Case 4 -

Primary Systemic Lambda Light Chain Amyloidosis James Stone Massachusetts General Hospital Boston, MA

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Clinical History The patient is a 37 year-old woman who presented with dyspnea on exertion, fatigue, orthopnea, and ankle edema. Chest X-ray revealed mild cardiac enlargement. An echocardiogram revealed biatrial enlargement, moderate left ventricular hypertrophy with mild global dysfunction, mild mitral regurgitation, moderate tricuspid regurgitation, moderate pulmonary hypertension, and a left ventricular ejection fraction of 45%. Coronary angiography revealed no significant coronary artery disease. A chest CT scan showed no evidence of pulmonary emboli or pericardial thickening. A right ventricular endomyocardial biopsy was performed. Case 4 - Figure 1 - Endomyocardial Biopsy, Hematoxylin & Eosin Stain. Case 4 - Figure 2 - Endomyocardial Biopsy, Congo Red Stain. Case 4 - Figure 3 - Endomyocardial Biopsy, Congo Red Stain with plane polarized light Pathologic Findings The endomyocardial biopsy revealed deposition of amorphous eosinophilic extracellular material in a predominantly interstitial and vascular distribution. Congo red stain revealed the material to show strong orange/pink staining with unpolarized light and green birefringence with polarized light, indicative of amyloid. Utilization of an immunofluorescence panel on fresh frozen tissue demonstrated the amyloid deposits to consist of lambda immunoglobulin light chain. Diagnosis: Primary Systemic Lambda Light Chain Amyloidosis Clinical Course Bone marrow biopsy revealed 6-8% plasma cells with amyloid deposition. Serum protein electrophoresis (SPEP) and immunofixation revealed a "low concentration" IgG-lambda monoclonal component. The patient was treated with high-dose melphalan chemotherapy with autologous stem cell transplantion and also cardiac allograft transplantation. A section from the explanted heart is provided for review. Discussion The amyloidoses are a group of disorders characterized by the formation of proteinaceous amyloid deposits in tissues. An amyloid deposit is composed of one specific amyloid-prone (amyloidogenic) protein along with several non-specific proteins. Over 20 distinct types of amyloid deposits are known to occur based on the specific amyloidogenic protein present. Both prognosis and specific therapies are based on the type of amyloid deposit present in the tissue. Pathologists are now facing and will continue to face increasing pressure from our clinical colleagues to subtype the amyloid deposits observed in tissue sections. Common: Some forms of amyloidosis affect > 90% of older patients Primary AL amyloidosis - annual incidence of 9/million/year Serious: Some forms have prognoses similar to the most aggressive malignancies. Primary AL amyloidosis with cardiac involvement - survival 6 months Treatable: Immunosuppression with Melphalan and Prednisone for AL amyloidosis. Liver transplantation for Familial Amyloidotic Polyneuropathy. Structure of Amyloid Deposits Disorder of protein folding, so-called "conformational disease" A particular protein adopts an extended beta-sheet conformation. 2-5 extended beta-sheets intertwine to form an amyloid fibril. The amyloid fibrils are insoluble and are deposited in tissue. The amyloid deposit (amyloid plaque) consists of amyloid fibrils (likely composed entirely of one protein) along with several non-specifically bound proteins. Non-specific proteins, present in most amyloid deposits include: Serum Amyloid P (SAP), Proteoglycans, Apolipoprotein E, Collagen IV Classification of Amyloidosis Classification based on the identity of the specific protein forming the amyloid fibrils. Over 20 such specific proteins now known. Systemic vs Localized Some localized forms have little to no clinical significance Acquired vs Hereditary Different clinical presentation Senile Systemic Amyloidosis vs Familial Amyloidotic Polyneuropathy Both result from transthyretin Impact on treatment options Factors That Trigger Amyloid Deposition Inheritance of an amyloidogenic mutation. Transthyretin - Familial Amyloidotic Polyneuropathy Cystatin C - Cerebral Amyloid Angiopathy Overproduction or accumulation of an amyloidogenic wild-type protein Serum Amyloid A - Chronic inflammatory conditions Beta-2 microglobulin - Chronic hemodialysis Ig light chains - monoclonal gammopathy An initial amyloid focus Deposition of wild-type transthyretin amyloid in the heart following liver transplantation for hereditary amyloidosis Infection Prion Diseases Animal model using serum amyloid A Unknown / Multifactorial Senile Systemic Amyloidosis Diagnosis of Amyloidosis H&E: Amorphous eosinophilic material May be difficult to distinguish from collagen Trichrome: Heterogeneous, grungy blue staining Modified Sulfated Alcian Blue: Amyloid-green, Collagen-red, Cells-yellow Useful for autopsies with extended post-mortem intervals Not entirely specific as mucins will stain green Congo Red: Orange or pink staining (intensity helpful but not diagnostic) Enhanced (over H&E stain) green birefringence with polarized light (considered diagnostic for amyloid) Primary Systemic Ig Light Chain ( AL ) Amyloidosis Accounts for 80% of systemic amyloidosis in North America 12 to 15% of patients with myeloma develop amyloidosis. 