Slide seminar SW and GC 03.10.2024 to be shared.pdf

Pathology

1. Slide Seminar-Molecular techniques
Gaurav Chatterjee & Sweta Rajpal
Associate Professor, Hematopathology laboratory
Tata Memorial Centre, Mumbai

2. •65 year-old male with seropositivity presented with neuropathy, subcutaneous nodules, and easy bruising.
•CBC
•Hb-8.1gm/dl,
•TLC-12.62 x109/L
•PLT-1,00 x109/L
•SPEP : M band
•IFE: IgM kappa by immunofixation.
Case 1

6. •What is your diagnosis?
•What next?
Case 1

7. Allele specific PCR for MYD88L265P
MYD88
FwprimerWT
Reverse
primer
Mutant
Reverse
primer

8. Case 4 MYD88 p.L265P -WILD TYPE

9. Case 4 MYD88 p.L265P Mutant type

10. What is your interpretation of the PCR?
Case 1

11. Case 2
•57/F
•c/o low grade fever on and off since 2 months
•Splenomegaly + (5 cm palpable)
•CBC: Hb -8.6 g/dL, TLC -35000/cumm, Platelets -56,000/cumm
•Differential count:
Blasts -15%
Neutrophils -29%
Lymphocytes -13%
Monocytes -42%
Eosinophils -01%

12. Bone marrow aspirate
Blasts 12%
Myelocytes 15%
Metamyelocytes08%
Neutrophils16%
Lymphocytes03%
Monocytes 10%
Erythroid32%

13. Case 2
What is your diagnosis?
What next?

14. Capillary electrophoresis NPM1& FLT3-ITD
4 bp insertion in
NPM1 exon 11
40 bp insertion in FLT3
Allelic ratio-20633/3864= 5.3
HIGH ALLELIC RATIO
Case 2

15. NPM1 & FLT3–BIOLOGICAL NEGATIVECase 2

16. Case 2
What is your diagnosis?
What next?

17. CASE 3
Mr PA, 55Y/M
Presented with fever and generalized weakness for 6 months
Recurrent transfusions for last 4 months
CBC
Hb 7.8 TLC 2.1 Plat 56

22. Chromosome07X0201
Gene EZH2SMC1AIDH1NRAS
Genomic
coordinate14850747653441941209113113115258747
Exon NM_004456:ex
on17
NM_006306:exo
n2
NM_005896:exon4NM_002524:exon2
Nucleotide
Change
C>TC>TG>AC>T
Codon Changec.G1978Ac.G287Ac.C394Tc.G35A
Protein
Change
p.G660Rp.R96Hp.R132Cp.G12D
VAF49.13%48.42%46.77%38.39%
Type of
Mutation
Non
synonymous
SNV
Non
synonymous
SNV
Non synonymous
SNV
Non synonymous
SNV
AMP Tier1212
Coverage849849849849
Next Generation
Targeted DNA
Sequencing
What is your
complete
diagnosis?

23. Case 4
•13-year-old boy. Presented with fever, bone pain.
•Hb 7.7 g/dL
•TLC 17.6 X 109/L
•Plt27 X 109/L
•Manual differential count done on PB revealed 81% blasts.
•BMA done at TMH provided. Please comment.

26. Cytochemical MPO

28. B-ALL dx is established.
How will you work up?

29. FISH and/or PCR for
BCR::ABL1
KMT2A-rearrangements
ETV6::RUNX1
TCF3::PBX1
Ploidy determination:
FISH for trisomy 4,10,18
Conventional karyotyping
FCM-basedploidy detection
Common diagnostic approach to risk-stratifyin B-ALL
Diploid
Negative
Any further workup?

30. Case 4: NGS-based copy number alteration
Can you make a diagnosis?

31. CASE 5

32. CASE 5 NGS
What is your diagnosis?
GeneGenomic
variant
Protein variantTranscript
Variant
ExonCOSMIC
database
reference
Variant
allele
frequenc
y (%)
Tier
PTPN1
1
NC_000012.11
:g.112888163
G>T
NP_002825.3:p.(
Gly60Val)
NM_002834.
5:c.179G>T
3COSM1302
8
42.61
SH2B3NC_000012.11
:g.111885287
G>A
NP_005466.1:p.(
Arg392Gln)
NM_005475.
3:c.1175G>
A
6COSM5840
118
22.82