Abstract
Background
Early life‐threatening cardiotoxicity and cardiac death have been reported after hematopoietic stem cell transplantation (HSCT). The purpose of the current study was to evaluate cardiac toxicity of conventional chemotherapy followed by HSCT with cardiac markers: heart‐type fatty acid binding protein (H‐FABP), glycogen phosphorylase BB (GPBB), high sensitive C reactive protein (hsCRP) cardiac troponin I, (cTnI), creatine kinase MB (CK‐MB mass) and myoglobin.
Methods
A total of 20 children who underwent HSCT for malignant and non‐malignant diseases were included in this study. Blood samples were collected from all patients in 0th, 7th and 21st day for evaluating these cardiac biomarkers. The patients’ echocardiography was assessment before and after one‐month of HSCT.
Results
Serum 21st H‐FABP level was significantly higher when compared with the 0th day H‐FABP level (P < 0.05) . 7th day hsCRP level was significantly higher than 0th and 21st day levels (P < 0.05). Interestingly, 7th day GPBB level was significantly lower than 0th and 21st day levels (P < 0.05). Myoglobin, CK‐MB mass and cTnI biomarkers remained within the reference range in all patients.
Conclusions
This study showed that H‐FABP and hsCRP both seem to be promising markers for evaluation of cardiotoxicity in HSCT process and probably superior to GPBB, cTnI, CK‐MB mass and myoglobin.
Keywords: H‐FABP, hsCRP, glycogen phosphorylase BB, chemotherapy, children
INTRODUCTION
Hematopoietic stem cell transplantation (HSCT) is the treatment of choice for defined malignant and non‐malignant hematological diseases in children 1. HSCT is a complex therapeutic procedure consisting of administration of high‐dose chemotherapy, immunosuppressive therapy, and/or radiotherapy, followed by intravenous infusion of hematopoietic stem cells to re‐establish marrow function 2. The main drawbacks of HSCT are early transplant‐related mortality and delayed complications, which interfere with patient's outcome, health condition, and quality of life. Early life‐threatening cardiotoxicity and cardiac death have been showed after HSCT. In comparison with other post‐transplant complications, cardiac, or cardiovascular outcomes appear to occur at a much lower frequency 1.
Although there are different techniques to detect cardiotoxicity, research are ongoing for practical, useful, noninvasive, and sensitive methods 3. Cardiac troponins (cTn) and myocardial izoenzyme of creatine kinase MB (CK‐MB) are cardiospecific markers that show structural injury of cardiomyocytes from various causes, including cardiotoxic effect of anticancer therapy. Heart‐type fatty acid binding protein (H‐FABP) and glycogen phosphorylase isoenzyme BB (GPBB) are new markers of myocardial ischemia and necrosis recently assessed in the diagnostics and risk stratification of acute coronary syndromes 4. High‐sensitivity C reactive protein (hsCRP) also plays a fundamental role in atherosclerosis, ischemic heart failure and coronary heart disease 5.
The purpose of the present study was to evaluate cardiac toxicity of conventional chemotherapy followed by HSCT with cardiac markers of H‐FABP, GPBB, hsCRP, cTnI, CK‐MB mass, and myoglobin. To our knowledge this is the first study evaluating these cardiac biomarker levels in children who underwent HSCT.
MATERIALS AND METHODS
Patients
A total of 20 consecutive children who underwent HSCT for malignant (n:12) and non‐malignant diseases (n:8) between the ages of 1–20 years at Ankara Children's Hematology and Oncology Hospital. All children included in this study had completed their 100 days after transplantation. Echocardiography was performed for all children before HSCT and 1 month after HSCT.
Written informed consent was obtained from all of the parents and/or subjects. This study was approved by the local ethics committee. The experiments were performed in accordance with the principles enumerated in the Helsinki Declaration of 1975.
Patients characteristic and demographic data of the children have been shown in Table 1.
Table 1.
