Diagnosis and monitoring of myeloma and AL amyloidosis

Diagnosis and monitoring of myeloma and AL amyloidosis Peter Mollee Haematology Department Cancer Services and Pathology Queensland Princess Alexandra Hospital Brisbane AL amyloidosis Amyloidosis and amyloid
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Diagnosis and monitoring of myeloma and AL amyloidosis Peter Mollee Haematology Department Cancer Services and Pathology Queensland Princess Alexandra Hospital Brisbane AL amyloidosis Amyloidosis and amyloid fibrils Disorder of protein folding Structurally diverse precursors adopt an abnormal common fibrillar conformation New properties: Bind Congo red and SAP Unusual stability Damage tissue structure, organ function Progressive and fatal without treatment Images courtesy Dr Hugh Goodman Amyloid type Other names Protein Mechanism AL Primary monoclonal Ig light chains AA Secondary / reactive serum amyloid A AB 2 M Dialysis related amyloidosis beta-2 microglobulin ATTRwt (wild type) ATTRm (mutated) Senile systemic (cardiac) amyloidosis Familial Amyloid Polyneuropathy Cardiomyopathy) transthyretin (wild type) transthyretin (mutated) AFib fibrinogen A- alpha chain acquired, abnormal, amyloidforming protein normal protein at supra-normal concentration normal protein at supra-normal concentration normal protein and concentration prolonged exposure hereditary, mutant protein hereditary, mutant protein ALys lysozyme hereditary, mutant protein AApoA1 and 2 ALect2 apolipoprotein s 1 and 2 leukocyte chemotactic factor 2 hereditary, mutant protein hereditary, mutant protein AGel Finnish; Meretoja syndrome gelsolin hereditary, mutant protein AL amyloidosis - when to suspect it? Nephrotic range proteinuria Cardiac failure with left ventricular hypertrophy in the absence of hypertension or aortic valve disease Sensorimotor peripheral neuropathy without obvious cause Hepatomegaly with a normal appearance on ultrasound or CT imaging Autonomic neuropathy Making the diagnosis of amyloidosis Needs (usually) a tissue biopsy Congo Red remains the gold standard Congo Red positivity birefringence and dichroism effects when examined under polarised light microscopy Can biopsy: Clinically involved organ Distant site (fat pad, marrow, rectum, salivary glands) Evaluation of newly diagnosed AL amyloidosis Histologically confirmed AL amyloidosis Define the plasma cell clone Other investigations Assess for symptomatic myeloma Assess organ involvement Define the plasma cell clone Serum protein electrophoresis + immunofixation Urine protein electrophoresis + immunofixation Serum free light chains (Bone marrow with assessment of plasma cell clonality) will detect 95% of plasma cell clones Which FLC assay should I use? Freelite TM assay Based on polyclonal anti-flc antibodies obtained from sheep immunised against human Bence Jones protein (Binding Site) Assay limitations Batch-to-batch variation Non-linearity Antigen excess Non-reactivity Overestimation Basis of most FLC clinical validation in AL N Latex FLC assay Based on a cocktail of monoclonal anti-flc antibodies (Siemens) Some analytical advantages over Freelite TM assay Introduced by several laboratories in Australia Little published validation in AL amyloidosis Results Analytical comparison Passing-Bablok regression line on a logarithmic scale Kappa Lambda Mollee. Clin Chem Lab Med Aug 10:1-8 Analytical comparison Bland-Altman difference plots Kappa 0.8 Log difference κ FLC (mg/l) (N Latex minus Freelite) Uninvolved FLC o Involved FLC Lambda Log difference λ FLC (mg/l) (N Latex minus Freelite) Mean κ FLC Logarithmic difference plot N = 94 Mean difference : [ to ] Mean λ FLC Logarithmic difference plot N = 94 Mean difference : [ to 0.131] Which FLC assay should I use? 1. Diagnostic sensitivity similar between N Latex and Freelite TM assays in AL amyloidosis Our study Italian study N Abnormal κ:λ ratio N Latex 79% 84% Freelite TM 85% 82% Overall diagnostic sensitivity (with serum and urine EPP) N Latex 98% 98% Freelite TM 98% 98% Clin Chem Lab Med. 2013;10:1 ASH 2012 Which FLC assay should I use? 2. Baseline dflc by both N Latex and Freelite TM assays predicts overall survival Our study Italian study N Median dflc N Latex 150 mg/l 165 mg/l Freelite TM 190 mg/l 180 mg/l Clin Chem Lab Med. 2013;10:1 ASH 2012 OS predicted by baseline