A 67-year-old male patient presented to Luohu People's Hospital of Shenzhen (Shenzhen, China) in August 2021 with the chief complaints of chest tightness, nocturnal dyspnea and a paroxysmal cough persisting for half a month. The patient had no family history of hematologic disorders or amyloidosis. Past medical history included hypertension (controlled) and no psychosocial risk factors. Between January 2019 and July 2021, the patient had sought medical attention at an external hospital (Shenzhen People's Hospital, Shenzhen, China) due to chest tightness and nocturnal dyspnea and was diagnosed with ‘chronic heart failure’. Prior interventions for heart failure in 2019 included diuretics, spironolactone tablets, furosemide tablets and sacubitril valsartan sodium tablets for 2 years, which provided transient symptom relief but no improvement in cardiac function. The patient suffered from the symptoms of chest tightness and orthopnoea at night, accompanied by paroxysmal cough, as well as numbness in the toes.

The patient also had symptoms such as difficulty breathing, edema, fatigue and atrial fibrillation in chronic heart failure. However, the patient's symptoms of chronic heart failure were different from those caused by traditional coronary heart disease and hypertension. The patient presented with the following: i) Postural hypotension and diastolic dysfunction; ii) special cardiac signs: a) Thickened ventricular wall but relatively preserved systolic function (ultrasound showed ‘granular’ myocardial echoes); b) low voltage on electrocardiogram (related to myocardial amyloid infiltration); iii) multiple systemic involvement manifestations (systemic deposition of amyloid protein): Peripheral neuropathy; numbness and pain in hands and feet; renal dysfunction and elevated creatinine levels; iv) poor response to traditional heart failure treatment: According to disease feedback, diuretics have limited efficacy in treating current patients Beta blockers and angiotensin-converting enzyme inhibitors are also not suitable as they can exacerbate hypotension. The main pathological manifestation of traditional chronic heart failure is myocardial systolic and diastolic dysfunction (13). Cardiac amyloidosis is an infiltrative cardiomyopathy and mainly characterized by amyloid deposition leading to myocardial stiffness and affecting diastolic function (14). Therefore, examinations for cardiac amyloidosis were necessary.

The blood routine examination indicated a white blood cell count of 5.68x10⁹/l, platelet count of 184x10⁹/l and hemoglobin concentration of 136 g/l, all within normal ranges. Cardiac enzyme tests revealed cardiac stress with a creatine kinase level of 48 U/l and an N-terminal pro-brain natriuretic peptide II level of 8,288 pg/ml. Cardiac infarction markers indicated myocardial injury with a troponin T level of 0.154 ng/ml and a myoglobin level of 96 ng/ml. The renal function panel showed a creatinine level of 256 µmol/l, a uric acid level of 521 µmol/l and a β2-microglobulin concentration of 14.7 mg/l, indicating impaired renal function. The Ig profile showed abnormal levels with results of IgG 19.5 g/l, IgA 0.60 g/l and IgM 9.98 g/l (Table I). Serum protein electrophoresis (Fig. S1; Data SI; Table SI) revealed albumin (ALB) at 49.2%, β1 globulin (β1G) decreased to 4.5% and β2G at 19.2%. Immunofixation electrophoresis (Data SI; Table SII) showed positive IgM-λ type M protein, an erythrocyte sedimentation rate of 59.0 mm/h, negative Bence Jones protein and a free light chain ratio (κ/λ ratio) of 0.26, with free κ and λ light chains at 132 and 507 mg/l, respectively. The urine analysis showed no protein and the 24-h urine protein measurement was 93.6 mg, which was within the normal range. The electrocardiogram showed low voltage in the limb leads, suggesting cardiac abnormalities (Fig. S2). Genetic mutation detection (Fig. S3; Data S1; Table SIII) was tested by droplet digital polymerase chain reaction based on droplet separation and single-molecule amplification techniques, which identified the presence of MYD88 innate immune signal transduction adaptor (MYD88) with positive L265P mutation with a mutation frequency of 2.89%. Noteworthy abnormalities included cardiac stress, myocardial injury, impaired renal function, abnormal Ig levels, positive IgM-λ type M protein, significant proteinuria and low voltage in the limb leads.

