Second-tier testing of newborns with an abnormal screening result for Krabbe disease
Follow-up testing after an abnormal newborn screening result for Krabbe disease
This test is used as a second-tier newborn screen for Krabbe disease (galactocerebrosidase deficiency) and includes both psychosine measurement and DNA analysis for the 30-kb deletion.
Krabbe disease (globoid cell leukodystrophy) is an autosomal recessive lysosomal storage disorder caused by an enzyme deficiency of galactocerebrosidase (GALC).
Although Krabbe disease is clinically variable, the most common and severe form of the disorder is early infantile onset that presents with rapid neurological regression and results in early death.
Second-tier testing reduces the number of false-positive results reported out.
Elevations in psychosine or the presence of a homozygous 30 kilobase deletion of the GALC gene support a diagnosis of Krabbe disease.
If the patient has abnormal newborn screening result for Krabbe disease, immediate action should be taken. Refer to the appropriate ACMG Newborn Screening ACT Sheet.(1)
Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS)/Polymerase Chain Reaction with Gel Electrophoresis
B-Galactosidase Galactosylceramide, Leukocytes
Beta-Galactosidase Galactosylceramide, Leukocytes
Cerebroside B-Galactosidase, WBC
Cerebroside Beta-Galactosidase(WBC)
Galactocerebrosidase
Galactocerebrosidase Deficiency
Galactosylceramidase Deficiency
Galactosylceramide
GALC Deficiency
Globoid Cell Leukodystrophy
Krabbe Disease
Krabbe's Disease
If the patient has abnormal newborn screening result for Krabbe disease, immediate action should be taken. Refer to the appropriate ACMG Newborn Screening ACT Sheet.(1)
Whole blood
1. Birth weight (grams)
2. Time of birth (24-hour time)
3. Gestational age (weeks)
| Question ID | Description | Answers |
|---|---|---|
| BG704 | Birth Weight (grams, XXXX) | |
| BG705 | Time of Birth (24hr Time, XX:XX) | |
| BG706 | Gestational Age (weeks, XX.X) |
Supplies: Card-Blood Spot Collection (Filter Paper) (T493)
Container/Tube:
Preferred: Blood Spot Collection Card
Acceptable: PerkinElmer 226 filter paper, Munktell filter paper, Whatman Protein Saver 903 paper, local newborn screening card, or blood collected in tubes containing heparin or EDTA and dried on filter paper.
Specimen Volume: 3 Blood spots
Collection Instructions:
1. Completely fill at least 3 circles on the filter paper card (approximated 100-microliters blood per circle).
2. Let blood dry on filter paper at ambient temperature in a horizontal position for a minimum of 3 hours.
3. Do not expose specimen to heat or direct sunlight.
4. Do not stack wet specimens.
5. Keep specimen dry.
Additional Information:
1. For collection instructions, see Blood Spot Collection Instructions.
2. For collection instructions in Spanish, see Blood Spot Collection Card-Spanish Instructions (T777).
3. For collection instructions in Chinese, see Blood Spot Collection Card-Chinese Instructions (T800).
1. New York Clients-Informed consent is required. Document on the request form or electronic order that a copy is on file. The following documents are available:
-Informed Consent for Genetic Testing (T576)
-Informed Consent for Genetic Testing-Spanish (T826)
2. Biochemical Genetics Patient Information (T602).
3. If not ordering electronically, complete, print, and send a Biochemical Genetics Test Request (T798) with the specimen.
2 Blood spots
| Blood spot specimen that shows serum rings or has multiple layers/applications | Reject |
| Insufficient specimen | Reject |
| Specimen Type | Temperature | Time | Special Container |
|---|---|---|---|
| Whole blood | Ambient (preferred) | 96 days | FILTER PAPER |
| Frozen | 96 days | FILTER PAPER | |
| Refrigerated | 96 days | FILTER PAPER |
Second-tier testing of newborns with an abnormal screening result for Krabbe disease
Follow-up testing after an abnormal newborn screening result for Krabbe disease
This test is used as a second-tier newborn screen for Krabbe disease (galactocerebrosidase deficiency) and includes both psychosine measurement and DNA analysis for the 30-kb deletion.
If the patient has abnormal newborn screening result for Krabbe disease, immediate action should be taken. Refer to the appropriate ACMG Newborn Screening ACT Sheet.(1)
Krabbe disease (globoid cell leukodystrophy) is an autosomal recessive disorder caused by a deficiency of galactocerebrosidase (GALC) leading to an accumulation of galactosylceramide and severe demyelination throughout the brain. Krabbe disease is primarily caused by variants in the GALC gene, and it has an estimated frequency of 1 in 100,000 births.
The clinical course of Krabbe disease can be variable, even within the same family. Eighty-five percent to 90% of patients present before the first year of life with central nervous system impairment, including increasing irritability, developmental delay, and sensitivity to stimuli. Rapid neurodegeneration, including white matter disease follows, with death usually occurring by 2 years of age. Late onset forms of the disease affect 10% to15% of individuals and are characterized by ataxia, vision loss, weakness, and psychomotor regression, typically presenting from age 6 months to the seventh decade of life.
Newborn screening for Krabbe disease has been implemented in some states. The early (presymptomatic) identification and subsequent testing of infants at risk for Krabbe disease may be helpful in reducing the morbidity and mortality associated with this disease. While treatment is mostly supportive, hematopoietic stem cell transplantation has shown some success if performed prior to onset of neurologic damage.
