Risk of Glioblastoma Multiforme in Patients Taking Ion Channel Blockers

Background Ion channels play a role in the development and progression of glioblastoma multiforme. This study investigates the association between the risk of developing glioblastoma multiforme in patients taking these medications. Methods A retrospective propensity score-matched analysis was performed using the TriNetX multinational electronic health record database for patients taking verapamil, digoxin, amiodarone, or diltiazem versus those not taking these medications. The outcome of interest was the incidence of glioblastoma multiforme. Results Verapamil users had an OR of 0.494 (p < 0.0001) of developing glioblastoma versus verapamil non-users. Patients on digoxin had an OR of 0.793 (p = 0.2393), patients on amiodarone had an OR of 0.600 (p = 0.0035), patients on diltiazem had an OR of 0.584 (p < 0.0001), and patients on verapamil, digoxin, amiodarone, or diltiazem had an OR of 0.641 (p < 0.0001) of developing glioblastoma versus patients not taking these medications. Conclusion In patients taking the ion channel blockers diltiazem, amiodarone, or verapamil, the odds of developing glioblastoma multiforme were lower than in patients not taking these medications.


Introduction
Glioblastoma multiforme (GBM) is a primary central nervous system (CNS) tumor that accounts for approximately 16% of all CNS neoplasms and causes approximately 15,000 deaths yearly in the United States [1,2]. The current school of thought suggests that this malignancy originates from the supporting cells of the CNS, known as glial cells. This tumor exhibits a particularly high affinity for invasion and spread into the surrounding brain parenchyma. Although current surgical and medical therapy is available for managing this tumor, the prognosis for patients remains dismal [3]. Only two factors have been shown to increase the risk of developing gliomas: high doses of ionizing radiation and certain inherited mutations [4]. However, several factors have been shown to worsen the prognosis of GBM, one of them being mutations in ion channels, specifically sodium, potassium, and calcium transporters, as well as the sodium/potassium-ATPase. Recent studies have shown that gliomas use these ion channels to foster their growth and invasion of the brain [5]. As such, it is possible to hypothesize that ion channel blockers could play a role in the development and progression of GBM.
This study investigates the association between the risk of developing GBM in patients taking ion channel blockers, specifically verapamil, diltiazem, digoxin, and amiodarone.

Materials And Methods
This study is a retrospective case-control study model using a multi-institutional healthcare database, the TriNetX research network, to collect data on patients diagnosed with GBM while taking verapamil, digoxin, diltiazem, or amiodarone. The TriNetX research network is a database that houses de-identified electronic medical records from several healthcare organizations spanning 57 academic medical institutions, and the information in this database is updated daily. This database contains information regarding patient demographics, diagnoses, medications, and outcomes. Since the database is federated, an Institutional Review Board approval for this study has been waived.
The TriNetX database was interrogated for patients who took the ion channel blockers verapamil, digoxin, amiodarone, or diltiazem. These were stratified into four different groups. The primary outcome of interest was the risk of GBM development in patients taking one of these drugs compared to patients who were not taking the drug. An analysis was performed for each of the drugs individually without patients taking any other ion channel blockers, as well as in a combined cohort where patients could be on any ion channel blocker. Chi-square analysis was used for categorical variables. The significance level was set as p-value ≤ 0.05.

Results
Tables 1, 2 show the baseline characteristics and measures of association for our patients taking verapamil. After matching, of the 512,098 patients using verapamil, 45 patients (0.009%) subsequently developed a glioblastoma. This is in comparison to the 512,098 patients not taking verapamil, of which 91 patients (0.018%) developed a glioblastoma (p = <0.0001, odds ratio (OR) = 0.494, 95% confidence interval (CI) = 0.346, 0.707).

