To date, no quantitative meta‐analysis has been published on this topic. In the current study, we investigated the associations between benzodiazepine use and the risk of cancer by using a meta‐analysis of observational epidemiologic studies such as case‐control studies and cohort studies.

In the meantime, observational epidemiological studies reported that there was no link between diazepam use and the risk 6 or progression 7 of breast cancer. However, recent cohort studies revealed that the use of temazepam, an intermediate acting benzodiazepine was associated with an increased risk of cancer, 8 and the benzodiazepine users were exposed to the risk of benign brain tumor about three times higher than the non‐benzodiazepine users. 9

Benzodiazepines are prescribed to handle a variety of medical conditions such as seizures, anxiety, insomnia, and panic disorder. 1 Previous in vitro laboratory and animal studies have reported controversial findings on the association between the use of benzodiazepine and the risk of cancer. Several animal studies reported that benzodiazepines increased the risk of thyroid cancer 2 or liver cancer. 3 Conversely, in vitro laboratory studies indicated that benzodiazepines might inhibit the proliferation of pituitary tumor cells 4 or have antitumor effects on colorectal and breast adenocarcinoma cells. 5

The pooled estimate calculated based on the fixed‐effect model was reported using the Woolf's (inverse variance) method when substantial heterogeneity was not found. When substantial heterogeneity was found, the pooled estimate calculated based on the random‐effects model was reported using the DerSimonian and Laird method. 12

To compute a pooled OR or RR with its 95% CI, we used the adjusted ORs or RRs and its 95% CIs in each study reporting the association between benzodiazepine use (highest use vs. never use) and the risk of cancer. We examined heterogeneity across studies using the Higgins, which measures the percentage of total variation across studies.was calculated as follows:whereis Cochran's heterogeneity statistic and df the degrees of freedom. Negative values ofwere set at zero; theresults are between 0% (no observed heterogeneity) and 100% (maximal heterogeneity).Anvalue >50% was considered to indicate substantial heterogeneity.

We investigated the associations between the use of benzodiazepine (use versus never use) and the overall risk of all cancers by utilizing adjusted data as a main analysis. We also performed subgroup analyses by type of study design (case‐control or cohort), type of cancer, gender, types of benzodiazepines, study region (US or Canada, Europe, and Asia), duration of benzodiazepine use, cumulative yearly dose, type of case‐control study (population‐based or hospital‐based), and methodological quality of study (high vs. low). To perform a dose‐response meta‐analysis, we categorized yearly cumulative doses into tertiles (low: temazepam <240 mg/year equal to <12DDDs (defined daily dose) sec/year, benzodiazepine <35 mg/year, and benzodiazepine 1–100 tablets/year; middle: temazepam 240–1640 mg/year equal to 12‐82DDDs/year, benzodiazepine 35–150 mg/year, and benzodiazepine 201–499 tablets/year; high: temazepam >1,640 mg/year equal to >82DDDs/year, benzodiazepine >150 mg/year, and benzodiazepine ≥500 tablets/year). The ATC (anatomical therapeutic chemical) code for temazepam is N05CD07.

The methodological quality of included studies was evaluated based on the Newcastle‐Ottawa Scale (NOS) for assessing the quality of case‐control studies and cohort studies in meta‐analyses. 10 A star system of the NOS ranges from 0 to 9 and is composed of three subscales: selection of studies, comparability, and exposure. We considered a study awarded stars of more than a mean score of each study type as a high‐quality study because the criteria for the high‐ or low‐quality of a study have not been established.

Three authors (Kim HB, Park YC, and Park BJ) independently evaluated the eligibility of all studies searched from the two databases. If there were disagreements on the selection of studies between investigators, they were resolved by discussion.

