The efficacy of aspirin and metformin combination therapy in patients with rectal aberrant crypt foci: a double-blinded randomized controlled trial

Cancer is a major health concern and the leading cause of death worldwide. The incidence and mortality of colorectal cancer (CRC) continues to increase worldwide [1]. Most cases of CRC originate from adenomas [2], and their removal has been shown to reduce the risk of future development of CRC and advanced adenoma [3, 4], thereby preventing CRC-related death [5]. However, patients with adenomas and CRC constitute a high-risk group for the development of metachronous colorectal adenoma and/or CRC [6]. Therefore, a paradigm shift from surveillance for the early detection of cancer or adenomas and polypectomy to new tactics for prevention, including chemoprevention, is required to lower the burden of this disease. Several large epidemiologic and clinical studies have evaluated the possible effects of more than 200 agents, including fiber, calcium, and non-steroidal anti-inflammatory drugs (NSAIDs), such as aspirin and selective cyclooxygenase-2 (COX-2) inhibitors, in protecting against CRC development [7]. Previous studies reported that aspirin suppressed the development of sporadic colorectal adenoma [8‐10]. However, the chemopreventive effect of aspirin is limited, and these studies also reported that aspirin increased gastrointestinal bleeding. In these patients, a post-aspirin chemoprevention drug is then needed to establish CRC chemoprevention.

Patients with type 2 diabetes who were prescribed metformin have been reported to be at a low risk of cancer development, including CRC), compared with those who were not treated with metformin [11, 12]. This evidence suggests that metformin might be a candidate agent for CRC chemoprevention in diabetic patients. In previous studies, we demonstrated the chemopreventive effect of metformin in two rodent models (a genetic model and a chemically-induced cancer model) and two human studies of colorectal carcinogenesis. We demonstrated that metformin inhibited the development of intestinal polyps in adenomatous polyposis coli mice, a murine model of familial adenomatous polyposis [13]; furthermore, we demonstrated that metformin inhibited azoxymethane-induced formation of colorectal aberrant crypt foci (ACFs) by activating AMP-activated protein kinase (AMPK) [14]. Both studies were conducted in nondiabetic mice, which suggested the direct chemopreventive potential of metformin per se. We also conducted a trial involving non-diabetic human patients and showed that oral low-dose metformin (250 mg/day) was safe and suppressed the formation of colorectal ACF. In the study conducted on nondiabetic human subjects, the drug was safe [15]. Based on these findings, we performed a randomized clinical trial (RCT) and showed that low dose metformin is safe and reduced the incidence of new polyps in patients after polypectomy of the colon [16]. In that trial, we showed the safety and chemopreventive effect of metformin on colorectal carcinogenesis. However, similar to aspirin, the chemopreventive effective was limited.

The current recommended treatment for hypertension or diabetes mellitus is multiple drug combination therapy, which requires a lower dose of each drug to achieve a maximum therapeutic effect while avoiding toxic effects. Here, we hypothesize that aspirin and metformin combination therapy is more effective than single use of these drugs for CRC prevention. To test this hypothesis, this proposed study will investigate whether the combined use of aspirin and metformin shows a stronger chemopreventive effect than aspirin or metformin alone. In CRC chemoprevention trials, in general, the incidence of adenomas or the cancer itself is set as the study endpoint. Although the incidence rate of CRC is the most reliable endpoint, the use of this endpoint would be unsuitable for chemoprevention trials because of the relatively low occurrence rate of CRC in the general population [17] and the long-term observation period that it would require. ACF are defined as lesions containing crypts that are larger in diameter and stain more darkly with methylene blue than normal crypts [18‐21]. They have been reported to be more prevalent in patients with cancer and adenomas, are reduced by the use of chemopreventive drugs such as NSAIDs, are considered a reliable surrogate biomarker of CRC [22, 23]. We previously reported the usefulness of ACF as a biological marker of CRC [24, 25], and conducted a chemoprevention trial for colorectal ACF [15, 26]. The advantages of chemoprevention trials using colorectal ACF as the primary endpoint are that long-term observation is not required to evaluate the drug effect, and ACF can be estimated quantitatively. Thus, we set ACF as a suitable endpoint for this trial. To the best of our knowledge, this is the first clinical trial to investigate aspirin and metformin combination therapy as a chemopreventive strategy in patients with colorectal ACF.

