New approaches in amyloidosis. Philip Hawkins National Amyloidosis Centre UCL & Royal Free Hospital, London

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1 New approaches in amyloidosis Philip Hawkins National Amyloidosis Centre UCL & Royal Free Hospital, London

2 Systemic Amyloidosis Fatal protein misfolding/aggregation disease caused by accumulation of fibrillar amyloid deposits in multiple organs Classified according to amyloid fibril protein SAA (AA), monoclonal Ig light chain (AL), transthyretin (ATTR) Causes at least 1 per 1000 deaths in UK Usually acquired - Most often AL type complicating subtle monoclonal gammopathy age >25 yr, usually >50 yr; M=F; kidneys, heart etc (>500 pts/yr) - Wild-type cardiac ATTR catching up: senile from 55yr; M>>F (>200 pts/yr) - AA amyloidosis complicating chronic inflammatory disorders increasingly rare But 5-10% hereditary, often without family history - Variant ATTR type most common (heart, nerves) - 1 in 20 black individuals possess TTR V122I variant (heart) - Many other rare familial types / proteins / mutations (kidneys, liver etc)

3 Types of amyloidosis in UK 7747 patients WT TTR, 624, 8% LECT 2, 28 Localised AL, 851, 11% Insulin 16, 0% AA, 810, 11% TTR, 563, 73% ApoA1, 48, 6% Hereditary, 772, 10% Fibrinogen A alpha chain, 122, 16% Gelsolin, 17, 2% Lysozyme, 22, 3% AL, 4646, 60% Hereditary All cases, 90% acquired >800 new amyloidosis patients per year currently seen at NAC

4 Clinical pathway Awareness, especially of senile cardiac ATTR type Red flags for cardiac amyloidosis Heart failure without cardiomegaly HFpEF, especially in males over 60 yr LVH on echo without a clear cause; ECG voltage may be normal or large in amyloid Periorbital purpura, macroglossia, neuropathy, proteinuria (AL) Heart failure in black patients >55 years (ATTR V122I variant; 1 in 20 possess gene) Heart, renal dysfunction etc in patients with paraprotein / MGUS (present in >10% over 70 yr) Recent CMR findings: asymmetric hypertrophy, traditionally associated with HCM, is the commonest pattern of ventricular remodelling in ATTR amyloidosis. Imaging vs traditional need for histology Investigation of underlying disorders, eg MGUS Typing, staging and monitoring the amyloid deposits Treatment pathways

5 New approaches Diagnosis Genetic testing Mass spectrometry / proteomics Imaging - DPD scintigraphy, Cardiac MRI (CMR) Treatment To reduce production of amyloid precursor proteins Complex chemotherapy options for AL type, adopting and evaluating plethora of new myeloma therapies RNA inhibitors for ATTR type Cytokine inhibitors for AA type To accelerate the otherwise very slow natural clearance of amyloid Anti-amyloid monoclonal antibodies

6 Amyloidosis 2 day diagnostic workup at NAC History include autonomic symptoms, bowel disturbance Clinical examination macroglossia, neuropathy, bruising etc 6 minute walk test; Postural blood 5 minutes Biomarkers of underlying clonal / inflammatory disease (FLC, immunofixation etc; SAA) track response to treatment Family history enquiry, low threshold for genetic testing Biomarkers of amyloidotic organ dysfunction (NTpro-BNP & Troponin T to stage, renal & liver function etc) Retrieve any previous histology samples for Congo red staining Immunohistochemistry & proteomics for amyloid fibril typing Echo, ECG, multiparametric cardiac MRI SAP and DPD (cardiac) scintigraphy Liver elastography Impedance measurements Consultation / MDT / management plan

7 123 I-labelled SAP scintigraphy Identify, quantify and monitor amyloid load in large solid organs AL AA AFib AGel

