CAG repeat length in an infertile male population of Irish origin

The androgen receptor (AR) gene, located on the X chromosome, is an important regulator of human spermatogenesis. In the past decade, the link between the CAG polyglutamine tract, situated on exon one of the AR gene, and reduced spermatogenesis has become a controversial one. Alterations in the length of the CAG polyglutamine tract have been associated with prostate cancer at a reduced intrinsic length and neuromuscular diseases at a CAG repeat length of 40. Minimal intermediate increases have been linked with depressed spermatogenesis in infertile males. Asian and Australian groups have published an association between increased CAG repeat length and reduced spermatogenesis while many European studies have found no such association. The aim of this study was to document the association between increased CAG repeat length and reduced spermatogenesis in a group of Irish infertile males and controls known to have fathered at least one child. The study employed the ABI 377 DNA sequencer to size the CAG repeat region of exon one of the AR gene in each group. Statistical analysis revealed no actual link between the length of the CAG tract and a reduction of spermatogenesis in a cohort of infertile patients (n = 66) of Irish ethnic origin when compared to a fertile control group (n = 77) (p = 0.599).     

CAG repeat polymorphism in androgen receptor gene is not directly associated with polycystic ovary syndrome but influences serum testosterone levels

Hyperandrogenemia has been the most consistent feature of polycystic ovary syndrome (PCOS). Androgens exert their effects through androgen receptors (ARs). The expansion of the codon CAG trinucleotide repeat polymorphism in exon 1 of the AR gene represents a type of genetic alteration associated with changes in the AR gene function. The purpose of this study was to establish a possible association of the AR gene CAG repeat length polymorphism with PCOS, and its influence on clinical and biochemical androgen traits. Two hundred and fourteen Croatian women with PCOS and 209 healthy control women of reproductive age were enrolled. Phenotypic hyperandrogenism, BMI and waist to hip ratio were recorded. Hormonal profiles, fasting insulin and glucose levels were measured on cycle days 3-5. Genotyping of the CAG repeat polymorphism in the AR gene was performed. We found no significant difference in the mean CAG repeat number between the PCOS patients and controls (22.1±3.4 vs. 21.9±3.2, P=0.286). There was a positive correlation between the CAG repeat length and total testosterone (TT) in the PCOS group (R=0.225, P=0.015). A multiple linear regression model using mean CAG repeat length, BMI, age and HOMA-IR as predictors explained 8.5% (adjusted R2) of the variability in serum TT levels. In this model the CAG repeat polymorphism was found to be a significant predictor of serum TT levels in PCOS patients (P=0.015). The logistic regression analysis revealed that the CAG repeat length is not a significant predictor of hirsutism and acne status (P=0.921 and P=0.437, respectively). The model was adjusted for serum TT, free testosterone, androstendione and DHEAS levels as independent variables, which were also not found to be significant predictors of hirsutism (P=0.687, P=0.194, P=0.675 and P=0.938, respectively) or acne status (P=0.594, P=0.095, P=0.290 and P=0.151, respectively). In conclusion, the AR CAG repeat polymorphism is not a major determinant of PCOS in the Croatian population, but it is a predictor of serum TT level variability in women with PCOS.     

Long CAG Repeat Sequence and Protein Expression of Androgen Receptor Considered as Prognostic Indicators in Male Breast Carcinoma

Background

The androgen receptor (AR) expression and the CAG repeat length within the AR gene appear to be involved in the carcinogenesis of male breast carcinoma (MBC). Although phenotypic differences have been observed between MBC and normal control group in AR gene, there is lack of correlation analysis between AR expression and CAG repeat length in MBC. The purpose of the study was to investigate the prognostic value of CAG repeat lengths and AR protein expression.

Methods

81 tumor tissues were used for immunostaining for AR expression and CAG repeat length determination and 80 normal controls were analyzed with CAG repeat length in AR gene. The CAG repeat length and AR expression were analyzed in relation to clinicopathological factors and prognostic indicators.

