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Presentation webinar March 3, 2010
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PLASMA CELL DISORDERS:NEW STRATEGIES FOR
SCREENING , DIAGNOSIS, PROGNOSIS AND MONITORING
Parameswaran Hari, M.D. Howard S. Robin, M.D.
March 3, 2010
IMPACT OF LABORATORY TESTING
• Diagnostic thinking• Therapeutic choice• Patient outcome• Prevention
BENCE JONES PROTEINThe First Tumor Marker
Multiple Myeloma
• Bence-Jones, Henry (1813-1873), British nephrologist and clinical pathologist.
• In 1848 Bence-Jones published an article announcing his discovery of a special protein in the urine of patients suffering from softening of the bones.
• The first physician to note the occurrence of xanthine in urine, Bence-Jones was also a recognized authority on diseases of the stomach and kidneys.
OSSEOUS MALADIES
MONOCLONAL GAMMOPATHIES.
Disorders Signs and symptoms• Bone pain or fractures• Elevated serum and/or
urine proteins• Anemia and fatigue• Renal dysfunction
1026 patients Mayo 1992
OBJECTIVES
1. Develop new strategies to screen for and diagnose monoclonal plasma cell gammopathies.
2. Evalulate new effective strategies for monitoring relapse and response to therapy of patients with plasma cell disorders.
3. Acquire knowledge regarding the prognostic value of current tests for monoclonal gammopathies.
CASE 1
• Previously healthy 75 yr old man• Physically active – runs 3 miles /day• Exertional dyspnea while running• Gave up running after a month• Chest discomfort & increasing SOB• Cardiopulmonary Tests:
– Stress test – No inducible ischemia– Catheterization – 40% LAD stenosis– CT chest – No Pulmonary embolism– Echocardiogram – Left Ventricular Hypertrophy
DYSPNEA OF UNKNOWN ORIGIN
• Clinical picture of increasing right heart failure– Elevated NT Pro BNP of 1500
• Repeat echocardiogram– Diastolic dysfunction and worsening septal hypertrophy
• Differential diagnosis – Restrictive cardiomyopathy– Cardiac amyloidosis
• Referral to hematology clinic• Serum Protein Electrophoresis (SPEP)• Immunofixation (IFE)• Urine Protein Electrophoresis (UPEP) – 24 hr urine All Negative for Monoclonal Spike
SERUM PROTEIN ELECTROPHORESIS
SERUM IMMUNOFIXATIONELECTROPHORESIS
PHYSIOLOGY OF LIGHT CHAINS ≤ 1 g/d
500mg/d from normal B and plasma cells:k l production 2:1
Lambda is dimeric and has longer T1/2T1/2 varies from hrs to 1-2 d (renal function)
Filtered freely by glomeruli then reabsorbed and metabolised by prox. tubules Tubules can absorb upto 10-30 gms per day.
IMMUNOELECTROPHORESIS
Urine IEP Urine IFE
TOOLS FOR EVALUATION OF MONOCLONAL GAMMOPATHIES
• SPEP• Immunofixation• UPEP on 24-hour urine• Serum free light chain assay
PLASMA CELLS PRODUCE WHOLE IMMUNOGLOBULIN AND FREE LIGHT CHAINS
Kappa Lambda
Intact Immunoglobulin
Exposed surface
Hidden surface
Kappa
Free Light Chain
Previouslyhidden surface
SERUM FREE LIGHT CHAIN ASSAY
SERUM FREE LIGHT CHAINS
FLC reference range:k 3.3 – 19.4 mg/L l 5.7 – 26.3 mg/L /k l ratio 0.26 - 1.65
