Prognostic value of tumor-stroma ratio in clear cell renal cell carcinoma after surgery

Introduction

Renal cell carcinoma (RCC) represents around 2–3% of all cancers. RCC is the most common solid kidney lesion and accounts for approximately 90% of all kidney malignancies (1). Clear cell renal cell carcinoma (ccRCC) is the predominant type accounting for 85–90% of cases (2). The incidence and mortality rates are increasing annually, the incidence has increased fivefold since 1970s and the mortality rate has increased twofold (3).

Radical nephrectomy is the gold treatment for RCC. Tumor stage, tumor grade, size, lymph node metastasis are definite prognostic factors. About 25–40% of patients may have recurrence after surgery, and the 5-year survival rate after metastasis is less than 10% (4). Clearly, in addition to pathological staging, there is still a need to explore new prognostic markers to optimize patient management and evaluate the prognosis of cancer patients.

The invasive growth of tumors is influenced by the mutual synergistic effects of tumor cells and tumor stroma in the tumor microenvironment. And tumor stroma is commonly tumor-type specific and being highly dynamic and heterogeneous. The tumor stroma ratio (TSR) has been reported in a variety of epithelial solid tumors, which can be a valuable predictor for evaluating the prognosis and treatment outcome of cancer patients (5). However, the prognostic value of TSR in ccRCC is still unclear. The aim of this study is to evaluate the prognostic value of TSR in patients with ccRCC after radical surgery. We present this article in accordance with the STROBE reporting checklist (available at https://tau.amegroups.com/article/view/10.21037/tau-23-666/rc).

Methods

Patients

A total of 569 patients with ccRCC undergoing nephrectomy or partial nephrectomy between January 2010 to December 2015 in Peking University First Hospital were retrospectively analyzed. Preoperative imaging examinations, such as chest X-ray, urinary system ultrasonography and computerized tomography (CT) or magnetic resonance imaging (MRI), were used routinely. Demographic and perioperative data as well as follow-up data were collected retrospectively. No distant metastasis was confirmed by preoperative imaging examination. Histologic subtyping was determined by at least two experienced pathologists. Histological diagnosis was determined according to the 2004 World Health Organization (WHO) classifications. The pathologic staging was determined according to the 2010 Tumor Node Metastasis (TNM) classification of American Joint Committee on Cancer (AJCC). The study was conducted in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by the Ethics Committee of Peking University First Hospital (No. 2023022) and individual consent for this analysis was waived due to retrospective nature of the study.

Pathological evaluation

The tumor tissues were stained with hematoxylin-eosin (HE). First, the most obvious area of tumor invasion was selected under 50× magnification, and the tumor cells and stroma could be seen. Then, the percentage of tumor cells in each field was evaluated at 100× magnification, and the remaining fraction was the percentage of stroma to calculate the TSR inside the tumor tissue. The presence of tumor cells at the boundary of the field of view taken was ensured. We evaluated TSR serially in a minimum of 10%, such as 10% for TSR between 0% and 10%, 20% for TSR between 10% and 20%, and so on until 100%. Five 100× visual fields were selected for evaluation, and the highest value was taken as the final TSR score of the patient. In order to simplify the operation and standardize the scoring criteria, we selected ten images as a reference for different tumor stroma proportion scores (Figure S1). Finally, the patients were divided into low-TSR group (TSR <50%) and high-TSR group (TSR ≥50%) according to the demarcation criteria proposed by Mesker et al. (6) (Figure 1). All pathological specimens were reviewed by a panel of experienced urological pathologists and any divergence in scoring was resolved through discussion with a third pathologist.

Figure 1 TSR evaluation in HE slides. (A) Low-TSR; (B) high-TSR. TSR, tumor stroma ratio; HE, hematoxylin-eosin.

Follow-up

Postoperative scheduled follow-up occurred every 3 months for 2 years and every 6 months for 3 years, and annually by telephone interview or clinical visit. Patients were followed up by routine blood and urine tests, biochemical tests and physical examination were performed at each visit. Imaging evaluations, including chest X-ray, abdominal ultrasound, abdominal CT or MRI were performed every 6 months for 5 years and then annually thereafter.

Statistical analysis

All statistical analysis was performed using SPSS software version 25.0 (IBM Corporation, Armonk, NY, USA). Categorical variables were shown as frequency. Numerical variables were shown as mean ± standard deviation (SD) or median. The Mann-Whitney U test was used for continuous variables. Pearson’s Chi-squared test or Fisher’s exact test was used for categorical data. Kaplan-Meier method with the log-rank test was used to compare survival curves. Univariate and multivariate Cox proportional hazards regression analyses were performed to determine the association between risk factors and prognosis. A value of P<0.05 was considered statistically significant.