We included observational epidemiological studies that meet all of the following criteria: (1) a case‐control study or a cohort study; (2) investigated the associations between the use of benzodiazepine and the risk of cancer; (3) reported outcome measures with adjusted odds ratios (OR) or relative risks (RR) and 95% confidence intervals (CI). If data were duplicated or shared in more than one study, the first published study was included in the analysis. Studies that were not published in peer‐reviewed journals or only presented in academic conferences were excluded.

We searched PubMed and EMBASE using common keywords linked with benzodiazepine use and the risk of cancer in January 2016. The keywords were as follows: “benzodiazepine,” “diazepam,” “alprazolam,” “clonazepam,” “temazepam,” and “oxazepam” for exposure factors; “cancer,” “tumor,” “carcinoma,” and “neoplasm” for outcome factors. Also, we reviewed the bibliographies of relevant articles to locate additional studies. The language of publication was not restricted.

A statistically significant dose‐response relationship was observed between benzodiazepine use and cancer risk ( p for trend <0.01). When compared with never use of benzodiazepine, the pooled OR/RR for the risk of cancer was 0.70 (95% CI: 0.55–0.88) in a low dose, 1.59 (95% CI: 1.26–2.00) in a middle dose, and 2.93 (95% CI: 2.45–3.52) in a high dose (Table 3 ).

Table 3 shows findings from subgroup meta‐analyses by various factors. Benzodiazepine use was consistently associated with the increased risk of cancer in the subgroup meta‐analyses by study region, type of case‐control study (population‐based or hospital‐based), and methodological quality. Subgroup meta‐analyses by gender revealed a significant positive association in the 13 studies including both gender (OR/RR = 1.20; 95% CI, 1.17–1.22), whereas no significant relationship was observed in nine studies with only female subjects. Regarding the type of benzodiazepines, intermediate‐acting benzodiazepines (alprazolam, oxazepam, temazepam, and lorazepam) showed a significantly increased risk of cancer (Table 3 and Fig. 3 ).

As shown in Figure 2 , benzodiazepine use was significantly associated with an increased risk of cancer in the random‐effects meta‐analysis of all 22 studies (pooled OR/RR, 1.19; 95% CI, 1.16–1.21). In the subgroup analyses by study design, both case‐control and cohort studies showed there was a significant positive association between benzodiazepine use and the risk of cancer: the pooled OR/RR was 1.18 (95% CI, 1.15–1.20) for 18 case‐control studies and 1.35 (95% CI, 1.24–1.47) for 4 cohort studies, respectively. No publication bias was found in the selected studies. (Begg's funnel plot was symmetric; Egger's test, p for bias = 0.39; not shown in Figure).

We assessed the methodological quality of studies included in the final analysis based on the Newcastle‐Ottawa Scale (NOS). The range of quality scores was 5 to 8; the average score was 6.8 for case‐control studies and 7.3 for cohort studies (Table 2 ). The high‐quality studies (scores of 7 or higher in case‐control studies or 8 or higher in cohort studies) included twelve case‐control study and two cohort studies.

A total of 22 studies published between 1982 and 2014 had 1,897,603 participants (213,823 patient cases and 1,683,780 controls). The mean age was 57.8 years (range, 18 to 95 years). Table 1 shows the general characteristics of the studies included in the final analysis. Eight studies 6 , 7 , 13 , 23 , 26 - 28 investigated breast cancer, three studies involved ovarian cancer, 14 , 19 , 21 two studies looked into colon cancer, 16 , 18 and five studies involved all types of cancer. 8 , 26 - 29 They were conducted in the following countries: US ( n = 12), 6 , 7 , 13 - 15 , 18 , 19 , 21 - 25 Canada ( n = 2), 13 , 23 Sweden ( n = 3), 16 , 17 , 20 Denmark ( n = 1), 26 France ( n = 1), 29 and Taiwan ( n = 3). 9 , 27 , 28 Among 18 case‐control studies, 13 studies 7 , 13 , 14 , 17 , 18 , 20 - 27 were population‐based studies, and five studies 6 , 13 , 15 , 16 , 19 were hospital‐based case‐control studies.

