Mos +21 in Newborn with 55% Blasts

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K H Ramesh

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Jul 29, 2020, 11:52:49 PM7/29/20
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Dear Colleagues:

Newborn with 55% blasts in PBL. Flow differential dx is Transient abnormal myelopoiesis (TMD) associated with Down Syndrome and/or Acute megakaryoblastic leukemia (AMKL).

Clinicians want us to r/o or confirm Trisomy 21 AND if +21 is limited to the Blasts only. As per Clinical Geneticist baby has absolutely no Down Syndrome features, and pediatric oncologists say baby has no organomegaly that would be associated with AMKL.

FISH RESULTS:

Interphase (only) form a Direct Harvest: 45% Trisomy 21

Interphase (only) from a 72 hour stimulated culture:  44% Trisomy 21

CHROMOSOME ANALYSIS (50 cell study):

UNSTIMULATED 24 AND 48 hour cultures: 95% Trisomy 21 ( no leukemia associated structural anomalies seen)

STIMULATED 72 hour Cultures: Only 5% Trisomy 21 and 95% Disomy 21
I am trying to understand the discrepancy between the stimulated and unstimulated chromosome analyses results.

I am also recommending FISH Study on a buccal swab specimen.  I have no way of letting the physicians know if the +21 is limited to the blasts cells based on the methodology used.

Like to know your thoughts on this .

Thank you all.

Best,

Ramesh

 

Dominiqu...@radboudumc.nl

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Jul 30, 2020, 6:20:37 AM7/30/20
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Hi Ramesh,

 

Interesting case!

The difference between unstimulated and stimulated blood cells is probably reflecting the difference between a very high B-cell content (blasts with a trisomy 21) in unstimulated cells and an increasing number of T-cells in the stimulated cultures since the phytohemagglutinin present in the medium typically  stimulates the T-lymphocytes to grow and not the B-cell population. For that reason you see much less trisomy 21 cells in metaphase cells because they come from dividing T-cells. So this emphasizes that the trisomy 21 is present only in B-cells and not in T-cells. To really exclude a congenital trisomy 21 the buccal swab FISH indeed is a good idea next to a skin biopsy.

Best ,

Dominique

Dr. D. Smeets, clinical laboratory geneticist
Theme Fertility and Pregnancy

Radboud University Medical Centre, Nijmegen

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K H Ramesh

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Jul 31, 2020, 1:26:11 AM7/31/20
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Hello:

Thank you. My understanding is that AMKL is a Myeloid Stem Cell-Megakaryocytic in origin and not B-cell lymphoid.

I will keep you all posted when I have the Buccal Swab FISH result.

Best,

Ramesh

 

From: cytogenetics-methods...@googlegroups.com <cytogenetics-methods...@googlegroups.com> On Behalf Of Dominiqu...@radboudumc.nl
Sent: Thursday, July 30, 2020 4:34 AM
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Subject: RE: Mos +21 in Newborn with 55% Blasts

 

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Pat Cammarata

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Jul 31, 2020, 1:26:40 AM7/31/20
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Good morning,

I have two thoughts:

1) If the trisomy 21 cells are in the myeloid cells, you can still have the myeloid blasts present in the stimulated culture. The T-cells are stimulated and will be more prevalent but the myeloid cells are still present. You can see dividing cells in a 72 hour culture in AML.

2) In the FISH slides, see if you can identify the lobulated nuclei of the myeloid cells and see if they are the trisomy 21 cells. Especially in the 72 hour stimulated culture.

Pat Cammarata

K H Ramesh

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Aug 4, 2020, 5:38:07 AM8/4/20
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HI Pat:

Thank you. I understand your points. The blasts have gone down from 55% to 30% now. Unable to find a Lab that offers Buccal Swab FISH Test to rule Tisomy 21. Mayo and Quest both said that they don’t offer this test.

Take care,

Ramesh

 

From: cytogenetics-methods...@googlegroups.com <cytogenetics-methods...@googlegroups.com> On Behalf Of Pat Cammarata
Sent: Thursday, July 30, 2020 9:12 AM
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Subject: RE: Mos +21 in Newborn with 55% Blasts

 

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Good morning,

Dominiqu...@radboudumc.nl

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Aug 4, 2020, 5:45:36 AM8/4/20
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It is also possible to perform a quick QF-pcr test on DNA isolated from a buccal swab. We do that all the time for prenatal DNA samples. If the trisomy 21 is not constitutional you will not trace it in the swab and the results are available within 24 hours.

Regards,

Dominique

K H Ramesh

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Aug 5, 2020, 1:01:38 AM8/5/20
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Thank you Dominique.

Ramesh

Sara Frias

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Aug 6, 2020, 7:22:41 AM8/6/20
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Hi Ramesh,
We have done FISH in bucal smears, here the methodology:
The buccal mucosa sample is received in a buffer solution of 0.01M trisHCl, 0.1M EDTA and 0.02M NaCl, pH7.0.
Centrifuge at 1500rpm for one minute to obtain a cell pack.
The slides are made and fixed with 80% methanol at 4 degrees C for 20 minutes.
Preheat the slides to 63 degrees C for 30 minutes.
Immerse the slides in a 300mg / mL pepsin solution at 37 degrees C, (test 10, 20 or 30 minutes), check in phase contrast that the cells are intact and without refringence.
Wash twice with PBS solution, then with water.
Fix in 4% paraformaldehyde for 20 minutes at 63 degrees C.
DO THE FISH.





Dra. Sara Frias Instituto Nacional de Pediatría México /Instituto de Investigaciones Biomédicas UNAM Insurgentes Sur 3700-C, CP 04530, Col. Insurgentes-Cuicuilco, Mexico D.F., México. Phone: Lab and FAX(5255)10845533, e-mail sara...@yahoo.com sara...@biomedicas.unam.mx


K H Ramesh

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Aug 6, 2020, 9:41:54 PM8/6/20
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Hi Sara:

Thank you.

Ramesh

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