Results

AR gene in many MBCs has long CAG repeat sequence compared with that in control group (P = 0.001) and controls are more likely to exhibit short CAG repeat sequence than MBCs. There was statistically significant difference in long CAG repeat sequence between AR status for MBC patients (P = 0.004). The presence of long CAG repeat sequence and AR-positive expression were associated with shorter survival of MBC patients (CAG repeat: P = 0.050 for 5y-OS; P = 0.035 for 5y-DFS AR status: P = 0.048 for 5y-OS; P = 0.029 for 5y-DFS, respectively).

Conclusion

The CAG repeat length within the AR gene might be one useful molecular biomarker to identify males at increased risk of breast cancer development. The presence of long CAG repeat sequence and AR protein expression were in relation to survival of MBC patients. The CAG repeat length and AR expression were two independent prognostic indicators in MBC patients.     

Androgen receptor CAG polymorphism and prostate cancer risk

Recent studies have suggested that polymorphisms of the androgen receptor gene ( AR) may influence the risk of prostate cancer (PC) development and progression. Here, we analyzed the length of the CAG repeat of the AR gene in 1363 individuals, including patients with PC, benign prostate hyperplasia (BPH), and population controls. There was a tendency for short CAG repeats to be associated with PC. The Odds Ratio (OR) for PC was 1.47 ( P=0.05) when individuals with short CAG repeats (18). CAG repeat length was not significantly associated with family history, disease stage, grade, age at diagnosis, prostate-specific antigen (PSA) level at diagnosis, or prognosis of the patients. Unexpectedly, short CAG repeats were significantly less common in patients with BPH compared with controls (OR=0.47, P=0.03). Our results suggest that the CAG polymorphism of the AR gene is unlikely to have a major role in the development or progression of PC in the Finnish population. The association of CAG repeats with the risk of BPH warrants further study.     

GGC and StuI polymorphism on the androgen receptor gene in endometrial cancer patients

Androgens have an anti-proliferative effect on endometrial cells. Human androgen receptor (AR) gene contains two polymorphic short tandem repeats of GGC and CAG, and a single-nucleotide polymorphism on exon 1 that is recognized by the restriction enzyme, StuI. Prior studies have shown that the lengths of the CAG repeat are inversely and linearly related to AR activity and associated with endometrial cancer. However, little is known about the GGC repeat and the StuI polymorphism of the AR gene. Thus, we investigated whether these AR polymorphisms are risk factors for endometrial cancer. To test this hypothesis, the genetic distributions of these polymorphisms were investigated in blood samples from endometrial cancer patients and healthy controls. The allelic and genotyping profiles were analyzed by polymerase chain reaction (PCR), PCR-restriction fragment length polymorphism (PCR-RFLP), and direct DNA sequencing, and analyzed statistically. The GGC repeat was significantly longer in endometrial cancer patients as compared to normal healthy controls. In general, an increased risk of endometrial cancer was found with increasing GGC repeat. The relative risk for the 17 GGC repeat was greater than 4, as compared to controls. However, the StuI polymorphism was not significantly different between patients and controls. The findings suggest that increased numbers of GGC repeat on the AR gene may be a risk factor for endometrial cancer.     

The androgen receptor CAG repeat length polymorphism associates with prostate volume in Finnish men with benign prostatic hyperplasia

The development of benign prostatic hyperplasia requires the presence of testicular androgens during prostate development, puberty, and ageing. We thus examined the association of three polymorphisms, namely, CYP3A5 6986A>G, CYP19A1 1531C>T, and androgen receptor (AR) gene CAG repeat length, which have previously been linked to the androgen pathway and with clinical characteristics of benign prostatic hyperplasia. Tissue samples from 262 consecutive prostate operations were used for genotyping. Prostate volumes and prostate-specific antigen values were collected from patient records. Linear regression analysis was performed to study the polymorphisms in an age-adjusted model. We did not find any association between the CYP3A5 6986A>G polymorphism and clinical characteristics of benign prostatic hyperplasia. Further, the previously published CYP19A1 1531C>T polymorphism association with an enlarged prostate could not be confirmed with this material. However, we detected an association between short AR gene CAG repeat length and a small prostate volume, which confirms a previous finding in the Finnish population. The data presented suggest a negligible role for the CYP3A5 6986A>G polymorphism in benign prostate enlargement in the Finnish population. However, the results presented do provide further evidence for potentially different genetic mechanisms behind benign prostatic hyperplasia in Finnish and other Caucasian populations. This is based on the conflicting results for AR gene CAG repeat length associations with benign prostatic hyperplasia found in published works.     