SCREENING FOR CARDIAC AMYLOIDOSIS
• Serum Free Light Chains• Free kappa – 530 mg/L• Free lambda – 16• Increased k: l ratio (33.13)
• Urine – 950mg/24h of albumin • Cardiac MRI scan with Gadolinum
MRI Heart
septum
ELECTRON MICROSCOPY OF FAT PAD ASPIRATE
TISSUE CONFIRMATION OF AMYLOID:Marrow Biopsy – 10% Plasma Cells
Kappa Light Chains restrictedFat Pad Aspirate - Positive
AL AMYLOIDOSIS:SUSPECT
Presence of Amyloid Related Syndrome SCREEN
Monoclonal plasma cell disorder PROVE
Positive Tissue Biopsy (Fat pad, Marrow, Organ) AL by typing – IHC, sequencing etc
SCREENING & DIAGNOSIS OF PLASMA CELL DISORDERS
• Plasma Cell Disorders are a spectrum of illnesses:– Myeloma (MM) – AL Amyloidosis– Light Chain Deposition– MGUS (Monoclonal Gammopathy of Unknown
Significance)– Smoldering MM– Plasmacytoma– Plasma Cell Leukemia
IDEAL SCREENING PANEL FOR MONOCLONAL GAMMOPATHY
• Serum Protein Electrophoresis ( SPEP) • SPEP + Serum Immunofixation (SIFE)• SPEP + SIFE + sFLC• SPEP + sFLC• SPEP + SIFE + sFLC + UPEP
– Most comprehensive
Condition N Abnormal FLC ratio %
Multiple myeloma (MM)
Symptomatic MM 1706 96
“Non secretory” MM 33 73
Light chain MM 252 100
MGUS 1262 33
AL amyloidosis 467 95
LCDD 28 93
Smoldering MM 345 90
Plasmacytoma 116 47
INCIDENCE OF ABNORMAL sFLC RATIO IN PLASMA CELL GAMMOPATHIES
Mead et al BJH 2004 Aug Drayson et al Blood 2001 Bradwell et al Lancet 2003
SCREENING FOR PLASMA CELL DISORDERS
Screening panels for detection of monoclonal gammopathies.Katzmann JA et al Clin Chem. 2009 Aug;55(8):1517-22.
NSPEP+sIFE+uIFE+FLC
SPEP+sIFE +uIFE
SPEP+sIFE+FLC
SPEP+FLC sIFE FLC
N 1877 1851 1821 1828 1770 1632 1395
ALL 98.6 97 97.4 94.3 87 74.3
MM 467 100 98.7 100 100 94.4 96.8
sMM 191 100 100 100 99.5 98.4 81.2
MGUS 524 100 100 97.1 88.7 92.8 42.4
Amyloid 581 98.1 94.2 97.1 96.2 73.8 88.3
Macroglobulinemia 26 100 100 100 100 100 73.1
Plasmacytoma 29 89.7 89.7 89.7 86.2 72.4 55.2
428 Patients with Monoclonal Urinary Protein100 80.8 85.7 93.5 99.5
0
20
40
60
80
100
Ra
te o
f P
os
itiv
e P
ati
en
ts,
%
Urine IFE
Serum PEP
Serum FLC
Serum PEP+IFE
Serum PEP+IFE+FLC
Serum PEP/IFE false negative 28 (6.5%):
Primary Amyloid, n = 19 Plasmacytoma, n = 3 MGUS, n = 3 Multiple Myeloma, n = 2Smoldering MM, n = 1
Serum PEP/IFE or FLC ratio 426 (99.5%) 1 Idiopathic BJ protein MGUS (?contamination)
RELATIVE SENSITIVITY OF ASSAYS
SERUM FREE LIGHT CHAIN ASSAY
• Serum assay– Eliminates need to collect urine specimen
• More sensitive than SPEP or SIFE• Indications:
– Diagnosis of Plasma Cell Disorders– Prognosis & Risk Stratification– Response Assessment on therapy– Disease Monitoring on follow up
CASE 2
• A 74 year old woman presented with elevated total protein August 2003– Routine Labs revealed high total protein 9.8 g/dL– Normal : CBC, Ca++, Bone X ray screen, Creatinine– SPEP & SIFE – IgG Kappa 3.2 g/dl– Marrow 23% Plasma cells kappa light chain clone
• Diagnosis – Smoldering MM (SMM)• Risk of Progression to MM?
Copyright ©2008 American Society of Hematology. Copyright restrictions may apply.