The echocardiography images (Fig. 1A and B) revealed left ventricular posterior wall hypertrophy with a thickness of 15.6 mm and interventricular septum of bilateral atrial enlargement with a thickness of 13.3 mm, which were both higher than normal values (both <12 mm). The thickened myocardium exhibited scattered sparkling granular echoes and a small amount of pericardial effusion. The heart valves were thick and rough and regurgitated. Fig. 1B shows that the left ventricular fractional shortening ratio was 22%, which is lower than the normal value (25-35%). The broadened main pulmonary artery diameter resulted in high pulmonary artery pressure. Fig. 1C shows that the left ventricular ejection fraction value of cardiac function was 44.3%, which is lower than the normal value (>50%). Fig. 1D shows that the left ventricular wall motion amplitude was slightly lower than normal, with a peak value of 61.6 cm/sec (normal range, 67-109 cm/sec). The aforementioned findings suggested that the left ventricular diastolic and systolic function were reduced, resulting in mild heart failure.

The bone marrow morphology (Data SI; Table SIV) revealed active proliferation with an increased proportion of lymphocytes, as shown in Fig. 2. Certain lymphocytes exhibited plasma-like differentiation, while lymphocytes and plasma-like lymphocytes exhibited focal aggregation. The lymphocytes were predominantly mature, with ~10% identified as abnormal lymphocytes. The abnormal lymphocytes had a small size, dense nuclear chromatin and a small cell mass, and part of the cell cytoplasm exhibited blue, foam-like, plasma cell-like changes or irregular cytoplasm edges with burrs. The proportion and morphology of erythrocytes were normal. Platelets were distributed in single or small clusters and were easily visible with a small number of abnormal platelets. The proportion of plasma cells was about 4%, which were mature plasma cells.

The flow cytometry of bone marrow (Data SI; Table SV) results showed that lymphocytes constituted 13.72% of nucleated cells (normal range, 10-20%), with T lymphocytes comprising 49.69% (normal range, 60-80%). The CD4+/CD8+ ratio was 0.37 (normal range, 1.4-2.0), reflecting a decreased ratio, yet no discernible phenotypic abnormalities were apparent. Among the lymphocytes, the proportion of CD19+ B cells was 32.78% (normal range, 5-15%; Fig. S4), exhibiting a phenotype characterized by CD19+, CD20+, Lambda++, Kappa-, CD5-, CD10-, CD3-, GD4-, CD8-, CD2- and CD7-. Granulocytes accounted for 68.96% (normal range, 50-60%), with a proportion within the normal range, and no notable abnormalities were observed in the proportion and phenotype of granulocytes at various stages. The CD34+ and CD117+ marrow cells had a low proportion of 0.47% (normal value, <1%), and no apparent phenotypic abnormalities were observed. Notably, abnormal mature B cells, displaying immunophenotypic abnormalities, made up ~4.94% (normal value, 0%) of nucleated cells. B-cell lymphoma excluding CD5- and CD10-could not be conclusively ruled out, necessitating further investigation to establish a definitive diagnosis.

Fig. 3A shows the bone marrow biopsy with immunohistochemistry and Congo red staining (Data S1), which was completed through biopsy of the posterior superior iliac spine. The results indicate that the proportion of red bone marrow decreased, the proportion of granulocytes, red cells and megakaryocytes in the red pulp was normal, and the morphology of each cell was relatively mature, with a few scattered plasma cells. In typical amyloidosis cases, amyloid substance is an amorphous extracellular eosinophilic substance, which is commonly identified by the normal Congo red staining method because with other staining, it is difficult to distinguish colors. The Congo red staining was positive, displaying in orange red, which indicated the existence of amyloid substance. Amyloid protein mass spectrometry was performed on abdominal fat biopsy (Congo red-positive) with 84% sensitivity (15), which is a recommendation surrogate biopsy tissue for most patients, and confirming AL type with predominant Ig λ C2 spectra (12%). Cardiac tissue biopsy was not feasible due to the patient's unstable condition. Fig. 3B shows the biopsy of abdominal wall fat tissue with Congo red staining; an apple green color originating from amyloid substance with double refraction was visible under a polarized light microscope, which was transformed through the amyloid substance. Therefore, the Congo red staining with orange red and apple green color transformation both confirmed cardiac amyloidosis in this case.

