High neutrophil‐to‐lymphocyte ratio as an early sign of cardiotoxicity in breast cancer patients treated with anthracycline

Abstract Background Cardiotoxicity, defined mainly as left ventricle (LV) dysfunction, is a significant side effect of anthracyclines (ANT) therapy. The need for an early simple marker to identify patients at risk is crucial. A high neutrophil‐to‐lymphocyte ratio (NLR) has been associated with poor prognosis in cancer patients; however, its role as a predictor for cardiotoxicity development is unknown. Objective Evaluating whether elevated NLR, during ANT exposure, plays a predictive role in the development of cardiotoxicity as defined by LV global longitudinal strain (LV GLS) relative reduction (≥10%). Methods and Results Data were prospectively collected as part of the Israel Cardio‐Oncology Registry. A total of 74 female patients with breast cancer, scheduled for ANT therapy were included. NLR levels were assessed at baseline (T1) and during ANT therapy (T2). All patients underwent serial echocardiography at baseline (T1) and after the completion of ANT therapy (T3). NLR ≥ 2.58 at T2 was found to be the optimal predictive cutoff for LV GLS deterioration. A relative LV GLS reduction ≥10% was significantly more common among patients with high NLR (50% vs. 20%, p = .009). NLR ≥ 2.58 at T2 increases the risk for LV GLS reduction by fourfold (odds ratio [OR]: 4.63, 95% confidence interval [CI]: 1.29–16.5, p = .02), with each increase of 1‐point NLR adding an additional 15% risk (OR: 1.15, 95% CI: 1.01–1.32, p = .046). Conclusions Our study provides novel data that high NLR levels, during ANT exposure, have an independent association with the development of LV dysfunction. Routine surveillance of NLR may be an effective means of risk‐stratifying.


| INTRODUCTION
Breast cancer is the most prevalent type of cancer in women. 1 While advancements in cancer therapy have led to a significant reduction in morbidity and mortality, the high burden of short-and long-term side effects brought about by modern medical therapy plays a significant role in patient outcomes. 2,3 Cardiotoxicity is the most significant complication and is manifested mainly as left ventricle (LV) dysfunction. 4,5 Anthracyclines (ANT), in particular doxorubicin, remains a cornerstone in chemotherapy regimens for the treatment of patients diagnosed with breast cancer, 5 and its cardiotoxic effects are characterized by dose-dependent irreversible LV dysfunction. 6,7 Early detection of ANT-related cardiac injury and clinician intervention is, therefore, paramount for the prevention of the development or worsening of LV dysfunction and heart failure (HF) in this patient population. 8 Currently, two-dimensional speckle-tracking echocardiography and the use of LV global longitudinal strain (LV GLS) is considered the optimal parameter for early detection of subclinical LV dysfunction in cancer patients. [9][10][11][12] The absolute neutrophil-to-lymphocyte ratio (NLR), a simple measure using basic cell counts, has been found to be positively associated with increased mortality in cancer patients as its number increases. 13,14 This has been suggested to be due to the fact that neutrophils are proinflammatory and can contribute to the progression of cancer, while lymphocytes have been found to act as tumor inhibitors 15 and an elevated NLR in cancer patients has been associated with poor prognosis. 14,16 Similarly, a number of studies have shown that elevated NLR in patients presenting with myocardial infarction is related to a major adverse cardiac event (MACE), as well as all-cause mortality, 17,18 further suggests a relation between an immune reaction and cardiovascular disease (CVD) outcomes.
However, data evaluating the role of NLR in predicting cardiotoxicity development in cancer patients is scarce.
The aim of our study was to evaluate whether elevated NLR, in patients diagnosed with breast cancer and treated with ANT, plays a predictive role in the development of cardiotoxicity, as defined by LV GLS reduction.

| Study population
The study population is part of the Israel Cardio-Oncology Registry (ICOR) -a prospective registry enrolling patients evaluated in the cardiooncology clinic at the Tel Aviv Sourasky Medical Center. All patients signed informed consent at the first clinic visit. The registry was approved by the local ethics Tel Aviv Sourasky committee (Identifier: 0228-16-TLV) and is registered on clinicaltrials.gov (Identifier: NCT02818517). Included in the cohort were female patients diagnosed with breast cancer and scheduled for doxorubicin therapy with a cumulative dose of doxorubicin ≥180 mg/m 2 . All patients performed at least two echocardiography evaluations; at baseline, before doxorubicin exposure (T1), and at the end of doxorubicin therapy (T3). The majority of the patients performed additional echocardiography evaluation following the third ANT cycle (T2). Blood samples, including NLR, were taken at baseline (T1) and during doxorubicin exposure (T2), in close proximity to the echocardiography exam (T2). Exclusion criteria included age below 18, male gender, reduced baseline LV function (LV ejection fraction [LVEF] < 53%), a history of cardiac disease (myocardial infarction, myocarditis, severe valvular diseases), past ANT therapy, dexrazoxane (cardioxane) therapy and no documentation of NLR values or LV GLS assessment. All echocardiography exams evaluated systolic and diastolic function, including LV GLS, as described in the echocardiography section. NLR levels, measured as the ratio between absolute neutrophil count to absolute lymphocyte count, were evaluated at baseline, before doxorubicin exposure (T1), and during doxorubicin exposure (T2), before echocardiography follow-up at the end of doxorubicin therapy (T3). The study population was stratified into two groups based on NLR values, according to the optimal predictive cutoff for LV GLS deterioration at T3, determined by a receiver operator characteristic (ROC) curve ( Figure 1). A significant reduction in LV GLS from T1 to T3 was defined as a relative reduction of ≥10%, as accepted by previous studies. 19