Dispenzieri, A. et al. Blood 2008;111:785-789
RISK OF PROGRESSION TO MYELOMA OR RELATED DISORDER IN 273 PATIENTS WITH SMOLDERING MYELOMA
Kapp
a FL
C
X 100 mg/L
Hb = 13.5 Hb = 13.0 Hb = 9.8
Symptomatic MyelomaFLC Ratio = 28.7
FLC Ratio = 27.8
FOLLOW UP OF PATIENT WITH SMM
24-moTertile (mg/L) N estimate1 (0.6-107) 99 85% 2 (108-746) 102 94%3 (762-71210) 100 73%
Years
TT3: iFLC Baseline Tertiles
Eve
nt
free
su
rviv
al,
%
1 or 2 vs 3, p<0.0011 vs 2, p=0.08
0
20
40
60
80
100
0 1 2 3
SERUM FLC AT DIAGNOSIS IS PROGNOSTIC IN MULTIPLE MYELOMA
Data from Arkansas group using involved FLC and EFS after therapy
TT3: INFERIOR SURVIVAL WITH TOP-TERTILE SFLC LEVEL AT BASELINE
0%
20%
40%
60%
80%
100%
0 12 24 36Months from Enrollment
Tertile 1 (0.06-10.7 mg/dL)Tertile 2 (10.8-74.6 mg/dL)Tertile 3 (76.2-7120 mg/dL)
Deaths / N9 / 99
4 / 10222 / 100
24-MonthEstimate
89% (81,96)94% (88,100)76% (67,86)
p-value Tertiles 1 and 2 vs. 3: <0.001, 1 vs. 2=0.14
Impact of Serum Free Light Chain (SFLC) on Survival
Years
rFLC N 5-yr survival (%)“Low” 46 82±9“High” 47 30±11
Ove
rall
surv
ival
,%
rFLC
P=0.0001
0
20
40
60
80
100
0 1 2 3 4 5 6 7 8 9
Kyrtsonis MC, et al. Br J Haeml. 2007;137:240-243
SERUM FLC RATIO AT DIAGNOSIS IS PROGNOSTIC IN MM
‘High rFLC’ >3.6 ( )k or < 0.02 ( )lYears
Risk OS,score N mo 0 73 51 1 169 39 2 199 30 3 135 22
ISS & rFLC
Ove
rall
surv
ival
,%
0
20
40
60
80
100
0 2 4 6 8 10 12 14
Snozek CL, et al. Leukemia
>32 (k) or <0.03 (l)
Data using FLC ratio
KEY PROGNOSTIC FACTORS FOR PROGRESSION OF MGUS
Prognostic factor Hazard ratio(95% C.I.)
p value
Abnormal FLC ratio(<0.26 or 1.65)
2.6 <0.001
Serum M protein size 2.4 <0.001
IgA, IgM, or biclonal IgA plus IgM
2.6 <0.001
MGUS: RISK STRATIFICATION MODEL
58%4. High-risk
All 3 factors abnormal
37%3. Hi/Intermediate-risk
Any 2 factors abnormal
21%2. Low/Intermediate-risk
Any 1 factor abnormal
5%1. Low-risk Serum M protein <1.5 gm/dL, IgG subtype,
normal FLC ratio
27%
18%
10%
2%
20 yr. risk of progression after
other causes of death
Absolute risk of progression at 20 yrs.
Risk Group
Copyright ©2006 American Society of Hematology. Copyright restrictions may apply.