Newborn screening can typically identify patients with Krabbe disease, even before onset of symptoms, as well as unaffected patients with GALC pseudodeficiency alleles. For these reasons, second-tier testing that includes both psychosine and 30-kilobase (kb) deletion analyses has been developed. Second-tier testing reduces the number of false-positive results and limits the identification of affected individuals to patients needing immediate follow-up.
Psychosine is one of 4 substrates degraded by GALC and is a neurotoxin at elevated concentrations.. It has been shown to be elevated in patients with active disease and, therefore, may be a useful biomarker for the presence of disease or disease progression.
The common 30-kb deletion spanning intron 10 through the end of the gene accounts for a significant proportion of disease alleles that contribute to infantile Krabbe disease. While enzyme activity alone is not predictive of age of onset, there are known genotype-phenotype correlations. Individuals who are homozygous for the deletion or compound heterozygous for the deletion and a second GALC genetic variant (with the exception of late-onset genetic variants) are predicted to have infantile Krabbe disease.
Although rare, a few infants with an early onset Krabbe disease phenotype due to deficiency of saposin A (SAP-A) have been identified. SAP-A is a sphingolipid activator protein that assists galactocerebrosidase in its action on galactosylceramide.
An interpretive report will be provided.
An interpretive report will be provided.
An elevation of psychosine is indicative of symptomatic Krabbe disease.
The presence of a homozygous 30-kilobase deletion is indicative of early onset Krabbe disease.
The absence of the 30-kilobase deletion in the GALC gene does not eliminate the possibility of positive-carrier status or the diagnosis of Krabbe disease. This assay does not include DNA sequencing of the GALC gene.
A Krabbe disease phenotype can also be caused by the absence of a physiologically active sphingolipid activator protein, saposin A.
Psychosine levels may be normal in patients who are not yet symptomatic or have late onset Krabbe disease.
1. ACMG Newborn Screening ACT Sheets. Accessed August 30, 2023. Available at www.acmg.net/ACMG/Medical-Genetics-Practice-Resources/ACT_Sheets_and_Algorithms/ACMG/Medical-Genetics-Practice-Resources/ACT_Sheets_and_Algorithms.aspx?hkey=9d6bce5a-182e-42a6-84a5-b2d88240c508
2. Turgeon CT, Orsini JJ, Sanders KA, et al. Measurement of psychosine in dried blood spots-a possible improvement to newborn screening programs for Krabbe disease. J Inherit Metab Dis. 2015;38(5):923-929
3. Orsini J, Morrissey M, Slavin L, et al. Implementation of newborn screening for Krabbe disease: Population study and cutoff determination. Clin Biochem. 2009;42(9):877-884
4. Orsini JJ, Escolar ML, Wasserstein MP, et al. Krabbe disease. In: Adam MP, Mirzaa GM, Pagon RA, et al, eds. GeneReviews [Internet]. University of Washington, Seattle; 2000. Updated October 11, 2018. Accessed August 31, 2023. Available at www.ncbi.nlm.nih.gov/books/NBK1238/
5. Wenger DA, Escolar ML, Luzi P, Rafi MA. Krabbe disease (globoid cell leukodystrophy). In: Valle DL, Antonarakis S, Ballabio A, Beaudet AL, Mitchell GA. eds. The Online Metabolic and Molecular Bases of Inherited Disease. McGraw-Hill; 2019. Accessed August 31, 2023.Available at https://ommbid.mhmedical.com/content.aspx?bookid=2709§ionid=225546481
6. Guenzel AJ, Turgeon CT, Nickander KK, et al. The critical role of psychosine in screening, diagnosis, and monitoring of Krabbe disease. Genet Med. 2020;22(6):1108-1118. doi: 10.1038/s41436-020-0764-y
Protocol 1:
Internal standard is added to a dried blood spot. The extract is evaporated and reconstituted prior to injection onto a liquid chromatography tandem mass spectrometry (LC-MS/MS). Following separation of the structural isomers glucopsychosine and psychosine by liquid chromatography, their concentrations are measured by MS/MS analysis in the multiple reaction monitoring positive mode to follow the precursor to product species transitions for psychosine (PSY). The ratio of the extracted peak area of PSY to internal standard as determined by LC-MS/MS is used to calculate the concentration of PSY in the sample.(Unpublished Mayo method)
Protocol 2:
A polymerase chain reaction-based assay is used to examine DNA for the presence of a 30-kilobase deletion encompassing exon 11 through the end of the GALC gene.(Unpublished Mayo method)
Monday through Saturday
This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. It has not been cleared or approved by the US Food and Drug Administration.
82542
81401
| Test Id | Test Order Name | Order LOINC Value |
|---|---|---|
| KD2T | Krabbe Disease 2ND Tier NBS, BS | 62309-0 |
| Result Id | Test Result Name |
Result LOINC Value
Applies only to results expressed in units of measure originally reported by the performing laboratory. These values do not apply to results that are converted to other units of measure.
|
|---|---|---|
| 48536 | Interpretation | 62309-0 |
| 48535 | Reviewed By | 18771-6 |
| BG704 | Birth Weight (grams, XXXX) | 8339-4 |
| BG705 | Time of Birth (24hr Time, XX:XX) | 57715-5 |
| BG706 | Gestational Age (weeks, XX.X) | 76516-4 |