Medical Policy

 

Subject: SensiGene® Fetal RhD Genotyping Test
Document #: GENE.00034 Publish Date:    03/29/2018
Status: Reviewed Last Review Date:    02/27/2018

Description/Scope

This document addresses the use of the SensiGene Fetal RhD Genotyping blood test (Sequenom, Inc., San Diego, CA) as a noninvasive diagnostic assessment tool to determine the fetal RhD genotype in an RhD negative mother. Pregnant women without the Rhesus (Rh) blood group, D antigen (RhD negative) may be at risk of producing antibodies if the fetus is RhD positive, a process known as alloimmunization. These antibodies can cross the placenta and cause hemolytic disease of the fetus and newborn, usually in subsequent pregnancies. If left undiagnosed and untreated, alloimmunization can result in serious fetal and neonatal morbidity and mortality.

Position Statement

Investigational and Not Medically Necessary:

Genetic testing to determine fetal RhD genotype using the SensiGene Fetal RhD Genotyping test is considered investigational and not medically necessary.

Rationale

Bombard (2011) conducted a prospective cohort study (n=236) to evaluate the feasibility and accuracy of the SensiGene test during the first and second trimesters of pregnancy in non-sensitized women. Of the 236 cases, 207 had sufficient deoxyribonucleic acid (DNA) extraction and a conclusive result. A blinded laboratory assessment of the SensiGene genotype assay was conducted. The first set of samples was obtained at 11 to 13 weeks of pregnancy (cohort 1) with documented fetal RhD serotype used as the reference standard. The second set of samples was obtained at 6 to 30 weeks of pregnancy using a laboratory reference standard provided by the manufacturer (blinded reference, cohort 2). In cohort 1, the SensiGene test correctly predicted the neonatal RhD phenotype in 201/207 samples, representing an accuracy of 97.1% (95% confidence interval [CI], 93.5-98.8). The sensitivity of the SensiGene test to accurately predict an RhD positive status was 97.2% (95% CI, 93.0-98.9). The corresponding specificity to accurately predict an RhD negative status was 96.9% (95% CI, 89.5-99.1). Finally, the positive predictive value (PPV) was 98.6% (95% CI, 94.9-99.6) and the negative predictive value (NPV) was 94.0% (5% CI, 85.6-97.6). In cohort 2, SensiGene demonstrated an accuracy of 99.5% when compared with the reference standard. Outcomes pertaining to analytical validity or clinical utility were not assessed. Study authors concluded that given the high rates of sensitivity, specificity, PPV and NPV using two different reference standards, the SensiGene test demonstrated high diagnostic accuracy to correctly identify the fetal RhD genotype in pregnant women. Despite these preliminary positive results, the study was hampered by a few limitations, including lack of blinding of participants and clinicians, and unclear methods of how laboratory personnel were blinded to study samples. Furthermore, there is a potential for funding or publication bias since this was a manufacturer supported study.

In a prospective cohort study, Tynan and colleagues (2011) evaluated the SensiGene test in pregnant, RhD negative women during the second trimester of pregnancy (n=150). Results of the SensiGene test were confirmed with serological and real-time polymerase chain reaction (PCR) testing of cord blood after birth. Of 150 samples, 148 (98.7%) were correctly classified, while 2 (1.3%) were determined to be inconclusive. A total of 86 (58.1%) and 62 (41.2%) were correctly identified as RhD positive and negative, respectively. One newborn was assessed as RhD positive by the SensiGene test and real-time PCR testing, but was serologically RhD negative. However, when serological testing was repeated at 18 months, the infant was confirmed as RhD positive. Limitations of this study include potential funding or publication bias due to manufacturer sponsorship and lack of blinded test assessments.

A prospective cohort study (Moise, 2013) evaluated fetal RhD status in RhD negative pregnant women who were not previously alloimmunized (n=123). The relative accuracy of the test was based upon gestational age and maternal blood samples were obtained during the first, second, and third trimesters of pregnancy. RhD status was initially confirmed by serological testing of neonatal cord blood. The overall accuracy of the SensiGene test to identify fetal RhD status was 99.1%, 99.1%, and 98.1% for the first, second, and third trimester, respectively. A total of 22 samples (6.3%) were determined to be inconclusive. The overall sensitivity of the SensiGene test was 99.6% (95% CI, 97.5%-100%). Of the 300 samples obtained, three false positives and one false negative were identified. Despite these preliminary promising results, the study was hampered by a few limitations, including lack of blinded assessors, participants and clinicians, and potential for conflicts of interest due to financial sponsorship by the manufacturer.

