Prognostic significance of neutrophil-to-lymphocyte ratio for long-term outcomes in patients with poorly differentiated thyroid cancer

Abstract

Poorly differentiated thyroid cancer (PDTC) is a distinct but rare type of thyroid cancer with intermediate biological behavior between differentiated and anaplastic thyroid cancers. PDTC was first defined in 2005 in Japan, but the diagnostic criteria changed in 2015, requiring the tumor to have more than 50% of poorly differentiated components for diagnosis. Because only six years have passed since the PDTC definition change, prognostic factors for long-term survival who meet the latest criteria have not been determined. Neutrophil-to-lymphocyte ratio (NLR) is a prognostic marker in various solid malignancies. However, its impact on PDTC remains unclear. This study aimed to evaluate the significance of NLR as a prognostic factor for patients with PDTC diagnosed based on the latest criteria. In total, 28 PDTC cases (4.4%) of 637 thyroid cancer patients who underwent surgery between 2002 and 2012 were retrospectively analyzed. The median follow-up period was 120 months (range, 7–216 months). Of the 13 deaths (46.4%), 9 patients (32.1%) died from PDTC. The median preoperative NLR was 2.7 (0.67–8.62), and the NLR cut-off value determined by the receiver operating characteristic curve was 2.88. Patients with a high NLR (>2.88) showed significantly worse disease-specific survival (hazard ratio [HR] 4.67, p = 0.036) and overall survival (HR 4.94, p = 0.007) than those with a low NLR (≤2.88). Multivariate analysis revealed that a high NLR independently predicted a worse prognosis (HR 6.06, p = 0.0087). In conclusion, NLR is a useful prognostic marker for patients with PDTC.

POORLY DIFFERENTIATED THYROID CANCER (PDTC) is a pathologically distinct type of thyroid cancer exhibiting intermediate biological behavior between differentiated thyroid cancer (DTC) and anaplastic thyroid cancer (ATC) [1-3]. PDTC is a relatively rare disease, accounting for 2 to 15% of all thyroid cancers [4, 5]; the prevalence variation is attributed to geographical (i.e., environmental) influences or differences in histopathological diagnosis criteria. In Japan, Ito et al. demonstrated that the PDTC frequency was 9.4% among 1942 patients diagnosed with papillary thyroid cancer (PTC) between 1987 and 1996 [6]. Despite its rarity, PDTC is a major cause of death from non-ATCs. Thus, PDTC is a clinically significant thyroid cancer type [7].

In Japan, PDTC was first documented as an independent pathological entity from DTC in the 2005 General Rules for the Japanese Society of Thyroid Surgery (JSTS) 6^th edition [8], following the 2004 World Health Organization (WHO) classification of tumors of endocrine organs [9]. In these rules, PDTC was defined as a tumor originating from follicular cells with poorly differentiated components (solid, insular, or trabecular growth pattern). Occupied areas with poorly differentiated components did not matter for PDTC diagnosis; tumors with even a small area of poorly differentiated components were diagnosed as PDTC according to the JSTS 6^th edition rules [8]. Subsequently, 2017 WHO classification required at least one of the following findings: increased mitotic activity (≥3 mitoses per 10 high-power fields), tumor necrosis, or convoluted nuclei without PTC nuclear features [10], as mentioned in the Turin proposal in 2006 [11]. Although the proportion of poorly differentiated components for PDTC diagnosis remains controversial, more than 50% of the compulsory proportion was designated for PDTC diagnosis in the JSTS 7^th edition, published in 2015 [12]. This transition in the diagnostic criteria caused a discrepancy in the patients diagnosed with PDTC before and after 2015. Namely, PDTC patients whose primary tumor contained less than 50% of poorly differentiated components before 2015 could no longer be considered as PDTC in Japan with the current criteria. Therefore, evaluating long-term survival and determining prognostic factors for PDTC patients who met the latest criteria has been difficult. The rarity of PDTC has also made prognostic factor evaluation difficult.

A growing body of evidence suggests that the prognosis of various solid malignancies is associated with systemic inflammation and the immunological status of patients [13]. Neutrophil-to-lymphocyte ratio (NLR), which is easily calculated by the ratio of total neutrophils to total lymphocytes in peripheral blood, reflects the balance between inflammation and immunity. An increase in neutrophils could result from cancer-related inflammation induced by cancer progression or cytokine release from tumor cells and tumor-associated immune cells. Nevertheless, it has been demonstrated that neutrophils can facilitate tumor growth and suppress anti-tumor immunity [14, 15]. In contrast, a lymphocyte decrease is associated with impaired innate and adaptive immunity against cancer [16, 17]. Thus, an elevated NLR represents a high inflammation and poor lymphocyte-mediated immune response to tumors and is associated with worse prognoses in various solid malignancies [18]. In PDTC patients, however, to date, no study has demonstrated a correlation between NLR and clinical outcomes. Therefore, the NLR clinical significance in patients with PDTC remains unclear.