Figure 1 shows a flow diagram of how we identified appropriate studies. A total of 796 articles were found by searching two databases and hand‐searching relevant bibliographies. We excluded 215 duplicated articles and additional 539 articles that did not meet the selection criteria. We reviewed the full texts of the remaining 42 articles. Among these, 21 articles were excluded because of the following reasons: not relevant to our analysis ( n = 7), insufficient data ( n = 3), using psychotropic medication other than benzodiazepine ( n = 6), data from pre‐existing cancer ( n = 3), using cancer mortality as a result, and using an acute biomarker as exposure. The remaining 21 studies including eighteen case‐control studies 6 , 7 , 13 - 27 and four cohort studies 8 , 9 , 28 , 29 were included in the final analysis (the study by Kaufman et al. 13 was considered as two separate case‐control studies). All of the included articles were written in English.

Discussion

In the current meta‐analysis of observational epidemiological studies, we found that benzodiazepine use was associated with an increased risk of cancer. Subgroup meta‐analyses by various factors also showed similar findings. Additionally, these associations were observed in a dose‐response manner.

There are several possible explanations for the increased risk of cancer with the use of benzodiazepine. First, some studies have linked use of benzodiazepine to numerous sources of infection that can increase the occurrence of cancer. Benzodiazepines can develop several viral infections that could increase the risk of cancer. In a prospective study of the AIDS Care Cohort and Vancouver Injection Drug Users Study (VIDUS) cohorts followed‐up from 1996 to 2013 in Canada,30 there was a significant relationship between the use of benzodiazepine and seroconversion of Hepatitis C virus (HCV). A long progress of chronic hepatitis C to liver cirrhosis by HCV gives rise to hepatocellular carcinoma (HCC).31 A 15‐year prospective cohort study of 925 participants who had HCV infection reported that a cumulative risk for HCC increased from 6.4% for low levels of HCV RNA to 14.7% for high levels of HCV RNA (p < 0.001).32

Another plausible agent connecting the use of benzodiazepine to the increased risk of cancer could be human immunodeficiency virus (HIV). When a total of 1682 subjects who had no HIV at first were followed for median of about 80 months,33 benzodiazepine use was significantly associated with an increased rate of HIV seroconversion (Adjusted Rate Ratio:1.50; 95% CI: 1.01–2.24). HIV infected people are more prone to get Human papilloma virus (HPV) infection.34 In addition, HIV might change the natural course of immune control associated with HPV and enhance the development of squamous intraepithelial lesions of cervix.34 In a study of 310,000 AIDS patients in the U.S. (257 605 males and 51 760 females), HIV infection tended to increase the risk of HPV related carcinomas such as anal cancer besides cervix cancer during the 10 years of follow‐up period.35

Benzodiazepine also seemed to promote bacterial infections that can raise the occurrence of various kinds of carcinomas. In a study of mice, diazepam aggravated a possibility of infection with Klebsiella pneumonia (K. Pneumoniae).36 In a 11‐year followed retrospective study of 2,294 subjects with pyogenic liver abscess in Taiwan, colorectal cancer incidence was 2.68 times more common (95% CI, 1.40–5.11) in patients with K. pneumoniae than patients without K. pneumoniae.37 In vitro administration of diazepam to adult hamsters showed a detrimental effect on host protection against Mycobacterium.38 Mycobacterium, an important respiratory pathogen, in turn could increase the development of lung cancer by DNA integration.39 Our meta‐analysis also demonstrated that lung cancer developed 20% more often among benzodiazepine‐users than among non‐users of benzodiazepine.

