationship between Telomere Length, TERT Genetic Variability and TERT, TP53, SP1, MYC Gene Co-Expression in the Clinicopathological Profile of Breast Cancer.

The molecular mechanisms of telomerase reverse transcriptase (TERT) upregulation in breast cancer (BC) are complex. We compared genetic variability within TERT and telomere length with the clinical data of patients with BC. Additionally, we assessed the expression of the TERT, MYC, TP53 and SP1 genes in BC patients and in BC organoids (3D cell cultures obtained from breast cancer tissues). We observed the same correlation in the blood of BC patients and in BC organoids between the expression of TERT and TP53. Only in BC patients was a correlation found between the expression of the TERT and MYC genes and between TP53 and MYC. We found associations between TERT genotypes (rs2735940 and rs10069690) and TP53 expression and telomere length. BC patients with the TT genotype rs2735940 have a shorter telomere length, but patients with A allele rs10069690 have a longer telomere length. BC patients with a short allele VNTR-MNS16A showed higher expression of the SP1 and had a longer telomere. Our results bring new insight into the regulation of TERT, MYC, TP53 and SP1 gene expression related to TERT genetic variability and telomere length. Our study also showed for the first time a similar relationship in the expression of the above genes in BC patients and in BC organoids. These findings suggest that TERT genetic variability, expression and telomere length might be useful biomarkers for BC, but their prognostic value may vary depending on the clinical parameters of BC patients and tumor aggressiveness.

Association of RANK and RANKL gene polymorphism with survival and calcium levels in multiple myeloma.

Multiple myeloma (MM) is a heterogeneous bone marrow cancer characterized by proliferation of malignant plasma cells in the bone marrow. One of its major symptoms are hypercalcaemia and bone lesions, which may result in pathologic bone fractures. Receptor activator for nuclear factor κB (RANK) and its ligand, RANKL, are part of an activation pathway for osteoclasts and are thus responsible for bone resorption. Furthermore, RANKL expression is increased in multiple myeloma. In the present study, we investigated the role of single nucleotide polymorphisms (SNPs) in the genes coding for RANK (rs1805034, rs8086340), RANKL (rs7325635, rs7988338), and TACI (rs34562254), a receptor for osteoclast-derived pro-survival factors. The study involved 222 patients and 222 healthy individuals, and the analysis included disease susceptibility, survival, bone lesions, calcium levels, and vascular endothelial growth factor levels. Patients with allele RANK rs1805034 C had higher survival (p = .003). This relationship was especially evident in women (p = .006). Furthermore, allele rs1805034 C was associated with slightly lower median age at diagnosis (64.0 vs. 65.5, p = .008). Allele RANKL rs7325635 A correlated with lower progression-free survival (p = .027), and with lack of early progression (p = .023). Additionally, women with allele rs7325635 G were found to have higher calcium blood concentration (p = .040). Allele TACI rs34562254 A was more common in MM patients in more advanced stages (II and III stage International Staging System) at diagnosis (p = .017), and the SNP showed a slight trend towards association in a multivariate analysis (p = .084). Taken together, our results suggest that RANK rs1805034 and RANKL rs7325635 may have a role in MM development and progression.

Heterogeneity of telomerase reverse transcriptase mutation and expression, telomerase activity and telomere length across human cancer cell lines cultured in vitro.

Promoter region of the telomerase reverse transcriptase gene (TERTp) constitutes a regulatory element capable to affect TERT expression (TE), telomerase activity (TA) and telomere length (TL). TERTp mutation status, TL, TA and TE were assessed in 27 in vitro cultured human cell lines, including 11 solid tumour, 13 haematological and 3 normal cell lines. C228T and C250T TERTp mutations were detected in 5 solid tumour and none of haematological cell lines (p = 0.0100). As compared to other solid tumour cell lines, those with the presence of somatic mutations were characterized by: shorter TL, lower TA and TE. Furthermore, cell lines carrying TERTp mutations showed a linear correlation between TE and TA (R = 0.9708, p = 0.0021). Moreover, haematological cell lines exhibited higher TE compared to solid tumour cell lines (p = 0.0007). TL and TA were correlated in both solid tumour (R = 0.4875, p = 0.0169) and haematological (R = 0.4719, p = 0.0095) cell lines. Our results based on the in vitro model suggest that oncogenic processes may differ between solid tumours and haematological malignancies with regard to their TERT gene regulation mechanisms.