Results

The clinicopathologic demographics of the patients are shown in Table 1. The mean age of 569 patients was 56.84±12.76 years old. There were 344 patients (60.5%) of 60 years old or below and 225 patients (39.5%) being over 60 years old. There were 401 males and 168 females. The pathological stage information was as follows, with 448 cases (78.7%) in T1 stage, 47 cases (8.3%) in T2, 74 cases (13.0%) in T3. According to the TSR, patients were divided into low-TSR group (n=333, 58.5%) and high-TSR group (n=236, 41.5%). There were significant differences in pathological stage, tumor grade, diameter, hemorrhage, cystic, necrosis and lymphovascular invasion between the two groups (P<0.05). There was no significant difference in age group, gender, obesity and tumor side (P>0.05). The median follow-up time was 67.0 months (interquartile range, 33.0–72.0 months). Of 569 patients, 39 patients died, of which 26 died due to the tumor, and 49 patients had distant metastasis. The 5-year overall survival (OS), cancer-specific survival (CSS) and metastasis-free survival (MFS) were 91.2%, 94.6% and 91.0%, respectively. The 5-year OS, CSS and MFS were 84.2%, 89.7% and 82.7% in the high-TSR group and 96.1%, 98.0% and 96.0% in the low-TSR group (P<0.05) (Figure 2).

Figure 2 Comparison of prognosis between the high-TSR group and the low-TSR group. (A) OS; (B) CSS; (C) MFS. TSR, tumor stroma ratio; OS, overall survival; CSS, cancer-specific survival; MFS, metastasis-free survival.

On univariate analysis, age >60 years, tumor side, tumor diameter, tumor stage and grade, necrosis, lymphovascular invasion and TSR were closely related to OS (P<0.05). Multivariate analysis showed that age >60 years [hazard ratio (HR) =2.455, 95% confidence interval (CI): 1.292–4.668, P=0.006], tumor grade (HR =6.580, 95% CI: 3.276–13.216, P<0.001) and TSR (HR =2.611, 95% CI: 1.265–5.387, P=0.009) were independent prognostic factors for OS (Table 2), We used Cox regression coefficients to generate a corresponding nomogram (Figure S2). Univariate analysis showed that tumor diameter, tumor stage and grade, lymphovascular invasion and TSR were closely related to CSS. Multivariate analysis showed that tumor stage (HR =3.213, 95% CI: 1.437–7.184, P=0.004), tumor grade (HR =6.102, 95% CI: 2.664–13.976, P<0.001) and TSR (HR =2.653, 95% CI: 1.063–6.621, P=0.03) were independent prognostic factors for CSS (Table 3). Univariate analysis showed that tumor diameter, tumor stage and grade, hemorrhage, cystic, lymphovascular invasion and TSR were closely related to MFS. Multivariate analysis showed that tumor stage (HR =4.805, 95% CI: 2.677–8.624, P<0.001), tumor grade (HR =6.423, 95% CI: 3.432–12.020, P<0.001), hemorrhage (HR =0.514, 95% CI: 0.265–0.996, P=0.049) and TSR (HR =2.370, 95% CI: 1.264–4.443, P=0.007) were independent prognostic factors for MFS (Table 4).

Discussion

Tumor tissue is composed of tumor cells and the surrounding complex stroma. Tumor microenvironment constitutes the immediate niche surrounding tumor tissues and plays the critical role of in tumorigenesis (7). As an essential element of the tumor microenvironment, the tumor stroma affects tumor biology and contributes to tumor initiation, progression, metastasis (8).

TSR was first applied to the prognosis evaluation of colon cancer by Mesker et al. in 2007 (6). In his study, TSR refers to the ratio of tumor cells to stroma in tumor tissue. At the same time, the measurement steps of TSR and TSR cut-off value were described in detail, which provided a basis for subsequent research. TSR can reflect the amount of stromal component around tumor cells, and can independently predict the prognosis of tumor. The TSR has been reported in a variety of epithelial solid tumors, such as colorectal carcinoma, breast cancer, gastric cancer, lung cancer, esophageal cancer, etc. (9).

Although the high-TSR is remarkably correlated with poorer survival outcomes, there been no reports on the role of TSR in ccRCC so far. In this study, we performed pathological image acquisition, score acquisition and prognostic analysis of HE slides of patients with ccRCC. The evaluation of TSR has the advantages of simplicity, no additional cost, and high reproducibility. If further research shows that TSR is indeed a new independent prognostic factor for various solid tumors, TSR can be used as a routine measurement indicator included in tumor pathological examination, so as to better evaluate the risk of patients and develop more practical and effective individualized treatment plans.