dflc N Latex Freelite TM Overall survival according to baseline dflc by N Latex FLC assay Overall survival according to baseline dflc by Freelite assay p = p = Overall survival 0.50 dflc 150mg/L Overall survival 0.50 dflc 190mg/L dflc 150mg/L dflc 190mg/L Months Months Monitoring AL Amyloidosis Updated Haematologic Response Criteria dflc is central to response assessment all clinical validation done with Freelite assay Criteria CR VGPR Response Negative SPEP/IFE Negative UPEP/IFE Normal FLC ratio dflc 40mg/l (or 90% reduction) PR dflc decrease 50% NR Less than PR Overall Survival Based on 6 Month Haematologic Response Palladini. JCO 2012 Which FLC assay should I use? 3. Monitoring utility of two FLC assays in AL amyloidosis appears similar but need further studies of N Latex FLC assay PR by both assays predicts overall survival Sample size too small to assess impact of other response categories Cut-offs for measureable disease and vgpr are based on Freelite TM values May not be appropriate for N Latex assay Results PR predicts OS by both assays N Latex Freelite TM Overall survival according to N Latex FLC determined FLC response Overall survival according to Freelite determined FLC response p = p = Partial response 0.75 Overall survival Less than partial response Overall survival Partial response Less than partial response Months Months Which FLC assay should I use? 4. N Latex FLC and Freelite TM assays have analytical differences in AL amyloidosis It is not clear which assay s result is more accurate Must use same assay on same platform in same laboratory when using FLC to monitor AL amyloidosis Further clinical validation studies of the N Latex assay in monitoring AL amyloidosis are required Myeloma Serum free light chain assay Reference intervals Screening MGUS Myeloma diagnosis Myeloma monitoring Flow cytometry based MRD Bone imaging Serum FLC assay: What is normal? Increased levels Increased FLC production Polyclonal inflammatory response Chronic liver disease Monoclonal light chain Reduced elimination Renal impairment Reduced levels Hypogammaglobulinaemia Screening for monoclonal gammopathies SPEP (with IFE) + UPEP SPEP (with IFE) + serum FLC Suitable screening panel for high tumour burden plasma cell disorders (e.g. myeloma, smoldering myeloma, Waldenstrom s macroglobulinaemia) Pragmatic reasons Pre analytical factors Analytical factors SPEP + IFE, UPEP and serum FLC required for AL amyloidosis and light chain deposition disease Dispenzieri, Leukemia 2009; Katzmann, Mayo Clin Proc 2006; Katzmann, Clin Chem 2009; Hill, Clin Chem 2009, Beetham, Ann Clin Biochem 2007. FLC assay in screening panels (Mayo Clinic) MM WM MGUS SMM SPEP Serum FLC Serum IFE UPEP/Urine IFE Plasmacytoma Extramedullary Plasmacytoma POEMS AL LCDD - Study limitations - focus on screening panel sensitivity but not specificity. - Screening population limited to patients with documented monoclonal gammopathies rather than general population with suspected MG. - Whether such proposed screening panels will prove useful in terms of sensitivity and specificity in general population needs to be assessed. FLC assay in MGUS Risk Group Low (n=449) Paraprotein 15g/L IgG subtype Normal FLC ratio 20yr risk of progression 20yr risk accounting for death 5% 2% Low-intermediate (n=420) 21% 10% Addition of FLC helps prognostication in MGUS One factor abnormal High-intermediate Low risk group (n=226) (~40% of MGUS) 37% need infrequent 18% Two follow-up factors abnormal (at 6 months then every 2-3 yrs IMWG High (n=53) guidelines (Kyle. Leukemia 58% 2010;24:1121) 27% All 3 factors abnormal Do they need any follow-up? Rajkumar. Blood. 2005;106:812 FLC assay in diagnosis of myeloma IMWG Recommendations (Blood 2011;117:4702) Assessment of the plasma cell clone SPEP and IFE Immunoglobulin quantification UPEP and IFE (24hr specimen) Serum free light chains Bone marrow FLC assay and symptomatic myeloma Initiation of therapy in myeloma predominantly based on presence of end organ damage i.e. CRAB criteria HyperCalcaemia Renal impairment Anaemia Bone disease Are there patients currently defined as asymptomatic myeloma who should be treated? Risk of asymptomatic MM progressing to symptomatic MM Factor 2yr progression to MM BM plasmacytosis 60% ~63% i/u FLC ratio 100 79% 1 MRI lesion 70% New IMWG criteria for