IN THE NAME OF ALLAH THE MOST BENEFICAL THE MOST MERCIFUL

1. IN THE NAME OF ALLAH THE MOST BENEFICAL THE MOST MERCIFUL

2. ASPM gene analysis in Pakistani families with primary microcephaly mapped to MCPH5 locus By Shamim Saleha PhD ScholarDepartment of Biotechnology and Genetic Engineering, KUST

3. Genetic Diseases: “A genetic disease is due to a faulty gene or group of genes. It occurs when recessive and/or dominant genes are passed down to the baby from generation to generation.” (Genetic Diseases Forum) • Genetic diseases are : • Congenital • Serious • Incurable • Treatable • Paramount importance • Family history • Genetic counseling • Currently about 6,000 genetic disorders known (Wade and Nicholas, 2006)

4. Neurological Inherited Disorders: • Large group of clinically heterogeneous diseases. (Klein, 2006) • Over 15 Neurological disorders are known to have a genetic cause. (NINDS, 2009) • Neurological disorders affect • Brain • Spinal cord • Muscles • Nerves (Hazama et al., 2009) • Rare and poorly understood(NINDS, 2009 )

5. Human Inherited diseases: • Monogenic disorders • disease of autosomal dominant, AD • disease of autosomal recessive, AR • disease of X-linked dominant inheritance, XD • disease of X-linked recessive inheritance, XR • disease of Y-linked inheritance • Polygenic diseases • Chromosomal diseases • Somatic inherited diseases (Valente et al., 2008)

6. Autosomal Recessive Diseases: • Among genetic disorders, that are strongly associated with consanguinity, are inherited only in autosomal recessive manner. (Hamamy et al., 2007) • AR diseases may emerge worldwide in a population as the prevalence of a deleterious gene or when degree of consanguinity increases. (Campbell et al., 2009) • In case of autosomal recessive inheritance, both parents and two thirds of surviving offspring are mark as carriers . (Roberts et al. 2002)

7. Consanguinity in World: • Only 1% : • Western Europe • North America • Australasia • Russia • 1% to 10% : • Iberian Peninsula • Japan • SouthAmerica • 20% to 50% : • North and sub-Saharan Africa • West, Central and South Asia (Bittles, 2008)

8. Consanguinity in Pakistan: • In Pakistan, approximately • 60% of marriages are consanguineous (Bittles, 2008) • Daily Times reported • 82.5% parents are first cousins • 6.8% are blood relatives • 6.3% belong to a same caste and family • only 4.4% are married out of their families. (Khan et al.,2008) • Practice of Consanguineous marriages • Economic • safety reasons • Ethnic groups • Religion (Campbell et al., 2009)

9. Genetic Disorders in Khyber PakhtoonKhwa: • Factors that contribute to the wide prevalence of genetic disorders, in this region, are: • High rate of consanguinity • Social trend to have more children until menopause • Practice of autogamy in Pathans • Lack of public awareness towards the early recognition and prevention of inherited disease. • People do not agree with medical explanations of a genetic mode of disease inheritance (Daily Times ,2008)

10. Objectives: • To trace genetic illnesses in those affected families that posses a pedigree records of Microcephaly • To determine phenotypes by clinical assessments and genotype/phenotype correlation by genetic analysis. • To identify and map diseased loci by genetic linkage studies. • To find the genetic complexity or heterogeneity of Microcephaly by mutation screening of diseased genes.

11. Materials & Methods: • Pedigree collection and Analysis • Blood Sampling • Human Genomic DNA Extraction • Linkage Studies by STR markers (http://www.uia.ac.be/dnalab/hhh/) • Amplification of DNA by polymerase chain reaction (PCR) • Polyacrylamide gel electrophoresis • Direct DNA sequencing

12. Microcephaly:

13. Microcephaly:

14. Clinical Heterogeneity of Microcephaly: • Autosomal recessive primary microcephaly (MCPH) is disorder of neurodevelopment. • Individuals with MCPH phenotype • reduced brain weight • reduced head circumference of at least 4 SD at birth • non progressive mental retardation of variable degree (Woods et al.,2005)

15. Prevalenceof Microcephaly: • More common in consanguineous populations (Nicholas et al.,2008; Kousar et al.,2009) • Incidence • 1 in 10,000 in Pakistan (Nicholas et al.,2008) • 1 in 1,000,000 in Caucasian (Woods et al.,2005) • 1 in 30,000 in Japan (Komai et al.,1955) • 1 in 250,000 in Holland (Van den Bosch,1959) • 1 in 2,000,000 in Scotland (Tomie etal.,1987)

16. Linkage Studies: Disease was mapped at MCPH5 locus, the gene in this region, is ASPM

17. ASPM pedigree with genotyping data:

18. Primers Sequences of ASPM:

19. Mutation Screening: • Common mutation in both families • 200 control was not observed • Mutation changes nucleotide at position 5684, results in a substitution of lysine with arginine at position 1985 of protein.

20. Outcomes: • The mapping of large number of families to MCPH5 locus and identification of common mutation in ASPM gene in families of Pakistani origin will enable us to formulate future strategies to control and prevent the disease to reduce the prevalence of MCPH in Pakistan. • genetic counseling • prenatal diagnosis • postnatal diagnosis • carrier testing

21. Any Question?

22. Thanks!