C-reactive protein with the survival outcomes of bladder cancer patients: a meta-analysis

Accumulating evidences suggested that serum C-reactive protein (CRP) was associated with the survival of bladder cancer patients. However, incongruent ndings have been reported. We comprehensively searched PubMed, Embase, and Web of science through August 2020 in order to nd all eligible studies on the association between CRP and the overall survival (OS), cancer-specic survival (CSS), disease-free survival (DFS) of bladder cancer patients. The pooled hazard ratios (HRs) together with their 95% condence intervals (CIs) were estimated by xed-effect model if the heterogeneity was low, and random-effect model if the heterogeneity was high. A series of subgroup meta-analysis were performed with regard to the specic characteristics of study design. of (HR = CI: Subgroup analyses supported the robust association between elevated CRP and CSS. increase had inferior


Conclusions
These ndings suggested that bladder cancer patients reported increase serum CRP had inferior prognostic outcomes.

Background
Bladder cancer is one of the most common cancer worldwide with 549, 393 new cases and 199, 922 deaths in 2018 [1]. Elderly men are predominant in bladder cancer's population with a median age for the primary diagnosis of 73 [2]. Non-muscle invasive bladder cancer (NMIBC) represents 75% of the cases but muscle-invasive bladder cancer (MIBC), which only represents the rest 25%, is more lethal [3,4]. Many studies have shown that smoking, occupational carcinogen exposure, dietary factors are potential risk factors of bladder cancer [2,5]. Despite considerable improvement in treatment of bladder cancer in the past decade, investigations on predictive capabilities of survival biomarkers are still of urgent need. It has been consistently mentioned that systemic in ammation plays a central role in cancer's initiation and progression, evidence has also been accumulated in supporting the connection between elevated systemic in ammation level and unfavorable survival of cancer patients [6].
C-reactive protein (CRP) is one of the most representative serum biomarkers for evaluating systemic in ammation [7]. Increased level of CRP has been linked to unfavorable prognosis of many cancers, like colorectal cancer [8], hepatocellular cancer [9], esophageal cancer [10], nasopharyngeal cancer, and cancers of urological system [11]. Several studies have been published in evaluating the prognostic role of CRP in bladder cancer [12][13][14][15], although most of them had concluded that elevated CRP was an independent predictor of inferior overall survival (OS) or cancer speci c survival (CSS) of bladder cancer patients, studies with insigni cant ndings also exist.
This incongruity in the association between CRP and the prognosis of bladder cancer indicates the necessity in thoroughly reviewing and synthesizing the results of existing studies. However, to our best knowledge, regarding to this topic, currently no exhaustive meta-analysis has been done. Therefore, the major aim of the current study is to perform a comprehensive meta-analysis regarding to the association between CRP and bladder cancer survival. We intend to simultaneously evaluate the in uence of CRP on multiple prognostic outcomes for bladder cancer patients, such as OS, CSS, and disease-free survival (DFS).

Searching strategy
In accordance with the PRISMA [16], eligible studies have systematically collected from 3 full-text databases (PubMed, Embase, Web of science) before August 11, 2020. We used the following words to preliminarily search for potentially relevant studies in the three databases mentioned above: ("bladder" OR "urothelial" OR "transitional cell") AND ("cancer" OR "tumor" OR "malignancy" OR "carcinoma" OR "neoplasm") AND ("CRP" OR "C-reactive protein" OR "in ammatory markers" OR "in ammatory factors" OR "in ammatory biomarkers") AND ("prognosis" OR "survival" OR "recurrence" OR "progression"). To guarantee the completeness of included studies, the reference lists of all potentially relevant studies were simultaneously checked and searched.

Inclusion and exclusion criteria
Inclusion criteria used to further screen for eligible studies were: (1) cohort design (either prospective or retrospective); (2) investigated the association between serum CRP and bladder cancer outcomes by using Cox proportional hazards model; (3) CRP was dichotomized by using speci c cut-offs; (4) report hazard ratios (HRs). The exclusion criteria were: (1) studies of case-control or cross-sectional design; (2) CRP was treated as continuous variable; (3) did not provide HR by using Cox model.