Patients Characteristic and Biochemical Marker Levels in Study Population
| n (%) | |
|---|---|
| Diagnosis of patients | |
| Acute myeloid leukemia | 1 (5) |
| Acute lymphoblastic leukemia | 4 (20) |
| Thalassemia Major | 2 (10) |
| Myelodysplastic syndrome | 2 (10) |
| Hodgkin Lymphoma | 2 (10) |
| Primary Hemophagocytic Lymphohistiocytosis | 1(5) |
| Fanconi aplastic anemia | 1 (5) |
| Hodgkin Lymphoma | 1 (5) |
| Hyperimmunoglobulin E syndrome | 1 (5) |
| Neuroblastoma | 1 (5) |
| Chediak Higashi syndrome | 1 (5) |
| Ataxia telangiectasia‐mix lineage leukemia | 1 (5) |
| Wiscott Aldrich syndrome‐NHL | 1 (5) |
| Severe combined immunodeficiency | 1 (5) |
| Stem cell source | |
| Bone marrow | 14 (70) |
| Peripheral stem cell | 6 (30) |
| Donors | |
| Sibling | 13 (65) |
| Father | 1 (5) |
| Mother | 1 (5) |
| Uncle | 1 (5) |
| Autologous | 4 (20) |
| Donor HLA | |
| HLA 10/10 compatible | 10 (50) |
| HLA 9/10 compatible | 4 (20) |
| HLA 8/10 compatible | 1 (5) |
| HLA 5/10 compatible | 1 (5) |
| Autologous | 4 (20) |
| Conditioning regimen | |
| Bu‐Cy | 3 (15) |
| TBI‐Etoposide | 3 (15) |
| Flu‐Cy‐ATG | 2(10) |
| Pesaro protocol 26 | 2 (10) |
| Bu‐Flu‐ATG | 2 (10) |
| Melphalan‐Etoposide‐Cy | 2 (10) |
| Melphalan‐Etoposide‐Carboplatin | 2 (10) |
| Bu‐Cy‐Etoposide | 1 (5) |
| Cy‐Melphalan | 1 (5) |
| Bu‐Cy‐Melphalan | 1 (5) |
| TBI‐Cy | 1 (5) |
| Outcome | |
| Alive without disease | 18 (90) |
| Died | 2 (10) |
| 0th day | 7th day | 21st day | |
|---|---|---|---|
| Biomarker | |||
| H‐FABP (pg/mL) | 588.5 ± 361.9 | 686.7 ± 326.3 | 739.5 ± 337.4* |
| GP‐BB (pg/mL) | 177.9 ± 68.9 | 125.7 ± 41.9* | 152.7 ± 49.7# |
| hsCRP (mg/mL) | 5.7 ± 5.8 | 20.1 ± 14.0* | 6.5 ± 7.7# |
NHL, Non‐Hodgkin lymphoma; Bu, Busulphan; Cy, Cylophosphamide; TBI, Total body irradiation; Flu, Fludarabine.
*P < 0.05 in comparison with 0th day levels; # P < 0.05 in comparison with 7th day levels.
Hematopoietic Stem Cell Transplantation
Conditioning regimens and GvHD prophylaxis were given according to the current protocols of different diseases. Twenty patients received 11 different conditioning regimens. Most patients (17/20) received myeloablative regimen. ATG at a dosage of 20–40 mg/kg body weight as part of the conditioning regimen was given to four patients. All patients received intravenous immunoglobulin (IVIG) at +1, +8, +15, +22 day, and IVIG was given to all patients whose Ig G level below 400 mg/dL after discharge from the hospital. Cyclosporine A combined with short‐term methotrexate was the most preferred protocol (14/20) for acute GvHD. The majority of patients (10/20) received full‐matched related HSCT while four patients received autologous HSCT. Only one patient received haploidentical HSCT for Severe Combined Immunodeficiency. Severity of oral mucositis was evaluated and daily graded according to adapted common toxicity criteria for stomatitis in patients 6. The presence and severity of acute GvHD were assessed daily by the attending HSCT physician/nurses and graded according to Glucksberg criteria 7. Steroid was used for patients with acute GvHD, chronic GvHD, and engraftment syndrome. Ciprofloxacin, flucanazole, and trimetoprim/sulfamethoxasole were used for antibacterial, antifungal, and Pneumocystis jiroveci prophylaxis; respectively. Neutrophil engraftment day was defined as the first of three sequential days in which the absolute neutrophil count was ≥500/mm3. Platelet engraftment day was defined as the first of three sequential days in which platelet count was ≥20,000/mm3 without transfusion for at least 7 days.
Determination of Biochemical Parameters
After an 8–12 hr overnight fast, the venipuncture was performed between 8:00 am and 9:00 am and blood samples were collected for obtaining serum on 0th day, 7th day, and 21st day after HSCT. Blood samples were centrifuged and serum samples were stored at –70°C until analysis. Measurements of H‐FABP, GPBB, and hsCRP were performed in ELISA reader using the commercially available ELISA kit. CK‐MB mass, Troponin I, and myoglobin were analyzed using the immunoassay kit. The samples were carried out together in the same experiment.
Statistical Analysis
Data were presented as mean values with standard deviation (SD) for continuous variable. For variables with normal distribution, repeated measures ANOVA was used. Friedman test followed by the Wilcoxon test was used for variables with non‐normal distribution. All statistical analyses were performed using the software SPSS 15.0 program. For all statistical tests, two‐tailed P value < 0.05 indicated the statistical significance of the results.