Dingli, D. et al. Blood 2006;108:1979-1983
Time to Progression
SOLITARY BONE PLASMACYTOMA
Overall Survival
Normal FLC ratio
Abnormal FLC ratio
Abnormal FLC ratio
Normal FLC ratio
FLC ratio is prognostic in solitary bone plasmacytoma
INVOLVED LC FLC<=60 mg/L FLC>60 mg/L
Hemoglobin(g/dL) N % N % p
<10 6 (10) 10 (26) 0.05
<11.5 11 (19) 20 (51) 0.0009
IgM (g/dL)
>4 3 (5) 13 (33) 0.0004
>7 1 (2) 3 (8) 0.30
B2M (>3 mg/L) 12 (20) 23 (59) 0.0002
WALDENSTROM MACROGLOBULINEMIA:HIGHER DISEASE BURDEN CORRELATES WITH SFLC
In WM patientsMedian sFLC > 60 mg/L was associated with lower hemoglobin and higher B2M
sFLC Lower involved FLC 20 mg/L vs. 36 mg/L (p=0.0003) in IgM-MGUS than WM patients Abnormal ratio in 23% vs. 76% (p<0.001)
Leleu et al Leuk Lymph 2008
PROGNOSTIC USES IN OTHER SITUATIONS
• Recovery of renal function in MM• Where monitoring for Light Chain elevation is
crucial– After renal transplant for Light chain deposition
disease in remission • Sensitive marker for NHL risk in HIV infection
2 -5 years prior to actual• Landgren et al J Clin Oncol. 2010 Feb 10;28(5):773-9
SERUM FREE LIGHT CHAIN ASSAY:IMPORTANT PROGNOSTIC MARKER
• Myeloma • Smoldering MM• MGUS• Amyloidosis• Plasmacytoma• Waldenstrom Macroglobulinemia
CASE 3
• 65 yr old man with symptomatic anemia• Urine light chains - >5gms in 24hrs of lambda light chains• SPEP & IFE – IgG lambda 2.2g/dl• Marrow – 62% Plasma Cells• SYMPOMATIC MYELOMA
• Treated with VAD chemotherapy with good response• Creatinine normalised• Complete Remission IFE negative
• Autologous Transplant - 10/2002• Continued in CR till 11/2003
PATIENT WITH MYELOMA NOW IN CR
• Patient presented to clinic with increasing fatigue• CBC:
– Hb 11.3, WBC 5.4, Plts 132– Creatinine 1.4 (baseline)– UPEP – no monoclonal bands– SPEP – no monoclonal protein
• Metastatic Bone Survey – NO new lesions• Serum free light chains: March 10 2006 April 3 2006
– FREE KAPPA CHAINS 12.7 mg/L 1.5
– FREE LAMBDA CHAINS 451.0 (H) mg/L 4430
– FREE KAPPA/LAMBDA RATIO 0.03 (L) 0.001
– Rising Free Light Chains – No Clinical Relapse (yet)!
No evidence of relapse by M spike or clinical criteria
CT Scan of Abdomen and PelvisCT SCAN OF CHEST/ ABDOMEN/PELVIS
Soft tissue plasmacytomas with light chain escape
TREATMENT OF LIGHT CHAIN SECRETING PLASMACYTOMAS
• Patient received D-PACE (Dexamethasone,Cisplatin, Adriamycin,Cytoxan, Etoposide) chemotherapy
• 2 cycles After cycle 1 ----------------- After cycle 2
• FREE KAPPA CHAINS <0.1 (L) mg/L too low• FREE LAMBDA CHAINS 741.0 (H) mg/L 2.7mg/L
• FREE KAPPA/LAMBDA RATIO 0.01 (L) ---• CT Scan – complete resolution of masses• FLC – resolution of involved LC elevation
Protein Half LifeIgG 20–25 daysIgA 6 daysIgM 6–8 daysFree Kappa 2–4 hoursFree Lambda 3–6 hours
Rapid clinical confirmation of effective therapy
FREE LIGHT CHAIN AS A MARKER OF RESPONSE IN MM
• Stored samples from a mature ECOG clinical trial (E9486) data• Assess serum FLC at baseline and after 2 months of alkylator
based therapy in 399 patients • FLC response after 2 months of therapy
– Superior to early M-protein measurement to predict overall response, but did not predict for survival endpoints
Dispenzieri A et al. Blood. 2008;111:4908-4915.
ROC OF FLC REDUCTION VS. M SPIKE RESPONSE
Early drop in FLC after 2 cyclesAbsolute FLC value orDelta FLC 40% decline in D FLC has :
77% sensitivity and 68% specificity for ultimate M spike reponse
Why use D FLC ?
