LABRAD : Vol 44, Issue 3 - December 2018

LABRAD

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12-2018

LABRAD : Vol 44, Issue 3 - December 2018

Aga Khan University Hospital, Karachi

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Aga Khan University Hospital, Karachi, "LABRAD : Vol 44, Issue 3 - December 2018" (2018). LABRAD. Book 28.

DECEMBER 2018 VOL. 44, ISSUE 3

2

A Publication of the Departments of Pathology & Laboratory Medicine and Radiology

CD20 Negative B-Cell Lymphomas 3

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Autologous Bone Marrow Transplantation at AKU 11 4XDQWL¿FDWLRQRI0RQRFORQDO3URWHLQ03URWHLQ 

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Judicious Use of Immunohistochemistry & Ancillary  Techniques in Precise Categorization of Lymphoproliferative Disorders. _{))'*3(7&7,PDJLQJLQ/\PSKRPDV } December 2018 Volume 44, Issue 3 Editor Dr Natasha Ali Associate Editor Dr Lena Jafri Editorial Committee

Department of Pathology and Laboratory Medicine Dr Nasir Ud Din 'U-RYHULD)DURRTL 'U=DKUD+DVDQ Dr Anila Rashid Radiology Dr Naila Nadeem Dr Dawar Khan /DEUDG$GPLQLVWUDWLRQ2I¿FH )DUKDQD$UVKDG

Department of Pathology and Laboratory Medicine $JD.KDQ8QLYHUVLW\+RVSLWDO 6WDGLXP5RDG32%R[ .DUDFKL3DNLVWDQ 7HO )D[ KRVSLWDOVDNXHGX.DUDFKLFOLQLFDOODERUDWRULHV

What is new in updated WHO Lymphoma &ODVVL¿FDWLRQ" 1HZXSGDWHG:+2O\PSKRPD&ODVVL¿FDWLRQDQG :+2EOXHERRNRQµ+DHPDWRO\PSKRLG¶XSGDWHG WKHGLWLRQZDVSXEOLVKHGLQ6RPHRIWKH VDOLHQWIHDWXUHVRIWKLVXSGDWHGFODVVL¿FDWLRQ LQFOXGH(QWU\RIµ'RXEOH([SUHVVLYHFP\F  EFOH[SUHVVLRQE\,+&¶DQGµ'RXEOH+LWFP\F  EFOWUDQVORFDWLRQE\),6+3&5¶LQµ$JJUHVVLYH %FHOOO\PSKRPDV¶¶,QDGGLWLRQXVHRI,+&IRU VXEFODVVL¿FDWLRQRI'/%&/LQWRµ*HUPLQDO&HQWUH *&%¶ $FWLYDWHG%FHOO$%&¶¶W\SHVLVDOVR made necessary. This is important not only from prognostic point of view but also for therapeutic UHDVRQVDVQHZDQGPRUHVSHFL¿FWKHUDSLHVDUH EHFRPLQJDYDLODEOH2QHRIWKHSURYLVLRQDO FDWHJRU\IURPLHµ+LJKJUDGH%FHOO lymphoma with features intermediate between

'/%&/ %/¶LVGURSSHG)RUPDWXUH7FHOO O\PSKRPDVµ)ROOLFXODU7KHOSHUFHOOV7)+¶IRXQG LQJHUPLQDOFHQWHUV±UHODWHGJHQHVLJQDWXUHVOLNH LQµ$QJLRLPPXQREODVWLF7FHOOO\PSKRPD$,7/¶ DUHHPSKDVL]HGDORQJZLWK3'3'/H[SUHVVLRQ DVQHZWKHUDSHXWLFVWUDWHJLHVLPPXQHFKHFNSRLQW LQKLELWRUVDUHEHFRPLQJDYDLODEOHWRWDUJHWWKHVH PROHFXOHV&ODVVL¿FDWLRQRIµFODVVLF+RGJNLQ O\PSKRPDVF+/¶KDVUHPDLQHGODUJHO\XQFKDQJHG DQGPRUHHPSKDVLVLVJLYHQRQLWVPLPLFNHUVOLNH µ3ULPDU\0HGLDVWLQDO%FHOOO\PSKRPDV30%&¶ 3URYLVLRQDOFDWHJRU\µ*UH\]RQHO\PSKRPDZLWK RYHUODSSLQJIHDWXUHVRIF+/ '/%&/¶SUHVHQWLQJ DVPHGLDVWLQDOPDVVLVUHWDLQHG¶ 6KDKLG3HUYH] Professor +LVWRSDWKRORJ\

From the Editor’s Desk

CD20 Negative B-Cell Lymphomas

Dr Arsalan Ahmed

+LVWRSDWKRORJ\

7KHVLQJOHPRVWFRPPRQO\XVHGPDUNHUWRLGHQWLI\ B-cell lymphomas is CD20. Certain types of

B-lineage lymphomas are characteristically negative for CD20. These include ALK+ diffuse large B-cell O\PSKRPDSODVPDEODVWLFO\PSKRPD++9 primary effusion lymphoma and B lymphoblastic O\PSKRPDOHXNHPLD/DFNRI&'LQRQHRI WKHVHKLJKJUDGH%FHOOO\PSKRPDVPD\OHDG the pathologist to consider a diagnosis other than %FHOOO\PSKRPD,QDGGLWLRQVRPH&'%FHOO lymphomas in patients who have been treated with ULWX[LPDEDOVREHFRPH&'QHJDWLYH

ALK+ Diffuse Large B-Cell Lymphoma

$/.GLIIXVHODUJH%FHOOO\PSKRPD'/%&/LV DUDUHDJJUHVVLYHODUJHFHOOO\PSKRPDZLWKXQXVXDO LPPXQRSKHQRW\SLFIHDWXUHVLQFOXGLQJH[SUHVVLRQ RI$/.SURWHLQDVZHOODVODFNRI&',WDIIHFWV patients over a broad range but patients are mostly \RXQJPHQZLWKDPDOHWRIHPDOHUDWLRRI7KHUH

is no association with immunosuppression. Disease LVXVXDOO\ZLGHVSUHDGLQYROYLQJO\PSKQRGHVDQG DOVRLQYROYLQJH[WUDQRGDOVLWHV*,WUDFWVRIWWLVVXH ERQHHWF7KHPRVWFRPPRQSDWWHUQRILQYROYHPHQW LVDGLIIXVHSUROLIHUDWLRQRIQHRSODVWLFFHOOVDOWKRXJK LQO\PSKQRGHVWKHUHLVRIWHQDVWULNLQJVLQXVRLGDO SDWWHUQRUDFRPELQDWLRQRIVLQXVRLGDODQGGLIIXVH involvement.

The neoplastic cells have the morphology of LPPXQREODVWVDQGRUSODVPDEODVWVVRPHWLPHV ZLWKPDWXUDWLRQWRSODVPDF\WLFFHOOV)LJXUH ,QIUHTXHQWO\WXPRUFHOOVDUHSOHRPRUSKLFDQGPD\EH multinucleated. The usual immunophenotype is ALK+ JUDQXODUF\WRSODVPLFVWDLQLQJ3DQ%FHOOPDUNHUV DUHQHJDWLYH&'ZHDNO\&'080 (0$&'3HUIRULQ++9DQG(%(57KH LPPXQRJOREXOLQPRVWRIWHQH[SUHVVHGLV,J$ The cytogenetic abnormality most characteristic RI$/.'/%&/LVWSTUHVXOWLQJ

4

O\PSKRPDDUH+,9DGXOWVZLWKDPHGLDQDJHLQWKH ¿IWKGHFDGHDQGDVWULNLQJPDOHSUHSRQGHUDQFH PBL is typically composed of a monotonous proliferation of large cells with the appearance RILPPXQREODVWVSODVPDF\WRLGLPPXQREODVWVRU SODVPDEODVWVZLWKYHVLFXODUQXFOHLDQGSURPLQHQW QXFOHROL)LJXUH,QRWKHUFDVHVWKHUHLV

SODVPDF\WLFGLIIHUHQWLDWLRQZLWKDVXEVHWRIFHOOV showing maturation toward plasma cells. PBL LVXVXDOO\SRVLWLYHIRUPDUNHUVDVVRFLDWHGZLWK SODVPDF\WLFGLIIHUHQWLDWLRQLQFOXGLQJ&'&' DQG080,5)DQGQHJDWLYHIRUPDUNHUVRI%FHOO GLIIHUHQWLDWLRQLQFOXGLQJ&'&'DQG3$; &'DQG(0$DUHIUHTXHQWO\H[SUHVVHG0<&LV H[SUHVVHGLQFDVHV,QVLWXK\EULGL]DWLRQIRU (%(5GHPRQVWUDWHV(%9LQDSSUR[LPDWHO\WZR WKLUGVRIDOOFDVHVDQGLQQHDUO\DOO+,93%/V +,9QHJDWLYHSDWLHQWVDSSHDUVRPHZKDWPRUHOLNHO\ WRKDYH(%9QHJDWLYH3%/++9LVQHJDWLYH,Q the majority of cases patients present with advanced stage disease. PBL is an aggressive lymphoma ZLWKDSRRUSURJQRVLVPHGLDQVXUYLYDO\HDULQ PXOWLSOHVWXGLHV+,9DVVRFLDWHG3%/DSSHDUVWR KDYHDZRUVHRXWFRPHWKDQ+,9DVVRFLDWHG'/%&/ DQG%XUNLWWO\PSKRPD0<&WUDQVORFDWLRQKDVEHHQ associated with worse outcome.

