13yrs male with ISOLATED neck lateral lymphadenopathy for ~2 months without any signs of disease/B symptoms.  
Biopsy of the lateral neck node (macro photo).  
 
SONOSCOPY: 22,8x18,4x9,5mm node: hypoechoic, with definite capsule. There is no definite hillus vissible, general vascularisation not prominent.  
Visual picture of specific lymphadenopathy.  
 
HISTO: Some large GCs with tumor population (oval), some "infiltrated" with disrupted FDC network (elongated) and interstitial interfollicular spread (as EBV IM) and diffuse zones with disrupted FDC networks with highly proliferating CB like "blastoid" cells in starry sky back (BL like picture/GCB IH, except MYC(-) and Bcl2/Mum1+ het. Normal reactive FCs at periphery (with weak CD10 and Bcl2(-)).  
 
IH: CD19+; CD22+ (weak); CD20/CD79a+ (heterog); Bcl6/CD10/LMO2+; Bcl2/CD43(-/+); Ig kappa+; cMYC(-); TdT(-); EBER/HHV8(-); MUM1+/-(focal); IgM+; CD30/CD138/IgD(-); Ki67 index 90%.  
 
 
FISH:  
MYC (8q24) break: NO.  
IRF4/DUSP22(6p25.3) translocation: NO.  
MYC/IGH t(8;14)(q24;q32) translocation: NO.  
Bcl2 break: NO.  
11q aberations: NO.  
 
MOLECULAR:  
CONVENTIONAL PCR: Not clonal IGH, IGK ir IGL and TCRB, TCRG ir TCRD locuses.  
NGS:  
Case was referred for molecular IG clonality assessment.  
DNA was extracted using Maxwell-16 FFPE Tissue LEV DNA Purification Kitl and subjected for IGH clonality assessment using BIOMED-2 protocol and PCR fragment analysis on ABI 3500 sequence analyzer. Both IGH FR1 (Tube A) and FR2 (Tube B) reactions detected clonal peak in the background yet with inconsistent sizing – in the different sides of the expected range interval. Therefore, additional Next generation sequencing analysis of the IGH FR2 region was performed (Invivoscribe kit, MiSeq instrument).  
Data were analyzed with Arrest/Interrogate, Vidjil and IMGT V-Quest tools.  
NGS data analysis revealed unproductive IGHV3-33*03 / IGHD5-12*01 / IGHJ4*02 rearrangement with <90% sequence identity to the IGHV3-33*03 suggesting considerable modification of germline sequence by AID and hypermutation. This may also explain inconsistent BIOMED-2 clonality assessment results .  
NGS sequencing data was confirmed with Sanger sequencing.  
 
 
PROPOSAL: Pediatric DLBCL (GCB type)(non clonal; without known FISH abberations) NOS (with folliculothropism).  
 
Thank You.

tzankov 2020-07-01 15:57	Thank you Ugnius,     very difficult case and - indeed - everything that was needed has been done. Since all translocations have been falsified and 11q aberrations too (congratulations!), my working hypothesis would be an explosive follicular hyperplasia with folliculolysis mimicking DLBCL. Indeed if you closely look at the diffuse areas, they perfectly fit to the dark zones of follicles and they rather outgrow the FDC than disrupting them. I have seen such types of reactions in younger individuals, especially in immunocompromized ones (HIV, DiGeorge, hyper-IgM etc.) - meaning that an thorough check by an experienced pediatric immunologists may be indicated. I am curious to see teh NGS, though even a PTEN or an EZH2 "mutation" would fit with a svere reactive process... please let me know how the results look like.
ugnius 2020-07-01 16:02	Thanx. I feel really BADLY. Clinicians wants to start, all folk alerted. I've never seen positive 11q yet. And diffuse zones are VOLUMINOUS. I will append NGS, if be done. I will resign as hematopathologist... :(
tzankov 2020-07-01 17:59	the scanned slide is disabled on my PC, so my impressions are based on the static images and the written reflects my personal opinion, which must not be right. if there woudl be any issues, I can only recommend submitting the case to an experienced center: Prof. Fend e.g. or Prof. Klapper... but let's wait for the NGS result.
ugnius 2020-07-02 07:44	Add in: we can perform only more sensitive CLONALITY at the moment (NGS). My additional argues are: Ig kappa monotype, CD20 dim and totally effaced difusse (partialy) high Ki67 population (without HHV8, EBER).
tzankov 2020-07-02 08:08	Dear Ugnius,     as I said my working hypothesis would be an explosive follicular hyperplasia with folliculolysis mimicking DLBCL and I would put all energy to verify/falsify this hypothesis.   1. How large was the LK?   2. What was the CT or US apperance of it?   3. Any idea if the capsule and/or the hilar structures have been effaced?   4. Late pre-plasmablastic cebtroblasts may be CD20dim...   5. Yes, go for a sensitive colanlity assay, yet - an explosive follicular hyperplasia may be be "clonal"/Light chian restricted.   6. Look for typcal mutations of pediatric lymphomas.
ugnius 2020-07-04 11:34	Thanx, Alex.     SOME notes: SONOSCOPY: 22,8x18,4x9,5mm node: hypoechoic, with definite capsule. There is no definite hillus vissible, general vascularisation not prominent.   Visual picture of specific lymphadenopathy.     DURATION for 2 months,     NGS clonality will be appended.   This is all I have (there is no accessible lymphoma panel on parafin NGS still: just trying PanCancer DNRseq RNRseq...).
tzankov 2020-07-05 12:51	"22,8x18,4x9,5mm node: hypoechoic, with definite capsule. There is no definite hillus vissible, general vascularisation not prominent.   Visual picture of specific lymphadenopathy." sounds like equivocal respecting malignat/benign;   back neck is rather typical for infectiosu agents...   well - "duration of 2 months" - depends on whether there has been any dynamics. Let's wait for the MGS clonality result.   I am sill not convinced of a malignant process, but as I said - this is may personal opninon
tzankov 2020-07-14 18:23	any news from the NGS analysis?
tzankov 2020-08-18 19:25	Dear Ugnius, are there any news, either molecular results or clinical dynamics of the above case? Please add any useful information! Thank you in advance.
ugnius 2020-09-11 14:22	Add ins (molecular):   Case was referred for molecular IG clonality assessment.   DNA was extracted using Maxwell-16 FFPE Tissue LEV DNA Purification Kitl and subjected for IGH clonality assessment using BIOMED-2 protocol and PCR fragment analysis on ABI 3500 sequence analyzer. Both IGH FR1 (Tube A) and FR2 (Tube B) reactions detected clonal peak in the background yet with inconsistent sizing – in the different sides of the expected range interval. Therefore, additional Next generation sequencing analysis of the IGH FR2 region was performed (Invivoscribe kit, MiSeq instrument).   Data were analyzed with Arrest/Interrogate, Vidjil and IMGT V-Quest tools.   NGS data analysis revealed unproductive IGHV3-33*03 / IGHD5-12*01 / IGHJ4*02 rearrangement with <90% sequence identity to the IGHV3-33*03 suggesting considerable modification of germline sequence by AID and hypermutation. This may also explain inconsistent BIOMED-2 clonality assessment results .   NGS sequencing data was confirmed with Sanger sequencing.
tzankov 2020-09-11 14:52	Dear Ugnius,     Thank you very much for this additional information.     With this in mind, I have to revise my opinion and suggest a descriptive diagnosis of atypical clonal lymphoproliferation, pediatric DLBCL possible.     Kind regards     Alex
ugnius 2020-09-17 10:26	Thanx, Alex. Never seen before.
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