Association of MICA-129Met/Val polymorphism with clinical outcome of anti-TNF therapy and MICA serum levels in patients with rheumatoid arthritis.

MHC class I polypeptide-related sequence A (MICA) is a stress-induced protein involved in activation of NK and T cells through interaction with NKG2D receptor. These molecules are atypically expressed in synovium of patients diagnosed with rheumatoid arthritis (RA). A total of 279 patients with RA, qualified to TNF-blockade therapy, were genotyped for MICA rs1051792 SNP. The effectiveness of anti-TNF agents was assessed with European League Against Rheumatism criteria. Significant relationship between MICA rs1051792 and outcome of TNF-blockade therapy has been found. The MICA rs1051792 GG genotype was overrepresented in patients non-responsive to anti-TNF drugs in comparison with other genotypes (p = 0.010). On the other hand, beneficial therapeutic response was more frequently detected among RA subjects possessing heterozygous genotype than those with homozygous genotypes (p = 0.003). Furthermore, increased MICA concentrations in serum were observed in patients possessing MICA rs1051792 GG genotype as compared with those with GA or AA genotypes (p = 1.8 × 10-5). The results from this study indicate the potential influence of MICA rs1051792 polymorphism on modulation of therapeutic response to TNF-blockade treatment in RA.

Analysis of IL-17A, IL-17F, and miR-146a-5p Prior to Transplantation and Their Role in Kidney Transplant Recipients.

Background/Objectives: The balance between regulatory and Th17 cells plays an important role in maintaining the immune tolerance after kidney transplantation (KTx) which is essential for transplantation success, defined as a long graft survival and an absence of organ rejection. The present study aimed to assess whether the pretransplant characteristics of IL-17A and IL-17F, their receptors, as well as miR-146a-5p, an miRNA associated with IL-17A/F regulation, can predict KTx outcomes. Methods: A group of 108 pre-KTx dialysis patients and 125 healthy controls were investigated for single nucleotide substitutions within genes coding for IL-17A, IL-17F, their IL-17RA/RC receptors, and miR-146a-5p. Genotyping was performed using LightSNiP assays. In addition, IL17-A/F serum concentrations were determined using ELISA while miR-146a-5p expression was analyzed by RT-PCR. Results: The IL-17F (rs763780) G allele prevailed in KTx recipients as compared to healthy individuals (OR = 23.59, p < 0.0001) and was associated with a higher IL-17F serum level (p = 0.0381) prior to transplantation. Higher miR-146a-5p expression before KTx was more frequently detected in recipients with an increased IL-17A serum concentration (p = 0.0177). Moreover, IL-17A (rs2275913) GG homozygosity was found to be associated with an increased incidence of deaths before KTx (OR = 4.17, p = 0.0307). T-cell or acute rejection episodes were more frequently observed among patients with the C allele of miR-146a-5p (rs2910164) (OR = 5.38, p = 0.0531). IL17-RA/-RC genetic variants (p < 0.05) seem to be associated with eGFR values. Conclusions: These results imply that IL-17F (rs763780) polymorphism is associated with the serum level of this cytokine and may be related to the risk of renal disease and transplant rejection together with miR-146a-5p (rs2910164), while the IL-17A (rs2275913) genotype may affect patients' survival before KTx.

MICB Genetic Variants and Its Protein Soluble Level Are Associated with the Risk of Chronic GvHD and CMV Infection after Allogeneic HSCT.

The aim of the present study was to determine the associations between the MICB genetic variability and the expression and the risk of development of post-transplant complications after allogeneic hematopoietic stem cell transplantation (HSCT). HSCT recipients and their donors were genotyped for two MICB polymorphisms (rs1065075, rs3828903). Moreover, the expression of a soluble form of MICB was determined in the recipients' serum samples after transplantation using the Luminex assay. Our results revealed a favorable role of the MICB rs1065075 G allele. Recipients with donors carrying this genetic variant were less prone to developing chronic graft-versus-host disease (cGvHD) when compared to recipients without any symptoms of this disease (41.41% vs. 65.38%, p = 0.046). Moreover, the MICB rs1065075 G allele was associated with a lower incidence of cytomegalovirus (CMV) reactivation, both as a donor (p = 0.015) and as a recipient allele (p = 0.039). The MICB rs1065075 G variant was also found to be associated with decreased serum soluble MICB (sMICB) levels, whereas serum sMICB levels were significantly higher in recipients diagnosed with CMV infection (p = 0.0386) and cGvHD (p = 0.0008) compared to recipients without those complications. A protective role of the G allele was also observed for the rs3828903 polymorphism, as it was more frequently detected among donors of recipients without cGvHD (89.90% vs. 69.23%; p = 0.013). MICB genetic variants, as well as serum levels of sMICB, may serve as prognostic factors for the risk of developing cGvHD and CMV infection after allogeneic HSCT.