HHV8+ Primary Effusion Lymphoma

3ULPDU\HIIXVLRQO\PSKRPD3(/LVDUDUH large B-cell lymphoma characterized by O\PSKRPDWRXVHIIXVLRQVLQYROYLQJSOHXUDO pericardial or peritoneal cavities unaccompanied by a solid mass. It is universally associated ZLWKKXPDQKHUSHVYLUXV++93(/DIIHFWV in a translocation involving the ALK gene on

chromosome 2 and clathrin gene on chromosome 17. 7KHSUHVHQFHRIWKHWOHDGVWRDIXVLRQSURWHLQ that shows a granular cytoplasmic pattern of staining with antibody to ALK.

ALK+ DLBCL is associated with poor prognosis DQGSDWLHQWVXVXDOO\SUHVHQWZLWKDGYDQFHGVWDJH,,, ,9ZLWKRYHUDOOPHGLDQVXUYLYDORIPRQWKV7KHVH tumors are usually negative of CD20 antigen and DUHWKXVLQVHQVLWLYHWR5LWX[LPDE%HWWHUWKHUDSHXWLF strategies are required for ALK+ DLBCL.

&UL]RWLQLEDQLQKLELWRURI$/.DQGW\URVLQHNLQDVH KDVVKRZQWKHUDSHXWLFHI¿FDF\LQVRPHFDVHVRI ALK+ DLBCL.

Plasmablastic Lymphoma

3ODVPDEODVWLFO\PSKRPD3%/LVDUDUHKLJK grade large B-cell lymphoma with distinctive clinical and pathological features including strong DVVRFLDWLRQZLWKLPPXQRGH¿FLHQF\SODVPDFHOO immunophenotype and frequent association with 0<&UHDUUDQJHPHQWDQG(%9

$OWKRXJK3%/ZDV¿UVWLGHQWL¿HGLQWKHRUDOFDYLW\ LWKDVVLQFHEHHQIRXQGLQRWKHUH[WUDQRGDOVLWHV LQFOXGLQJSDUDQDVDOVLQXVHVJDVWURLQWHVWLQDOWUDFW ERQHOLYHUDQGVSOHHQWHVWHVVNLQDQGVRIWWLVVXH and less often in lymph nodes. Although a majority RIFDVHVRFFXULQWKHVHWWLQJRI+,9LQIHFWLRQ3%/ FDQDOVRRFFXULQ+,9QHJDWLYHLQGLYLGXDOVPRVW frequently in the setting of immunosuppression for organ transplantation or in elderly patients. PBL affects patients from early childhood to advanced DJHKRZHYHUPRVWSDWLHQWVZLWKWKLVW\SHRI

Figure 1: ALK+ DLBCL. The neoplasm is arranged in sheets and consist of large cells with vesicular nuclei, prominent cherry red nucleoli and abundant eosinophilic cytoplasm. Inset: The neoplastic cells show granular cytoplasmic positivity for ALK immunostain.

Figure 2. Plasmablastic Lymphoma. The neoplastic cells exhibit immunoblastic and plasmablastic morphology with large vesicular nuclei and prominent nucleoli.

young and middle-aged adults with males much more often affected than females. Nearly all SDWLHQWVDUH+,9SRVLWLYHWKH\SUHVHQWODWH LQWKHFRXUVHRI+,9LQIHFWLRQDQGDUHRIWHQ profoundly immunodeficient at the time they SUHVHQWZLWKO\PSKRPD2QO\RFFDVLRQDO SDWLHQWVDUH+,9QHJDWLYHWKH\DUHRIWHQROGHU DGXOWVIURP++9HQGHPLFDUHDV3(/KDVD YHU\SRRUSURJQRVLVDOWKRXJKDPRQJ+,9 SDWLHQWVWKHRXWFRPHPD\EHEHWWHUIRUWKRVH UHFHLYLQJDQWLUHWURYLUDOWKHUDS\$57'HDWK LVGXHWRO\PSKRPDVRPHWLPHVFRPSOLFDWHGE\ RSSRUWXQLVWLFLQIHFWLRQRU.DSRVL¶VVDUFRPD7KH neoplastic cells may be large plasmablastic and LPPXQREODVWOLNHRUSOHRPRUSKLFZLWKDQDSODVWLF morphology. Binucleated or multinucleated IRUPVPD\EHSUHVHQW6RPHQHRSODVWLFFHOOV PD\UHVHPEOH5HHG6WHUQEHUJFHOOV1HRSODVWLF FHOOVH[SUHVV&'&'DQG080,5) DQGW\SLFDOO\ODFN&'3$;&'D%&/ DQGLPPXQRJOREXOLQ&'LVRIWHQH[SUHVVHG $EHUUDQWH[SUHVVLRQRI7FHOODVVRFLDWHGDQWLJHQV LVUHODWLYHO\FRPPRQ7KH++9WXPRUFHOOV DUHXVXDOO\FRLQIHFWHGE\(%9 &DVHVRI++9O\PSKRPDZLWKPRUSKRORJLF immunophenotypic and genetic features similar to WKRVHRI3(/EXWSURGXFLQJVROLGWXPRUOHVLRQVLQ O\PSKQRGHVRULQH[WUDQRGDOVLWHVKDYHDOVREHHQ GHVFULEHG7KHVHKDYHEHHQFDOOHGH[WUDFDYLWDU\ 3(/&RPSDUHGWRFODVVLF3(/H[WUDFDYLWDU\ ++9DVVRFLDWHGODUJH%FHOOO\PSKRPDDSSHDUV VOLJKWO\PRUHOLNHO\WRH[SUHVVSDQ%FHOODQWLJHQV

such as CD20 and CD79a and also monoclonal immunoglobulin.

B-Cell Lymphomas Treated with Rituximab

Many patients with CD20+ B-cell lymphoma WUHDWHGZLWKULWX[LPDEXVXDOO\LQFRQMXQFWLRQZLWK FRPELQDWLRQFKHPRWKHUDS\DFKLHYHDVXVWDLQHG complete remission. If relapses occur or if the O\PSKRPDVSHUVLVWWKH\DUHXVXDOO\DOVR&' %FHOOO\PSKRPDVEXWRFFDVLRQDOO\UHFXUUHQWRU persistent disease shows loss or downregulation RI&',QPRVWFDVHVWKH&'QHJDWLYH lymphoma is of the same histologic type as the RULJLQDOO\PSKRPD,QVRPHLQVWDQFHVKRZHYHUD CD20+ low-grade B-cell lymphoma treated with ULWX[LPDEFDQSURJUHVVWRD&'QHJDWLYHGLIIXVH ODUJH%FHOOO\PSKRPD7KHPHFKDQLVPIRUODFNRI &'LVQRWNQRZQLQDOOFDVHV)RUFDVHVWUHDWHG DVKRUWWLPHEHIRUHLWLVWKHRUHWLFDOO\SRVVLEOHWKDW recurrent or persistent CD20-negative lymphoma is GXHWRPDVNLQJRUUHPRYDORI&'PROHFXOHVE\ ULWX[LPDE,QRWKHUFDVHV&'QHJDWLYLW\KDVEHHQ DWWULEXWHGWRGHOHWLRQRIWKH&'JHQHPXWDWLRQV within the CD20 coding region or to epigenetic mechanisms. 7KHODFNRI&'PD\OHDGWRFRQVLGHUDWLRQWKDWWKH SDWLHQWKDVGHYHORSHGDVHFRQGXQUHODWHGQHRSODVP 7KHUHIRUHIDPLOLDULW\ZLWKWKLVSKHQRPHQRQ DORQJZLWKVWDLQLQJIRURWKHUPDUNHUVNQRZQWREH H[SUHVVHGE\WKHO\PSKRPDLQFOXGLQJRWKHUSDQ% FHOOPDUNHUVLVXVHIXOLQHVWDEOLVKLQJDGLDJQRVLV