Polymorphisme du gène du récepteur des androgènes: recherche de mosaïques tissulaires et relation avec l'infertilité masculine

Androgens are male steroid hormones necessary for initiation and maintenance of spermatogenesis. They act via a specific receptor which belongs to the nuclear receptor superfamily. The transactivation N-terminal domain of this receptor is characterized by a CAG repeat polymorphism in the first exon of the gene coding for polyglutamines. An increased CAG repeat length is involved in Kennedy's disease, a neurodegenerative disease associated with infertility and impaired virilization. Some recent studies have shown a correlation between the number of CAG repeats and male fertility. The aim of this study was to define this correlation and to determine whether or not infertile men presented a tissue mosaicism between blood and sperm, as described in several diseases involving CAG repeats. The length of CAG repeats of blood and testicular cells was measured in 36 oligospermic or azoospermic patients and 15 controls. An inverse correlation was found between CAG repeat length and total number of ejaculated spermatozoa. However, no tissue mosaicism between blood and sperm was observed in our population.     

Polymorphic GGC repeats in the androgen receptor gene are associated with hereditary and sporadic prostate cancer risk

Androgen receptor (AR) has long been hypothesized to play an important role in prostate cancer etiology. Two trinucleotide repeat polymorphisms (CAG and GGC repeats in exon 1 of the AR gene) have been investigated as risk factors for prostate cancer in several studies. However, the results are inconclusive, probably because of the variations of study designs, characteristics of study samples, and choices of analytical methods. In this study, we evaluated evidence for linkage and association between the two AR repeats and prostate cancer by using the following comprehensive approaches: (1) a combination of linkage and association studies, (2) a test for linkage by parametric analysis and the male-limited X-linked transmission/disequilibrium test (XLRC-TDT), (3) a test for association by using both population-based and family-based tests, and (4) a study of both hereditary and sporadic cases. A positive but weak linkage score (HLOD=0.49, P=0.12) was identified in the AR region by parametric analysis; however, stronger evidence for linkage in the region, especially at the GGC locus, was observed in the subset of families whose proband had < or = 16 GGC repeats (HLOD=0.70, P=0.07) or by using XLRC-TDT ( z'=2.65, P=0.008). Significantly increased frequencies of the < or = 16 GGC repeat alleles in 159 independent hereditary cases (71%) and 245 sporadic cases (68%) cases compared with 211 controls (59%) suggested that GGC repeats were associated with prostate cancer ( P=0.02). Evidence for the association between the < or = 16 GGC repeats and prostate cancer risk was stronger with XLRC-TDT ( z'=2.66, P=0.007). No evidence for association between the CAG repeats and prostate cancer risk was observed. The consistent results from both linkage and association studies strongly implicate the GGC repeats in the AR as a prostate cancer susceptibility gene. Further studies on this polymorphism in other independent data sets and functional analysis of the GGC repeat length on AR activity are warranted.     

In vivo expansion of trinucleotide repeats yields plasmid and YAC constructs for targeting and transgenesis

Production of mouse models of inherited neurodegenerative diseases is an important step towards understanding the mechanism of neurotoxicity and for testing potential therapies. We are interested in creating a mouse model for X-linked spinal and bulbar muscular atrophy (SBMA), a neuromuscular disorder caused by expansion of a CAG repeat within the androgen receptor (AR) gene. To permit generation of mice that will show a SBMA phenotype within their life span, we decided to obtain a yeast artificial chromosome (YAC) carrying the AR gene and introduce CAG repeat mutations numbering 100 or more triplets. SBMA patients with more than 70 CAGs have never been observed; therefore, we chose to expand a 59 CAG repeat tract in vivo in Escherichia coli. Although we set out to expand this repeat tract using a recombination paradigm involving two plasmid co-propagation, we did not observe large expansions. We were instead able to incrementally generate repeat tracts from 100 to 200 CAGs in a yeast integrating plasmid vector by taking advantage of replication instability. In the course of our experiments that yielded these CAG repeat tracts, we evaluated the role of repeat orientation, vector co-propagation, and recA function on the expansion process. We then used one of the yeast integrating vectors to successfully produce an AR YAC construct carrying 100 CAG repeats. AR YAC CAG100 will serve as a valuable reagent for the production of a SBMA mouse.     