symptomatic myeloma BM plasmacytosis 60% i/u FLC ratio 100 (Freelite assay) 1 MRI lesion (at least 5mm in size) What about rapidly rising paraprotein? FLC monitoring useful to diagnose light chain escape Serum FLC in monitoring of myeloma LC escape My occurs own preference ~10% myeloma is to monitor that present with both with FLC an intact Use the same immunoglobulin (Brioli Blood and 2014;123:3414) 24 hour urine IMWG Recommendations (Blood 2011;117:4702) method for disease Diagnosis often delayed as UPEP but often not done Poor correlation between 24hr BJP monitoring and FLC FLC response didn t predict PFS or OS Measurable paraprotein SPEP (preferred) Quantitative immunoglobulin (esp if co-migrating in beta region) Light chain myeloma 24 hour urine Non-secretory and oligosecretory myeloma FLC (Dispenzieri. Blood 2008;111:4908) but improved monitoring is required Progress in myeloma therapy Regimen Complete 1yr OS remission Melphalan + prednisolone (MP) 2% 70% Autologous stem cell transplantation ~25% 80% MP + thalidomide 16% 90% MP + bortezomib 32% 92% MP + lenalidomide 24% 100% Lenalidomide + dexamethasone ~15% 94% Carfilzomib, lenalidomide + 79% 100% dexamethasone Improving overall survival - older patient data Liwing. BJH 2013 epub Monitoring myeloma - Stringent CR IMWG 2006: new response category scr = CR + normal FLC ratio (Durie. Leukemia 2006) scr did not predict EFS or OS in: 34 patients in CR post-autosct (Blood 2009;114:4954) 52 patients (post-allosct) in CR (Blood 2010;115:3413) 203 patients (ClinChem 2009;55:1510) 260 patients (JCO 2011;29:1627) Abnormal FLC ratio associated with oligoclonal bands (Blood 2009;114:4954) IMWG 2011 scr = CR + normal FLC ratio + absence of clonal plasma cells by IHC or flow cytometry (Rajkumar. Blood 2011) Monitoring myeloma - Flow MRD IMWG 2011: new response category Immunophenotypic CR = scr + absense of phenotypically aberrant plasma cells (minimum 10 6 cells) Phenotypically aberrant plasma cells Lack of CD19 Strong CD56 Weak CD27 Weak CD45 The demonstration of phenotypically abnormal plasma cells is more sensitive and specific for the detection of MRD than clonality assessment by IHC and/or flow cytometry. Haematologica 2008;93(3):431 Monitoring myeloma - Bone imaging Evaluation of the depth of response beyond the conventionally defined CR level is required Need to detect: persistence focal bone lesions (potentially harbouring non-secretory MM cells) sites of active disease outside the medullary cavity Skeletal survey Not useful for the assessment of response to treatment or during the follow-up phase, because healing of lytic lesions is a rare event, even in patients who achieve sustained CR (Acta Medica Scandinavica, 212, 385). Monitoring myeloma - MRI Resolution of focal lesions (MRI-CR) post-transplant Confers superior OS Especially in patients with 7 focal lesions at baseline (JCO 2007;25:1121) Post-ASCT number of focal lesions predicted for OS (Haematologica 2012) Beware: Focal lesions may remain hyper-intense in both responder and non-responder patients for several months after therapy Possibly due to treatment-induced necrosis and inflammation False diffuse bone marrow pattern may appear, due to the use of growth factors Monitoring myeloma FDG PET/CT Promising tool to monitor response, due to ability to distinguish between active disease and fibrotic lesions PET-CR predictive of PFS and OS in multiple studies PET-negativity preceded the achievement of conventionally defined CR (Blood 2009;114:2068) Persistence of high tumour metabolism after induction treatment is an early predictor of worse PFS (Blood 2011;118:5989) PET/CT negativity after ASCT retained prognostic significance for the risk of progression or death even among patients achieving conventionally-defined CR (Blood 2011;118:5989) No standardisation; no intra-observer reproducibility studies AL amyloidosis Screening panel should include SPEP, UPEP and FLC dflc is central to disease monitoring Myeloma SPEP remains cornerstone of diagnosis and monitoring FLC assay important in diagnostic assessment and monitoring response, particularly in light chain and oligosecretory myeloma FLC also has a role to diagnose light chain escape Need improved techniques to monitor response MRD assessment Bone imaging
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