Prognostic Value of Serum Neutrophil to Lymphocyte Ratio and SUVmax in Stage I and II Colon Cancer

Objective: This study aimed to evaluate the correlation between maximal standardized uptake value (SUVmax) of primary colon cancer and serum neutrophil-to-lymphocyte ratio (NLR), and to assess the prognostic value of SUVmax and serum NLR in stage I and II colon cancer patients. Methods: In this retrospective study a total of 128 patients with pathologically confirmed stage I and II colon cancer diagnosed between January 2014 and December 2017 were included. All patients underwent F-18 Fluorodeoxyglucose (FDG) positron emission tomography (PET)/computed tomography (CT) and differential white blood cell (WBC) counts before surgery. The correlations between SUVmax and NLR were assessed. The prognostic value of SUVmax and NLR for predicting recurrence-free survival (RFS) was investigated. Results: The mean NLR was 2.2 ± 1.2, and the mean SUVmax of primary tumor was 15.2 ± 7.9. There was significant correlation between NLR and SUVmax (rho=0.2, p=0.02). Mean follow-up period was 59.8 ± 19.2 months and 12 patients experienced a recurrence. In univariable analysis, NLR (p=0.0084, HR=5.0223, 95% CI=1.5117-16.6853), C-reactive protein (CRP) (p=0.021, HR=4.1115, 95% CI=1.2380-13.6551), carbohydrate antigen 19-9 (CA19-9) (p=0.0134, HR=4.2683, 95% CI=1.3519-13.4766), and Kirsten ras sarcoma viral oncogene (KRAS) mutation (p=0.0338, HR=3.4703, 95% CI=1.0998-10.9499) were significant prognostic factors for the recurrence. In multivariable analysis, NLR (p=0.0256, HR=4.1155, 95% CI=1.1887-14.2490) and CA19-9 (p=0.0257, HR=4.139, 95% CI=1.1880-14.4200) were independent prognostic factors for the recurrence. Conclusions: Significant correlation was observed between SUVmax of primary colon cancer and serum NLR. Furthermore, in the multivariable analysis conducted on early colon cancer cases, NLR and CA19-9 were found to be independently associated with RFS. This suggested that NLR could be used as a supplementary tool for identifying patients at high risk of recurrence in early colon cancer. However, SUVmax was not associated with prognosis, suggesting that it cannot be used for predicting prognosis in early colon cancer.


Introduction
In 2020, approximately 150,000 new cases of colorectal cancer were diagnosed in the United States, resulting in 52,000 deaths from the disease, making it the third leading cause of death (Siegel et al., 2022).Due to increased use of screening colonoscopies, the incidence and mortality rates have been decreasing since 2000.(Siegel et al., 2020) Approximately 40% of colon cancer patients are diagnosed at stage I or II (localized disease), and the 5-year survival rate for these patients is around 89%.Unfortunately, within this subgroup, 11% of patients

RESEARCH ARTICLE
Prognostic Value of Serum Neutrophil to Lymphocyte Ratio and SUVmax in Stage I and II Colon Cancer (Greten and Grivennikov, 2019).Among the inflammatory markers, serum neutrophil-to-lymphocyte ratio (NLR) calculated from differential white blood cell (WBC) counts has been reported as a prognostic factor in several solid tumors (Templeton et al., 2014;Ocana et al., 2017).
F-18 Fluorodeoxyglucose (FDG) is a glucose analogue that accumulates in tumor and inflammatory cells via GLUT protein (Mochizuki et al., 2001).F-18 FDG positron emission tomography/computed tomography (PET/CT) is useful for diagnosing tumors, staging, monitoring therapy response, and detecting recurrence in colon cancer cases (Chowdhury et al., 2010;Lee et al., 2017).F-18 FDG PET/CT is also useful for evaluating inflammatory diseases (Hess et al., 2014).The degree of FDG uptake in cancer lesions is reflected by tumor microenvironment, which consists of tumor cells, tumor stromal cells, endothelial cells, immune cells, and non-cellular components (Baghban et al., 2020).Several previous studies have reported the correlation between maximal standardized uptake value (SUVmax) of primary cancer and NLR in head & neck, esophageal, pancreas, cholangiocarcinoma, and lung cancers (Sürücü et al., 2015;Jeong et al., 2017;Guo et al., 2019;Mirili et al., 2019;Seo et al., 2019;Araç et al., 2020;Werner et al., 2021;Bojaxhiu et al., 2022).However, only two studies have investigated the relationship between SUVmax and NLR in colon cancer, (McSorley et al., 2018;Xu et al., 2019) and none have specifically focused on stage I and II colon cancer.Therefore, we intended to evaluate these factors and assess their prognostic value in stage I and II colon cancer.The aim of this study was to evaluate the correlation between SUVmax of primary colon cancer and serum NLR, and to assess the prognostic significance of SUVmax and serum NLR in stage I and II colon cancer patients.

