Accuracy of the determination of fetal RhD from maternal plasma DNA

• Grant number: 12/00401-2
• Support Opportunities: Regular Research Grants
• Start date: June 01, 2012
• End date: May 31, 2014
• Field of knowledge: Health Sciences - Medicine - Medical Clinics
• Principal Investigator: José Eduardo Levi
• Grantee: José Eduardo Levi
• Host Institution: Instituto de Medicina Tropical de São Paulo (IMT). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Associated researchers: Adolfo Wenjaw Liao ; Karen Nogueira Chinoca Ziza

Abstract

The discovery, in 1997 of cell free fetal DNA on the maternal plasma widened the possibilities of non-invasive prenatal diagnosis, since this genetic material of fetal origin may be obtained by safe and simple collection procedures. Qualitative analysis of these nucleic acids are routinely performed on those cases where it is possible to distinguish maternal from fetal DNA, i.e. genes that are previously known not to be present in the mother to be genome. The most spread among these tests is the one towards fetal gender determination, routinely offered worldwide, Brazil included. The presence of sequences from the Y chromosome in the maternal plasma indicates a male fetus pregnancy, since normal women would never bear such sequences in their plasma. Apart from the recreational use, this test has obvious applications on sex-linked inherited diseases such as hemophilia and congenital adrenal hyperplasia.Another relevant application, already routinely applied in the United Kingdom, France, Holland and other European countries is the RhD fetal determination, of importance for mothers RhD negative due to the well known consequences of maternal alloimunization by the fetal red cells carrying the RhD antigen, the Hemolytic Disease of the Fetus and the Newborn (HDFN).Like the fetal gender model, RhD negative mothers have a complete deletion of the RhD gene, thus, when sequences from it are found in the maternal plasma it indicates that the fetus is RhD positive. This antenatal diagnosis allows an early intervention, when needed, and saves doses of anti-D immunoglobulin. It also spares the mothers to be from serial collections of blood for antibody monitoring plus ultrasounds and clinical visits, and the anxiety involved on that. The successful use of RhD antenatal genotyping stems from the fact that is a non-invasive risk free method, easily performed. In Brazil, it has not been applied systematically in part due to the mixed ethnic background of our population, in whom, alleles that contain part of the RhD gene are found on phenotypically RhD negative individuals. In the proposed study we aim to verify the accuracy of a Real Time PCR method that overcomes the problems arising from the rare alleles above mentioned, in our population. We will recruit RhD negative pregnant women and compare the results of the molecular test against the phenotype on cord blood or after birth, obtaining sensitivity and specificity values. We hope to achieve a method of nearly 100% accuracy and provide directions to its routine application on clinical practice. (AU)