A meta-analysis conducted by Zhu and colleagues (2014) included 41 publications representing 11,129 samples from non-invasive Rh genotyping of cell-free fetal DNA (CffDNA) obtained from maternal blood. A total of 352 samples were excluded due to inconclusive results. The overall diagnostic accuracy from the remaining samples was 98.5%, and sensitivity and specificity were 99% and 98%, respectively. Diagnosis in the first trimester showed the highest accuracy at 99% and 30 studies reported 100% diagnostic accuracy of fetal RhD genotyping. It is not clear how many of the studies chosen for inclusion used the SensiGene test but based on this analysis, this non-invasive testing method warrants further investigation into its clinical utility and impact on perinatal outcomes.

Additional larger prospective (Moise, 2016) and retrospective (Vivanti, 2016) observational studies have been published with promising results on the sensitivity, specificity, PPV and NPV of the SensiGene. Although the test continues to demonstrate a high degree of accuracy both publications disclose conflicts of interest with the manufacturer of the test.

The American Congress of Obstetrics and Gynecology (ACOG), has not issued specific clinical practice guidelines, recommendations, or position statements regarding fetal RhD genotyping using the SensiGene Genotyping test.

The SensiGene RhD Genotyping test is not subject to federal regulation by the Food and Drug Administration (FDA). Genetic tests are regulated under the Clinical Laboratory Improvement Amendments (CLIA) Act of 1988. Premarket approval by the FDA is not required provided the test is performed in a laboratory facility that observes CLIA regulations.

In summary, the results from studies consistently demonstrate a high diagnostic accuracy for the SensiGene test in identifying fetal RhD genotype in RhD negative pregnant women. The sensitivity, specificity, PPV, and NPV are comparable to those derived from invasive diagnostic assessments using amniocentesis. However, all studies to date are either small or of low quality. In particular, the available individual studies were characterized by several weaknesses, including the potential for conflicts of interest due to manufacturer sponsorship or involvement, and the occurrence of test failures due to invalid or insufficient sampling. There was no direct evidence to allow definitive conclusions with conventional diagnostic assessments. Additionally, there is no evidence assessing the clinical utility of the SensiGene test, and it is unclear whether its use will lead to improved health outcomes.

Questions persist regarding the clinical utility of the SensiGene test in helping pregnant women avoid unnecessary anti-D immune globulin treatment or avoid adverse events associated with invasive diagnostic procedures. Also, despite the relatively low false-negative rate associated with the SensiGene test, a certain percentage of women will still become alloimmunized to an RhD positive fetus. Additional research is needed to ascertain the test’s relevance and usefulness in clinical practice.

Background/Overview

The Rh blood group system includes a variety of surface markers or antigens, the most common of which is RhD. In general, the presence of the RhD antigen on red blood cells is referred to as “RhD positive” and the absence as “RhD negative.” The prevalence of RhD negative blood type generally varies by race and ethnicity. Approximately 15% of Caucasians, 5% to 8% of African Americans, and 1% to 2% of Asians and Native Americans are RhD negative, respectively (ACOG, 2006).

Alloimmunization is a term that refers to the development of antibodies in an RhD negative mother if a sufficient number of red blood cells from an RhD positive fetus enter maternal circulation. Alloimmunization is often the result of fetomaternal hemorrhage during delivery, which has been reported in 15% to 50% of all births. Fetomaternal hemorrhage may be the result of miscarriage, pregnancy termination, trauma, or invasive procedures, such as amniocentesis. Alloimmunization against the RhD antigen is the most common cause of hemolytic disease of the fetus and newborn (HDFN) in subsequent pregnancies, symptoms of which can vary from mild to severe. If left undiagnosed and untreated, alloimmunization can result in serious perinatal morbidity and mortality. However, the introduction of postpartum administration of anti-D prophylaxis immune globulin to RhD negative women has significantly reduced the morbidity and mortality of HDFN over the last 40 years (ACOG, 2006; Daniels, 2009).

