B- cell lymphoproliferative disorders Flashcards
List the differential diagnosis for CD5+ LPDs and how to differentiate between them
1) CLL.
- Most common CD5+ LPD
- Classic morphology: small, mature, monomorphic lymphocytes with clumped nuclear chromatin, round nucleus, high N:C ratio and mildly basophilic cytoplasm.
- Focal nodular or interstitial pattern of infiltration on trephine. Diffuse infiltration and expanded proliferation centres in high risk disease.
- Classic immunophenotype: CD5+ CD19+ B cells. Weak CD20, CD22 and SIg. CD79a weak or absent. CD23 and CD200+
- FMC-, LEF1+
2) MCL.
- Second most common CD5+ LPD
- Classic morphology: small, mature, monomorphic lymphocytes with condensed nuclear chromatin a high N:C ratio and mildly basophilic cytoplasm. Nuclear clefts commonly seen. Many variants (small cell, marginal, pleomorphic, blastic)
- Nodular, paratrabecular, interstitial or diffuse infiltration on trephine.
- Classic immunophenotype: CD5+, CD19+ B cells that are CD20, CD22, CD79a and SIg positive. CD23 and CD200 negative .
- CCND1 and SOX11 positive in classic cases.
- t(11;14) translocation identified on FISH.
3) B-PLL
- CD5 positivity seen in 20-30%
- Monomorphic population of prolymphocytes present (intermediate sized cells, moderately diffuse nuclear chromatin with prominent nucleolus, medium to high N:C ratio with mildly basophilic cytoplasm)
- >55% of lymphocytes must be prolymphocytes.
- Immunophenotype: CD19+, CD20+, CD22+, CD79a+, CD23 in 10-20%. CD200 usually negative. Strong SIg.
- Complex karyotypes, del(17)p/ TP53 mutations common.
4) Marginal zone
- CD5 positivity seen in up to 20% of cases of MZL
- Monomorphic population of intermediate sized lymphoid cells with mature nuclear chromatin and a high N:C ratio. Bipolar cytoplasmic fimbriae in SMZL.
- Nodular or diffuse infiltration on trephine. Can form a germinal centre. Intrasinusoidal in SMZL.
- Immunophenotype not specific: positive for CD19, CD20, CD22, CD79a and SIg. CD10- and usually negative for CD23 and CD200.
- No specific diagnostic features. Often a diagnosis of exclusion. del(7q) seen in a subset and is rare in other LPDs.
5) LPL
- CD5 positivity seen in up to 20% of cases of LPL
- Plasmacytic differentiation present: plasmacytoid lymphocytes present as well as plasma cells (with normal, mature morphology). Increased mast cells (with normal morphology) on aspirate sample.
- Interstitial or diffuse infiltration on trephine. Plasma cells admixed.
- Immunophenotype not specific: positive for CD19, CD20, CD22, CD79a and SIg. CD10- and usually negative for CD23 and CD200.
6) CD5+ DLBCL
- CD5 positivity in 5-10% of DLBCL, usually ABC phenotype.
- Large immature lymphoid cells. Can be pleomorphic.
- High forward scatter on flow cytometry. CD19+, CD20+, CD22+, CD79a+. CD10+/-. High Ki67.
- MYC, BCL2, BCL6 overexpression, translocation variable.
List the immunophenotypic/ genetic features of hairy cell leukaemia
- Flow cytometry: high side scatter. Bright sIg, CD20, CD22 and CD11c. CD25, CD103, CD123 and CD200 positive. FMC7 positive. Usually CD10 negative
- Annexin 1 positive (most specific marker)
- B-RAF V600E
- TBET.
Discuss monoclonal B lymphocytosis
- Refers to a clonal lymphocyte count <5x10^9 in the peripheral blood without any clinical symptoms or signs (cytopenia, lymphadenopathy, hepatosplenomegaly) of an underlying LPD.
- Divided into those with a CLL phenotype vs those with a non-CLL phenotype (associated with a worse prognosis, must exclude an underlying LPD)
- Divided into low count (<0.5x10^9) and high count (>0.5x10^9) MBL
- In those with a CLL MBL, low count is associated with a good prognosis with a very low rate of clinical progression.
- High count CLL MBL is identical to CLL Rai stage 0/ Binet stage A and has a rate of progression to CLL that is 1-2% per year.
