Jerry A' Katzmann, Ph'D' Division of Clinical Biochemistry

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Jerry A. Katzmann, Ph.D. Division of Clinical
Biochemistry ImmunologyDepartment of Lab Med
PathSeptember 18, 2008

• Serum FLC Quantitation Impact on Urine Testing

Jacksonville, Florida
Scottsdale, Arizona
Rochester, Minnesota
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Serum Protein Electrophoresis Immunofixation
Electrophoresis
Normal Serum
Polyclonal Hypergammaglobulinemic Serum
Multiple Myeloma Serum
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Serum Protein Electrophoresis Immunofixation
Electrophoresis
Light Chain MM Serum
Light Chain MM Urine
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Monoclonal Serum ProteinsNewly Diagnosed
Multiple Myeloma (n1027)
Kyle et. al., Mayo Clin Proc 2003
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(No Transcript)
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PEL Need for Serum Urine Samples
SERA
URINES
MGUS with Serum M-spike
MM with Serum M-spike Urine M-spike
LCMM with Urine M-spike
AL with no Serum or Urine M-spike
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Free Light Chain Assay Specificity
The Binding Site
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FLC reference range k 3.3 19.4 mg/L l 5.7
26.3 mg/L k/l ratio 0.26 - 1.65
100,000
Normal lt70 yrs
10,000
Normal gt70 yrs
1,000
Kappa
LCMM
Lambda
100
LCMM
Serum FLC, lambda, mg/L
Nonsecretory
myeloma
10
Polyclonal
1
1
10
100
1,000
10,000
100,000
Serum FLC, Kappa, mg/L
Drayson, et al, Blood, 2001
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Dysproteinemias FLC(Monoclonal Gammopathies)

• Diagnosis
• Monoclonal vs. polyclonal
• Serum PEL, IFE, FLC (NSMM, AL, LCDD)
• Prognosis
• Progression
• MGUS M-spike size (PEL), isotype (IFE), FLC
  ratio
• SMM M-spike size (PEL), BMPC, FLC ratio
• Plasmacytoma M-spike persistence (PEL), FLC
  ratio
• Survival
• MM b2M, Albumin, FLC ratio
• Disease Monitoring
• If serum and urine M-spikes are not measurable
  (lt1 g/dl lt 200 mg/24 h respectively), use FLC
  concentration if ratio is abnormal and
  Involved FLC gt10 mg/dL
• Empirical recommendation
• use involved FLC minus uninvolved FLC

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Serum FLC Quantitation Impact on Urine Testing

• Are urine studies needed for initial monoclonal
  gammopathy screen?
• When doing urine studies, is sample concentration
  necessary?
• Can serum FLC be used for disease monitoring (and
  how should it be interpreted)?

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Prospective Review of 1020 sequential patients
PEL, IFE, FLC
Diagnosis n abn FLC MM (intact Ig)
330 95 SMM 72
88 MGUS 114
44 NSMM 20 70 LCDD
7 100 AL 110
91
Katzmann et al, Clin Chem, 2005
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Diagnostic Sensitivity in AL
110 sequential, newly diagnosed AL pts with
orders for serum PEL, IFE, FLC plus urine PEL
IFE
Assay (s) Sensitivity Serum IFE
69 Urine IFE 83 Serum IFE Urine
IFE 95 Serum FLC k/l Ratio
91 FLC k/l Ratio Urine IFE 91 FLC
k/l Ratio Serum IFE 99 All 3 assays
99
Katzmann et al, Clin Chem, 2005
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Diagnoses for 428 Patients Selected for Urine
Monoclonal Protein
All 428 patients are untreated have a positive
urine IFE, and concurrent serum assays for PEL,
IFE, FLC
Diagnosis of pts. () MM
148 (34.6) Primary Amyloid 123
(28.7) MGUS 69 (16.1) Smoldering
Myeloma 59 (13.8) Plasmacytoma 10
(2.3) Osteosclerotic Myeloma 5
(1.2) Macroglobulinemia 3
(0.7) Lymphoproliferative Disease 4
(0.9) Light Chain Deposition Disease 2
(0.5) Smoldering Macroglobulinemia 2
(0.5) Plasma Cell Leukemia 2
(0.5) Cryoglobulinemia 1 (0.2)
Katzmann, et al, Mayo Clinic Proc, 2006
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Serum PEL IFE Results 428 Patients with
Monoclonal Urinary Protein