8 NAC histology experience Congo red stain for amyloid Carpel tunnel 2% SKIN 4% Soft Tissue 2% Lung 3% Head and Neck 3% Cardiac 7% Renal/ urology 19% GI 11% Spleen /Lymph node 1% BMT 21% Fat Aspirates 21% Connective Tissues/muscle /fat 2% Fat aspirates 329 Renal 244 Bone marrow 317 Cardiac 70 Rectal 50 Skin 58 Other 216 Total 1284 cases More than half external pts Using NAC Congo Red stain as the gold standard, referred biopsies have: False positive rate 24% overall False negative rate 12% overall

9 Diagnostic pitfalls Presence of plasma cell dyscrasia does not prove amyloid is of AL type 10-20% of older population have a clonal plasma cell dyscrasia using the sensitive assays required in work-up of AL amyloidosis Immunohistochemistry has limitations 30% AL cases cannot be definitively confirmed Variable, often low penetrance in hereditary amyloidosis there is often no family history

10 Proteomics - Laser dissection microscopy and mass spectrometry Amyloid fibril protein identified by mass spectrometry Presence of amyloid corroborated by presence of 2 of 3 signature proteins SAP, ApoE, ApoA4, which are universal nonfibrillar constituents of amyloid. Important development Tiny amounts of tissue required

11 Fibril protein and amyloid signature non-fibrillar protein constituents Mascot data

12 Cardiac amyloidosis Almost always AL or ATTR type Restrictive cardiomyopathy, preserved ejection fraction ATTR senile cardiac type under-diagnosed

13 Echocardiography in amyloidosis A thick walled heart, but how much of it is amyloid? Strain Tracks the longitudinal movement of the heart muscle. Apical sparing typical in amyloid.

14 Cardiac MRI in amyloidosis CMR provides more accurate measurements of volume, mass and wall thickness than echocardiography Enables myocardial tissue characterisation Characteristic patterns of late gadolinium enhancement (LGE) T1 mapping through the measurement of the native (non-contrast) myocardial T1, post-contrast T1 and extracellular volume (ECV) T1 (longitudinal relaxation time) markedly elevated in the presence of cardiac amyloid Evaluation of perfusion coronary amyloid angiopathy and more

15 Cardiac MRI: Diffuse transmural late gadolinium enhancement in a patient with cardiac ATTR amyloidosis

16 EELGE in Q-CMR Equilibrium contrast MRI to measure ECV Gd Gd Gd Gd Vd (b) Gd Gd Vd (m) Gd Gd in blood Eequal

17 ECV elevated in disease: fibrosis and amyloid Sado DM, Heart 2012

18 Repurposed bone scintigraphy: Tc-DPD scan in cardiac ATTR amyloid Negative Positive Fused SPECT/CT image

19 Grading 99m Tc-DPD uptake in the heart Distinction between Grade 1 and Grade 2 is key Grade 1 Grade 2 Grade 3 Bone Cardiac Grade 1 = Mild cardiac uptake, less than bone; no attenuation of bone uptake Grade 2 = Moderate cardiac uptake equal or greater than bone plus some bone attenuation Grade 3 = Strong cardiac uptake with little or no bone signal

20 99m Tc-DPD scintigraphy in systemic AL amyloidosis 189 patients All had cardiac involvement Grade 1 57 (76%) All patients 189 DPD Positive 75 DPD negative 114 Grade 2 13(17%) Grade 3 5(7%) A quarter of positive scans in AL amyloidosis have grade 2 uptake

21 Heart failure with echocardiogram and/or cardiac magnetic resonance imaging (CMR) suggesting/indicating cardiac amyloid Bone scintigraphy with DPD Grade 0 Grade 1 Grade 2 to 3 Serum immunofixation + Urine Immunofixation + serum free light chain assay No Yes Yes No Yes No Cardiac Amyloidosis unlikely Cardiac AL Amyloidosis Histological confirmation and typing required Need specialized assessment for Diagnosis : Histological confirmation and typing of amyloid Variant ATTR Cardiac ATTR amyloidosis TTR genotyping Wild-Type TTR