This study aimed to evaluate the association between the NLR and the long-term survival of PTDC patients who met the latest criteria. We pathologically re-evaluated PDTC cases diagnosed before 2015 based on the General Rules for JSTS 6^th edition [8] and excluded those that did not meet the current diagnostic criteria. We retrospectively investigated the preoperative NLR values of PDTC patients and analyzed the correlation between these values and patient outcomes.

Materials and Methods

Patients and study design

In this retrospective study, 50 PDTC patients were identified according to the General Rules for the JSTS 6^th edition (used at that time) [8] among 637 thyroid cancer patients who underwent primary surgery at Shinshu University Hospital from 2002 to 2012. All PDTC cases were pathologically re-assessed by the pathologist (T.U.), who was blinded to the clinical information, to determine if they met the 2015 General Rules for the JSTS 7^th edition [12]. The rules required that tumors be at least 50% of poorly differentiated components with at least one of the following findings: increased mitotic activity (≥3 mitoses per 10 high-power fields), tumor necrosis, or convoluted nuclei without PTC nuclear features. Twenty patients were excluded, as well as two patients without detailed laboratory data for NLR calculations. Finally, 28 patients were included in the study.

Surgical treatment

Hemithyroidectomy was performed in patients with a tumor within 1 cm in size and localized in the hemilobe of the thyroid. Patients with tumors >5 cm in size, multifocal tumors, a tumor with extrathyroidal extension, or distant metastasis underwent total thyroidectomy. Except for these patients, surgical procedure (hemithyroidectomy or total thyroidectomy) was determined considering other clinical features, including patient’s age, performance status, and involvement of neck lymph nodes. Central neck dissection was performed for patients with tumors <1 cm in size or follicular neoplasms. Unilateral modified radical neck dissection was performed for patients with clinically apparent lymph node metastases, a tumor ≥1 cm in size, or a tumor with extrathyroidal extension. Bilateral modified radical neck dissection was performed if bilateral lymph node metastases were suspected.

Postoperative therapy and follow-up

Patients with distant metastases or post-surgical residual tumors in the neck (R1: microscopic residual disease or R2: gross residual disease) were treated with 100 mCi of radioactive iodine-131 (RAI) per unit time ranging from 2 to 10 times. External radiation therapy (50–60 Gy) was also administered to the neck of patients with R2. All patients were followed-up with continued suppression of serum thyroid-stimulating hormone levels (<0.01 μIU/mL). Serum thyroglobulin levels were monitored at least twice per year, and ultrasonography or computed tomography (or both) was performed once every one to two years to screen for locoregional or distant recurrence.

Data collection

Clinicopathological information, including age, sex, surgical procedure, tumor size, neck lymph node metastases, extrathyroidal extension, and the presence of distant metastases at diagnosis, were retrospectively collected from the patients’ medical records.

Disease-specific survival (DSS) was defined as the period from the day of the initial surgery to the day of death due to PDTC. Overall survival (OS) was defined as the primary surgery to the date of death from any cause.

The preoperative neutrophil-to-lymphocyte ratio calculation

The NLR values were calculated using the routine blood examination results obtained within one month before the initial surgery by dividing the total neutrophil count by the total lymphocyte count [19]. The receiver operating characteristics (ROC) curve was analyzed to determine the best NLR cut-off values for OS.

Ethics statement

This study was approved by the local clinical investigation ethics committee of Shinshu University (no. 5036). Because this was a retrospective study of anonymized data, the informed consent requirement was waived. This study complied with the provisions of the Declaration of Helsinki (64^th WMA General Assembly, Fortaleza, Brazil, October 2013).

Statistical analyses

Variables were analyzed using the chi-square or Fisher’s exact test (categorical) and the Mann-Whitney U test (continuous). DSS and OS were estimated using the Kaplan-Meier method, and significant differences in survival were assessed using the log-rank test. Univariate and multivariate analyses with the Cox proportional hazards model were performed to determine the significant factors. Multivariate analysis was performed for parameters with p < 0.05 in the univariate analysis. All statistical analyses were performed using GraphPad Prism 8.0.2, and statistical significance was set at p < 0.05.