4XDQWL¿FDWLRQRI0RQRFORQDO3URWHLQ

(M-Protein)

The M-protein or paraprotein is an abnormal immunoglobulin fragment that is produced in H[FHVVE\DQDEQRUPDOPRQRFORQDOSUROLIHUDWLRQ RISODVPDFHOOVW\SLFDOO\LQPXOWLSOHP\HORPD 000RVWSDWLHQWVZLWKXQWUHDWHG00KDYH high M-protein levels in their blood and urine. (DFK0SURWHLQFRQVLVWVRIWZRKHDY\FKDLQV ȖĮȝįRUܭDQGWZROLJKWFKDLQVțRUȜ although occasionally just light chains or heavy

FKDLQVDUHVHFUHWHGDQGUDUHO\QRQHDWDOO7KH initial laboratory evaluation of the monoclonal gammopathies includes serum and urine protein HOHFWURSKRUHVLV63(3DQG83(3UHVSHFWLYHO\ 2QFHDPRQRFORQDOJDPPRSDWK\KDVEHHQ LGHQWL¿HGWKHTXDQWL¿FDWLRQRIWKH0SURWHLQLV DJRRGVXUURJDWHPDUNHUIRUPRQLWRULQJWKH MM. 'U+DIVD0DMLGDQG'U,PUDQ6LGGLTXL Chemical Pathology



M-proteins on SPEP and UPEP Reports

$WWKHVHFWLRQRI&KHPLFDO3DWKRORJ\$JDURVH gel electrophoresis is performed for screening RI0SURWHLQDQGODWHULPPXQR¿[DWLRQ

electrophoresis is performed to determine the W\SHRILPPXQRJOREXOLQ7KHVHPLTXDQWL¿FDWLRQ of M-proteins is done by separating the proteins E\HOHFWURSKRUHVLVVWDLQLQJWKHJHOZLWKDG\H scanning the gel to determine the percent of various protein fractions and determining the serum total protein concentration by a separate assay to convert these percentages into a semi-TXDQWLWDWLYHFRQFHQWUDWLRQ)LJXUH7KHDEVROXWH TXDQWL¿FDWLRQRILPPXQRJOREXOLQVLVGRQH

E\QHSKHORPHWU\7KH,0:*¶V,QWHUQDWLRQDO 0\HORPD:RUNLQJ*URXSUHVSRQVHFULWHULDIRU PRQLWRULQJ00LVEDVHGPDLQO\RQPHDVXULQJ free light chain ratios and M-protein quantitation on protein electrophoresis.

Limitations of Immunoglobulin measurement

Guideline recommends to use the serum or XULQH0SURWHLQFRQFHQWUDWLRQZKHUHDYDLODEOH IRUPRQLWRULQJDVLWLVPRUHVSHFL¿FWKDQ DEVROXWHLPPXQRJOREXOLQTXDQWL¿FDWLRQGRQH E\QHSKORPHWU\DVLWLQFOXGHVPRQRFORQDODV well as polyclonal immunoglobulins. In case 0SURWHLQLVSUHVHQWLQEHWDUHJLRQWKHFKDQFHV of overestimating is high with nephelometric PHDVXUHPHQWDVWKHQRUPDOLPPXQRJOREXOLQV are often present in greater quantities than the 0SURWHLQLWVHOILQEHWDUHJLRQ$OVRWKHUHDUH FHUWDLQOLPLWDWLRQVRI0SURWHLQPHDVXUHPHQWWKDW DWDYHU\ORZFRQFHQWUDWLRQRI0SURWHLQV63(3 overestimates and underestimates at very high FRQFHQWUDWLRQVGHK\GUDWLRQHIIHFWRIJHO 7RFRQFOXGH0SURWHLQTXDQWL¿FDWLRQE\ HLWKHUPHWKRGUHPDLQVWKHFRUQHUVWRQHRI00 monitoring techniques. The two techniques for immunoglobulin concentration are complementary WRHDFKRWKHUKRZHYHULWLVLPSRUWDQWWRQRWH WKDWLQPRQLWRULQJ00SDWLHQWV0SURWHLQ FRQFHQWUDWLRQRQ63(3GHQVLWRPHWU\VKRXOG RQO\EHFRPSDUHGWR63(3DQGQHSKHORPHWU\ values should only be compared to nephelometry YDOXHVDVWKHVH¿JXUHVVKRXOGQRWEHXVHG

interchangeably.

Figure 1: M-Protein Concentration on SPEP.

6HUXPSURWHLQ(OHFWURSKRUHVLV % Fraction (g/dl) 60&  

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Electrophoresis (IFE)

'U6LEWDLQ$KPHG Chemical Pathology

Monoclonal gammopathies are characterized by SUHVHQFHRIDEQRUPDOSURWHLQVDQWLERGLHVLQWKH blood which are produced from plasma cells in the bone marrow. Thus the occurrence of monoclonal gammopathies ususally point towards a clonal H[SDQVLRQRISODVPDFHOOVRUPDWXUH%O\PSKRF\WHV 'LVHDVHVVXFKDVPXOWLSOHP\HORPD:DOGHQVWURP¶V PDFURJOREXOLQHPLDO\PSKRSUROLIHUDWLYHGLVHDVH SULPDU\V\VWHPLFDP\ORLGRVLVOLJKWFKDLQGHSRVLWLRQ GLVHDVHDVZHOODVWKHSUHPDOLJQDQWGLVRUGHUVRI smoldering myeloma and monoclonal gammopathy

RIXQGHWHUPLQHGVLJQL¿FDQFH0*86DUH

FODVVL¿HGXQGHUWKHEURDGHUKHDGLQJRIPRQRFORQDO gammopathies.

Patiens suspected of Monoclonal gammopathies usually undergo a serum protein electrophoresis 63(3ZKLFKUHYHDOVDFKDUDFWHULVWLFPRQRFORQDO EDQG0VSLNHPRUHRIWHQLQWKHJDPPDJOREXOLQ UHJLRQDQGOHVVIUHTXHQWO\LQWKHEHWDRUDOSKD UHJLRQV$EQRUPDOLWLHVLGHQWL¿HGRQ63(3VKRXOG be immunotyped to endorse and characterize the

PRQRFORQDOSURWHLQ,PPXQR¿[DWLRQHOHFWURSKRUHVLV ,)(LVWKHSURFHVVWKDWLVXWLOL]HGIRUWKHSXUSRVH of immunotyping of monoclonal proteins. This test is done in order to identify the monoclonal LPPXQRJOREXOLQKHDY\FKDLQJDPPDDOSKDPX GHOWDRUHSVLORQDQGRUOLJKWFKDLQW\SHNDSSDRU ODPEGD

A review of guidelines from multiple societies UHFRPPHQGHGWKDW63(3LQFRQMXQFWLRQZLWK,)( should be used as a screening panel. Keeping the IDFWLQPLQGWKDW,)(LVPRUHVHQVLWLYHWKDQ63(3LQ addition to its recommendation as part of the initial screening practice it is also used by most clinicians for following up response to therapy.