SERUM FLC VS. UPEP 24 H FOR FOLLOW UP
Dispenzieri A et al. Blood. 2008;111:4908-4915.
Low correlation between change in M spike by UPEP and FLC changeFOR FOLLOW UP – still need 24 hr UPEP
PR CR sCR Prog
No measurable disease1
AL iFLC ↓ 50%Nl rFLC & CR by IFE & BM
NDiFLC ↑ 50% (>
100 mg/L)
MMdFLC ↓
50%Nl rFLC & CR by IFE & BM
↑ 50% dFLC
Measurable disease1
MMNl rFLC & CR by IFE & BM
1 Measurable includes serum M-protein ≥ 10 g/L or a urine M-protein ≥ 200 mg/day for myeloma patients (100 mg/day for AL patients).
Gertz MA et al. Am J Hematol. 2005;79:319-328.Durie BG et al. Leukemia. 2006;20:1467-1473.
RESPONSE CRITERIA IN MM AND AL INCORPORATE FLC
CAVEATS
• Stringent CR – not well validated for minimal residual disease and long term outcomes
• SFLC Ratio abnormal despite good remission• Oligoclonal reconstitution• de Larrea et al Blood. 2009 Dec 3;114(24):4954-6• Temporal discordance when measured as rate of drop
or rise of FLC is not concurrent with M spike– Can lead to erroneous conclusions
CONCORDANCE WITH IFE – DEPENDS ON WHEN MEASURED
Serial Measurements of FLC in MM can be discordant with immunofixation results.
For e.g – patients undergoing therapy when the FLC normalizes before IFERelapse when FLC becomes abnormal prior to IFE change
Singhal et al. Blood July 2009; 114 , 382648 serial samples from 122 patients in varying stages of therapy.Serum IFE positive in 88% samplesFLC Ratio abnormal in 62% samples
FLC Ratio discordant with FLC in 874 samples
Lower Limit for FLC Ratio
Urine ElectrophoresisNeg --------------------------------------------- Positive
Abnormal FLC ratio
SFLC ESCAPE FROM PLATEAU
• FLC escape precedes clinical relapse
• FLC Ratio abnormal 16 mo prior to UPEP turning positive
• Clinical Relapse vs. Biochemical progression
Time UPEP of positivity
Lambda LC elevation
ON AVERAGE : 6 MOS. EARLIER INDICATION OF RELAPSE THAN SIFE
6-mo Earlier Indication of Relapse
Mosbauer et al. Haematologica 2007:92:275-276
, ( m
g/L)
0
20
40
60
80
100
0 20 40 60 80 100 120
Ove
rall
su
rviv
al (
%)
Months
FLC CR? No. DeathsYes Yes 28 0Yes No 13 1No Yes 14 2No No 32 4
FLC RESPONSE IS BETTER PREDICTOR OF OS THAN IS IFE RESPONSE IN AL
Dispenzieri A, et al. Blood. 2006;107:3378-3383
SERUM SFLC ASSAY• SCREENING for PLASMA CELL DISORDERS• DIAGNOSIS of PLASMA CELL DISORDERS
• PROGNOSTIC VALUE
– Monoclonal gammopathy of undermined significance– Smoldering myeloma– Symptomatic myeloma– Plasmacytoma– AL amyloidosis
• HEMATOLOGIC RESPONSE
– AL amyloidosis– “Non-secretory” myeloma– Stringent complete response in multiple myeloma ( not fully validated)– Light chain deposition disease
• EARLY RESPONSE TO THERAPY
• RARE SITUATIONS – Free Light Chain escape – HIV and NHL– Waldenstrom
SERUM FLC ASSAY RATIONALE
• Simple test• Predicts outcome• Predicts response to therapy• Correlates with current prognostic markers• Compatible with practice
CLINICIAN’S PERSPECTIVE ON LABORATORY TESTING
• Screening
• Diagnosis
• Prognosis
• Predictive
• Monitoring
GOALS OF PLASMA CELL DYSCRASIA TESTING
• Improve predictive value of screening
• Identify candidates for biopsy
• Identify candidates to treat
• Determine optimal therapy
• Predict response to therapy