Genotype-Phenotype Heterogeneity in Haemophilia

Haemophilia was previously regarded as a classical example of Mendelian inheritance, with mutation in only a single gene (F8 or F9) causing the disease phenotype. The disease manifests complete penetrance. Studies, however, revealed the striking genetic and phenotypic heterogeneities of the disease. With further sophistication of clinical and molecular techniques, the disease was also found to have allele heterogeneity, phenotypic plasticity and variation in expressivity. The variations are more pronounced in F9 variants with five distinct phenotypes. All these phenomena advocate a rather complex genotype-phenotype relationship for the disease. A keen insight into the matter may unveil new avenues of therapeutics

Sensitive detection of pre-existing BCR-ABL kinase domain mutations in CD34+ cells of newly diagnosed chronic-phase chronic myeloid leukemia patients is associated with imatinib resistance: implications in the post-imatinib era.

BACKGROUND: BCR-ABL kinase domain mutations are infrequently detected in newly diagnosed chronic-phase chronic myeloid leukemia (CML) patients. Recent studies indicate the presence of pre-existing BCR-ABL mutations in a higher percentage of CML patients when CD34+ stem/progenitor cells are investigated using sensitive techniques, and these mutations are associated with imatinib resistance and disease progression. However, such studies were limited to smaller number of patients. METHODS: We investigated BCR-ABL kinase domain mutations in CD34+ cells from 100 chronic-phase CML patients by multiplex allele-specific PCR and sequencing at diagnosis. Mutations were re-investigated upon manifestation of imatinib resistance using allele-specific PCR and direct sequencing of BCR-ABL kinase domain. RESULTS: Pre-existing BCR-ABL mutations were detected in 32/100 patients and included F311L, M351T, and T315I. After a median follow-up of 30 months (range 8-48), all patients with pre-existing BCR-ABL mutations exhibited imatinib resistance. Of the 68 patients without pre-existing BCR-ABL mutations, 24 developed imatinib resistance; allele-specific PCR and BCR-ABL kinase domain sequencing detected mutations in 22 of these patients. All 32 patients with pre-existing BCR-ABL mutations had the same mutations after manifestation of imatinib-resistance. In imatinib-resistant patients without pre-existing BCR-ABL mutations, we detected F311L, M351T, Y253F, and T315I mutations. All imatinib-resistant patients except T315I and Y253F mutations responded to imatinib dose escalation. CONCLUSION: Pre-existing BCR-ABL mutations can be detected in a substantial number of chronic-phase CML patients by sensitive allele-specific PCR technique using CD34+ cells. These mutations are associated with imatinib resistance if affecting drug binding directly or indirectly. After the recent approval of nilotinib, dasatinib, bosutinib and ponatinib for treatment of chronic myeloid leukemia along with imatinib, all of which vary in their effectiveness against mutated BCR-ABL forms, detection of pre-existing BCR-ABL mutations can help in selection of appropriate first-line drug therapy. Thus, mutation testing using CD34+ cells may facilitate improved, patient-tailored treatment

Biallelic variants in LINGO1 are associated with autosomal recessive intellectual disability, microcephaly, speech and motor delay

Purpose: To elucidate the novel molecular cause in two unrelated consanguineous families with autosomal recessive intellectual disability. Methods: A combination of homozygosity mapping and exome sequencing was used to locate the plausible genetic defect in family F162, while only exome sequencing was followed in the family PKMR65. The protein 3D structure was visualized with the University of California–San Francisco Chimera software. Results: All five patients from both families presented with severe intellectual disability, aggressive behavior, and speech and motor delay. Four of the five patients had microcephaly. We identified homozygous missense variants in LINGO1, p.(Arg290His) in family F162 and p.(Tyr288Cys) in family PKMR65. Both variants were predicted to be pathogenic, and segregated with the phenotype in the respective families. Molecular modeling of LINGO1 suggests that both variants interfere with the glycosylation of the protein. Conclusion: LINGO1 is a transmembrane receptor, predominantly found in the central nervous system. Published loss-of-function studies in mouse and zebrafish have established a crucial role of LINGO1 in normal neuronal development and central nervous system myelination by negatively regulating oligodendrocyte differentiation and neuronal survival. Taken together, our results indicate that biallelic LINGO1 missense variants cause autosomal recessive intellectual disability in humans

Patients’ characteristics.

<p>Ph+<b> = </b>Philadelphia chromosome positive.</p><p>PEM = pre existing mutations.</p

Comparison of the frequencies of pre-existing BCR-ABL KD mutations and mutations detected after manifestation of imatinib resistance in CML patients.

<p>Comparison of the frequencies of pre-existing BCR-ABL KD mutations and mutations detected after manifestation of imatinib resistance in CML patients.</p

Clinical, cytogenetic, and molecular follow-up studies of CML patients with and without BCR-ABL PEMs who received imatinib treatment.

<p>N: number of patients, PEMs: pre-existing mutations; IM: imatinib; CHR: complete hematological response; PHR: partial hematological response; CCyR: complete cytogenetic response; MCyR: major cytogenetic response; minor CyR: minor cytogenetic response; MMR: major molecular response.</p

Sequences of ASO primers and corresponding annealing temperatures (bold nucleotides in the primers denote nucleotide changes corresponding to mutations).

*<p>Substitutions of amino acids; positions according to GenBank no. AAB60394for ABL type 1a.</p>**<p>Changes of nucleotide; positions according to GenBank no.M14752.</p>$<p>L (bp) = Primer length in base pairs.</p>#<p>A Tm =  Annealing temperature in degree Celsius.</p