Confirmatory Testing Considerations

1. Confirmatory Testing Considerations Beth Vogel, MS, CGC Genetic Counselor Newborn screening program Wadsworth center New york state department of health

2. Overview • Implementation • Short-term follow-up • Case review meetings • Diagnostic testing

3. Implementation: Guidance • Partnership with clinicians • New York State SCID Consortium • 9 external group members • National recommendations • Newborn Screening Translational Research Network, CLSI Guidelines

4. Implementation: Guidance • Discussion points for NBS and clinician group • Minimum standard confirmatory tests • Short-term follow-up • Diagnostic categories • Transplants • Algorithm changes • Detection of non-SCID disorders

5. Implementation: Education • Target hospitals and primary care providers • TREC assay is not well known! • Developed educational materials • Letter to hospitals • Phone consultations • NBS and Specialty Care Centers WHAT IS A TREC?!

6. Short-term Follow-up Process • Eight Specialty Care Centers • Referral is called to primary care provider and specialty care center (SCC) • Diagnostic form completed by clinician

7. Case Review Meetings • Identify topics for Consortium conference calls • Unusual cases • Identify need for new diagnostic categories • Algorithm changes • Correlation of laboratory findings and clinical outcomes

8. Case Review Meetings • Maintain consistency in case closure • Incidence • Algorithm changes • Long-term follow-up

9. Diagnostic Categories • From NBSTRN website: • SCID • Leaky SCID/Omenn syndrome • Variant SCID • Syndromes with T cell impairment • Secondary T cell lymphopenia other than preterm alone • Preterm alone

10. SCID • Classic phenotype • <300 autologous T cells/μL • Emergent treatment required (transplant, gene therapy, enzyme replacement therapy)

11. Leaky SCID/Omenn Syndrome • Low T cell count • 3oo to 1500 T cells/μL • Requires treatment (transplant, gene therapy, enzyme replacement therapy) • Omenn syndrome • Erythrodermia, hepatosplenomegaly, eosinophilia, elevated IgE, restricted TCR diversity of T cells

12. Variant SCID • Moderately decreased T cell counts • May not require transplant

13. Syndromes with T Cell Impairment • DiGeorge syndrome/22q11.2 deletion syndrome • CHARGE syndrome • Jacobsen syndrome • RAC2 defect • DOCK8 deficiency • Ataxia telangiectasia • VACTERL association • Barth syndrome • TAR syndrome • Down syndrome • Ectrodactyly ectodermal dysplasia syndrome • Other

14. Secondary T Cell LymphopeniaOther Than Preterm Alone • Birth defects • Gastroschisis, intestinal lymphangiectasia, congenital heart defects, cardiac surgery +/- thymectomy • Metabolic disorder, degenerative neuromuscular disease

15. Preterm Alone • <37 weeks gestation • TREC copy number typically improves over time • If TRECs are undetectable, then full work-up warranted regardless of gestational age

16. Idiopathic T-cell Lymphopenia • Non-SCID • No congenital anomalies • Low T-cells that require clinical monitoring

17. Diagnostic Testing: Complete Blood Count • Lymphocytes • T cells = 70% of circulating lymphocytes • Proposed as a method to screen for SCID prior to current technologies • “ALC (absolute lymphocyte count) <2500/uL in early infancy requires further evaluation” • R. Buckley, MD

18. Diagnostic Testing:Flow Cytometry • Evaluation of lymphocyte subpopulations • Normal ranges vary by age

19. Diagnostic Testing:Flow Cytometry • CD = Cluster of differentiation

20. Diagnostic Testing: Flow Cytometry

21. Diagnostic Testing: Flow Cytometry

22. Diagnostic Testing:Lymphocyte Proliferation to Mitogens • Test of T-cell function • Less sensitive, but more specific test of T-cell function • Culture human peripheral blood mononuclear cells with plant lectin mitogens) • phytohemagglutinin (PHA), pokeweed mitogen (PWM), concanavalin (conA) • Response highest in newborns and decreases with age

23. Diagnostic Testing: Mitogens

24. Diagnostic Testing:Naïve and Memory T-cell Count • Not typically included as part of standard flow cytometry • CD45RA+ = Naive • CD45RO+ = Memory

25. Genetic Testing • Multiple genes • Mutation info often not needed before transplant • May be important if diagnosis is unclear • Important for genetic counseling • 45% of SCID cases are X-linked (IL2RG-related) • Remainder of the cases are recessive

26. Diagnostic Form

27. Diagnostic Form

28. References • Hicks MJ, Jones JK, Thies AC, et al: Age-related changes in mitogen-induced lymphocyte function from birth to old age. Am J ClinPathol 1983;80:159-163 • Baker M, Grossman W, Laessig R, et al. Development of a routine newborn screening protocol for severe combined immunodeficiency. J Allergy ClinImmunol. 2009; 124: 522-527 • Puck M, et al. Population-based newborn screening for severe combined immunodeficiency: Steps toward implementation. J Allergy ClinImmunol. 2007; 120 (4): 760-768. • Chan K, Puck J. Development of population-based newborn screening for severe combined immunodeficiency. J Allergy ClinImmunol. 2005; 115: 391-398. • Comeau A, Hale J, Pai S, et al. Guidelines for implementation of population-based newborn screening for severe combined immunodeficiency. J Inherit Metab Dis. 2010 • https://www.nbstrn.org/sites/default/files/SCID%20National%20Monthly%20March%202012.pdf • Buckley RH. The long quest for neonatal screening for severe combined immunodeficiency. Clinical Reviews in Allergy and Immunology. 2012 • Adeli MM and Buckley RH. Why newborn screening for severe combined immunodeficiency is essential: a case report. Pediatrics. 2010; 126(2):e465-e469. • Chase NM, Verbsky JW, Routes JM. Newborn screening for SCID: three years of experience. Ann. N.Y. Acad. Sci. 2011; 1238: 99-105. • Buckley RH, Transplantation of hematopoietic stem cells in human severe combined immunodeficiency: longterm outcomes. Immunol Res. 2011; 49: 25-43.

29. Thank you Questions?