7KLVWHFKQLTXHLVXVHGIRUWKHLGHQWL¿FDWLRQRI SURWHLQVZLWKLQFRPSOH[PL[WXUHVDIWHUVHSDUDWLRQ by either conventional zone electrophoresis or isoelectric focusing. Most commonly antigens ZKLFKDUHRIWHQLPPXQRJOREXOLQVDUHVHSDUDWHG by electrophoresis followed by precipitation with VSHFL¿FDQWLERGLHVLQVLWX,PPXQR¿[DWLRQDVWKH QDPHGH¿QHVFRQVLVWVRIDQHOHFWURSKRUHVLVSKDVH

DQGD¿[DWLRQSKDVH,WLVDWHFKQLTXHWKDWLQYROYHV anchoring a protein in situ after electrophoresis. To EHJLQZLWKWKH¿UVWVWHSLVDJDURVHJHOHOHFWURSKRUHVLV LQRUGHUWRVHSDUDWHWKHSURWHLQV6XEVHTXHQWO\ DQWLVHUXPLVVSUHDGRQWRWKHJHOVXUIDFHLWGLVSHUVHV LQWRWKHJHODQGUHDFWVZLWKWKHDQWLJHQ7KH¿UVW SDWWHUQLVWUHDWHGZLWKWKH¿[DWLYHVROXWLRQWKH second instead with a pentavalent antiserum FRQWDLQLQJLPPXQRJOREXOLQV,J*,J$,J0DQG DQWL.DSSDDQG/DPEGDFKDLQVIUHHDQGERXQG Then washing and unprecipitation of the gel is HQVXHGIXUWKHUPRUHVROXEOHSURWHLQVDUHUHPRYHG NHHSLQJWKHDQWLJHQDQWLERG\FRPSOH[FRQ¿QHG ZLWKLQWKHJHOPDWUL[)LQDOO\YLVXDOL]DWLRQRI SUHFLSLWDWHGSURWHLQFRPSOH[HVZLWKDSURWHLQVWDLQ LVSHUIRUPHG,)(FDQHLWKHUUHYHDODQRUPDOSDWWHUQ or identify a monoclonal protein or a polyclonal LPPXQRJOREXOLQSDWWHUQ)LJXUHVKRZVDQ,)( gel with the monoclonal protein as an IgG antibody SDUDSURWHLQZLWKDNDSSDOLJKWFKDLQDUURZ $QRUPDOSDWWHUQDVGHSLFWHGLQ¿JXUHVKRZVD GDUNHULPPXQRJOREXOLQ*,J*ODQHDOLJKWHU LPPXQRJOREXOLQ$,J$DQDEVHQWLPPXQRJOREXOLQ 0,J0DQGDGHQVHUNDSSDFRPSDUHGWRODPEGDODQH

Figure 1: IFE- showing IgG Kappa monoclonal gammopathy Figure 2: IFE-showing no evidence of monoclonal gammopathy

B-Cell Prolymphocytic Leukemia

Dr Anila Rashid

+DHPDWRORJ\ 7UDQVIXVLRQ0HGLFLQH

%FHOOSURO\PSKRF\WLFOHXNHPLD%3// is a very rare B cell neoplasm comprised of SURO\PSKRF\WHVW\SLFDOO\ZLWKLQYROYHPHQWRI WKHSHULSKHUDOEORRGERQHPDUURZDQGVSOHHQ %\GH¿QLWLRQWKHSURO\PSKRF\WHVFRPSULVHPRUH WKDQSHUFHQWRIWKHFHOOVLQWKHEORRGDQGERQH PDUURZ%3//LVDQH[WUHPHO\UDUHGLVHDVHOHVV WKDQRQHSHUFHQWRI%FHOOOHXNHPLDV%3// mainly affects older adults with a mean age at SUHVHQWDWLRQRI\HDUV0HQDQGZRPHQ appear to be equally affected. The majority of patients are Caucasian.



Patients typically present with a rapidly rising white EORRGFHOOFRXQWDQGPDVVLYHVSOHQRPHJDO\DQHPLD DQGWKURPERF\WRSHQLDDUHSUHVHQWLQDERXWSHU FHQWUHVSHFWLYHO\6\VWHPLF%V\PSWRPVLHIHYHUV QLJKWVZHDWVZHLJKWORVVDUHFRPPRQ

Peripheral blood and bone marrow morphology:

%\GH¿QLWLRQPRUHWKDQSHUFHQWRIWKH circulating cells in the peripheral blood are SURO\PSKRF\WHVPRUHW\SLFDOO\WKHSHUFHQWDJH of prolymphocytes is greater than 90 per cent. Peripheral blood prolymphocytes are medium-sized cells with moderately condensed chromatin DQGDVLQJOHSURPLQHQWYHVLFXODUQXFOHROXV 7KHQXFOHXVLVW\SLFDOO\URXQGRURYDODQGWKH

cytoplasm is usually moderately abundant and VOLJKWO\EDVRSKLOLF>¿JXUH@7KHERQHPDUURZLV LQ¿OWUDWHGLQDQLQWHUVWLWLDORUQRGXODUSDWWHUQE\ prolymphocytes. Immunophenotype:%3//W\SLFDOO\H[SUHVVEULJKW VXUIDFH,J0,J'EULJKWVXUIDFH,JNDSSDRU ODPEGDOLJKWFKDLQEULJKW&'DQG&'&' &'DDQG)0&7KLVLVLQFRQWUDVWWRFKURQLF O\PSKRF\WLFOHXNHPLDZKLFKJHQHUDOO\KDVGLP H[SUHVVLRQRIVXUIDFH,JDQG&'=$3DQG &'DUHH[SUHVVHGLQDERXWSHUFHQWRIFDVHV ZKLOH&'DQG&'DUHH[SUHVVHGLQDERXWRQH third of cases.

Differential diagnosis: The differential diagnosis of

B-PLL includes other chronic lymphoid neoplasms ZLWKDOHXNHPLFSUHVHQWDWLRQDQGLQFOXGHVWKH following:

- T- cell PLL

 &KURQLFO\PSKRF\WLFOHXNHPLD - Mantle cell lymphoma

 )ROOLFXODUO\PSKRPD - Lymphoplasmacytic lymphoma  +DLU\FHOOO\PSKRPD Prognosis:6XUYLYDOLVXVXDOO\WKUHHWR¿YH\HDUV GHVSLWHWKHUDS\3URJQRVWLFPDUNHUVKDYHEHHQ GLI¿FXOWWRGHWHUPLQHEXWDQHPLDWKURPERF\WRSHQLD DGYDQFHGDJHDQGWKHSUHVHQFHRI73PXWDWLRQV appear to predict a poor outcome.

Figure 1: B-cell prolymphocytic leukemia, showing monomorphic prolymphocytes with condensed chromatin, prominent nucleolus, and scanty basophilic cytoplasm.

3URWHLQHOHFWURSKRUHVLVDQGLPPXQR¿[DWLRQDUH commonly employed for the diagnosis and prognosis of certain hemato-lymphoid malignancies and are subject to a number of interferences that may affect SDWLHQW¶VGLDJQRVHV$QDO\WLFDOLQWHUIHUHQFHVDUHRI WZRW\SHVZKLFKPD\EHHQGRJHQRXVRUH[RJHQRXV

Endogenous interferences Fibrinogen

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in Serum Protein Electrophoresis and

,PPXQR¿[DWLRQ

'U)DU\DO+XVQDLQ Chemical Pathology )LEULQRJHQLVQRWXVXDOO\SUHVHQWLQVHUXP VSHFLPHQV+RZHYHULWPD\EHSUHVHQWLQVHUXP RISDWLHQWVZLWKFRDJXODWLRQGLVRUGHUVSDWLHQWV UHFHLYLQJDQWLFRDJXODWLRQWKHUDS\RUZKHQDSODVPD sample is wrongfully provided instead of a serum VDPSOH:KHQ63(LVSHUIRUPHGRQWKHVHVDPSOHV ¿EULQRJHQPLJUDWHVWRWKHEHWDJDPPDUHJLRQ and it may be misinterpreted as a monoclonal LPPXQRJOREXOLQ,)(IRUFRQ¿UPDWLRQLVXVXDOO\ the solution. Although not routinely performed

LQGLDJQRVWLFSUDFWLFH,)(ZLWKDQWL¿EULQRJHQ antibodies can help remove any interference caused E\¿EULQRJHQ

Protein electrophoresis of serum sample with ¿EULQRJHQ$$GGLWLRQDOEDQGRIDSSUR[LPDWHO\ J/LQGLFDWHGE\EOXHDUURZLVREVHUYHGZLWK63( ORFDWHGLQWKHȕȖIUDFWLRQ%,)(ZLWKDQWLȖĮȝ țDQGȜVHUDLOOXVWUDWHVWKDWWKHIRFDOEDQGLQWKHȕȖ UHJLRQLVQRWDPRQRFORQDOLPPXQRJOREXOLQ&,)( ZLWKDQDQWLERG\DJDLQVW¿EULQRJHQFRQ¿UPVWKDWWKH EDQGLVFDXVHGE\¿EULQRJHQLQWHUIHUHQFH7DNHQ IURP5HFRJQLWLRQDQGPDQDJHPHQWRIFRPPRQ UDUHDQGQRYHOVHUXPSURWHLQHOHFWURSKRUHVLV DQGLPPXQR¿[DWLRQLQWHUIHUHQFHV&KULVWRSKHU 50F&XGGHQD-RDQQHV&OLQLFDO%LRFKHPLVWU\9ROXPH -DQXDU\3DJHV Hemolysis

The main interference mechanisms are spectral interference from high concentrations of

KHPRJORELQVSHFWUDOLQWHUIHUHQFHOHDGVWRDORVVRI absorbance signal or a change in absorbance signal that is due to the interfering factor and not the VDPSOHLWVHOIDQGGLUHFWUHOHDVHRIDQDO\WHVIURP UHGEORRGFHOOVKHPRJORELQSRWDVVLXPPDJQHVLXP LURQSKRVSKDWHODFWDWHGHK\GURJHQDVHDQGDVSDUWDWH

DPLQRWUDQVIHUDVH,Q63(KHPRJORELQDQG

KHPRJORELQFRPSOH[HVVKRZXSDVGLVFUHWHEDQGVLQ the alpha-2 and beta regions.