- Some MBLs may be transient/ self-limited
What are some of the features of unmutated CLL
- Unmutated CLL is thought derive from an antigen naïve B cell that has not experienced the germinal centre
- Associated with more aggressive disease, chemotherapy resistance and reduced overall survival.
- Advanced stage (C, III, IV)
- Lymphocyte doubling time <12 months
- High ZAP70
- High CD38
- High B2MG
- High-risk cytogenetics (del(17p), complex karyotype)
- Higher rates of clonally related Richter’s transformation
List the poor prognostic features of CLL
1) Clinical
- Age
- Male sex
- CIRS score >6
- Clinical stages (lymphadenopathy, splenomegaly)
2) Lab/ Morphology
- Thrombocytopenia (<100)
- Anaemia (<100- 110)
- Lymphocyte doubling time <12 months
- Increased LDH, B2M
- Atypical morphology
- Diffuse pattern of BM infiltration
- Expanded proliferation centre
- Clonally related Richter’s transformation
3) Immunophenotype
- ZAP70 positive
- CD38 positive
- CD49d positive
3) Molecular
- Del(17p)/ TP53 mutations, complex karyotype
- Unmutated IGHV
- NOTCH1, MYC mutations
What is Richter’s transformation. What different types are there?
- Transformation of CLL to an aggressive diffuse large B cell lymphoma or Hodgkin’s lymphoma
1) DLBCL= most common (2-8% of CLL patients)
- Clonally related= most common= worse prognosis
- Most common in those with unmutated IGHV.
- Associated with mutations in TP53, MYC, CDKN2A and NOTCH1
- Clonally unrelated= least common= same prognosis as de novo DLBCL
- Most common in those with mutated IGHV.
2) Hodgkin’s Lymphoma
- 50% clonally unrelated
- Thought to arise out of immunosuppression due to CLL and therapy
- More common in those with mutated IGHV.
- Poorer prognosis than de novo Hodgkins lymphoma
List the mutations most commonly seen in CLL and the associated prognosis
1) Del(13q)
- 25-40%
- Good prognosis. Average survival >15 years.
2) Trisomy 12
- 15-20%
- Associated with atypical morphology.
- Shorter treatment free survival.
3) Del(11q)
- 20-25%.
- Shorter time to progression.
4) Del(17p)
- 5- 10%
- Associated with unmutated IGHV, TP53 mutations, clonal evolution, complex karyotype.
- Associated with resistance to chemotherapy.
List the Rai and Binet stages in CLL
Rai staging: Stages 0-IV - 0= isolated lymphocytosis - 1= + lymphadenopathy - 2= and/or spleen/liver - 3= and/or anaemia - 4= and/or thrombocytopenia OS ranges from >15yrs to 3-5yrs.
Binet staging:
- A isolated lymphocytosis
- B >3 lymphoid areas
- C anaemia/ thrombocytosis
How is a diagnosis of atypical CLL made?
- Atypical morphology: cleaved forms, prolymphocytes, large/ immature cells present and comprise >15% of total lymphocytes.
- Atypical immunophenotype: can be CD5-, CD23-, Sig+ (strong) and FMC7+. Usually CD200 positive.
- Associated with trisomy 12
What is grade 3 follicular lymphoma?
Defined by the presence of more than 15 centroblasts per HPF
- Grade 3A: centrocytes also present.
- Grade 3B: centrocytes absent
Ki67 often >20%
What are the hallmark features of follicular lymphoma?
How do these features differ across the different grades of follicular lymphoma?
- Germinal centre B cell that has undergone VDJ rearrangement
- BCL2 over-expression with t(14;18) in 85-90%
- CD10 positive (rare in other small cell lymphomas)
- Aberrations in chromatin modifiers: KMT2D, CREBBP, EZH2
- Grade 3 follicular lymphoma, especially grade 3B, is more likely to be CD10-, BCL2-, MUM1/IRF4+ and BCL6+ than grade 1/2 FL
List the prognostic features of follicular lymphoma
- FLIPI: age >60, Hb <120, stage III or IV, >4 nodal sites, raised LDH
- Grade 3 disease
- Aberrations in TP53, MYC translocations.