Patients
This retrospective study included a total of 128 patients diagnosed with stage I or II colon cancer between January 2014 and December 2017.All patients underwent F-18 FDG PET/CT scans prior to surgery.Differential WBC counts, serum carcinoembryonic antigen (CEA), serum carbohydrate antigen 19-9 (CA19-9), serum C-reactive protein (CRP), and serum albumin tests were conducted within 30 days before surgery.Patients with a history of cancer were excluded from the study.The pathologic stages of the colon cancer patients were determined according to the American Joint Committee on Cancer Manual, 8 th edition.(Amin MB, 2017) After curative surgery, adjuvant chemotherapy was performed in stage II patients with high-risk features (Baxter et al., 2022).All patients underwent routine follow-up evaluation including laboratory tests, colonoscopy, CT, or PET/CT every 3-12 months.Patients who did not experience recurrence were censored at the date of their last follow-up.

FDG PET/CT imaging
For all patients included in the study, oral intake and intravenous glucose injection were prohibited for at least 6 h before the PET/CT scan.Blood glucose levels of the patients were measured before administering F-18 FDG, and PET/CT scans were only conducted if the blood glucose level was below 200 mg/dl.A whole-body scan, covering the area from head to thigh (torso) was performed 60 minutes after intravenous injection of approximately 370 MBq of F-18 FDG.The PET/CT examinations were performed using PET/CT scanners (Discovery STE or Discovery 690; GE Healthcare, Milwaukee, WI, USA).The CT images were acquired using a multidetector CT equipment with a standard protocol that consisted of 140 kV, 60-80mA, and a section thickness of 3.75 mm.Emission PET data were acquired for 2 minutes per bed position.PET images were reconstructed using an ordered-subset expectation maximization iterative reconstruction algorithm with three iterations, 18 subsets, a matrix size of 256x256, and a transaxial field-of-view of 50 cm.Subsequently, the PET images were fused with the CT images.

Image evaluation
Semi-quantitative analysis was performed independently by 2 experienced nuclear medicine physicians who were blinded to the clinicopathologic results.For semi-quantitative analysis, a three-dimensional volume of interest (3D VOI) was drawn on the primary tumor, and SUVmax was calculated for the pixels within the 3D VOI.

Statistical analysis
Statistical analyses were conducted using MedCalc for Windows, version 20.210 (MedCalc Software, Ostend, Belgium).Receiver operating characteristic (ROC) curve analysis was performed to evaluate the diagnostic performance of NLR for differentiating recurrence group and non-recurrence group.Based on the optimal cutoff for NLR, patients were assigned to either low NLR group or high NLR group.The recurrence-free survival (RFS) was defined as the time from colon cancer surgery to recurrence.The Kaplan-Meier survival curve was used to assess RFS rates, and RFS curves were compared using the log-rank test.Univariate and multivariate analyses were performed, and hazard ratios (HR) with 95% confidence interval (CI) were obtained.For all statistical comparisons, P-value less than 0.05 was considered statistically significant.

Characteristics of the study population
The characteristics of the patients included in the study are shown in Table 1.A total of 128 patients were included in this study (68 male and 60 female patients; mean age, 66.5 ± 11.5 years).The pathologic staging of colon cancer was stage I in 46 patients (36%), and stage II in 82 patients (64%).None of the stage I patients received adjuvant chemotherapy.While 46 patients (56%) of stage II received adjuvant chemotherapy (FOLFOX or capecitabine).Kirsten ras sarcoma viral oncogene (KRAS) mutation was found in 39 patients (30%).High microsatellite instability (MSI-H) was found in 22 patients (17%).The mean number of harvested lymph nodes was 30.0 ± 11.5.The mean levels of CEA were 4.8 ± 9.3, and the mean levels of CA19-9 were 15.6 ± 42.8.