RESULTS
Patients and Transplantation Course
A total of 20 children (six girls and 14 boys) with the mean age of 9.5 ± 4.5 years (1–20 years) comprised the study group. Of these 20 children, 16 received allogeneic and four received autologous HSCT. The mean number of CD34 (+) stem cell given to the patients was 4.1 ± 2.6 × 106/kg (1.1–8.7 × 106/kg) and the mean number of nucleated cells was 5.6 ± 2.5 × 108/kg (2.5–11.0 × 108/kg). Neutrophil engraftment was achieved at median 14 days (11–21) and platelet engraftment, at median 19 days (13–36). Among 20 children who underwent HSCT, 18 had complications related to HSCT after the procedure (15 had febrile neutropenia, 13 had oral mucositis, two had acute GvHD, two had veno‐occlusive disease, one had engraftment syndrome, one had hemorrhagic cystitis). The median follow‐up duration was 376 days (120–576). The patients’ echocardiography was normal before (Ejection fraction: 74.2 ± 4.6%) and after 1 month of HSCT (Ejection fraction: 70.2 ± 3.8). No patient manifested clinical cardiotoxicity with symptoms of heart failure in the first 100 day after HSCT.
Biochemical Results
In this study, hsCRP levels at 7th day have been significantly increased in comparison with hsCRP levels at 0th day. Moreover, hsCRP level at 21st day has returned to 0th day's levels. If H‐FABP levels at 0th and 7th, days were compared; H‐FABP levels at 7th day were increased and this increase continued for H‐FABP levels at 21st day; whereas only the difference between H‐FABP levels at 0th and 21st day was statistically significant. If GPBB levels at 0th, 7th, and 21st days were compared; GPBB levels were decreased at 7th day and those increased at 21st day that reached to baseline level at 0th day. Myoglobin, CK‐MB mass, and cTnI biomarkers remained within the reference range in all patients. H‐FABP, GPBB, and hsCRP levels at 0th, 7th, and 21st days have been shown in Table 1.
DISCUSSION
Cardiac toxicity is a well known and potentially serious complication during treatment of hemato‐oncology patients 8. Thus, we decided to evaluate carditoxicity of children who underwent HSCT using cardiotoxicity markers of H‐FABP, GP‐BB, hsCRP, cTnI, CK‐MB mass, and myoglobin.
H‐FABP is a 14.5‐kDa cytosolic protein present in the myocardium and is a key cytosolic transporter of fatty acids 9. Even in the absence of irreversible myocardial necrosis, H‐FABP which is more sensitive and specific than troponin I and CK‐MB in the early diagnosis of acute coronary syndrome is released during myocardial ischemia 10. Our findings also, suggest that circulating level of H‐FABP is increased at 21st day after HSCT in children who underwent HSCT. It might be associated with myocardial injury manifested by release of H‐FABP from cardiomyocytes.
GP‐BB is a glycolytic enzyme that plays an essential role in the regulation of carbohydrate metabolism. GPBB isoenzyme is not limited to human heart and brain, and small amounts have been described in numerous other tissues including aorta, leukocytes, testis, digestive tract, spleen, kidney, bladder, and liver 11.
Studies has showed that GP‐BB was the most sensitive biomarker to detect myocardial infarction when compared to myoglobin, CK‐MB, and cTnT at first hour 12. Horacek et al. found a significant correlation between elevation in GPBB and diastolic LV dysfunction on echocardiography during HSCT 13. Their other study also showed that GPBB seems to be a promising marker for evaluation of anthracycline‐induced cardiotoxicity and probably superior to cTnI and cTnT 14. Interestingly, our results indicate that GPBB levels were decreased at 7th day and those increased at 21st day that was comparable to the level at 0th day. Since, most of our patients received myeloablative conditioning regimen and their leukocyte count was very low at 7th day after HSCT. Furthermore, median neutrophil engraftment day was 14 day after HSCT (11–21 days). Thus, GPBB does not seem to be of value in the detection of cardiotoxicity during HSCT process in children.
A number of prospective research have established that hsCRP is an independent predictor of future risk for cardiovascular events among healthy individuals and among patients with acute coronary syndromes 15. In this study, hsCRP levels at 7th and 21st days have been significantly increased in comparison with hsCRP levels at 0th day.
Horacek et al. evaluated acute and chronic cardiotoxicity of anthracyclines in acute leucemia patients with NT‐proBNP, cTnT, and CK‐MB mass. They showed that early after anthracyclines administration, even in higher cumulative doses, no patient had a measurable cTnT and increased CK‐ MB mass concentrations 16. Similarly, our results demonstrated troponin I, CK‐MB mass, and myoglobin do not seem to be of value in the detection of early cardiotoxicity in children who underwent HSCT.
In conclusion, this study showed that both H‐FABP and hsCRP seem to be promising markers for evaluation of cardiotoxicity in children who underwent HSCT and probably are superior to cTnI, CK‐MB mass, myoglobin, and GPBB. Taken all together, these findings could be considered to be a sign of acute subclinical cardiotoxicity. Further studies involving larger numbers of individuals would be valuable to improve our understanding of these cardiac biomarkers in children who underwent HSCT. It is not clear whether these acute changes will have predictive value for development of treatment‐related cardiotoxicity in the future. It should be evaluated during prospective follow‐up studies.
CONFLICT OF INTEREST
The authors report no conflict of interest.
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