Shagun Aggarwal , S. Jamal Mohamed Nurul Jain ...

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1- Department of Medical Genetics, 3- Department of Pathology, Nizam's Institute of Medical Sciences,2- Diagnostics Division, Centre for DNA Fingerprinting ...
Molecular autopsy identifies recombinant GBA gene in a case of Gaucher disease with Ichthyosis phenotype Shagun

1,2 Aggarwal ,

S. Jamal Mohamed Nurul

2 Jain ,

Ashwani

3 Tandon ,

Aneek D.

2 Bhowmik ,

Ashwin

2 Dalal

1- Department of Medical Genetics, 3- Department of Pathology, Nizam’s Institute of Medical Sciences,2- Diagnostics Division, Centre for DNA Fingerprinting and Diagnostics, Hyderabad

Introduction Molecular workup forms an important part of the perinatal autopsy and helps in confirming the diagnosis of a suspected genetic disease in a fetus with suggestive gross and microscopic findings. This case report illustrates the identification of perinatal lethal Gaucher disease in a fetus with ichthyosis, a dermatosis known to be genetically heterogeneous.

Case report Third degree consanguineous couple. History of previous pregnancy termination due to antenatal detection of cardiomegaly and polyhydramnios Present pregnancy: USG at 16 weeks- Echogenic bowel, USG at 27 weeks- Bilateral clenched hands, Micrognathia, cardiomegaly, growth restriction Generalised skin thickening. Taut, thick, glistening skin on hands, feet and face Short digits with nodular joints and telephalyngy, labial thickening

Autopsy findings

Thick ear lobules, thick lips with eclabium, thick, upturned nares with flat, short nose, ectropion, severe micrognathia

Structure of GBA genes and recombinants 16 Kb GBA GBAP

Intrathoracic examinationcardiomegaly with thick looking ventricular walls Intra-abdominal examinationHepatosplenomegaly

Molecular study

55bp deletion in D409H in normal control normal control L444P,A456P,V460V in normal control

References

Mother’s sample No 55bp deln, D409H Heterozygous L444P, A456P,V460V present

Conclusion

Simple PCR leads to coamplification of GBA and pseudogene

A456P

7765 bp long PCR product

V460V

Reciprocal and Nonreciprocal Recombination at the Glucocerebrosidase Gene Region: Implications for Complexity in Gaucher Disease. Am. J. Hum. Genet. 72:519–534, 2003. Type 2 Gaucher disease: the collodion baby phenotype revisited. Arch Dis Child Fetal Neonatal Ed 2000;82:F163–F166. Novel Mutations in the Glucocerebrosidase Gene of Brazilian Patients with Gaucher Disease. JIMD Reports DOI 10.1007/8904_2012_174.

Molecular testing forms an important part of fetal autopsy, Confirms clinical diagnosis Identifies underlying mutation, Can elucidate interesting genetic mechanisms, Improves genotype-phenotype correlation

L444P

A456P

V460V

Father’s sample No 55bp deln, D409H Heterozygous L444P, A456P,V460V present

L444P

Abdominal wall

Back

No blood or fibroblasts from fetus available Beta glucosidase enzyme assay on parental leucocytes Mother: 4.4 ,Father : 5.1,Normal : 6– 9 (nmol/hr/mg) Control sample No 55bp deln,D409H No L444P,A456P,V460V

V460V (G/C)

Dorsum of hand

Arm

Biochemical study

Long PCR leads to selective amplification of GBA gene

55bp Rec 1 (NciL)- L44P,A456P,V460V deletion D409H L444P A456P (G/C) (T/C) (G/C) Rec 2 (TL)- D409H,L444P,A456P,V460V Highly Homologous regions prone for Rec 3- 55bp deln, D409H,L444P,A456P,V460V recombination events

L444P

Primers for Long PCR

Histopathology- Skin findings suspicious of Ichthyosis Autolytic changes in liver,spleen and cardiac tissue

A456P

V460V

Discussion Both parents found to be heterozygous for the Rec Ncil recombinant allele Fetus retrospectively confirmed to be perinatal lethal Gaucher disease Rec Ncil allele: most common recombinant of GBA gene Presumed to occur from gene fusion/crossover event between GBA and GBAP at intron 9 or intron 9/exon 10 junction Homozygous state of RecNcil allele-Two cases of Afghani and Lebanese origin with perinatal lethal Gaucher disease Compound heterozygous state leads to complete phenotypic spectrum of Gaucher disease