Publication quality assessment
Newcastle-Ottawa Scale (NOS) was used to access the quality of included publications in our study. The scores of NOS were ranged from 0 (lowest score) to 9 (highest score). A study with a NOS score higher than 5 was considered high-quality [17]. We only included studies with NOS scores higher than 5 in this metaanalysis.

Data extraction
First author's name, year of publication, country of origin, type of bladder cancer, cut-off value, HRs with their corresponding 95% con dence intervals (CIs), covariates in multivariate model, and p-values were extracted by two investigators independently. The extraction results were compared, and any discrepancy was solved by a third researcher.

Statistical analysis
We used STATA 15.0 software to perform all statistical analyses. The outcomes of interest in the present study were OS, CSS and DFS. The pooled HRs with 95% CIs were used to measure the association between CRP and the survival of bladder cancer patients. I-squared statistic ( ) >50% was used to de ne the statistical heterogeneity: the xed-effect model was used when heterogeneity was low, while the random-effect model was used when heterogeneity was high.
We performed the subgroup meta-analyses based on the cut-offs of CRP, types of bladder cancer, different sample sizes and origins of the studies. Funnel plot, together with Egger's and Begg's tests were used to test for publication bias.

Study selection
The owchart for studies selection process was presented in Figure 1. Initially, 683 articles were found after searching in the three databases. Two hundred and four (204) duplicated articles were excluded. For the rest 479 articles, after titles and abstracts review, 406 were further excluded (reasons for exclusion are: not bladder cancer, editorial review, did not analyze CRP, reviews or meta-analysis, published in other languages rather than English, animal studies, conference abstracts, not prognostic studies). Then 73 articles were reviewed in full text to carefully assess the eligibility, 60 of them were further excluded (reasons for exclusion are: other types of urological cancer patients mingled with bladder cancer patients, did not provide measurements of CRP, did not investigate the outcomes of study interest, case-control or cross-sectional design studies, did not use Cox proportional hazards model). In the end, 13 studies were included into our meta-analysis [12][13][14][15][18][19][20][21][22][23][24][25][26].

Study characteristics
The characteristics of included studies were summarized in Table 1 CRP with bladder cancer survival A total of 8 included studies investigated OS, high heterogeneity was observed among them ( =97%, < 0.001), therefore a random-effect model was tted. The pooled results indicated that, an elevated CRP was in general associated with poor OS of bladder cancer patients (HR=2.24, 95% CI: 1.16-4.34, p=0.017). We also combined studies which reported prognostic signi cance of CRP in CSS and DFS of bladder cancer by using xed-effect model, we found that the elevated CRP was also signi cantly associated with poor CSS (HR=1.53 95% CI: 1.36-1.72, p<0.001) and DFS (HR=2.07, 95% CI: 1.24-3.35, p=0.005) of bladder cancer (Figure 2).
Considering that different cut-offs of CRP may in uence the combined results, we performed a meta-analysis by only including studies which adopted the cutoff of 0.5mg/dl or close. Altogether 6 and 3 studies were combined for CSS and OS, and the results of random-effect models revealed that, an elevated CRP level was only prominently associated with CSS (HR=1.53, 95%CI: 1.26-1.86, p<0.001) (Figure 3).