Results

Patient clinicopathological characteristics

The clinicopathological characteristics of the 28 included patients are presented in Table 1. Twenty patients (71.4%) were aged over 55 years. Fifteen patients (53.6%) were men, and 13 patients (46.4%) were women. Eighteen patients (64.3%) were preoperatively diagnosed as PTC, while 6 patients (21.4%) were diagnosed as follicular neoplasms by fine-needle aspiration cytology (FNA). Four patients (14.3%) were suspected as PDTC or poorly differentiated carcinoma from unknown origin by FNA, and thus, core-needle biopsy was performed, resulting in the diagnosis of PDTC. As for clinical stage according to the General Rules for JSTS 8^th edition (2019) [20], 12 patients (42.8%), 4 patients (14.3%), 6 patients (21.4%), and 6 patients (21.4%) had stage I, stage II, stage III, and stage IVb disease, respectively. Twenty-two patients (78.6%) were treated with total thyroidectomy, and 6 patients (21.4%) underwent hemithyroidectomy. Neck dissection was performed in 26 patients (92.9%). There were 12 patients with tumors ≤2 cm (42.8%), 8 with tumors between 2.1 and 4.0 cm (28.6%), and 8 with tumors >4.0 cm (28.6%). Lymph nodes were involved in 14 patients (50.0%), and extrathyroidal extension was detected in 17 patients (60.7%). Distant metastases were present in 7 patients (25.0%) at the time of diagnosis (lung: n = 7, 25.0%; bone: n = 2, 7.1%). For the pathological stage determined by the General Rules for JSTS 8^th edition (2019) [20], 11 (39.3%) had stage I disease, 4 (14.3%) had stage II, 7 (25.0%) had stage III, and 6 (21.4%) had stage IVb. Fifteen patients (53.6%) underwent curative resection (R0), and 13 patients had residual tumors (microscopic residual disease, R1: n = 9, 32.1%; gross residual disease, R2: n = 4, 14.3%). Thirteen patients (46.4%) had tumors with more than 75% poorly differentiated components. After surgery, 16 patients (52.7%) were treated with RAI therapy. Eight patients (28.6%) underwent external radiation therapy of the neck.

Table 1 Clinicopathological characteristics of patients and comparison of high NLR and low NLR patients

Variables	Total n = 28 (%)	High NLR n = 13 (%)	Low NLR n = 15 (%)	p value
Age
≤55	8 (28.6%)	3 (23.0%)	5 (33.3%)	0.54
>55	20 (71.4%)	10 (77.0%)	10 (67.7%)
Sex
Male	15 (53.6%)	7 (53.8%)	8 (53.3%)	0.97
Female	13 (46.4%)	6 (46.2%)	7 (46.7%)
Preoperative diagnosis
PTC	18 (64.3%)	7 (53.8%)	11 (73.3%)	0.41
Follicular neoplasms	6 (21.4%)	3 (23.0%)	3 (20.0%)
PDTC	4 (14.3%)	3 (23.0%)	1 (6.6%)
cStage
I	12 (42.8%)	5 (38.5%)	7 (46.7%)	0.67
II	4 (14.3%)	2 (15.4%)	2 (13.3%)
III	6 (21.4%)	2 (15.4%)	4 (26.7%)
IVa	0 (0%)	0 (0%)	0 (0%)
IVb	6 (21.4%)	4 (30.8%)	2 (13.3%)
Surgical procedure
Total thyroidectomy	22 (78.6%)	11 (84.6%)	11 (73.3%)	0.65
Hemithyroidectomy	6 (21.4%)	2 (15.4%)	4 (26.7%)
Neck dissection
No	2 (7.1%)	2 (15.4%)	0 (0%)	0.11
Yes	26 (92.9%)	11 (84.6%)	15 (100%)
Tumor size (cm)
≤2.0	12 (42.8%)	5 (38.5%)	7 (46.7%)	0.55
2.1–4.0	8 (28.6%)	3 (23.0%)	5 (33.3%)
>4.0	8 (28.6%)	5 (38.5%)	3 (20.0%)
pN stage
0	11 (39.3%)	5 (38.5%)	6 (40.0%)	0.62
1	14 (50.0%)	5 (38.5%)	9 (60.0%)
x	3 (10.7%)	3 (23.0%)	0 (0%)
Extrathyroidal extension
No	11 (39.3%)	2 (15.4%)	9 (60.0%)	0.15
Yes	17 (60.7%)	11 (84.6%)	6 (40.0%)
M status
0	21 (75.0%)	9 (69.2%)	12 (80.0%)	0.67
1	7 (25.0%)	4 (30.8%)	3 (20.0%)
pStage
I	11 (39.3%)	4 (30.8%)	7 (46.7%)	0.68
II	4 (14.3%)	2 (15.4%)	2 (13.3%)
III	7 (25.0%)	3 (23.0%)	4 (26.7%)
IVa	0 (0%)	0 (0%)	0 (0%)
IVb	6 (21.4%)	4 (30.8%)	2 (13.3%)
Residual tumor
R0	15 (53.6%)	6 (46.2%)	9 (60.0%)	0.46
R1	9 (32.1%)	3 (23.0%)	6 (40.0%)
R2	4 (14.3%)	4 (30.8%)	0 (0%)
Poorly differentiated component
≥50%, <75%	15 (53.6%)	6 (46.2%)	9 (60.0%)	0.46
≥75%	13 (46.4%)	7 (53.8%)	6 (40.0%)
RAI
No	12 (42.8%)	5 (38.5%)	7 (46.7%)	0.71
Yes	16 (57.2%)	8 (61.5%)	8 (53.3%)
External radiation therapy
No	20 (71.4%)	8 (61.5%)	12 (80.0%)	0.41
Yes	8 (28.6%)	5 (38.5%)	3 (20.0%)