22 The database is accessible at Dr Dorota Rowczenio

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25 Treatment strategies in amyloidosis Precursor protein Reversion to normally folded protein Amyloid Fibril formation Stabilise amyloidforming proteins Reduce supply of amyloid precursor protein β sheet breakers Enhance removal of existing amyloid Immunotherapy SAP depletion

26 Increasing choice of chemotherapy regimens for AL amyloidosis Options Oral Melphalan-Dex Cyclo-Thal-Dex Velcade based regimens Cyclo-Vel-Dex Lenalidomide containing regimens High dose melphalan with autologous stem cell rescue (~20% of pts) Anti-CD38 and new IMiDs & proteasome inhibitors in trial Factors to consider Organ involvement, stage Speed of clonal response Depth of clonal response Duration of clonal response Anticipated side effects Anticipated treatment related mortality Response to previous treatments Aim is sustained >90% reduction in serum free light chain

27 Survival is improving in AL amyloidosis Bortezomib Revlimid Thalidomide

28 Amyloid can regress slowly when amyloid precursor protein supply is substantially suppressed, but not in the heart? AA AL Hawkins et al. Arthritis Rheum 1993;36: Hawkins et al. N Engl J Med 1990;323:

29 Cardiac MRI follow-up after chemotherapy in AL amyloidosis 57 yrs man with cardiac AL amyloidosis Chemotherapy achieved sustained complete remission (100% FLC response) Baseline: EF 55%, strain -13%, NTpro-BNP 1100, ECV 47% 1 yr Follow-up: EF 65%, strain -23%; NTpro-BNP 264, ECV 32% (normal) No change in wall thickness or mass more muscle, less amyloid

30 Monoclonal antibody therapies for systemic amyloidosis Several different antibody therapy approaches in development Targeting underlying clonal disease in AL Daratumumab - human monoclonal anti-cd38 90-Yttrium-labelled anti-cd66 marrow conditioning for autologous stem cell transplantation Targeting the amyloid deposits Chimeric fibril-reactive antibody 11-1F4 in AL Amyloidosis Prothena NEOD001 for AL Amyloidosis targeting misfolded light chain proteins Phase III GSK/NAC CPHPC + anti-sap mab targeting the amyloid bound SAP present in all amyloid deposits Phase II

31 Anti-SAP Day 1 post antibody (in mice) Congo red F4/80 (global macrophage marker)

32 Clearance of AL amyloid 6 weeks after anti-sap therapy Major regression of liver AL amyloid 6 weeks after one course of CPHPC + anti-sap 3-D imaging enables quantification of tracer uptake Reduction in liver stiffness and ECV on MRI

33 Phase II CPHPC + anti-sap in cardiac amyloidosis ATTR and AL cardiac amyloidosis Repeat dosing x 6 cycles over 6 months Multiparametric cardiac MRI endpoints to measure clearance of amyloid

34 Prospects for treatment of ATTR amyloidosis Liver transplantation for patients with early symptoms associated with TTR Met30. Amyloidogenic property of wild-type TTR limits this approach in patients with cardiac involvement (i.e. most patients!) TTR stabilizing drugs (diflunisal, tafamidis) show some promise, though limited clinical efficacy demonstrated to date Reducing production of TTR Silencing RNA therapy (Alnylam) Antisense oligonucleotide therapy (Ionis) Reduction of TTR synthesis in liver by >80% Phase III completed halts / reverses neuropathy

35 Patisiran 3 weekly i.v. sirna open label study in hereditary ATTR amyloidosis Sustained ~80% TTR knockdown % Mean TTR Knockdown Relative to Baseline (SEM) Study Day N Available to Graph S Dose 1 (pre- and post-dose) Dose 3 (pre-dose) Dose 5 (pre-dose) Days Since First Visit Dose 9 (pre-dose)

36 Summary New era of: Awareness, esp cardiac ATTR senile type Diagnostic non-invasive imaging Novel therapies to reduce amyloid protein production Antibody therapies to enhance amyloid clearance High cost drugs to deal with!