+HPRO\]HG samples must EHLGHQWL¿HG and sampling should be repeated if YLDEOHUH sampling LVXQOLNHO\ to help in patients ZLWKGLI¿FXOWGUDZVVXFKDVWKRVHUHFHLYLQJ FKHPRWKHUDS\EXWLWLVDYLDEOHRSWLRQZKHUHEORRG FROOHFWLRQLVSHUIRUPHGLPSURSHUO\ (IIHFWRIKHPRO\VLVRQ63(5HGDUURZVGHQRWH KHPRJORELQKDSWRJORELQFRPSOH[HVFDXVHGE\ JURVVKHPRO\VLV7DNHQIURP5HFRJQLWLRQDQG PDQDJHPHQWRIFRPPRQUDUHDQGQRYHOVHUXP SURWHLQHOHFWURSKRUHVLVDQGLPPXQR¿[DWLRQ interferences-Christopher R.McCuddenaJoannes &OLQLFDO%LRFKHPLVWU\9ROXPH-DQXDU\ 3DJHV

HAAAs and Heterphile antibodies

+$$$VKXPDQDQWLDQLPDODQWLERGLHVDQG heterophile antibodies interfere with laboratory testing by binding to the antibody reagents used in assays. This can cause:

D)DOVHO\HOHYDWHGUHVXOWVEULGJLQJRIVDQGZLFK DQWLERGLHVLQDVLWHDVVD\ E)DOVHO\GHFUHDVHGUHVXOWVSUHYHQWLQJIRUPDWLRQRI EULGJHVWKHUHE\GHFUHDVLQJWKHPHDVXUDEOHVLJQDO )XQGDPHQWDOO\+$0$DQGKHWHURSKLOLFDQWLERGLHV GHWHFWDEOHE\,)(DPRXQWWRPRQRFORQDO JDPPRSDWKLHVRIXQGHWHUPLQHGVLJQL¿FDQFH 0*86,QWKHVHFDVHVWKHUHLVDUHDOPRQRFORQDO DQWLERG\EXWLWLVQRWDVVRFLDWHGZLWKFOLQLFDOO\ VLJQL¿FDQWGLVHDVHSODVPDFHOOG\VFUDVLD

Polyclonal increases in IgG4

,Q,J*UHODWHGGLVHDVH,J*5'WKHUHLV

D(OHYDWHGVHUXPFRQFHQWUDWLRQVRISRO\FORQDO,J* E7XPRUOLNHVZHOOLQJRIWKHLQYROYHGRUJDQV F$O\PSKRSODVPDF\WLFLQ¿OWUDWHHQULFKHGZLWK polyclonal IgG4-positive plasma cells

G$YDULDEOHGHJUHHRI¿EURVLV

The focal band detected by electrophoresis in sera IURPSDWLHQWVZLWK,J*5'FDQEHFRQ¿UPHG DVSRO\FORQDOE\LPPXQR¿[DWLRQ.DSSDODPEGD VNHZLQJFDQRFFXULQWKHSRO\FORQDO,J* A B T G A M K Ȝ T Fib C

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fraction of a minority of the IgG4-RD patients. )DFWRUVWKDWFRXOGVXSSRUWWKHLGHQWL¿FDWLRQRI WUXH,J*0SURWHLQVLQFOXGHDGLVFUHWH0VSLNH VWULFWO\DVVRFLDWHGZLWKRQO\RQHOLJKWFKDLQ and a suppressed concentration of polyclonal immunoglobulins in multiple myeloma patients. Isoelectric focusing can be helpful to differentiate between polyclonal and monoclonal.

(OHFWURSKRUHVLVSDWWHUQREVHUYHGLQVHUXPZLWK HOHYDWHGSRO\FORQDO,J*$6HUXPSURWHLQ electrophoresis of a patient with IgG4-RD with an ,J*VXEFODVVYDOXHRIPJG/UHIHUHQFHYDOXH ±PJG/7KHDUURZLQGLFDWHVDFKDUDFWHULVWLF IRFDOEDQGEULGJLQJWKHȕȖIUDFWLRQRIWKHVSHFWUXP %,PPXQR¿[DWLRQHOHFWURSKRUHVLVZLWKDQWLȖĮȝ țDQGȜVHUDLOOXVWUDWHVWKDWWKHIRFDOEDQGLQWKHȕȖ region consists mainly of IgG antibodies. Connecting

OLJKWFKDLQVLQWKLVSDWLHQWDUHPDGHXSRIERWKNDSSD and lambda which stresses the polyclonal nature RIWKH,J*DQWLERGLHV&,VRHOHFWULFIRFXVLQJFDQ be helpful to differentiate between polyclonal and monoclonal IgG. The polyclonal pattern of a normal FRQWUROODQHDQGDSDWLHQWZLWK,J*5'ODQH LVFRQWUDVWHGE\PRQRFORQDOSDWWHUQVREVHUYHGLQ SDWLHQWVZLWK,J*PRQRFORQDOJDPPRSDWKLHVODQHV

DQG$UURZKHDGVVKRZWKHPRQRFORQDOµIURQW¶ of the banding characteristic of IgG M-proteins RQLVRHOHFWULFIRFXVLQJ7DNHQIURP5HFRJQLWLRQ DQGPDQDJHPHQWRIFRPPRQUDUHDQGQRYHO VHUXPSURWHLQHOHFWURSKRUHVLVDQGLPPXQR¿[DWLRQ interferences-Christopher R.McCuddenaJoannes &OLQLFDO%LRFKHPLVWU\9ROXPH-DQXDU\ 3DJHV Exogenous Interferences Contrast dyes Capillary zone electrophopresis is based on an ultraviolet detection at 200 nm via WKHSHSWLGHERQGVUDGLR opaque agents absorbing at the same wavelength can be observed by 63(:KHQDEORRG sample is collected after performing a contrast dye injection imaging WHVWDQDGGLWLRQDO VSLNHDGLVWRUWLRQ RUPRGL¿FDWLRQ interference occurs in WKHĮJOREXOLQIUDFWLRQ or less frequently the ȕJOREXOLQIUDFWLRQ The result can usually EHFRQ¿UPHGZLWK,)(

A B C

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Antifungals and antibiotics

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,QWHUIHUHQFHRI)OXRURF\WRVLQHDWWKHHQGRIWKH HQGRIWKHȖJOREXOLQIUDFWLRQ%,PPXQR¿[DWLRQ shows that no monoclonal component can

H[SODLQWKHRFFXUUHQFHRIWKLVVPDOOVSLNH& &KHPLFDOVWUXFWXUHRI)OXRURF\WRVLQH7DNHQ IURP5HFRJQLWLRQDQGPDQDJHPHQWRIFRPPRQ UDUHDQGQRYHOVHUXPSURWHLQHOHFWURSKRUHVLV DQGLPPXQR¿[DWLRQLQWHUIHUHQFHV&KULVWRSKHU 50F&XGGHQD-RDQQHV&OLQLFDO%LRFKHPLVWU\9ROXPH -DQXDU\3DJHV Rare/Novel Interferences

Gelatin-based plasma substitutes

Gelatin-based plasma substitutes have been reported

IUDFWLRQVLPXODWLQJDPRQRFORQDOFRPSRQHQW6HYHUDO DQWLELRWLFVFDQDOVRSURGXFHDGGLWLRQDOVSLNHVLQ Capillary zone electrophoresis patterns:

 D &HIWULD[RQHFDQLQGXFHDVPDOOGLVWLQFWSHDN at the anodal site of the prealbumin fraction  E 6XOIDPHWKR[D]ROHFDQSURGXFHDVPDOOSHDNDW   WKHUDSHXWLFFRQFHQWUDWLRQVFORVHUWRWKH   DOEXPLQSHDNWKDQFHIWULD[RQH  F3LSHUDFLOOLQDVVRFLDWHGZLWKWD]REDFWDPLVDOVR   OLNHO\WRLQGXFHDQDGGLWLRQDOVSLNHEHWZHHQ   WKHĮWKHȕJOREXOLQIUDFWLRQ WRLQGXFHDQLQFUHDVHRIWKHȖJOREXOLQIUDFWLRQ ZLWKDSRO\FORQDOSDWWHUQVKLIWHGWRWKHȕJOREXOLQ fraction. Hydroxocobalamin +\GUR[\FREDODPLQLVJLYHQLQF\DQDLGHSRLVRQLQJ and although a rare cause of interference it can cause DQDGGLWLRQDOVSLNHZLWKLQWKHĮJOREXOLQDOEXPLQ fraction. Monoclonal therapies 5LWX[LPDEDQG%HYDFL]XPDELQWKHUDSHXWLFGRVHV FDQSURGXFHDYLVLEOH0SURWHLQRQ63(3,)( 6LOWX[LPDE'DUDWXPXPDEDQG(ORWX]XPDEDOO,J* NDSSDPRQRFORQDODQWLERGLHVFDQDOVRDSSHDUDVD VPDOO0SURWHLQPRVWRIWHQLQFRQFHQWUDWLRQVXSWR J/

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,QDQDXWRORJRXVERQHPDUURZWUDQVSODQWSDWLHQW¶V own stem cells are collected and stored in blood EDQN$IWHUWKLVSDWLHQWLVWUHDWHGZLWKKLJKGRVHV RIF\WRWR[LFFKHPRWKHUDS\ZKLFKGHVWUR\VWKH PDOLJQDQWFHOOV+RZHYHUWKHVHF\WRWR[LFDJHQWV DOVRNLOOQRUPDOKHPDWRSRLHWLFVWHPFHOOVOHDGLQJ WROLIHWKUHDWHQLQJP\HORVXSSUHVVLRQ7KHUHIRUH LQRUGHUWRDFKLHYHWKHQRUPDOKHPDWRSRLHVLVWKH collected hematopoietic stem cells are reinfused into the patient. These stem cells after “homing” into the marrow cavity start generating normal blood cells.

There are number of indications for autologous KHPDWRSRLHWLFVWHPFHOOWUDQVSODQWDWLRQKRZHYHU more than 90 per cent procedures are performed in PXOWLSOHP\HORPD+RGJNLQ¶VDQGQRQ+RGJNLQ¶V lymphoma. Multiple myeloma is an incurable PDOLJQDQF\KRZHYHUPHGLFDOOLWHUDWXUHUHYHDOVD VXUYLYDODGYDQWDJHRIPRQWKVLQWKLVFRKRUW RISDWLHQWV1RZDGD\VWKHPRVWFRPPRQLQGLFDWLRQ of autologous transplant is multiple myeloma. $SSUR[LPDWHO\SHUFHQWSDWLHQWVDUHFXUHGRI +RGJNLQ¶VO\PSKRPDZLWKFKHPRUDGLDWLRQWKHUDS\ )RUWKHUHPDLQLQJSHUFHQWFDVHVDXWRORJRXVERQH marrow transplant is a potentially curative treatment

Autologous Bone Marrow Transplantation at AKU

'U0RKDPPDG8VPDQ6KDLNK +DHPDWRORJ\ RSWLRQ6LPLODUO\GLIIXVHODUJH%FHOOO\PSKRPD LVWKHFRPPRQHVWQRQ+RGJNLQ¶VO\PSKRPDLQ WKHZRUOGDVZHOODVLQ3DNLVWDQ5LWX[LPDEZLWK &+23DQG&+23OLNHFKHPRWKHUDS\RIIHUVFXUHLQ DSSUR[LPDWHO\SHUFHQWRIWKHSDWLHQWSRSXODWLRQ ,QWKHUHPDLQLQJSHUFHQWDXWRORJRXVERQH marrow transplant is an attractive remedy of cure. :HHVWDEOLVKHGD%RQH0DUURZ7UDQVSODQWXQLWDW $.8+LQ,WRSHQHGDQHUDRIQHZKRSHQRW only for the people of our country but for the region as well. Initially it was a two bedded unit which

subsequently was doubled in capacity owing to increased number of patients. The Bone Marrow Transplant Unit is fully equipped according to international standards with all intensive care facilities and KLJKHI¿FLHQF\ particulate air +(3$¿OWHUV along with strict policy for infection FRQWURO:HDUH offering bone marrow transplant facility including autologous WUDQVSODQW7LOOGDWHERQHPDUURZWUDQVSODQW SURFHGXUHVKDYHEHHQGRQHRXWRIZKLFKDUH DXWRORJRXVPDUURZWUDQVSODQWVZLWK+RGJNLQ¶V O\PSKRPDQRQ+RGJNLQO\PSKRPDDQGPXOWLSOH myeloma constituting the major proportion. ,QFRQFOXVLRQKLJKGRVHFKHPRWKHUDS\IROORZHG by autologous stem cell transplant is an effective WUHDWPHQWRSWLRQLQSDWLHQWVZLWKUHODSVHG

refractory lymphoma and myeloma allowing further consolidation of response attained by salvage therapy.

)ROOLFXODU/\PSKRPD)/LVDFRPPRQ%FHOO lymphoma of germinal center origin with classically DVVRFLDWHGKLVWRORJLFLPPXQRSKHQRW\SLFDQGJHQHWLF IHDWXUHV,QWKHFXUUHQW:+2&ODVVL¿FDWLRQRI +HPDWRO\PSKRLGQHRSODVPVVRPHFKDQJHVKDYHEHHQ PDGHLQ)ROOLFXODUO\PSKRPDZKLFKDUHKLJKOLJKWHG LQWKLVDUWLFOH,QVLWXIROOLFXODUQHRSODVLD,6)1 IRUPHUO\FDOOHG)/LQVLWXGXRGHQDOW\SH)/DQG GLIIXVHYDULDQWRI)/KDYHEHHQUHFRJQL]HGDVQHZ RI¿FLDOYDULDQWVRI)/3HGLDWULF)ROOLFXODU/\PSKRPD IRUPHUO\DYDULDQWRI)/LVQRZDVHSDUDWHHQWLW\ LQ:+2&ODVVL¿FDWLRQDQGDUHQRZNQRZDV 3HGLDWULFW\SH)ROOLFXODU/\PSKRPD37)//DUJH %FHOOO\PSKRPDZLWK,5)UHDUUDQJHPHQWLVDQHZ provisional entity which helps to distinguish from 3HGLDWULFW\SH)ROOLFXODU/\PSKRPDDQGRWKHU'LIIXVH Large B-cell lymphoma.

,QVLWXIROOLFXODUQHRSODVLD,6)1LVGH¿QHGDV partial or total colonization by clonal B cells carrying WKH%&/WUDQVORFDWLRQFKDUDFWHULVWLFRI)/LQDQ RWKHUZLVHUHDFWLYHO\PSKQRGH,6)1KDYHDORZ rate of progression but are often associated with prior RUV\QFKURQRXVRYHUWO\PSKRPDVWKHUHIRUHQHHGLQJ additional clinical assessment. The neoplastic FHOOVRI,6)1H[SUHVV&'DQG%&/)LJXUH %VLPLODUWRFRQYHQWLRQDO)/,6)1VKRXOGEH distinguished from lymph nodes showing only partial LQYROYHPHQWE\)/3)/ZKLFKDUHPRUHOLNHO\WR SURJUHVV7KHPDOLJQDQWIROOLFOHVRI3)/DUHODUJHU

&KDQJHVLQ:+2&ODVVL¿FDWLRQRI

Hematolymphoid Neoplasms Related to

Follicular Lymphoma

Dr Arsalan Ahmed +LVWRSDWKRORJ\

than germinal centers of reactive lymph nodes which H[KLELW,6)1)LJXUHDDQGWKH\KDYHDWHQGHQF\WR LQYROYHRQHDUHDRIO\PSKQRGH8QOLNH,6)1WKH QHRSODVWLFSUROLIHUDWLRQLQ3)/FDQEHVHHQRXWVLGH WKHJHUPLQDOFHQWHU7KH,6)1DUHDOVRDVVRFLDWHG ZLWKWDQGGHOS3DWLHQWVZKRDUH LQFLGHQWDOO\GLDJQRVHGZLWK,6)1DQGKDYHQRRWKHU FOLQLFDOHYLGHQFHRI)/KDYHDYHU\ORZULVN SHUFHQWRIGHYHORSLQJVXEVHTXHQW)/ 'XRGHQDOW\SH)/LVDWUXO\ORFDOL]HGDQGLQGROHQW %O\PSKRSUROLIHUDWLYHGLVRUGHUVKDULQJPDQ\RI WKHKLVWRORJLFLPPXQRSKHQRW\SLFDQGF\WRJHQHWLF IHDWXUHVRIFRQYHQWLRQDOORZJUDGH)/7KLV O\PSKRPDSUHVHQWVDVPXOWLSOHVPDOOSRO\SV typically in the second part of the duodenum. 'XRGHQDOW\SH)/LVFRPSRVHGRIQHRSODVWLFIROOLFOHV LQWKHVXEPXFRVDVXEPXFRVDDQGWKHVHIROOLFOHVDUH FRPSRVHGSUHGRPLQDQWO\RIFHQWURF\WHV)LJXUH ZLWKDORZ.LSUROLIHUDWLRQLQGH[,WLVGLI¿FXOW to differentiate this entity from conventional nodal )/LQWKHDEVHQFHRIFRPSOHWHFOLQLFDOLQIRUPDWLRQ 7KHVHSDWLHQWVKDYHDQH[FHOOHQWSURJQRVLVDQGZDWFK and wait strategy is adopted for most patients.