PTC, papillary thyroid cancer; PDTC, poorly differentiated thyroid cancer; R0, no residual disease; R1, microscopic residual disease; R2, gross residual disease; RAI, radioactive iodine therapy; NLR, neutrophil-to-lymphocyte ratio

The median follow-up period was 120 months (range, 7–216 months). Thirteen patients (46.4%) died during the follow-up period, including 9 patients (32.1%) who died from PDTC. The median preoperative NLR was 2.7 (0.67–8.62). None of the patients exhibited anaplastic transformation through clinical courses.

Associations between NLR and patient outcomes

The optimal NLR cut-off value determined by the ROC analysis was 2.88 [(area under the curve [AUC] = 0.74, sensitivity/specificity = 0.75)] (Fig. 1). The patients were segregated into high and low NLR groups according to this cut-off value. There were no differences in the clinicopathological features between the high and low NLR groups, including age, sex, preoperative diagnosis, the clinical stage, surgical procedure, the presence of neck dissection, tumor size, lymph node metastases, extrathyroidal extension, the presence of distant metastases, the pathological stage, residual tumors, the proportion of poorly differentiated components, and the presence of postoperative RAI or external radiation therapy (Table 1).

Fig. 1

ROC curve for OS according to the NLR value. AUC = 0.74.

ROC, receiver operating characteristic; OS, overall survival; NLR, neutrophil-to-lymphocyte ratio; AUC, area under curve

To evaluate the association between the NLR value and patient outcomes, we compared DSS and OS in the high and low NLR groups. The 5-year DSS rate was 86.6% in the low NLR group and 69.2% in the high group. The 10-year DSS rates in patients with low and high NLR were 86.6% and 46.1%, respectively. The high NLR patients showed significantly shorter DSS than the low NLR patients (hazard ratio [HR] 4.67, 95% confidence interval [CI] 1.13–19.3, p = 0.036) (Fig. 2a). Similar to DSS, the 5- and 10-year OS rates were lower in patients with a high NLR than those with a low NLR (5-year OS, low NLR = 86.6%, high NLR = 61.5%; 10-year OS, low NLR = 80.0%, high NLR = 38.4%), and the OS was significantly worse in the high NLR group than the low NLR group (HR 4.94, 95% CI 1.94–14.9, p = 0.007) (Fig. 2b).

Fig. 2

Kaplan–Meier curves for DSS (a, HR 4.67, 95% CI 1.13–19.3; p = 0.036) and OS (b, HR 4.94; 95% CI 01.94–14.9; p = 0.007) according to NLR. DSS, Disease-specific survival; HR, hazard ratio; CI, confidence interval; OS, overall survival; NLR, neutrophil-to-lymphocyte ratio

Univariate and multivariate analyses of OS

Univariate analysis showed that more than 75% of poorly differentiated components, and a high NLR were significantly associated with poor OS (more than 75% of poorly differentiated components: HR 3.64, 95% CI 1.08–12.2, p = 0.036, high NLR: HR 5.03, 95% CI 1.35–18.6, p = 0.015). Multivariate analysis using the Cox hazard model showed that high NLR was an independent factor for poor OS (HR 6.06, 95% CI 1.57–23.3, p = 0.0087) (Table 2).