In cases of pregnant women without RhD hemolytic antibodies, knowledge that the fetus is RhD negative is useful in determining the need for prenatal and postnatal anti-D prophylaxis immune globulin. In the case of alloimmunized women with RhD antibodies, knowledge that the fetus is RhD negative may reduce need for intensive prenatal monitoring to predict and treat fetal anemia and continuously monitor high risk pregnancies. In addition, in alloimmunized women who require invasive prenatal testing for prenatal diagnosis of genetic abnormalities, knowledge of the fetal RhD genotype may be clinically useful when deciding whether to conduct first trimester chorionic villus sampling (CVS) or amniocentesis, both of which heighten the risk of worsening maternal sensitization and fetal HDFN (Daniels, 2009). It should be noted that even with the presence of RhD incompatibility between mother and fetus, about 50% of alloimmunized pregnancies could be identified with fetal RhD genotyping as not being at increased risk, which would reduce or eliminate the need for unnecessary examinations and treatment with anti-D prophylaxis immune globulin, while also reducing parental anxiety (Bombard, 2011).

The diagnosis of RhD alloimmunization is based upon detection of anti-RhD antibodies in maternal serum performed at the first prenatal visit. In RhD negative women with an initially negative result and uncomplicated pregnancy, this antibody screen should be repeated at 28 weeks of pregnancy, and again at delivery. The test most commonly used for diagnostic purposes is the indirect Coombs test, a technique used to determine antibody titers (antibodies in the plasma).

Alternatively, advances in fetal DNA testing have allowed the prediction of the fetal RhD phenotype. Conventional analysis of fetal DNA has relied upon invasive methods of sampling fetal tissues, such as amniocentesis and CVS, both of which are associated with a risk of fetal harm (Lo, 2012). Clinical guidelines published by the ACOG (2006) support the use of amniocentesis as the primary modality used to determine fetal blood type, while CVS is discouraged due to increased risk of fetomaternal hemorrhage.

Noninvasive alternatives to determine prenatal fetal RhD genotype are currently being investigated. One example is the SensiGene Genotyping test, a noninvasive prenatal blood test developed to determine the fetal RhD genotype in an RhD negative mother. The test is based upon current research demonstrating the presence of intact fetal cells and cell-free fetal nucleic acids (cffNA) that have crossed the placenta and enter maternal circulation (Tounta, 2011). The SensiGene test detects circulating cffDNA extracted from a mother’s blood sample during the first or second trimester of pregnancy, and uses a real-time polymerase chain reaction (PCR) to amplify the RhD genotype.

According to the manufacturer, the SensiGene Genotyping test can be performed in the first trimester at 10 weeks of gestation. The test is intended for use in pregnant RhD negative alloimmunized women whose partners are either RhD positive or have an unknown RhD status. It is also intended to identify fetal RhD status when maternal antibody titers are unclear.

Definitions

Clinical utility: Results of a diagnostic or prognostic test provide clinically relevant information about diagnosis, treatment, management, or prevention of a particular disease. As a result of the test, changes in medical management lead to clinically useful improvements in health outcomes.

Genotype: All or part of the genetic constitution of an individual or group, inherited from parents.

Isoimmunization: Development of specific antibodies as a result of antigen stimulation from the red blood cells (RBCs) of another species; synonym to alloimmunization.

Phenotype: The physically manifested properties of an organism, its physiology, morphology, and behavior.

Coding

The following codes for treatments and procedures applicable to this document are included below for informational purposes. Inclusion or exclusion of a procedure, diagnosis or device code(s) does not constitute or imply member coverage or provider reimbursement policy. Please refer to the member's contract benefits in effect at the time of service to determine coverage or non-coverage of these services as it applies to an individual member.