The amyloid protein mass spectrometry spectrum of abdominal adipose tissue was used to determine the amyloidosis type (Data S1), as shown in Fig. 4. Fibrinogen α chain (Fibα), the Ig light chain constant (Igλ C2), the Ig heavy chains Igµ, Igγ1 and Igα C1, Ig light chain κ C, lysozyme (lys) and gelsolin (gel) were detected. Among the currently known sub-typing proteins in Fig. 4, Fibα has the highest relative abundance in spectra, but Fibα<β+γ indicates that Fibα comes from blood contamination (α 218, β 294, γ 188) and is not a sedimentary protein (16). Compared with heavy chains, light chains (especially λ type) are more likely to form amyloid fibrils due to their genetic sequence and high secretion characteristics (13). Thus, light chains are more prone to deposit in the cardiac position, leading to cardiac amyloidosis. Excluding Fibα, among other typing proteins, Igλ C2 had the highest relative abundance in the spectrum, κ and λ values are significantly higher than the normal range (Table I), indicating that the main type is AL amyloidosis. Therefore, based on the positive Congo red staining and abdominal adipose tissue mass spectrometry analysis, the diagnosis was cardiac amyloidosis of the AL type.

Based on the comprehensive characteristics of the case and auxiliary examination results, the diagnosis aligns with the criteria for LPL/WM, IgM-λ type, high-risk, accompanied by cardiac AL amyloidosis. The patient's prior diagnosis of chronic heart failure made in 2019 was attributed to non-specific causes, and no amyloidosis or hematologic malignancy was suspected until 2021, which highlights the importance of considering amyloidosis in refractory heart failure. The diagnosis of WM was confirmed through abnormal IgM (9.98 g/l) levels, λ-free light chains (507 mg/l), the existence of small B lymphocytes, plasma cell-like lymphocytes, plasma cells, M protein and MYD88 with L265P mutation. The cardiac amyloidosis was identified through cardiac biomarkers (N-terminal pro-B-type natriuretic peptide: 8,288 pg/ml), cardiac echocardiography (sparkling echoes), positive Congo red staining of bone marrow and apple green color transformation of abdominal fat biopsy, and mass spectrometry peaks of Ig λ C2 and µ heavy chain as the dominant amyloid protein with AL type.

In August 2021, the patient initiated the BR regimen, which consisted of Bendamustine (0.15 g intravenously on days 2-3) and Rituximab (0.6 g intravenously on day 1), following a 28-day chemotherapy cycle. A second course of the BR regimen was administered in September 2021, maintaining the same regimen and dosage as the initial course. The disease was subsequently evaluated as stable disease. In October 2021, the patient underwent a third-line treatment with the BRD regimen. This regimen included subcutaneous injections of 2 mg bortezomib on days 1, 8, 15 and 22, intravenous infusion of 0.5 g rituximab on the same days, and intravenous infusion of 10 mg dexamethasone on days 1, 8, 15 and 22, with treatments administered once a week within a 28-day chemotherapy cycle. After the treatment, the patient was determined to show a partial response (PR). The patient adhered to treatment with no adverse events and at the six-month follow-up, the patient showed sustained hematological PR but progressive diastolic dysfunction on echocardiography. The patient voluntarily left the hospital due to economic difficulties. The PR was mainly based on hematological criteria: >50% reduction in serum IgM (from 9.98 to 4.79 g/l), >50% reduction in the difference of serum free light chain (dFLC) and a decrease of cardiac injury markers (troponin T: 0.119 ng/ml). Cardiac function (ejection faction: 50%) remained stable but did not improve due to irreversible amyloid deposition. The follow-up was terminated as the patient did not return to the hospital for treatment after discharge.