NOD2/CARD15 polymorphisms (P268S, IVS8+158, G908R, L1007fs, R702W) among Kuwaiti patients with Crohn's disease: A case-control study.

Background: Nucleotide-binding oligomerization domain-containing two (NOD2/CARD15) gene polymorphisms are implicated in the pathogenesis of Crohn's disease (CD). Aim: To describe the allelic frequency of NOD2/CARD15 gene variants among Kuwaiti patients with CD and investigate potential genotype/phenotype associations. Methods: Adult Kuwaiti citizens with an established diagnosis of CD and healthy controls were enrolled from October 2018 to May 2020. Three common NOD2/CARD15 polymorphisms (R702W, G908R, and L1007fs) and P268S and IVS8+158 polymorphisms were screened by polymerase chain reaction/restriction analysis length polymorphism (PCR/RFLP). Results: Ninety adult Kuwaiti patients with CD and 210 healthy subjects (as controls) were recruited. P268S, IVS8+158, G908R, and R702W minor alleles were identified in 38.9%, 21.1%, 12.2%, and 4.4% of CD patients, respectively. NOD2/CARD15 polymorphisms coexisted in 35 healthy controls (16.7%) and 21 CD patients (23.3%). Individuals with either a single or multiple polymorphism were approximately two times more likely to have CD than those with no polymorphism. Patients with multiple polymorphisms had significantly more stricturing and penetrating disease. Conclusion: NOD2/CARD15 gene polymorphisms were significantly associated with an increased risk of disease and aggressive phenotypes among the Kuwaiti CD population.

Mutation spectrum analysis of Duchenne/Becker muscular dystrophy in 68 families in Kuwait: The era of personalized medicine.

Duchenne and Becker muscular dystrophies (DMD/BMD) are X-linked recessive neuromuscular disorders characterized by progressive irreversible muscle weakness and atrophy that affect both skeletal and cardiac muscles. DMD/BMD is caused by mutations in the Dystrophin gene on the X chromosome, leading to the absence of the essential muscle protein Dystrophin in DMD. In BMD, Dystrophin is partially functioning with a shorter protein product. Recent advances in molecular therapies for DMD require precise genetic diagnoses because most therapeutic strategies are mutation-specific. Hence, early diagnosis is crucial to allow appropriate planning for patient care and treatment. In this study, data from DMD/BMD patients who attended the Kuwait Medical Genetic Center during the last 20 years was retrieved from a Kuwait neuromuscular registry and analyzed. We combined multiplex PCR and multiplex ligation-dependent probe amplification (MLPA) with Sanger sequencing to detect Dystrophin gene mutations. A total of 35 different large rearrangements, 2 deletion-insertions (Indels) and 4 substitution mutations were identified in the 68 unrelated families. The deletion and duplication rates were 66.2% and 4.4%, respectively. The analyzed data from our registry revealed that 11 (16%) of the DMD families will benefit from newly introduced therapies (Ataluren and exon 51 skipping). At the time of submitting this paper, two cases have already enrolled in Ataluren (Tranlsarna™) therapy, and one case has been enrolled in exon 51 skipping therapy.

The genomic architecture of NLRP7 is Alu rich and predisposes to disease-associated large deletions.

NLRP7 is a major gene responsible for recurrent hydatidiform moles. Here, we report 11 novel NLRP7 protein truncating variants, of which five deletions of more than 1-kb. We analyzed the transcriptional consequences of four variants. We demonstrate that one large homozygous deletion removes NLRP7 transcription start site and results in the complete absence of its transcripts in a patient in good health besides her reproductive problem. This observation strengthens existing data on the requirement of NLRP7 only for female reproduction. We show that two other variants affecting the splice acceptor of exon 6 lead to its in-frame skipping while another variant affecting the splice donor site of exon 9 leads to an in-frame insertion of 54 amino acids. Our characterization of the deletion breakpoints demonstrated that most of the breakpoints occurred within Alu repeats and the deletions were most likely mediated by microhomology events. Our data define a hotspot of Alu instability and deletions in intron 5 with six different breakpoints and rearrangements. Analysis of NLRP7 genomic sequences for repetitive elements demonstrated that Alu repeats represent 48% of its intronic sequences and these repeats seem to have been inserted into the common NLRP2/7 primate ancestor before its duplication into two genes.

Impact of 226C>T MSH2 gene mutation on cancer phenotypes in two HNPCC-associated highly-consanguineous families from Kuwait: emphasis on premarital genetic testing.

Lynch syndrome or hereditary nonpolyposis colorectal cancer (HNPCC) is one of the commonest cancer susceptibility syndromes. It is characterized by early onset colon cancer and a variety of extracolonic tumours. Germline mutations in the DNA mismatch repair genes (MLH1, MSH2, MSH6, PMS1, and PMS2) are responsible for this disorder. Identifying an affected individual depends on the tumour histopathology, family history that fulfils the Amsterdam and/or Bethesda criteria, tumour immunohistochemistry, microsatellite instability, and finally molecular analysis of an affected member. It is a laborious, time consuming and expensive procedure, which needs the effort of a multi-disciplinary team. However, once the diagnosis is established and germline defect is identified, other high risk pre-symptomatic carriers could be offered intensive surveillance and management as a preventive measure against cancer development. Here, we present two large highly consanguineous HNPCC-families from Kuwait in whom a founder MSH2 mutation was identified. The relationship between this mutation and cancer expressivity in two large consanguineous families harbouring other genetic defects is discussed. Moreover, we shed light on the challenges pertaining to diagnosis, screening, premarital counselling of couples and prenatal diagnosis of offspring with biallelic MSH2 gene mutation.