When services are Investigational and Not Medically Necessary:

CPT

 

81403

Molecular pathology procedure, Level 4 (eg, analysis of single exon by DNA sequence analysis, analysis of >10 amplicons using multiplex PCR in 2 or more independent reactions, mutation scanning or duplication/deletion variants of 2-5 exons) [when specified as the following]:

  • RHD (Rh blood group, D antigen) (eg, hemolytic disease of the fetus and newborn, Rh maternal/fetal compatibility), deletion analysis (eg, exons 4, 5 and 7, pseudogene), performed on cell free fetal DNA in maternal blood

 

 

ICD-10 Diagnosis

 

O36.0110-O36.0999

Maternal care for anti-D (Rh) antibodies

Z31.82

Encounter for Rh incompatibility status

References

Peer Reviewed Publications:

  1. Bombard AT, Akolekar R, Farkas DH, et al. Fetal RHD genotype detection from circulating cell-free fetal DNA in maternal plasma in non-sensitized RhD negative women. Prenat Diagn. 2011; 31(8):802-808.
  2. Daniels G, Finning K, Martin P, Massey E. Noninvasive prenatal diagnosis of fetal blood group phenotypes: current practice and future prospects. Prenat Diagn. 2009; 29(2):101-107.
  3. Lo YM, Chiu RW. Genomic analysis of fetal nucleic acids in maternal blood. Annu Rev Genomics Hum Genet. 2012; 13:285-306.
  4. Moise KJ Jr, Boring NH, O’Shaughnessy R, et al. Circulating cell-free fetal DNA for the detection of RHD status and sex using reflex fetal identifiers. Prenat Diagn. 2013; 33(1):95-101.
  5. Moise KJ Jr, Gandhi M, Boring NH, OʼShaughnessy R, et al. Circulating cell-free DNA to determine the fetal RHD status in all three trimesters of pregnancy. Obstet Gynecol. 2016; 128(6):1340-1346.
  6. Tounta G, Kolialexi A, Papantoniou N, et al. Non-invasive prenatal diagnosis using cell-free fetal nucleic acids in maternal plasma: progress overview beyond predictive and personalized diagnosis. EPMA J. 2011; 2(2):163-171.
  7. Tynan JA, Angkachatchai V, Ehrich M, et al. Multiplexed analysis of circulating cell-free fetal nucleic acids for noninvasive prenatal diagnostic RHD testing. Am J Obstet Gynecol. 2011; 204(3):251.e1-6.
  8. Vivanti A, Benachi A, Huchet FX, et al.Diagnostic accuracy of fetal rhesus D genotyping using cell-free fetal DNA during the first trimester of pregnancy. Am J Obstet Gynecol. 2016; 215(5): 606.e1-606.e5.
  9. Zhu YJ, Zheng YR, Li L, et al. Diagnostic accuracy of non-invasive fetal RhD genotyping using cell-free fetal DNA: a meta analysis. J Matern Fetal Neonatal Med. 2014; 27(18):1839-1844.

Government Agency, Medical Society, and Other Authoritative Publications:

  1. American Congress of Obstetrics and Gynecology (ACOG). ACOG Practice Bulletin. No. 4. Clinical Management Guidelines for Obstetrician-Gynecologists. Prevention of RhD alloimmunization. 1999; 66(1):63-70.
  2. American Congress of Obstetrics and Gynecology (ACOG). ACOG Practice Bulletin No. 75: Management of alloimmunization during pregnancy. Clinical Management Guidelines for Obstetrician-Gynecologists. 2006; 108(2):457-464.
  3. Royal College of Obstetrics and Gynecology (RCOG). Non-invasive prenatal testing for chromosomal abnormality using maternal plasma DNA (Scientific Impact Paper 15). Published March 04, 2014. Available at: http://www.rcog.org.uk/noninvasive-prenantal-diagnosis-using-cell-free-dna-maternal-blood. Accessed on February 05, 2018.
Index

Genotyping
RhD genotyping
Rhesus blood group
Rh factor
SensiGene

The use of specific product names is illustrative only. It is not intended to be a recommendation of one product over another, and is not intended to represent a complete listing of all products available.

Document History

Status

Date

Action

Reviewed

02/27/2018

Medical Policy & Technology Assessment Committee (MPTAC) review. Updated header language from “Current Effective Date” to “Publish Date.” Updated References section.

Reviewed

02/02/2017

MPTAC review. Updated Rationale, Background/Overview and References sections.

Reviewed

02/04/2016

MPTAC review. Updated Rationale, Background/Overview and Reference sections. Removed ICD-9 codes from Coding section.

Reviewed

02/05/2015

MPTAC review. Updated Description/Scope, Rationale and References sections.

 

10/01/2014

Updated Coding section with CPT Tier 2 genetic testing code changes.

New

02/13/2014

MPTAC review. Initial document development.