| Echocardiography
Follow-up echocardiographic examinations were performed using the same strict protocol, personnel, and equipment (General Electric system, model Vivid S70). Routine LV echocardiographic parameters BARUCH ET AL. | 329 included LV diameters and LVEF. 20 Early transmitral flow velocity (E), late atrial contraction (A) velocity, and early diastolic mitral annular velocity (septal and lateral e′) were measured in the apical fourchamber view to provide an estimate of LV diastolic function. 20 The peak E/peak e′ (E/e′) ratio was calculated (septal, lateral, and average mitral E/e′ ratio), and the deceleration time of the E wave was measured. The left atrium volume index was calculated using the biplane area length method at end-systole. Images were acquired using a high frame rate (>50 frames/s), 21 and thereafter stored digitally for offline analysis. LV GLS was measured using EchoPac STE software and tracking within an approximately 5 mm wide region of interest. An end-systolic frame was used to initialize LV boundaries which were then automatically tracked throughout the cardiac cycle.
Manual corrections were performed to optimize boundary tracking as needed. Optimization of images for endocardial visualization through adjustment of gain, compress, and time-gain compensation controls were done before acquisition.

| Statistical analysis
Categorical variables were expressed as frequency and percentages. χ 2 test was used to evaluate the association between these variables. The distribution of continuous variables was assessed using the Kolmogorov-Smirnov test. Normally distributed continuous variables were described as mean and standard deviation, these variables were compared using the independent samples t-test. Non-normally distributed continuous variables    0.51-0.77, p = .046 ( Figure 1). According to this analysis, the optimal cutoff value was ≥2.58, with 79% sensitivity and 52% specificity.

| Baseline parameters
All patients were female with a mean age of 50 ± 12 years. None of the patients had a history of coronary artery disease. Cardiac risk factors were not frequent and ranged from 7% to 22% (Table 1) Table 1.

| Laboratory parameters
The median time elapsed from the first doxorubicin treatment to NLR evaluation at T2 was 44 (IQR: 28-64) days, which in the majority of the patients was following the third cycle of ANT with a cumulative dose of  (Figure 2). As displayed in Table 1, no difference was observed between study groups for hemoglobin or platelet levels, neither in T1 nor T2. White blood cell counts were significantly higher in the high NLR group only in T2 (p < .001). Serum creatinine levels were similar between both groups at both time points.

| DISCUSSION
We evaluated, for the first time to our knowledge, the association between elevated NLR values and the development of cardiotoxicity among patients with breast cancer. We found that NLR ≥ 2.58, during ANT exposure (T2), was associated with a fourfold increased risk for developing significant LV GLS deterioration, with incremental increases of 1-point NLR adding an additional 15% risk.
The use of NLR measurements as a surrogate marker of poor outcomes is gaining popularity, both in the field of cancer and CVD. Past studies have shown that high baseline NLR values are associated with poor overall survival in different types of cancer, including breast cancer. 13,14 This may be explained 14,22,23  has been shown to promote apoptosis. 18 While the role of high NLR as an independent predictor for poorer outcomes in both cancer and CVD patient populations has been well studied, its role in predicting cardiotoxicity development in cancer patients is yet unknown. Our study showed that high NLR, during ANT therapy (T2) was significantly associated with LV GLS deterioration, while baseline NLR (T1) was not, which is likely explained by the significant NLR increase from T1 to T2 (Figure 2) following ANT exposure. Of note, patients were also treated with granulocytescolony stimulating factor (GCSF) during ANT therapy which may have caused an increase in NLR. However, we observed that the change in NLR in our study was mainly a result of a reduction in the absolute lymphocyte count rather than an elevated absolute neutrophil count that is seen with GCSF exposure. Furthermore, based on the timing of the NLR increase, it is suggested that it was more likely a result of ANT exposure rather than a baseline inflammatory state from cancer. Importantly, there were no differences in baseline clinical characteristics or blood tests between the groups that may confound or falsely elevate the measured NLR or act as a risk predictor of LV GLS deterioration. Antiaggregation, n (%) 5 (6.8) 1 (3.3) 4 (9.1) .64 Laboratory data at T1 Hemoglobin (g/dl), mean ± SD 12.9 ± 1.0 12.8 ± 1.0 12.9 ± 1.0 .61 White blood cell count (10 9 /L), mean ± SD 6.7 ± 2.4 6.3 ± 2.

| CONCLUSIONS
In summary, our study provides novel data that high NLR, during ANT exposure, has an independent association with the development of LV GLS deterioration, a parameter of LV dysfunction. Routine surveillance of NLR values may be an effective and rapid means of risk-stratifying and predicting early systolic dysfunction among patients with breast cancer following ANT therapy. T A B L E 3 Multivariate binary regression models for prediction of LV GLS reduction Note: Bold values are significant p < .05.