This is the first study proposed to evaluate the chemopreventive effect of aspirin and metformin combination therapy in patients with rectal ACF. Given its role as an analgesic, antipyretic and agent for cardiovascular prophylaxis, aspirin has become one of the most commonly used drugs. Many studies have provided considerable evidence demonstrating its potential for the prevention of CRC. Aspirin exerts its anticancer effects through several interconnected mechanisms, including prostaglandin synthesis and catabolism in epithelial cells [30, 31]; inhibition of WNT β-catenin signalling [32, 33]; inactivation of platelets [34, 35] and the host immune response [31, 36]. Aspirin may also act through several other unknown mechanisms. Furthermore, aspirin may has been hypothesized to act synergistically with other agents [37]. Further study is needed to clear the chemopreventive effect of aspirin. Metformin (1,1-dimethylbiguanide hydrochloride) is a biguanide derivative that is widely used for the treatment of diabetes mellitus [38]. Metformin activates AMPK, which inhibits the mammalian target of rapamycin (mTOR) pathway [39]. The mTOR pathway plays an important role in the regulation of cellular protein translational machinery and cell proliferation [40]. The best-characterized downstream effector of mTOR is S6 kinase, which regulates the initiation and elongation phases of translation [41]. Activation of the mTOR pathway has been shown to accelerate cell cycle progression from G1 to S in CRC DLD-1 cells [42]. Therefore, AMPK activation may inhibit cell growth and proliferation by suppressing protein synthesis, thereby having a potent antiproliferative effect. Recent evidence indicates that metformin has a suppressive effect on tumorigenesis and cancer cell growth [43‐45]. In one study, metformin was demonstrated to activate AMPK and consequently decrease cellular proliferative activity, to produce a general decrease in protein synthesis in vitro in human breast carcinoma cells [43]. Metformin was also shown to inhibit the proliferation of human prostate cancer cells [45]. In addition, we performed a RCT and we showed that metformin reduced the incidence of new polyps in patients after polypectomy of the colon [16]. However, similar to aspirin, the chemopreventive effective was limited. In this study, we aim to evaluate the chemopreventive effect of aspirin and metformin combination therapy in patients with rectal ACF and determine whether the combined use of aspirin and metformin produces a stronger chemopreventive effect than each drug alone. Because we hypothesize that both aspirin and combination therapy may have a chemopreventive effect, a higher number of baseline ACFs (> 10) was needed to detect a difference between the two treatment effects. We previously reported a mean number of ACFs in adenomas of 6.2 ± 7.0 [24], which is why we set > 10 ACFs as an inclusion criterion. Consequently, we expect the trial to take longer because of the small number of patients who meet the inclusion criteria.

This trial may have the following limitations. First, ACFs are considered as a reliable surrogate biomarker of CRC [22], although their biological significance still remains controversial. However, setting study endpoint as ACF has large merit to reduce efforts because ACFs are quantitative and it is possible to observe changes in a short period of time. Thus, we devised a trial using ACF as the primary endpoint to evaluate the chemopreventive effects of aspirin and metformin combination therapy. Second, an intervention period of 8 weeks may be too short to allow the reliable detection of differences between the groups. However, we showed in a previous study that oral administration of metformin for 1 month suppressed the formation of colorectal ACF in humans. Other reports show that ACFs decrease in number in 8 weeks with the use of NSAIDs [23]. Therefore, we think that an intervention period of 8 weeks would be sufficient to evaluate the changes in the number of ACF. We previously conducted a short-term chemoprevention trial of metformin for colorectal ACF, and we showed the suppressive effect of the drug on the formation of ACF. Thereafter, we conducted a long-term metformin chemoprevention trial for colorectal polyps. We propose to repeat the same steps for the chemoprevention trial to investigate the combined use of aspirin and metformin. Third, our study lacks dose-response data. However, low-dose aspirin (100 mg/d) is used worldwide with reported chemopreventive effects on CRC and adenoma. Furthermore, we previously reported the effect of low-dose metformin (250 mg/d) on colorectal adenoma and ACF. Finally, our study lacks a metformin alone arm and double placebo arm, while the use of aspirin alone has not been established to suppress to the formation of ACFs. However, aspirin is one of the most efficiacious chemopreventive agents for the treatment of colorectal adenoma and CRC, which is why we used aspirin alone as the control arm in the present study. A four-arm study using a double placebo is needed in the future to investigate the combination effect of metformin and aspirin.

If this combination therapy was found to be more effective for the prevention of CRC, the impact would be significant. Therefore, we consider it of interest to determine whether the combined use of aspirin and metformin shows a stronger chemopreventive effect on the formation of human colorectal ACFs than either aspirin or metformin alone.