The outcomes of the present study pointed that TSR was significantly associated with OS, CSS and MFS in univariate and multivariate survival analyses. TSR was shown to be correlated with the aggressive clinicopathological characteristics of ccRCC patients. Increased stromal proportion in ccRCC was also associated with poor prognosis and increased metastasis risk. Our study is the first to confirmed that TSR is an independent prognostic factor for ccRCC patients.

Carcinoma is composed of cancer cells and the surrounding stroma derived from normal tissues. In normal tissue, the stroma serves as a barrier by inhibiting the invasion and proliferation of tumor cells, but stromal components can promote tumor progression in tumor tissue (10). Tumor cells may change the properties of the stromal tissues by exchanging some enzymes and cytokines with its surrounding stroma. This process leads to the formation of a microenvironment conducive to tumorigenesis and invasion (11). Tumor cells invade the stroma by penetrating the basement membrane, which eventually leads to the mesenchymal transition activated by the normal stroma. This process promotes fibroblast proliferation and extracellular matrix deposition, which expands the tumor stroma (12). The interactions between stromal cells and direct or indirect interactions with tumor cells not only acquire abnormal phenotypes or functions, but also change the function of tumor stroma and promote tumor development through various molecular signaling pathways (8). Tumor stroma is composed of a series of cells, including endothelial cells, cancer-associated fibroblasts (CAFs), immune cells, glial cells, epithelial cells, extracellular matrix and extracellular molecules. CAFs, as main cellular components of the tumor microenvironment, can promote the proliferation of epithelial cells and regulate the biological activity of tumor cells through cytokine-mediated signal transmission and extracellular matrix remodeling. So it plays an important role in tumor occurrence, invasion and metastasis (5). Activated CAFs can secrete special growth factors to induce normal cells to undergo malignant transformation. Stromal cell derived factor, vascular endothelial cell derived growth factor and cytokines can promote angiogenesis and lead to the occurrence of tumors. In addition, osteoblasts can attract cancer cells into the bone marrow and promote malignant tumor cells bone metastasis. Adipocytes can affect cancer metabolism and are involved in tumor establishment and progression. Macrophages are another key component of the tumor stroma, etc. (13).

The TSR can provide tumor microenvironment information in addition to TNM stage and pathological grade to the prognosis evaluation of ccRCC patients during routine pathological examination, which is beneficial to improve the risk stratification, tumor management and prognosis evaluation of tumor patients. TSR is suitable for clinical pathological diagnosis of ccRCC due to its advantages of easy acquisition, low cost, convenient evaluation and unified standard (9). Besides, evidence supporting the use of robot-assisted partial nephrectomy for various complex scenarios of RCC has continuously grown over the past decade (14). But limitations remain, most studies being retrospective, often lacking a detailed and long-term reporting of functional and oncological outcomes. In light of this study’s focus on the prognostic value of TSR in ccRCC, it might be beneficial to help improve some prognosis information for the treatment of complex renal masses.

There are some limitations in this study. The retrospective study was performed in a single medical center, which may cause a potential selection bias. Most of the patients in this study had localized tumors, and few patients had advanced or metastatic tumors, therefor, the correlation between TSR and lymph node and distant metastasis has not been evaluated. We will continue to advance this study and continue to enroll a larger sample of patients to minimize limitations. We are optimistic that this initial study can serve as a foundation for developing effective predictors for ccRCC. A larger number of series are needed to confirm these results and achieve further evidence about the clinical effect.

Conclusions

TSR is a new independent prognostic risk factor for ccRCC patients. The assessment of TSR is simple and cost-effective, and it is a useful supplement added to the pathological evaluation system. More trials and longer follow-up are needed for further confirmation.

Acknowledgments

Funding: This study was supported by Beijing Municipal Natural Science Foundation (grant number 7222190).

Footnote

Reporting Checklist: The authors have completed the STROBE reporting checklist. Available at https://tau.amegroups.com/article/view/10.21037/tau-23-666/rc

Data Sharing Statement: Available at https://tau.amegroups.com/article/view/10.21037/tau-23-666/dss

Peer Review File: Available at https://tau.amegroups.com/article/view/10.21037/tau-23-666/prf

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tau.amegroups.com/article/view/10.21037/tau-23-666/coif). The authors have no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. The study was conducted in accordance with the Declaration of Helsinki (as revised in 2013). The study was approved by the Ethics Committee of Peking University First Hospital (No. 2023022) and individual consent for this analysis was waived due to retrospective nature of the study.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

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