• Lab Test Abnormal ()
• Urine IFE 428 (100)
• Serum PEL 346 (80.8 )
• Serum IFE 400 (93.5)
• Serum PEL/IFE missed 28 pts (6.5)
• Primary Amyloid, n 19
• Multiple Myeloma, n 2
• Plasmacytoma, n 3
• Smoldering MM, n 1
• MGUS, n 3

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Laboratory Results 428 Patients with Monoclonal
Urinary Protein

• Lab Test Abnormal ()
• Urine IFE 428 (100 )
• Serum IFE 400 (93.5 )
• Serum FLC K/L ratio 367 (85.7 )
• Serum PEL/IFE or FLC ratio 426 (99.5)
• 1 Idiopathic lambda Bence Jones Proteinuria (?FLC
  MGUS), urine M-spike 83 mg/24 hrs
• 1 MGUS (?sample contamination urine IgA kappa,
  serum neg, subsequent urine neg)

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Monoclonal Gammopathies Diagnostic Screening
Algorithm

• Medicare cost
• serum FLC 38.00
• urine PEL/IFE 71.04
• Theoretical savings 33
• Because only a fraction of the serum samples also
  have an accompanying urine sample, this savings
  will not be fully realized.
• The diagnostic sensitivity should increase with
  the serum FLC as a surrogate for urine IFE.

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Urine Diagnostic Tests Sample Concentration

• To achieve high sensitivity, current protocols
  concentrate urine up to 200X (depending on
  protein content)
• Our Lab rejects 3-4 of all urine samples due to
  inadequate volume to allow specimen concentration
• The Sebia UPEP and IFE protocols for
  unconcentrated urine is 96 sensitive compared to
  our concentrated urine protocol

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Urine IFE Concentrated and Unconcentrated Samples

• Protein Conc. (mg/dL)
• 0-16 17-50 gt50
• Concordant
• Positive (n53) 20 14 19
• Negative (n56) 34 2 20
• Discordant 2 0 0

Roden, A., et al, AJCP, in press
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Urine M-Spike Comparison Concentrated vs.
Unconcentrated
Unconcentrated
Concentrated
Roden, A., et al, AJCP, in press
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Urine Testing

• If serum PEL, IFE, FLC are the initial
  algorithm, urine studies not required as part of
  the screen
• Urine studies for urine TP, protein distribution,
  and M-spike quantitation are still needed for
  final diagnosis and monitoring but these do not
  require concentrated urine

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Disease MonitoringPublished Response Criteria
for FLC
PR Partial Response, CR Complete Response, sCR
stringent Complete Response 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. 200579319-328. Dur
ie BG et al. Leukemia. 2006201467-1473.
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Appraisal of Immunoglobulin Free Light Chain as a
Marker of Response

• Data and stored samples from a mature ECOG
  clinical trial (E9486) data set to assess serum
  levels of immunoglobulin free light chains FLC at
  baseline and after 2 months of alkylator based
  therapy in 399 patients
• FLC response after 2 months of therapy was
  superior to early M-protein measurement to
  predict overall response, but did not predict for
  overall or progression free survival

Dispenzieri A et al. Blood. 20081114908-4915.
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Best-fit Early FLC Reduction for Overall
Hematologic Response
N317
Dispenzieri A et al. Blood. 20081114908-4915.
ECOG 1988-1992, n399
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Serum FLC Quantitation Impact on Urine Testing

• Diagnosis
• Monoclonal vs. polyclonal
• Serum PEL, IFE, FLC eliminate urine from
  clonality screening
• Prognosis
• Disease Monitoring
• No need to concentrate urine
• If serum and urine M-spikes are not measurable
• dFLC (involved FLC minus uninvolved FLC)
• 50 reduction is best cut-point for response
• Serum FLC needed in addition to serum/urine IFE
  for stringent CR in MM