'LIIXVH)/YDULDQWW\SLFDOO\SUHVHQWVDVDEXON\ localized mass in the inguinal region. The lymph

Figure 1: ISFN. (A) Benign appearing germinal center with preserved mantle zone. The neoplastic B-cells within germinal center show strong expression of BCL2 (B) as compared to faint staining in mantle zone.

14

Figure 3: Diffuse FL variant. Effacement of lymph node architecture by diffuse neoplastic proliferation of small mature appearing lymphocytes. No nodular architecture seen. The neoplastic cells show an immunophenotypic SUR¿OHVLPLODUWRFRQYHQWLRQDO)/

Figure 4: Pediatric type FL. The germinal center is populated

predominantly be blastoid follicular cells. Inset shows neoplastic cells are negative for BCL2 immunostain.

QRGHLVHIIDFHGE\GLIIXVHSUROLIHUDWLRQRIVPDOO PDWXULQJDSSHDULQJO\PSKRF\WHV)LJXUH7KH

immunophenotype is characteristic of conventional )/LQDGGLWLRQWKHQHRSODVWLFFHOOVH[SUHVV&' LPPXQRVWDLQ+RZHYHUXQOLNHFRQYHQWLRQDOORZ JUDGH)/WKLVYDULDQWLVQHJDWLYHIRUWKHW WUDQVORFDWLRQEXWVKRZVGHOHWLRQRIS'HOHWLRQ RISLVDOVRVHHQLQFRQYHQWLRQDO)/ZKHUHLWLV associated with adverse outcome.

3HGLDWULFW\SH)ROOLFXODU/\PSKRPD37)/ presents as high grade but low stage disease with DQH[FHOOHQWSURJQRVLV37)/LVORFDOL]HGLQYROYHV SULPDULO\O\PSKQRGHVRIKHDGDQGQHFNUHJLRQ DQGVKRZVPDUNHGPDOHSUHGRPLQDQFH7KHO\PSK node is effaced by pure follicular proliferation FKDUDFWHUL]HGE\ODUJHH[SDQVLOHKLJKO\SUROLIHUDWLYH follicles that contain prominent blastoid follicular center cells rather than classic centroblasts RUFHQWURF\WHV)LJXUH7KHFKDUDFWHULVWLF LPPXQRSKHQRW\SLFSUR¿OHRIWKLVO\PSKRPDVKRXOG

EHSRVLWLYHH[SUHVVLRQRI&'&'DQG%&/ DQGDUHQHJDWLYHIRU%&/7KH\ODFN%&/%&/ ,5)DQG0<&UHDUUDQJHPHQWV1HDUO\DOOFDVHVDUH localized and may not require treatment other than H[FLVLRQ7KHFULWHULDRISHGLDWULFW\SH)/KRZHYHU must be strictly applied to avoid underdiagnosing FRQYHQWLRQDOJUDGHWKUHH)/ZLWKSDUWLFXODUFDXWLRQ UHTXLUHGEHIRUHPDNLQJWKLVGLDJQRVLVLQDQDGXOW /DUJH%FHOOO\PSKRPD/%&/ZLWK,5) rearrangement occurs most commonly in children and young adults and is considered a distinct new SURYLVLRQDOHQWLW\LQ:+2&ODVVL¿FDWLRQ 7KHVHO\PSKRPDVPRVWW\SLFDOO\RFFXULQ:DOGH\HU ULQJDQGRUFHUYLFDOO\PSKQRGHVDQGDUHORZVWDJH 7KH\PD\KDYHDIROOLFXODUIROOLFXODUDQGGLIIXVH RUSXUHGLIIXVHJURZWKSDWWHUQUHVHPEOLQJ)/JUDGH 3B or a DLBCL. The neoplastic lymphocytes are a monotonous population composed of medium to large cells with vesicular nuclei and small basophilic QXFOHROLDQGWKH\ODFNVWDUU\VN\SDWWHUQ)LJXUH$ DQG%2QLPPXQRSKHQRW\SLQJWKHVHO\PSKRPDV H[SUHVV&'&'%&/DQG%&/EXWPRVW LPSRUWDQWO\VKRZVWURQJH[SUHVVLRQIRU,5) 080DQGKLJKSUROLIHUDWLYHLQGH[)LJXUH&DQG

Figure 2: Duodenal-type FL. (A) Large follicles present beneath small intestinal mucosa populated predominantly by small lymphocytes and surrounded by thin rim of mantle zone. The neoplastic cell show positive expression for CD10 (B) and BCL2 (C).

Figure 5: LBCL with IRF4 rearrangement. (A) Large back to back neoplastic follicles which contain montonous population of medium to large lymphocytes with vesicular nuclei and small nucleoli (B). The neoplastic lymphocytes show positive expression for MUM1 (C) and BCL6 (D).

'7KHJHQHWLFSUR¿OHVKRZVWKDWPRVWFDVHVKDYH ,*,5)UHDUUDQJHPHQWVVRPHWLPHVWRJHWKHUZLWK %&/UHDUUDQJHPHQWEXWWKH\XQLIRUPO\ODFN%&/ rearrangements. This lymphoma is considered to EHPRUHDJJUHVVLYHWKDQRWKHU37)/EXWSDWLHQWV when treated have shown favourable outcome. These cases must be distinguished from the CD10-,5)080)ROOLFXODUO\PSKRPDVZKLFKDUH often associated with DLBCL and occur in older LQGLYLGXDOV7KHUHIRUHLQDSURSHUFOLQLFDOVHWWLQJ FDVHVZLWKFRH[SUHVVLRQRI&'%&/DQG,5) 080VKRXOGEHVFUHHQHGIRU,5)UHDUUDQJHPHQWV

,PPXQRKLVWRFHPLFDO,+&ZRUNXSLVDQ integral part of almost all lymphoid neoplasms & its judicious use may not only save both time DQGPRQH\EXWLVDNH\WRPDNHDGH¿QLWLYH GLDJQRVLV)LUVWVWHSLQGHHGLVDZHOO¿[HG ZHOOSURFHVVHGDQGRSWLPDOO\FXW+ (VWDLQHG VHFWLRQ6WDUWLQJFULWLFDOHYDOXDWLRQRIDO\PSKRLG OHVLRQDWYHU\ORZPDJQL¿FDWLRQOLNH;DQG JUDGXDOO\PRYLQJWRKLJKHUPDJQL¿FDWLRQV ensure recognition of the overall architecture as effacement of the normal architecture is the essence of most lymphoproliferative disorders. )LUVWEDWWHU\RI,+&WREHUHTXHVWHGGHSHQGV on morphological differential diagnoses. In some cases where even epithelial malignancies FDQQRWEHUXOHGRXWLQLWLDOSDQHOPD\LQFOXGH MXVW/&$DORQJZLWK&\WRNHUDWLQIRULQVWDQFH nasopharyngeal carcinoma may closely mimic ODUJHFHOOO\PSKRPD,I1+/LVPRUHOLNHO\¿UVW SDQHOPD\MXVWLQFOXGH&')LJXUH&'DQG .L,IPRUSKRORJ\LVRIVPDOOFHOOW\SH&' ZLOOEHDFUXFLDOPDUNHUDVGXDOSRVLWLYLW\DORQJ

Judicious Use of Immunohistochemistry &

Ancillary Techniques in Precise Categorization

of Lymphoproliferative Disorders.