Molecular and clinical evaluation of primary congenital glaucoma in Kuwait.

PURPOSE: To report the spectrum of the CYP1B1 mutation in Kuwaiti patients with primary congenital glaucoma (PCG). DESIGN: Clinical diagnosis of PCG and laboratory based experimental study. METHODS: Polymerase chain reaction-restriction polymorphism length fragment (PCR-RPLF) and direct sequencing of exon 2 and the coding region of exon 3 of CYP1B1 gene were the methods used for screening 17 PCG patients, their families, and 105 health individuals from the same ethnicity. RESULTS: Four different mutations were detected in CYP1B1 in 70.6% of the screened patients. The most common one (47%) was homozygote Gly61Glu mutation, previously described in Saudi Arabia, Turkey, and Morocco; all patients were products of consanguineous marriages. The second common mutation was a novel missense (Ala388Thr) mutation found in three patients (17.6%) as compound heterozygote with Arg368His in one patient, and with Gly61Glu in another one while the second mutation in third patient was not detected in the CYP1B1 gene. One patient (5.8%) was homozygote for Cyt280X mutation previously reported in only one Japanese family. In addition to these mutations, a novel Val422Gly polymorphic site was found in three of the PCG patients and in 18 of the 210 tested chromosomes of healthy volunteers. CONCLUSIONS: The CYP1B1 mutation spectrum of Kuwaiti PCG patients is similar to that detected in the neighboring countries. No clear genotype-phenotype correlation detected in patients showed different types of CYP1B1 mutation.

Molecular characterization of glucose-6-phosphate dehydrogenase gene defect in the Kuwaiti population.

CONTEXT: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is one of the most common human enzyme deficiencies. More than 130 different molecular abnormalities have been described worldwide, with considerable variation in the enzyme among various racial groups. Data from Kuwaiti populations are scarce, and the studies available are the result of screening male blood donors who may not be truly representative of the Kuwaiti population. OBJECTIVE: The objective of this study was to investigate the mutation spectrum of the G6PD gene among Kuwaiti Arabs. DESIGN: DNA was extracted from 82 G6PD-deficient Kuwaiti subjects (75 men and 7 women) and screened for gene mutations using polymerase chain reaction/restriction fragment length polymorphism and polymerase chain reaction/single-strand conformation polymorphism followed by direct sequencing. A total of 1209 randomly selected Kuwaiti adult subjects of both sexes were then screened for any characterized mutation. RESULTS: G6PD Mediterranean(563C-->T), and A-(202G-->A,376A-->G) genotypes were characterized as the most common variants among the G6PD-deficient population, representing 0.742 and 0.124 allele frequencies, respectively. The 2 previously described mutations, G6PD Chatham(1003G-->A) and Aures(143T-->C), were found at lower frequencies (0.101 and 0.034, respectively). The allele frequencies for these 4 G6PD variants among the randomly selected Kuwaitis were 0.035, 0.0074, 0.0046, and 0.0023 for Mediterranean, A-, Chatham, and Aures, respectively. CONCLUSION: This study has characterized the molecular heterogeneity of G6PD variants among ethnic Kuwaitis. The findings suggest that gene flow from the Indian subcontinent, sub-Saharan African, and other parts of the Mediterranean may have contributed to the observed G6PD mutations seen in the Kuwaiti population.

CTG repeat number at the myotonic dystrophy locus in healthy Kuwaiti individuals: possible explanation of why myotonic dystrophy is rare in Kuwait.

BACKGROUND: Myotonic dystrophy is caused by unstable (CTG)(n) repeat expansion. On normal chromosomes, this repeat is highly polymorphic, with a copy number ranging from 4 to 38. Myotonic dystrophy is considered more prevalent in Western European and Japanese populations but less prevalent, rare, or even absent in others. It has been proposed that the expanded (CTG)(n) alleles originated from the group of the large normal alleles. OBJECTIVE: To determine whether there is a lower prevalence of the large alleles in the Kuwaiti population. DESIGN AND PARTICIPANTS: We determined the size distribution of the CTG repeats by means of polymerase chain reaction in blood DNA derived from 185 healthy Kuwaiti individuals representing the 5 Kuwaiti provinces. RESULTS: We found a total of 17 (CTG)(n) alleles, with a range of 5 to 37 repeats. The (CTG)(5) allele was the most frequent single allele (100/370 [27.0%]), whereas the (CTG)(10-13) was the most frequent class of alleles (161/370 [43.5%]). Using 18 repeats as the cutoff point, chi(2) analysis showed a statistically significant lower frequency of greater than 18 alleles in the Kuwaiti population compared with the European population (chi(2) = 12.7; P<.001). CONCLUSIONS: These data may explain the rare occurrence of myotonic dystrophy in the Kuwaiti population. Further study of healthy families within the high-normal repeat range is in progress to investigate the possible instability of the (CTG)(>18) alleles in our area.