Telomere length and hTERT genetic variants as potential prognostic markers in multiple myeloma.

Telomere dysfunction is a notable event observed in many cancers contributing to their genomic instability. A major factor controlling telomere stability is the human telomerase reverse transcriptase catalytic subunit (hTERT). Telomere shortening has been observed in multiple myeloma (MM), a plasma cell malignancy with a complex and heterogeneous genetic background. In the present study, we aimed to analyse telomere length and hTERT genetic variants as potential markers of risk and survival in 251 MM patients. We found that telomere length was significantly shorter in MM patients than in healthy individuals, and patients with more advanced disease (stage III according to the International Staging System) had shorter telomeres than patients with less advanced disease. MM patients with hTERT allele rs2736100 T were characterized with significantly shorter progression-free survival (PFS). Moreover, allele rs2736100 T was also found to be less common in patients with disease progression in response to treatment. hTERT rs2853690 T was associated with higher haemoglobin blood levels and lower C-reactive protein. In conclusion, our results suggest that telomere length and hTERT genetic variability may affect MM development and can be potential prognostic markers in this disease.

Pro- and anti-inflammatory cytokines and growth factors in patients undergoing in vitro fertilization procedure treated with prednisone.

Embryo implantation is a key moment in pregnancy. Abnormal production of pro- and anti-inflammatory cytokines, their receptors and other immune factors may result in embryo implantation failure and pregnancy loss. The aim of this study was to determine the profile of selected pro- and anti-inflammatory factors in the blood plasma of patients undergoing in vitro fertilization (IVF) and control women who achieved pregnancy after natural conception. The examined patients were administered steroid prednisone. We present results concern the plasma levels of IFN-ɣ, BDNF, LIF, VEGF-A, sTNFR1 and IL-10. We found that IVF patients receiving steroids differed significantly from patients who were not administered such treatment in terms of IFN-γ and IL-10 levels. Moreover, IVF patients differed in secretion of all tested factors with the fertile controls. Our results indicated that women who secrete at least 1409 pg/ml of sTNFR1 have a chance to become pregnant naturally and give birth to a child, while patients after IVF must achieve a concentration of 962.3 pg/ml sTNFR1 in blood plasma for successful pregnancy. In addition, IVF patients secreting VEGF-A above 43.28 pg/ml have a greater risk of miscarriage or a failed transfer in comparison to women secreting below this value. In conclusion, fertile women present a different profile of pro- and anti-inflammatory cytokines, and growth factors compared to patients with recurrent implantation failure (RIF).

Assessment of Telomerase Reverse Transcriptase Single Nucleotide Polymorphism in Sleep Bruxism.

INTRODUCTION: Sleep bruxism (SB) is a widespread masticatory muscle activity during sleep and affects approximately 13.2% of the general population. Telomerase reverse transcriptase (TERT) plays a role in preventing the shortening of the telomere. This prospective, observational study aimed to investigate the relationship between single nucleotide polymorphism (SNP) of TERT and the severity of SB and to identify the independent risk factors for SB. METHODS: A total of 112 patients were diagnosed by performing one-night polysomnography based on the guidelines of the American Academy of Sleep Medicine. TERT SNP was detected by real-time quantitative polymerase chain reaction (qPCR). RESULTS: Statistical analysis showed the lack of relationship between the rs2853669 polymorphism of TERT and severity of SB (p > 0.05). However, the study showed that patients with allele T in the 2736100 polymorphism of TERT had a lower score on the phasic bruxism episode index (BEI). Based on the receiver operating characteristic (ROC) curve, the value of phasic BEI was 0.8 for the differential prediction for the presence of allele T in the locus. The sensitivity and specificity were 0.328 and 0.893, respectively. The regression analysis showed that lack of TERT rs2736100 T allele, male gender, and arterial hypertension are the risk factors for the higher value of phasic BEI. CONCLUSION: The SNP of the TERT gene affects phasic SB intensity. The absence of TERT rs2736100 T allele, male sex, and arterial hypertension are independent risk factors for phasic SB.