CAG repeat variants in the POLG1 gene encoding mtDNA polymerase-gamma and risk of breast cancer in African-American women

The DNA polymerase-gamma (POLG) gene, which encodes the catalytic subunit of enzyme responsible for directing mitochondrial DNA replication in humans, contains a polyglutamine tract encoded by CAG repeats of varying length. The length of the CAG repeat has been associated with the risk of testicular cancer, and other genomic variants that impact mitochondrial function have been linked to breast cancer risk in African-American (AA) women. We evaluated the potential role of germline POLG-CAG repeat variants in breast cancer risk in a sample of AA women (100 cases and 100 age-matched controls) who participated in the Women's Circle of Health Study, an ongoing multi-institutional, case-control study of breast cancer. Genotyping was done by fragment analysis in a blinded manner. Results from this small study suggest the possibility of an increased risk of breast cancer in women with minor CAG repeat variants of POLG, but no statistically significant differences in CAG repeat length were observed between cases and controls (multivariate-adjusted odds ratio 1.74; 95% CI, 0.49-6.21). Our study suggests that POLG-CAG repeat length is a potential risk factor for breast cancer that needs to be explored in larger population-based studies.     

Polymorphism of CAG repeats in androgen receptor of carnivores

Androgen effect is mediated by the androgen receptor (AR). The polymorphism of CAG triplet repeat (polyCAG), in the N-terminal transactivation domain of the AR protein, has been involved either in endocrine or neurological disorders in human. We obtained partial sequence of AR exon 1 in 10 carnivore species. In most carnivore species, polyglutamine length polymorphism presented in all three CAG repeat regions of AR, in contrast, only CAG-I site polymorphism presented in primate species, and CAG-I and CAG-III sites polymorphism presented in Canidae. Therefore, studies focusing on disease-associated polymorphism of poly(CAG) in carnivore species AR should investigate all three CAG repeats sites, and should not only consider CAG-I sites as the human disease studies. The trinucleotide repeat length in carnivore AR exon 1 had undergone from expansions to contractions during carnivores evolution, unlike a linear increase in primate species. Furthermore, the polymorphisms of the triplet-repeats in the same tissue (somatic mosaicism) were demonstrated in Moutain weasel, Eurasian lynx, Clouded leopard, Chinese tiger, Black leopard and Leopard AR. And, the abnormal stop codon was found in the exon 1 of three carnivore species AR (Moutain weasel, Eurasian lynx and Black leopard). It seemed to have a high frequency presence of tissue-specific somatic in carnivores AR genes. Thus the in vivo mechanism leading to such highly variable phenotypes of the described mutations, and their impact on these animals, are worthwhile to be further elucidated.     

No CAG repeat expansion of polymerase gamma is associated with male infertility in Tamil Nadu,South India

Mitochondria contains a single deoxyribonucleic acid (DNA) polymerase, polymerase gamma (POLG) mapped to long arm of chromosome 15 (15q25), responsible for replication and repair of mitochondrial DNA. Exon 1 of the human POLG contains CAG trinucleotide repeat, which codes for polyglutamate. Ten copies of CAG repeat were found to be uniformly high (0.88) in different ethnic groups and considered as the common allele, whereas the mutant alleles (not -10/not -10 CAG repeats) were found to be associated with oligospermia/oligoasthenospermia in male infertility. Recent data suggested the implication of POLG CAG repeat expansion in infertility, but are debated. The aim of our study was to explore whether the not -10/not -10 variant is associated with spermatogenic failure. As few study on Indian population have been conducted so far to support this view, we investigated the distribution of the POLG CAG repeats in 61 infertile men and 60 normozoospermic control Indian men of Tamil Nadu, from the same ethnic background. This analysis interestingly revealed that the homozygous wild type genotype (10/-10) was common in infertile men (77% - 47/61) and in normozoospermic control men (71.7% - 43/60). Our study failed to confirm any influence of the POLG gene polymorphism on the efficiency of the spermatogenesis.     