Table 2 Univariate and multivariate Cox proportional hazards regression analysis for overall survival

Variables	Univariate			Multivariate
	p value	HR	95%CI	p value	HR	95%CI
Age (≥55 vs. <55)	0.11	5.56	0.71–43.2
Sex (male vs. female)	0.45	0.64	0.20–2.04
M status (M1 vs. M0)	0.13	2.39	0.75–7.59
Residual tumor (R1, R2 vs. R0)	0.26	1.32	0.60–6.07
Poorly differentiated component (≥75% vs. ≥50%, <75%)	0.036	3.64	1.08–12.2	0.018	4.55	1.28–16.1
NLR (high vs. low)	0.015	5.03	1.35–18.6	0.0087	6.06	1.57–23.3

R0, no residual disease; R1, microscopic residual disease; R2, macroscopic residual disease; RAI, radioactive iodine therapy; NLR, neutrophil-to-lymphocyte ratio; HR, hazard ratio; CI, confidence interval

Discussion

This study showed that a high preoperative NLR was significantly associated with shorter survival in PDTC patients diagnosed according to the currently approved criteria in Japan. To our knowledge, this is the first study to identify NLR as a prognostic factor for long-term outcomes in PDTC patients.

Although NLR has been recognized as a prognostic marker in various solid malignancies [18], its prognostic impact on thyroid cancer remains controversial. Indeed, Kim et al. indicated that high NLR was associated with poor DFS in patients with DTC who underwent surgery [21], while no association between NLR and patients outcomes in DTC was found in the several studies [22, 23]. Thus, the significance of NLR as a prognostic factor has not been virtually demonstrated in DTC yet; however, afterwards, the significance of NLR as a prognostic factor has been indicated in a subset of DTC with distant metastases and RAI refractory (RR)-DTC. Fukuda et al. recently demonstrated that a high NLR was correlated with poor OS in patients with RR-DTC treated with lenvatinib, a multiple tyrosine kinase inhibitor (TKI) that became a standard therapy for locally recurrent unresectable and metastatic RR-DTC [24]. They also showed that an NLR decrease during lenvatinib treatment was associated with a better response in RR-DTC [24]. More recently, Ito et al. reported that a high NLR negatively impacted the prognosis of DTC with distant metastasis [25]. These results indicate that high NLR is correlated with the aggressive behavior of DTC.

Generally, NLR is higher in patients with clinically aggressive tumors than those with indolent tumors because rapid tumor progression is likely to cause a systemic inflammatory response, resulting in elevated neutrophil counts [21, 23]. In line with this notion, the previous studies demonstrated that high NLR was associated with large tumor volume, presence of extrathyroidal extension, and lymph node metastases in patients with DTC [17, 26]. These findings and the results of this study indicate that NLR represents disease activity and can be a marker for predicting disease progression and prognosis of follicular cell-derived thyroid cancer with aggressive features, such as PDTC and advanced DTC, including those with distant metastasis.

Immune checkpoint inhibitors (ICIs) have brought a paradigm shift in various cancer treatments. Recent studies demonstrated that ICIs show considerable promise in treating advanced DTC and ATC [27, 28], and ICIs are anticipated to be approved for thyroid cancer treatment in near future. Accumulating evidence in various solid malignancies indicates that patients with a high local immune response, represented by the abundant infiltration of lymphocytes into tumors, are likely to show favorable responses to ICIs because lymphocyte infiltration is needed to obtain the respectable ICI effects [29]. On the other hand, previous studies demonstrated that the density of tumor-infiltrating lymphocytes (TILs) inversely correlates with NLR [30, 31], and patients with abundant TILs tended to have a lower NLR. Thus, patients with a low NLR are likely to respond to ICIs, and, indeed, patients with a low NLR had prolonged survival after anti-PD-1 therapy (a representative ICI) in non-small cell lung cancer patients [32, 33]. Hence, like TKIs, NLR might become a predictive marker for ICI treatment in patients with PDTC if they are allowed for use.

This study had several limitations. First, it was a single-center, retrospective cohort study. Second, only a small number of patients were enrolled. Therefore, large-scale studies are needed to validate our results. The optimal cut-off value of NLR for PDTC also remains elusive. In this study, we used 2.88 as the cut-off value determined by the ROC curve. However, previous studies investigating the clinical impact of NLR in thyroid cancer used various cut-off values (1.6–10) [34-36]. Therefore, an optimal cut-off value should be determined in future studies. One strength of this study was that we re-evaluated the histopathological findings of the tumors diagnosed as PDTC by the pre-2015 criteria and excluded cases that did not meet the current guidelines. This re-evaluation enabled us to investigate the long-term outcomes of PDTC patients.

In conclusion, this study determined that NLR is a prognostic marker in PDTC patients. Our results underscore the importance of systemic inflammatory responses and immunity in PDTC progression.

Acknowledgement

We would like to thank Editage (www.editage.com) for English language editing.

Disclosure Statement

None of the authors have any potential conflicts of interest associated with the research.

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