Genetic Variants of the TERT Gene and Telomere Length in Obstructive Sleep Apnea.

Introduction: Obstructive sleep apnea (OSA) is a worldwide breathing disorder that has been diagnosed globally in almost 1 billion individuals aged 30−69 years. It is characterized by repeated upper airway collapses during sleep. Telomerase reverse transcriptase (TERT) is involved in the prevention of telomere shortening. This prospective, observational study aimed to investigate the relationship between single nucleotide polymorphisms (SNPs) of TERT and the severity of OSA, taking into account hypertension and diabetes prevalence. Methods: A total of 149 patients with OSA were diagnosed using one-night video-polysomnography based on the American Academy of Sleep Medicine guidelines. The TERT SNPs and telomere length (TL) were detected using real-time quantitative polymerase chain reaction. Results: Statistical analysis showed that there is no relationship between the rs2853669 and rs2736100 polymorphisms of TERT, and the severity of OSA (p > 0.05). Moreover, no relationship between TL and the severity of OSA was observed. The G allele in the locus of rs2736100 TERT was associated with hypertension prevalence and was more prevalent in hypertensives patients (46.00% vs. 24.49%, p = 0.011). The prevalence of hypertension was higher in patients with the C allele in the locus of rs2853669 than in patients without this allele (50.79% vs. 30.23%, p = 0.010). Moreover, a lower prevalence of diabetes was observed in homozygotes of rs2736100 TERT than in heterozygotes (5.63% vs. 15.38%, p = 0.039). Conclusion: This study showed no relationship between OSA and TERT SNPs. However, SNPs of the TERT gene (rs2736100 and rs2853669) were found to affect arterial hypertension and diabetes prevalence.

TERT genetic variability and telomere length as factors affecting survival and risk in acute myeloid leukaemia.

Acute myeloid leukaemia (AML) is a neoplasm of immature myeloid cells characterized by various cytogenetic alterations. The present study showed that in addition to the FLT3-ITD and NPM1 mutation status, telomere length (TL) and telomerase reverse transcriptase (TERT) gene polymorphisms may affect risk and overall survival (OS) in AML. TL was longer in healthy controls than in AML patients and positively correlated with age in the patients, but not in healthy subjects. TL was found to be independently affected by the presence of the FLT3-ITD mutation. As for the TERT gene polymorphism, AML patients with the TERT rs2853669 CC genotype were characterized by significantly shorter OS than patients carrying the T allele. Another observation in our study is the difference in TL and OS in patients belonging to various risk stratification groups related to the FLT3-ITD and NPM1 mutation status. Patients with adverse risk classification (mutation in FLT3-ITD and lack of mutation in NPM1) presented with the shortest telomeres and significantly worse OS. In conclusion, OS of AML patients appears to be affected by TERT gene variability and TL in addition to other well-established factors such as age, WBC count, or FLT3-ITD and NPM1 mutation status.

TERT-Regulation and Roles in Cancer Formation.

Telomerase reverse transcriptase (TERT) is a catalytic subunit of telomerase. Telomerase complex plays a key role in cancer formation by telomere dependent or independent mechanisms. Telomere maintenance mechanisms include complex TERT changes such as gene amplifications, TERT structural variants, TERT promoter germline and somatic mutations, TERT epigenetic changes, and alternative lengthening of telomere. All of them are cancer specific at tissue histotype and at single cell level. TERT expression is regulated in tumors via multiple genetic and epigenetic alterations which affect telomerase activity. Telomerase activity via TERT expression has an impact on telomere length and can be a useful marker in diagnosis and prognosis of various cancers and a new therapy approach. In this review we want to highlight the main roles of TERT in different mechanisms of cancer development and regulation.

Polymorphisms within the RANK and RANKL Encoding Genes in Patients with Rheumatoid Arthritis: Association with Disease Progression and Effectiveness of the Biological Treatment.