90% of Patients with AL amyloidosis have a monoclonal gammopathy. May present locally in association with plasmacytoma lambda light chains are more amyloidogenic than kappa light chains (3:1). Particular gene rearrangements are more highly associated with amyloid formation. Almost any organ can be involved. Treated with immunosuppression, stem cell transplantation, and cardiac transplantation. When cardiac involvement present, survival ~6 months. Senile Systemic Amyloidosis Deposition of wild-type transthyretin Etiology Unknown 25% of people over age 80 at autopsy Clinically significant involvement much less common Accounts for ~20% of cases of amyloidosis diagnosed by endomyocardial biopsy Most patients diagnosed on biopsy alive after 2 years. Heart most severely involved, lungs frequently involved, and subtle involvement of blood vessels in other organs. Formerly Senile Cardiac Amyloidosis Sub-typing Amyloid Deposits Establishing the Presence of a Monoclonal Gammopathy (SPEP / Immunofixation) ~10% of patients with a monoclonal gammopathy and systemic amyloidosis actually have a hereditary amyloidosis, mostly either fibrinogen or transthyretin. Monoclonal gammopathies are not uncommon in the elderly and most are not associated with amyloid deposition. For many of the Hereditary Amyloidoses, multiple mutations have been observed in the culprit protein, some of which are of low penetrance and late onset. Over 80 amyloidogenic mutations in transthyretin identified. Ile122 mutation present in 4% of American blacks. Newer Serum Free light Chain Analyses may improve sensitivity and specificity over traditional SPEP/immunofixation. Direct Sub-Typing of Amyloid Deposits with Immunologic Approaches Immunohistochemistry (IHC), Immunofluorescence (IF), Immunoelectron microscopy (IEM) Pitfalls and challenges of immunologic techniques Not widely utilized with cardiac tissue. Very high background staining leading to high false positive rates (IHC). Some are prohibitively expensive for routine diagnostic use (IEM). The final processed amyloid fragment may lack the antigenic site leading to false negatives. Clinical application of an immunofluorescence panel for the subclassification of amyloid in endomyocardial biopsy specimens. Ancillary Approaches for Sub-Typing Amyloid Deposits Genetic approaches May be useful for hereditary amyloidoses. An indirect approach that does not definitively subtype the amyloid deposits in the tissue. Biochemical approaches Amyloid fibril extraction and identification by western-blotting and/or sequencing possible but laborious. Newer mass spectrometry technologies promising for direct identification of amyloid components. Hurdles to achieve routine accurate quantitation. Table 1: Systemic Acquired Amyloidoses Amyloidosis Culprit Protein Clinical Setting Organs Involved Primary Systemic (AL) (AH) Immunoglobulin light chains (rarely heavy chains) Monoclonal gammopathy, myeloma Kidney, Heart > other organs, most organs affected Senile Systemic (ATTR) Transthyretin 25% of people over age 80, occasionally clinically significant Heart, Lungs, and blood vessels Reactive / Secondary (AA) Serum Amyloid A Chronic infections or chronic inflammatory diseases Spleen > Liver > other organs ABeta2M Beta2-Microglobulin Chronic Hemodialysis Joints, blood vessels, heart Table 2: Systemic Hereditary Amyloidoses Protein Disease, Organs Affected Transthyretin Over 80 Mutations known Val-122-Ile, 4% American blacks, late onset Val-30-Met Familial Amyloidotic Polyneuropathy, N. Europeans Heart, Kidney, Nerves, other (depending on specific mutation). Apolipoprotein A-I Liver, kidney, heart Apolipoprotein A-II Kidney, heart Gelsolin Finnish Hereditary Amyloidosis, Lattice Corneal Dystrophy Type II, Cornea, nerves Lysozyme Kidney > other organs Fibrinogen alpha chain Kidney > other organs Cystatin C Icelandic Hereditary Cerebral Amyloid Angiopathy Table 3: Localized Acquired Amyloidoses (Selected) Organ Protein Features Cardiac Atria Atrial Natriurtetic Factor (ANF) 80% autopsies over age 80 16% of resected atrial appendages Cardiac Valves unknown 50% of resected valves Significance uncertain Aortic Media Lactadherin / medin >90% people over age 50, Significance uncertain Islets of the pancreas Islet Amyloid Polypeptide / Amylin >90% of patients with type 2 diabetes, Insulinomas Significance uncertain Thyroid Calcitonin Associated with medullary carcinoma Brain Prion Protein CJD, transmissible Brain A-beta protein sporadic Alzheimer's disease Table 4: Localized Hereditary Amyloidoses Organ Protein Features Brain A-beta protein Cerebral Amyloid Angiopathy, Dutch type Familial Alzheimer's disease Brain Prion Protein Familial CJD Brain ABri Protein British Familial Dementia Cornea Beta-ig-h3 Lattice Corneal Dystrophy, type I References Merlini G, Bellotti V. 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