List the 4 subtypes of follicular lymphoma and the important features of these subtypes
1) Testicular follicular lymphoma
- Children and young adults
- Cytologically high grade (usually 3A)
- Lack t(14;18)
2) In situ follicular neoplasia
- Colonisation of germinal centres by B cells with t(14;18)
- If incidentally found: <5% risk of progression to FL
- Higher rates of progression if there are high numbers of FL like cells in the PB.
3) Duodenal type follicular lymphoma
- t(14;18) present within clonal B cells
- Low grade with few centroblasts present
- Good prognosis
4) Paediatric FL
- Children and young adults
- Head and neck lymph nodes. Single site of disease common
- Lack t(14;18)
- Appear high grade, high proliferation rate
- Excellent prognosis
Marginal zone lymphoma can be a diagnosis of exclusion. List the flow cytometry and immunohistochemical markers that can be performed to exclude other low grade LPDs.
- Flow cytometry: CD5-, CD10-, CD23-, CD200-, CD103-. Strong expression of SIg and the B cell markers.
- IHC: BCL6-, CCND1-, SOX11-, LEF1- and annexin A1 negative.
Discuss the genetic profile of MALT
- IGHV rearranged
- Biased IGHV gene usage
- Chromosomal translocations involving involving MALT1 on chromosome 18 and IgH on chromosome 14 are present in a subset
- t(11;18): chimeric protein BIRC3-MALT1
- t(1;14): transcriptional deregulation of BCL10
- t(14;18): transcriptional deregulation of MALT1
- t(3;14); transcriptional deregulation of FOXP1
What immunohistochemical stains, if positive, can be helpful in the diagnosis of MZL?
IRTA1 and MNDA
List the lymphomas that can demonstrate an intrasinusoidal pattern of bone marrow infiltration
- Splenic marginal zone lymphoma
- Nodal marginal zone lymphoma
- Splenic diffuse red pulp small B cell lymphoma
- Hairy cell variant
- Intravascular lymphoma
- Hepatosplenic T cell lymphoma
- T-cell LGL
How is the immunophenotype of hairy cell variant the same and how is it different from hairy cell leukaemia?
- Similarities: Pan B cell expression, CD11c, CD103 and strong SIg positive.
- Differences: CD25, CD200, CD123 and annexin A1 negative. Bright CD72.
What lymphomas can show plasmacytoid differentiation?
- Lymphoplasmacytic lymphoma/ Waldenstroms Macroglobulinaemia
- Marginal zone lymphoma
- Follicular lymphoma
- Angioimmunoblastic T cell lymphoma
List the poor prognostic markers in LPL/ WM
- Age
- Cytopenia
- Elevated LDH and/ or B2M
- High serum paraprotein
- CXCR4 mutations (seen in 30%-40%)
- TP53 aberrations
What are the diagnostic features of cold agglutin disease?
- Monoclonal IgM kappa
- Nodular pattern of infiltration without a significant plasmacytic cytology or reactive mast cell population.
- Plasma cells surrounding aggregates and within the interstitium rather than admixed with the lymphoid population.
- Median infiltration ~10%
- Flow cytometry pattern: kappa light chain restricted B lymphocytes CD20+, IgMs+/IgDs+, CD27+, CD5-/+, CD11c-, CD23-, CD38-.
- Highly restricted IGHV and light chain usage that differs from that seen in WM)
- KMT2D and CARD11 mutations
- No MYD88
What are the features of leukaemic non-nodal mantle cell lymphoma?
- Isolated spleen, bone marrow and peripheral blood involvement
- No nodal involvement
- Non-blastoid or pleomorphic morphology
- CCND1 positive, SOX11 negative
- Low Ki67 <10%
- Mutated IGHV genes
What are some of the poor prognostic features seen in mantle cell lymphoma?
- Age
- Raised LDH
- Raised WCC
- ECOG >1
- Ki67 >30%
- Unmutated IGHV
- Blastoid or pleomorphic morphology
- TP53 aberrations/ del(17p)
- SOX11 positivity (in some but not all studies)
What is the Hans algorithm and why is it used?
- Gene expression profiling in DLBCL has shown three distinct subtypes of DLBCL with different prognosis
- Germinal centre phenotype: good prognosis 80% 3 yr survival
- Activated B cell phenotype: poor prognosis: 45% 3 year survival
- Unclassified.