Inconsistency of the results regarding the genetic variability within genes coding for receptor activator of nuclear factor κB (RANK) and its ligand (RANKL) in rheumatoid arthritis (RA) prompted us to study the RANK and RANKL polymorphisms as potential biomarkers associated with disease predisposition and response to anti-TNF treatment in a group of Polish patients with RA. This study enrolled 318 RA patients and 163 controls. RANK (rs8086340, C > G; rs1805034, C > T) and RANKL (rs7325635, G > A; rs7988338 G > A) alleles were determined by real-time PCR with melting curve analysis and related with clinical parameters. In addition, RANKL serum levels were measured by ELISA. The RANK rs8086340-G allele was overrepresented among patients as compared to controls (OD = 1.777, p = 0.038). C-reactive protein (CRP) levels were significantly (p < 0.05) associated with RANK rs8086340 polymorphism and were higher in the CC-homozygotes at the baseline while lower in the GG-carriers at the 12th week of the treatment. At the latter time point RANKL rs7325635-GG-positive patients also showed significantly lower CRP concentrations. Higher alkaline phosphatase levels before induction of anti-TNF therapy were observed in RANK rs8086340 and RANK rs1805034 CC homozygotes (p = 0.057 and p = 0.035, respectively). The GG homozygosity of both RANKL single nucleotide polymorphisms was significantly associated with the number of swollen joints (rs7988338 and rs7325635, before and at the 12th week of therapy, respectively, p < 0.05 in both cases). These results imply that polymorphisms within the RANK and RANKL genes affect RA susceptibility and anti-TNF treatment outcome.

Genetic basis of sleep bruxism and sleep apnea-response to a medical puzzle.

Sleep bruxism (SB) and obstructive sleep apnea (OSA) are co-occurring sleep conditions. The study aimed to evaluate the association of selected single-nucleotide polymorphisms (SNPs) occurring within the genes of the serotonin and dopamine pathways in SB and OSA and investigate the relationship between them. The study group included 100 Caucasian patients. SB and OSA were diagnosed in 74 and 28 patients, respectively. In addition, 125 unrelated Caucasian healthy blood donors served as randomly selected controls to enable comparison of polymorphisms. The following SNPs were analyzed: rs2770304 and rs6313 within the serotonin receptor encoding gene (HTR2A), rs4680 polymorphism of the catechol-O-methyltransferase (COMT) gene, and rs686 within the dopamine receptor (DRD1) encoding gene. The prevalence of the DRD1 rs686 G variant (GG homozygosity) was found to be high in the study group compared to the control group. Bruxism episode index (BEI) was found to be significantly increased in the HTR2A rs6313 TT homozygotes compared to the heterozygous patients. Moreover, within a group of the HTR2A rs2770304 TT homozygous cases, a statistically significant correlation was observed between BEI and apnea-hypopnea index. These results indicate that DRD1 rs686 may potentially affect predisposition to SB, that HTR2A rs6313 SNP may be involved in SB pathogenesis, and that HTR2A rs2770304 polymorphism might contribute to the association between SB and OSA. This suggests a possible genetic contribution to the etiology of primary SB.

Variability within the human TERT gene, telomere length and predisposition to chronic lymphocytic leukemia.

Background: The human telomerase reverse transcriptase (TERT) gene encodes the catalytic subunit of telomerase that is essential for maintenance of telomere length. We aimed to find out whether variability within the TERT gene could be associated with telomere length and development of the disease in non-treated patients with chronic lymphocytic leukemia (CLL). Materials and methods: Telomere length, rs2736100, rs2853690, rs33954691, rs35033501 single-nucleotide polymorphisms, and variable number of tandem repeats (VNTR-MNS16A) were assessed in patients at diagnosis. In addition, blood donors served as controls for the polymorphism studies. Results: The minor rs35033501 A variant was more frequent among CLL patients than in healthy controls (OR=3.488, p=0.039). CLL patients over 60 years of age were characterized with lower disease stage at diagnosis (p=0.001 and p=0.008, for the Rai and Binet criteria, respectively). The MNS16A VNTR-243 short allele was more frequent in patients with a low disease stage (p=0.020 and p=0.028, for the Rai and Binet staging system) and also among older patients having longer telomeres (p=0.046). Patients with Rai 0-I stage were characterized with longer telomeres than those with more advanced disease (p=0.030). This relationship was especially pronounced in patients carrying the rs2736100 C allele, independently of the criteria used, ie, Binet (p=0.048) or Rai (p=0.001). Conclusion: Our results showed that the genetic variation within the TERT gene seems to play a regulatory role in CLL and telomere length.