3URIHVVRU6KDKLG3HUYH] +LVWRSDWKRORJ\ ZLWK%PDUNHUOLNH&'RU&'ZLOOSUDFWLFDOO\ UHVWULFWGLIIHUHQWLDOGLDJQRVHVWRWZRHQWLWLHVLH µVPDOO%FHOOO\PSKRF\WLFO\PSKRPDOHXNHPLD¶ %6// µPDQWOHFHOOO\PSKRPD0&/¶ )LJXUH,IPRUSKRORJ\LVEODVWRLGSDUWLFXODUO\ LQFKLOGUHQ&'$VKDOOUHSODFH&'DVPRVW SUHFXUVRU%O\PSKREODVWLFO\PSKRPDOHXNHPLD GRQ¶WH[SUHVV&',QDGGLWLRQLQUHODSVHGFDVHV

 RIPDWXUH%FHOOO\PSKRPDVOLNHµGLIIXVHODUJH %FHOOO\PSKRPD'/%&/¶WUHDWHGZLWKDQWL &'5HWX[LPDE&'H[SUHVVLRQPD\EH ORVWDQG3$;ZLOOEHDEHWWHU%FHOOPDUNHUWR FRQ¿UP%FHOOGLIIHUHQWLDWLRQ,IDO\PSKRPDLV /&$SRVLWLYHEXWERWK&' &'LVQHJDWLYH µDQDSODVWLFODUJHFHOOO\PSKRPD$/&/¶DQG myeloprolifertive disorder should be suspected ZLWKIXUWKHUWHVWLQJE\&'&' 032 ,IRQPRUSKRORJ\µ+RGJNLQ/\PSKRPD+/¶ LVPRUHOLNHO\&'&'3D[ /03RU (%(5DUHOLNHO\WRFRQ¿UPWKHGLDJQRVLV.L KDVQRYDOXHLQWKHGLDJQRVLVRI+/6SHFLDO VWDLQVOLNH3$6DUHSDUWLFXODUO\KHOSIXOWRH[FOXGH QRQ+RGJNLQ¶VO\PSKRPD1+/¶LQWKHVHWWLQJ RIµURXQGEOXHFHOOWXPRUµDVDEXQGDQWLQWUD F\WRSODVPLFJO\FRJHQZLOOEHKLJKO\XQOLNHO\LQ DO\PSKRPDDQGZLOOIDYRUµ(ZLQJVDUFRPD¶RU µ5KDEGRP\RVDUFRPD¶2WKHUDQFLOODU\WHFKQLTXHV OLNH),6+WHVWLQJIRUWLQVXVSHFWHG%XUNLWW

lymphoma or Double hit large B-cell lymphoma )LJXUHPD\EHRIVLJQL¿FDQFH%DQG7FHOO receptor gene rearrangements are also useful tools WRFRQ¿UPWKHFORQDOLW\RIWKHQHRSODVWLFO\PSKRLG FHOOV)RU%FHOOO\PSKRPDVNDSSDRUODPEGDOLJKW FKDLQUHVWULFWLRQGHPRQVWUDWHGE\,+&RU,6+LQ VLWXK\EULGL]DWLRQLVIUHTXHQWO\XVHG+RZHYHU IRU7FHOOFORQDOLW\WKHUHLVQRVXFKPDUNHU

hence T cell receptor gene rearrangement studies are used to distinguish benign from malignant T-lymphoid proliferations. Benign reactive T lymphoid proliferations do not harbor monoclonal UHDUUDQJHPHQWVZKHUHDVPDOLJQDQWO\PSKRLG WXPRUVKDUERUFORQDOUHDUUDQJHPHQWV)LJXUH +RZHYHULQVSLWHRILWVLQFUHGLEOHSRZHUUHVXOWVRI clonality assays should be interpreted with caution as false-positive as well as false-negative results are possible. In summary a methodical and step by step approach is the best guarantee to reach a conclusive diagnosis in a judicious cost-effective manner.

Figure1B diffuse strong membrane positivity with CD20 by immunohistochemistry.

Figure 2: Flow cytometric analysis of a case of small B-cell lymphocytic lymphoma/leukemia. Note double positivity of neoplastic lymphoid cells to CD19 & CD5.

Figure 3A: FISH IGH/MYC/CEP8 Negative for translocation (8;14), two green two orange signals represent normal copies of chromosome 8 and 14

Figure 3B: FISH IGH/MYC/CEP8 Positive for translocation (8;14), two yellow fusion signal represents translocation (8;14). Furthermore an enumeration control probe that binds to centromeric region of chromosome 8 gives aqua color signal.

7F5ȕVDPSOHVKRZLQJFORQDOVLQJOHSHDN

7F5ȕWXEH$SRVLWLYHFORQDOFRQWUROVKRZLQJFORQDOVLQJOH peak

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Figure 4: Gene scan of two samples showing clonal and polyclonal peaks with controls.

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7F5ȕWXEH$SRVLWLYHSRO\FORQDOFORQDOFRQWUROVKRZLQJ polyclonal (several) peaks

TEST SAMPLES

POSITIVE CONTROL

 ,Q+RGJNLQ¶V/\PSKRPDSDUWLFXODUO\LQWKHWULDOV FRQGXFWHGLQDGROHVFHQWDJHJURXS3(7&7KDV KHOSHGLQGHFUHDVHGWR[LFLWLHVRIFKHPRWKHUDS\ EDVHGRQWKHUHVSRQVHVHHQRQWKHVFDQVZLWKRQO\ DPLQRULW\UHTXLULQJLQWHQVL¿FDWLRQRIWUHDWPHQW )RU1RQ+RGJNLQ¶V/\PSKRPDV3(7&7LV helpful in the older age group where decisions are needed regarding escalating treatment in patients ZKLFKDUHSRRUUHVSRQGHUVWRLPSURYHWKHRYHUDOO outcome.

3(7&7KDVEHHQVWDQGDUGL]HGE\HQUROOLQJ a large number of patients. The parameters of standardization include quality assurance and reporting. A five point Deauville criteria is used for reporting of images. A score of RQHRUWZR”QRUPDOPHGLDVWLQDOXSWDNH is equivalent to de-escalating treatment or complete metabolic response. There were several trials which were concerned about over treatment of patients for which scores of one to WKUHH”QRUPDOOLYHUXSWDNHKDYHLQFUHDVLQJO\ been used to diagnose complete metabolic response.

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recurrences as well in patients with lymphoma. It is now currently used as up-front imaging technique in patients who are diagnosed with +RGJNLQ¶VRU1RQ+RGJNLQ¶V/\PSKRPD $WSUHVHQWHDUO\UHVSRQVHWRWUHDWPHQWIRU /\PSKRPDVLVEHVWDVVHVVHGE\3(7&7 Lymphomas are malignant neoplasms

characterized by the abnormal proliferation of B or T lymphocytes1. Malignant lymphomas encompass a wide variety of distinct disease entities. It is common in developed and developing FRXQWULHV7KH(DVW$VLDUHJLRQKDVRQHRIWKH lowest incidence rates of malignant lymphoma. The incidence of malignant lymphoma around the world has been increasing at a rate of three WRIRXUSHUFHQWRYHUWKHODVWIRXUGHFDGHVZKLOH some stabilization has been observed in developed countries in recent years.

The management and treatment of lymphomas starts from histopathological diagnosis to staging the disease which involves imaging techniques DQGERQHPDUURZELRSV\,QUHFHQW\HDUVLPDJLQJ WHFKQLTXHVKDYHHYROYHGDQG3(7&7KDVEHFRPH integral part of management protocol. Positron HPLVVLRQWRPRJUDSK\3(7ZLWKGHR[\ >ÀXRULQH@ÀXRUR'JOXFRVH_))'*LV

rge tracer responsible for the increased glucose XSWDNHDQGJO\FRO\VLVRIPDOLJQDQWZKLFKLVDEOH to demonstrate metabolic abnormalities before PRUSKRORJLFDOFKDQJHVRFFXU))'*3(7&7 DFTXLUHV3(7DQG&7GDWDLQWKHVDPHLPDJLQJ session and anatomically localizes lesions detected RQWKH))'*3(7VFDQ6LPLODUO\3(7&7 has allowed better characterization of staging in patients with lymphomas leading to decreased cycles of chemotherapy and smaller volumes of radiotherapy accordingly.

18

_{F-FDG PET/CT Imaging in Lymphomas}

Dr Natasha Ali +DHPDWRORJ\