Discussion
These results suggest that in patients using the ion channel blockers verapamil, amiodarone, or diltiazem, the odds of developing GBM were lower than in patients not taking these drugs. These results suggest a similar pattern for digoxin, albeit statistically insignificant. Furthermore, this association persisted when all patients were analyzed in a general group.
GBM originates from the brain's supporting cells, and these cells express a myriad of ion channels, including sodium, potassium, and anion channels [6]. Genomic analysis of mutations present in GBM has shown the presence of mutations in the genes encoding these ion channels in 90% of the glioblastoma samples examined [7]. Research suggests that mutations in these ion channels harbor a poor prognostic factor for patients by promoting proliferation, migration, and invasion of normal brain tissue by GBM. This effect is primarily believed to be mediated by the action of ion channels in promoting progression through the cell cycle [8].
Studies have shown that different types of Ca2+ selective ion channels are upregulated in GBM, where they confer a host of pro-survival benefits to the tumor, including promoting tumor invasiveness, proliferation, and resistance to apoptosis [9]. For example, diltiazem and verapamil primarily block the L-type voltagegated calcium channels. This specific Ca2+ channel is expressed in several tumor cells, and blockage of this channel inhibits cancer cell invasion. This effect is primarily mediated by inhibiting the role of these channels in the development of filopodia, thereby preventing tumor cell migration and invasion of nearby healthy tissue [10]. Furthermore, verapamil has been shown to inhibit the T-type Ca2+ channels, and inhibition of this channel has been shown to induce apoptosis in glioblastoma cells [11]. As such, these Ca2+ channel blockers may prevent tumorigenesis via myriad mechanisms, including prevention of cell cycle progression, induction of apoptosis, and prevention of aberrant migration of malignant cells.
The anti-tumorigenic effects of cardiac glycosides have been previously established [12]. In addition, in vitro studies have shown that digoxin can exhibit antiproliferative and pro-apoptotic effects in GBM [13].
Although the mechanism of action has not yet been elucidated, the current school of thought suggests that inhibition of sodium currents might be a mechanism by which digoxin exerts its anti-tumor effects. Digoxin primarily acts by inhibiting the Na+/K+ ATPase, an energy-dependent transporter that plays a role in maintaining homeostatic levels of potassium and sodium in cells. Inhibition of this channel has been shown to independently induce cell death in GBM and increase tumor cells' sensitivity to chemotherapy [14]. As such, it is plausible that digoxin can play a role in preventing the development and progression of GBM.
K+ channels also contribute to the proliferation and apoptosis resistance exhibited by GBM. Specifically, GBM overexpresses certain voltage-dependent K+ channels, which are reportedly involved in signaling pathways that promote proliferation and inhibit apoptosis [15]. Some of these effects are caused by the role of K+ channels in establishing the resting membrane potential. Changes to this baseline can alter cell-cycle progression, promoting a pro-tumorigenic profile. Clinical studies have shown that the use of inhibitors of these channels is associated with better survival in patients with GBM, again emphasizing the role of these channels in the development and progression of GBM [16]. High expression of a specific subtype of the potassium channel (Kv10.1) in GBM cells is associated with a more dismal prognosis [17]. Amiodarone is an anti-arrhythmic that can block voltage-gated potassium, calcium, and sodium channels. This drug has also been shown to reduce glioblastoma growth in vivo by exhibiting direct anti-cancer effects and antiangiogenic activity [18,19]. As such, some anti-tumorigenic effects of amiodarone are likely due to its inhibition of ion channels, which inhibit tumor cell proliferation and migration and its effect on angiogenesis.
Thus, the effect of these drugs on the development of GBM is probably due to a mixture of the various mechanisms aforementioned, including delayed progression across the cell cycle, inhibition of cell proliferation, and induction of apoptosis in de-novo malignant cells.
Several limitations exist in this study. Firstly, and most importantly, this analysis was primarily retrospective; hence, this investigation is limited to the constraints of such studies. Secondly, some information about the medication history could not be obtained from the TriNetX database. Specifically, the dosage of each medication, the indication in each patient, and the duration of usage of these medications could not be obtained. Furthermore, information about the stage and grade of each patient's GBM diagnosis could not be retrieved. The isocitrate dehydrogenase (IDH) mutation status and O6-methylguanine-DNA methyltransferase (MGMT) promoter methylation status of the tumors were unknown. The International Classification of Diseases, Tenth Revision (ICD-10) codes are primarily used for billing purposes. Finally, due to the nature of database studies, some misidentification is always present.

Conclusions
These findings suggest that in patients taking the ion channel blockers diltiazem, amiodarone, or verapamil, the odds of developing GBM were lower than in patients not taking these drugs. The same relationship was seen in patients taking digoxin; however, this association was not statistically significant. Ion channels play a fundamental role in the development and progression of GBM. Therefore, inhibition of these channels could serve as a therapeutic target for the management of GBM.