Subgroup analysis
In order to test for robustness of the combined results, we performed a series of subgroup analysis based on different characteristics of included studies for CSS, the most popular outcome of interest among included studies. Different cut-offs for CRP (0.5mg/dl or close VS. 1 mg/dl, Figure 4A), different types of bladder cancer (Unspeci ed VS. MIBC, Figure 4B), different sample sizes of studies (dichotomized by the mean, <236 VS. ≥236, Figure 4C), different origins of studies (Europe VS. Asia, Figure 4D) were used sequentially to perform subgroup analyses. Results were generally robust with regard to different dichotomization characteristics.
In ammation has been identi ed a major cause of tumor progression [27]. The following underlying mechanisms may be involved. Firstly, in ammatory cells can produce microenvironment for tumor growth, promote angiogenesis and favor neoplastic spread and metastasis [28]. Sui et al. in their review noted that in ammatory microenvironment plays a critical role in the initiation and the progression of bladder cancer: pro-in ammatory cells (such as macrophages, suppressor cells, regulatory T cells, dendritic cells, mast cells, neutrophils and lymphocytes) and cytokines (such as tumor necrosis factor-α and interleukins) collectively contribute to bladder cancer formation and progression [29]. Moreover, the disease of cancer itself can initiate a series of systemic in ammatory responses which include hormonal disorder, and changes in blood indicators like CPR, under the in uence of neuroendocrine metabolism, hematopoietic function, and energy metabolism [30].
CRP is a highly sensitive biomarker of acute and chronic in ammation [31], and it is easy to measure in clinical practice [32]. After interleukin-6 mediated release by hepatocytes, CRP promotes the proliferation of cancer cells [33]. Therefore, elevated serum CRP goes with the creation of the tumor microenvironment, becomes a critical component of the host's response to the tumor [34]. Shrotriya et al. in their systematic review remarked signi cantly different levels of CRP between cancer patients and the healthy individuals, despite the ambiguity of its etiological role [35]. In addition, several studies have shown a strong relationship between elevated level of CRP with the poor prognosis and the progression of the disease in a variety of malignancies, including urological cancers [35,36]. CRP level has a signi cant impact on both NMIBC and MIBC. For NMIBC patients who underwent transurethral resection (TURBT), they have a high risk to progress to MIBC after the treatment [37], and CRP seems participated in that oncologic progression [30]. Moreover, radical cystectomy (RC) is the standard treatment for MIBC, and the CRP level before the surgery was associated with the survival outcome of the patients [3].
Therefore, pre-operative CRP level may to some extent de ne the eventual prognosis of bladder cancer patients. With this regard, regulate CRP level might be an option to gain survival bene ts in this group of patients. Currently, the effect of nonsteroidal anti-in ammatory drugs (NSAIDs) and cyclooxygenase-2 (COOX-2) inhibitors had already been discussed in cancer chemoprevention studies [38,39]. Parada et al. used rat bladder cancer model to evaluate the effect of celecoxide on COOX-2, and demonstrated that celecoxide had an inhibitory effect on carcinogenesis of bladder cancer, because it signi cantly reduced the serum CRP level [40]. With regard to the scarcity of available studies, whether the suppression of serum CRP by administrating NSAIDs or COOX-2 inhibitors can exhibit prognosis signi cance for bladder cancer patients remains unknown, future clinical trials are needed to investigate this important issue.
The current study is the rst thorough meta-analysis regarding to CRP and the multiple prognostic outcomes of bladder cancer. Even so, several limitations should be noticed. First of all, in consideration of statistical e ciency, when synthesizing the results, we included as many eligible studies as possible, therefore the heterogeneity of studies is apparent for some pooled estimations. Second, although in the subsequent analyses, we tried to control for heterogeneity by using subgroup analysis, we can only perform subgroup analysis based on very limited characteristics provided by the original studies, the in uence of other potential confounding factors could not be effectively discussed.

Conclusion
In summary, this meta-analysis demonstrated that elevated CRP is signi cantly associated with multiple survival outcomes of bladder cancer patients. This nding suggests that the regulation of serum CRP might be an option for treating bladder cancer patients. Future longitudinal studies of high quality are warranted to corroborate our ndings, and to evaluate the prognostic e cacy of CRP regulation treatments.

Declarations
Ethics approval and consent to participate Not applicable.

Consent for publication
Not applicable.

Availability of data and materials
All data generated in this analysis are available from the corresponding author.

Competing interests
The authors declare that they have no competing interests.