- Gene expression profiling is expensive and available only in specialised laboratories.
- The Hans algorithm uses immunohistochemistry to determine cell of origin. It is used as a surrogate for GEP studies.
- CD10 positive cases as classified as GCB
- CD10-, BCL6+, MUM1/IRF4- cases are classified as GCB
- CD10-, BCL6- are classified as non-GCB
- CD10-, BCL6+, MUM1/IRF4+ are classified as non-GCB
- 86% concordance with GEP. Sensitivity of 85-90% for GCB subtype.
List the four genetic subtypes of DLBCL and the key features of the different subtypes
1) MCD: MYD88 and CD79b mutations
- Seen predominately in ABC subtype
- High reliance on chronic active B-cell receptor signalling
- Poor outcome. 5yr OS 26%
2) BN2: BCL6 fusions and Notch2 mutations
- Equally distributed across ABC and unclassified subtypes
- Also rely on chronic active B-cell receptor signalling via NF-KB
- Associated with a better outcome. 5yr OS 65%
3) N1: Notch1 mutations
- Seen predominately in the ABC subtype.
- Associated with a poor outcome: 5yr OS 36%
4) EZB: EZH2 and BCL2 translocations
- Seen predominately in the GCB phenotype.
- Associated with a favourable outcome: 5yr OS 68%
What is double hit/ triple hit DLBCL
- DLBCL with CMYC and BCL2 and/ or BCL6 gene rearrangements.
- Poorer prognosis when compared to non DH/ TH DLBCL: more intensive treatment used (DA-EPOCH-R vs R-CHOP)
- Prognosis of CMYC/ BCL6 DH debated: some data suggests that the prognosis is not as poor as CMYC/BCL2 rearranged although more recent data (Lunenburg analysis) suggests that prognosis is just as poor.
- CMYC/ BCL2 occur almost exclusively in GCB phenotype
- CYMC/BCL6 occur almost exclusively in ABC phenotype
- CMYC occurs equally in both subtypes
What are the approaches to FISH testing in DLBCL
1) Screen all DLBCL treated with curative intent ( ESMO guidelines)
2) Screen all GCB DLBCL: will miss ~1/3rd of cases (most of which will be BCL6/CMYC, NCCN)
3) Screen all double expressor DLBCL: will miss ~1/3rd of cases
4) Screen all for CMYC, proceed to BCL2 and BCL6 if positive
Cannot use high Ki67 or high clinical risk factors.
List the poor prognostic features in DLBCL
- High IPI score
- Age >60
- ECOG >1
- > 1 extra nodal sites
- Stage III or IV
- LDH elevated
- Male sex
- Concordant bone marrow involvement
- CNS involvement
- ABC phenotype (likely MCD and N1)
- CD5+
- CMYC with an IGH partner
- Double expressor
- (Double hit)
What is the immunophenotype of PMBCL
- CD30+, CD23+, MUM1/IRF4+, CD15-
- Should not have CMYC, BCL2, BCL6 rearrangements
What is the immunophenotype of BL
- Germinal centre B cell origin: Strong SIg expression with light chain restriction and expression of pan B-antigens.
- Expression of GC markers CD10 and BCL6
- Overexpression of CMYC
- Ki67 ~100%
- BCL2 negative (distinguishes from DLBCL)
- TDT negative (distinguishes from B-ALL)
Discuss the genetics of BL
- Characterised by the MYC translocation: most commonly t(8;14): with IgH
- Other translocations involve lambda light chain
(22q) or less commonly kappa (2p)
- Other translocations involve lambda light chain
- Simple karyotype
- Gain of 1q= most common additional karyotypic abnormality
- In addition to being translocated, MYC is the most common gene mutated in BL
- ID3 and TCF3 mutations are common in sBL: result in antigen independent signalling through the BCR
- Cell cycle genes such as CCND3 and CDKN2A mutations are also commonly seen
- TP53 aberrations in 55% of sBL
What is Burkitt like lymphoma with 11q aberrations?
- Clinical, morphologic, immunophenotypic, gene expression and prognostic profile of BL BUT lack a demonstrable MYC translocation by FISH
- Characterized by an interstitial 11q23 gain immediately followed by a telomeric loss at 11q24
- Can have more complex karyotypes than classical BL