Evaluation of the effects of androgen receptor gene trinucleotide repeats and prostate-specific antigen gene polymorphisms on prostate cancer

The number of trinucleotide repeats [CAG (coding for polyglutamine), GGC (coding for polyglycine)] in the first exon of the androgen receptor (AR) gene and prostate-specific antigen (PSA) gene androgen response element I A/G polymorphism are both related to prostate cancer prognosis. We investigated whether these genomic changes occur in the AR and PSA genes, which are usually found in individuals with prostate cancer, of Turkish patients and to find out their distribution in the population. We used PCR and PCR-RFLP assays for AR and PSA genes, respectively, to detect molecular changes in 44 prostate cancer patients. Our findings indicate that individuals with prostate cancer tend to have around 18 CAG trinucleotide repeats. We observed significant differences between 22 controls, 33 benign prostate hyperplasia (BPH) patients and 44 adenocarcinoma patients for long CAG repeats. However, we did not find any significant differences in GGC repeats between controls, BPH and adenocarcinoma patients (P = 0.408). We also did not observe significant differences in the PSA A/G polymorphism frequency between controls, BPH and adenocarcinoma patients (P = 0.483). In conclusion, CAG and GGC repeats in the AR and PSA gene polymorphisms may be associated with prostate cancer risk and BPH in the Turkish population.     

Variation in CAG repeat length of the androgen receptor gene predicts variables associated with intrasexual competitiveness in human males

An expanding body of research suggests that circulating androgens regulate the allocation of energy between mating and survival effort in human males, with higher androgen levels promoting greater investment in mating effort. Because variations in the number of CAG codon repeats in the human androgen receptor (AR) gene appear to modulate the phenotypic effects of androgens - with shorter repeat lengths associated with greater androgenic effects per unit androgen - polymorphisms in this gene may predict trait-like individual differences in the degree to which men are calibrated toward greater mating effort. Consistent with this, men in the present study with shorter CAG repeat lengths exhibited greater upper body strength and scored higher on self-report measures of dominance and prestige, all of which are argued to be indices of mating effort. Repeat length failed to predict sociosexual orientation (i.e. pursuit of short-term mating relationships), however, suggesting that the traits correlated with this polymorphism may be primarily associated with intrasexual competitiveness in the service of long-term mating effort. None of these measures of mating effort was related to baseline testosterone concentrations (either as main effects or as interactions with CAG repeat length), implying that long-term androgen exposure associated with AR gene polymorphisms may account for more variance in some androgen-dependent traits than does current testosterone concentration. These findings provide further evidence for the importance of the CAG repeat polymorphism in the AR gene in explaining a broad range of individual differences in human males.     

Modification of BRCA1-associated breast cancer risk by the polymorphic androgen-receptor CAG repeat

Compared with the general population, women who have inherited a germline mutation in the BRCA1 gene have a greatly increased risk of developing breast cancer. However, there is also substantial interindividual variability in the occurrence of breast cancer among BRCA1 mutation carriers. We hypothesize that other genes, particularly those involved in endocrine signaling, may modify the BRCA1-associated age-specific breast cancer risk. We studied the effect of the CAG repeat-length polymorphism found in exon 1 of the androgen-receptor (AR) gene (AR-CAG). AR alleles containing longer CAG repeat lengths are associated with a decreased ability to activate androgen-responsive genes. Using a sample of women who inherited germline BRCA1 mutations, we compared AR-CAG repeat length in 165 women with and 139 women without breast cancer. We found that women were at significantly increased risk of breast cancer if they carried at least one AR allele with >/=28 CAG repeats. Women who carried an AR-CAG allele of >/=28, >/=29, or >/=30 repeats were given a diagnosis 0.8, 1.8, or 6.3 years earlier than women who did not carry at least one such allele. All 11 women in our sample who carried at least one AR-CAG allele with >/=29 repeats had breast cancer. Our results support the hypothesis that age at breast cancer diagnosis is earlier among BRCA1 mutation carriers who carry very long AR-CAG repeats. These results suggest that pathways involving androgen signaling may affect the risk of BRCA1-associated breast cancer.