Discussion
In the patients with stage I and II colon cancer, the overall 5-year survival rate is relatively high at approximately 89%.However, 11% of patients experience mortality within 5 years (Siegel et al., 2022).It is important to identify patients with a low risk of relapse who may suffer a recurrence, for proper patient management.In this study, SUVmax showed a weak association with NLR.Both NLR and CA19-9 were significantly associated with the RFS of stage I and II colon cancer.obstruction, bowel perforation, lymphovascular invasion, perineural invasion, inadequate lymph node sampling (<12 nodes), and poorly differentiated histology (G3) (Rebuzzi et al., 2020).Although NLR is not considered a risk factor for adjuvant chemotherapy in these patients, patients with high NLR have a significantly higher rate of recurrence compared to those with low NLR.Therefore, NLR could help identify patients with high risk of recurrence who require more careful follow-up.
In many types of cancers, SUVmax of primary cancer has been identified as a significant factor associated with prognosis (Paesmans et al., 2010;Ghooshkhanei et al., 2014;Zhu et al., 2017;Wang et al., 2019;Lee et al., 2021b).However, majority of the studies on colon cancer have reported no significant association between SUVmax of primary colorectal cancer and prognosis.These studies often included a small number of patients with poorly differentiated tumors (Ogawa et al., 2015;Lee et al., 2017;Baik et al., 2018;Yin et al., 2021).In the present study, there was also no significant association between SUVmax and RFS in stage I and II colon cancer patients (p=0.4500).Generally, SUVmax correlates with tumor size and differentiation (Na et al., 2009).In several cancers, poorly differentiated tumors tend to have higher SUVmax than well or moderately differentiated tumors (Vesselle et al., 2008;Kidd et al., 2009;Ekmekcioglu et al., 2013).Previous studies reported that patients with a higher number of poorly differentiated clusters tend to have a poorer prognosis compared to those with a lower number of poorly differentiated clusters in early colon cancer (Ueno et al., 2012;Kinoshita et al., 2015).The small number of patients with poorly differentiated tumors in the present study may explain the lack of a significant relationship between SUVmax and prognosis (Table 1).Future studies with a larger sample size, specifically including more patients with poorly differentiated tumors are needed.
In the context of F-18 FDG PET/CT, the SUV of bone marrow and spleen may be a better indicator of systemic inflammation than the SUV of primary cancer.Previous studies have reported that bone marrow to liver ratio and spleen to liver ratio were significantly associated with prognosis of colon cancer (Lee et al., 2018;Lee et al., 2021a).However, in early colon cancer, no studies have investigated the association between prognosis and glucose metabolism of the bone marrow or spleen on F-18 FDG PET/CT.Therefore, further evaluation is necessary.
Our study had some limitations.Firstly, the number of patients with early stage colon cancer who had recurrence was small, and we did not perform an overall survival analysis.Thus, further studies with larger sample size are required.Secondly, we did not evaluate volumetric parameters such as metabolic tumor volume (MTV) and total lesion glycolysis (TLG).Although SUVmax was easy to calculate, it only represented the highest SUV within tumors and therefore does not provide volumetric information.MTV and TLG, which represented the metabolically active volume, would be more relevant as markers of systemic inflammation, and further evaluation using these parameters is warranted.
In conclusion, there was weak correlation between Chronic inflammation has been recognized as significant factor in the development of cancer (Greten and Grivennikov, 2019).The inflammatory process associated with cancer can be viewed from two perspectives.Firstly, in terms of anti-tumorigenic immune response, lymphocytes play an important role in adaptive immune responses.Cytotoxic T lymphocytes produce tumor related antigens and cytokines, that plays important role in antitumor immunity.Tumor infiltrating lymphocytes are related to improved clinical outcome in many cancers.(Dunn et al., 2004) Secondly, in the aspect of pro-tumorigenic immune response, neutrophils, specifically N1 tumor associated neutrophils, play a crucial role in cancer growth and progression by releasing various chemokines, cytokines, and enzymes.They promote tumor angiogenesis by secreting proangiogenic factors such as vascular endothelial growth factors (Galdiero et al., 2013;Shaul and Fridlender, 2018).

Figure 2 .
Figure 2. ROC Curve of NLR for Predicting Recurrence in Stage I, II Colon Cancer Patients.AUC, area under curve; NLR, neutrophil to lymphocyte ratio; ROC, receiver operating characteristic curve.

Figure 3a .
Figure 3a.Recurrence-Free Survival Curves for Stage I, II Colon Cancer Patients According to NLR.NLR, neutrophil to lymphocyte ratio