Second possible biological mechanism is inflammation. Inflammation can be an important factor as a linkage between the use of benzodiazepine and the risk of cancer risk because chronic inflammation might be associated with use of benzodiazepine. Alprazolam administration can cause chronic inflammation induced by “cotton wool granuloma”.40 Also, benzodiazepines might increase the levels of inflammation mediators such as histamine, prostaglandins or cytokines.41 Administration of midazolam intravenously and diazepam per oral before undergoing a surgery could lead to inhibition of neutrophil apoptosis by affecting the depolarization of mitochondrial membrane.42 Neutrophil apoptosis plays an essential role in retaining homeostasis of the immune system and prevent the damages of host organs by facilitating the immune response. Another experimental report of adult male rats which received alprazolam and clonazepam for 4 weeks showed that the inflammatory toxic effects of benzodiazepine could emerge through decline of anti‐SRBC (Sheep RBC) antibody titers and interleukin‐2 (IL‐2) levels.43 Anti‐SRBC antibodies are used for a sensitive tool to evaluate the degree of inflammatory changes. Natural killer (NK) cells and T‐lymphocyte cells produce IL‐2, and decreased levels of IL‐2 indicate that suppression of the immune system could be progressing. In an Ehrlich tumor‐bearing mice study, diazepam injection diminished the amount of leukocytes and the macrophage function44 in a dose‐response manner. Besides the gamma‐aminobutyric acid A (GABA‐A) receptor complex existing in central nervous system, peripheral‐type binding sites (PBRs) of benzodiazepine have been found both in immune cells and carcinoma cells.45 The association between PBRs density and progression of cancer cells might exist. Inflammation is a crucial factor for cancer.46 Increased levels of proinflammatory cytokines such as interleukin‐1 (IL‐1) and tumor necrosis factor (TNF) could aggravate the course of disease in non‐Hodgkin's lymphoma and gastric cancer, respectively.47, 48 Additionally, tumor associated macrophages (TAMs), which are assembled during the inflammatory reaction, are markers of a poor prognosis when they exist in the tissue of cancer.49 Therefore, inflammation due to benzodiazepines might lead to the development of cancer.

Third, regarding esophageal cancer, benzodiazepines can loose the tone of lower esophageal sphincter (LES) by relaxation of smooth muscles in esophagus.50 In addition to myogenic influences on LES pressure, anticholinergic effects of benzodiazepine on vagal nerve could lead to a decrease of LES pressure.51 The LES plays a critical role in preventing gastric acid from flowing backward to esophagus. This might affect the possibility of developing esophagus carcinoma.

Another reason other than biological mechanism is that cancer patients suffer from psychiatric conditions such as insomnia, anxiety, and depression, which are frequently accompanied by the use of benzodiazepines.52 For example, more than one third of cancer patients state anxiety.53 While the frequency of major depressive disorders in general is 3–4%,54 it increases to 5–26% in advanced tumor patients.55 As a result, cancer patients are more likely to use benzodiazepines than general populations.56

Our study has important limitations. First, a small portion of the included studies in our analysis adjusted tobacco smoking and alcohol drinking as a confounding factor, both of which are well known important factors for developing cancer. Out of the 22 included studies, only eight studies8, 17-20, 22, 23, 29 adjusted tobacco smoking as a confounding factor, and seven studies8, 17, 18, 20, 22, 23, 29 adjusted alcohol intake as a confounding factor. Thus, we are unable to exclude the confounding effect of these important factors such as smoking or alcohol drinking regarding the association between benzodiazepine use and the risk of cancer. Another important limitation is that we only included observational epidemiological studies such as case‐control studies and cohort studies because few randomized controlled trials have been published on this topic so far. In general, case‐control studies are more susceptible to biases, such as selection bias and recall bias than cohort studies. These biases might lead to spurious associations. Also, cohort studies have a lower level of evidence than randomized controlled trials. Thus, our meta‐analysis does not provide the high level of evidence.

In conclusion, our meta‐analysis of observational epidemiological studies found that benzodiazepine use was associated with an increased risk of cancer. Further large randomized controlled trials providing a higher level of evidence should be conducted to confirm our findings.