Drug repurposing in oncology: a systematic review of anticancer effects of Lanatoside C at the molecular level

Abstract

Cancer continues to pose a major global health burden, with rising incidence and mortality rates. The protracted timelines, and high costs associated with traditional drug development highlight the urgent need for alternative drug development strategies. Drug repurposing, which involves identifying new anticancer uses for existing FDA-approved drugs, offers a promising and cost-effective approach. Lanatoside C, a cardiac glycoside approved for heart conditions, has recently gained attention for its potential anticancer properties. This systematic review consolidates preclinical evidence on the anticancer effects of Lanatoside C, focusing on its molecular mechanisms of action across various cancer types in both in vitro and in vivo models. A systematic search was conducted to identify preclinical studies assessing Lanatoside C’s effects on cancer cell lines and animal models. Studies were included if they evaluated anticancer efficacy and elucidated molecular mechanisms. Data extraction and methodological quality assessment were performed independently by multiple reviewers, using the Toxicological Data Reliability Assessment Tool (ToxRTool). Eighteen studies met inclusion criteria. Lanatoside C consistently inhibited cancer cell proliferation, induced apoptosis, and caused cell cycle arrest (primarily at the G2/M phase) in a dose-dependent manner. Mechanistically, Lanatoside C modulated key signaling pathways, like Wnt/β-catenin, PI3K/AKT/mTOR, MAPK, JAK/STAT, and ER stress/GRP78. Additional mechanisms included ferroptosis induction in lung cancer, and TRAIL-mediated apoptosis in glioblastoma. Both in vitro and in vivo studies demonstrated significant antitumor effects, supporting the translational potential of Lanatoside C as a repurposed anticancer agent. In conclusion, available Preclinical evidence indicates that Lanatoside C exerts broad-spectrum anticancer effects via modulation of different molecular pathways. These findings demonstrate its clinical utility as a monotherapy and in combination regimens, while emphasizing the need for rigorous translational and safety studies to facilitate its integration into oncology practice.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12885-025-15062-3.

Introduction

Cancer remains one of the pressing global health challenges, with epidemiological data revealing a concerning trajectory. The 2022 Global Cancer Statistics reported approximately 20 million newly diagnosed cancer cases worldwide and 9.7 million cancer-related deaths. This corresponds to an estimated lifetime risk of developing cancer in one of every five individuals globally, with one in nine men and one in twelve women dying from the disease [1]. Importantly, low- and middle-income countries bear 70% of cancer-related deaths, a disparity exacerbated by limited access to advanced therapies and preventive care [2]. Projections indicate that the global cancer burden will rise to 35 million cases by 2050, representing a 77% increase from the 20 million cases estimated in 2022 [1]. Given this alarming trajectory, there is an urgent need for innovative treatment paradigms to better prepare for these projections. Furthermore, the rise of treatment resistance of cancer with existing therapeutic approaches, coupled with the exorbitant costs and extended time required for novel drug development, has created an urgent need for a cost-effective strategy [3–5]. One such strategy for identifying potential anticancer agents is drug repurposing.

The traditional method of developing new drugs usually begins with preclinical studies, followed by a series of clinical trials. During the preclinical studies, scientists assess how a potential drug behaves by examining its toxicity, efficacy, pharmacodynamics, and pharmacokinetics in laboratory-grown human cancer cells and different animal models. If the initial results from the preclinical studies are promising, the drug progresses to clinical testing in humans. The clinical trials are in three main stages: Phase I to assess safety, Phase II to assess effectiveness, and Phase III for comparative evaluation with existing drugs. This entire development process is both time-intensive and costly, typically spanning 10 to 15 years and requiring an estimated investment of $1 to $2 billion. Despite this substantial commitment, fewer than 1% of candidate compounds advance to clinical trials, and an even smaller fraction ultimately receive regulatory approval [6–9].

However, drug repurposing focuses on exploring new therapeutic applications for drugs already approved by Food and Drug Administration (FDA). This approach takes advantage of drugs initially developed for one condition but later found to have benefits in treating other diseases. In recent years, this strategy has been gaining prominence [10]. Kirtonia et al., discussed this drug repurposing strategy and its potential transformative impact in the field of oncology [11]. Interest in drug repurposing has risen in recent years. Some of the successful pharmaceuticals, such as chlorambucil and busulfan, were originally derived from mustard gas, a toxic compound which was used in chemical warfare in the 20th century. Though these compounds were originally designed as alkylating agents, they were later discovered to be effective in the treatment of leukemia after decade-long research [12]. Cardiac glycosides, including Lanatoside C (Lan C), approved by the FDA for the treatment of heart conditions (such as cardiac arrhythmias and hypotension, primarily by inhibiting the Na+/K+-ATPase), exemplify this repurposing potential [13–17]. This inhibition, crucial for cardiac function, has also been implicated in various cellular processes relevant to cancer development and progression, making these compounds attractive candidates for drug repurposing.

Recent in vitro and in vivo studies have shown that certain cardiac glycosides, such as ouabain, digoxin, lanatoside C, and digitoxin, have anticancer properties. These drugs have been shown to kill senescent cells, trigger apoptosis, and dissociate clusters of circulating tumor cells into individual cells, thereby helping to prevent cancer metastasis [15, 18, 19].

The literature exploring the potential of Lanatoside C, an FDA-approved cardiac glycoside, in cancer treatment is extensive; however, no systematic synthesis of in vitro and in vivo studies elucidating the molecular mechanisms and pathways targeted by Lanatoside C is available. This systematic review seeks to address these gaps by consolidating evidence on Lanatoside C’s anticancer efficacy across in vitro and in vivo models. It further maps conserved and cancer-specific molecular mechanisms of Lan C across different cancer types and evaluates its translational potential, including combination therapy approaches and safety considerations.

Methodology

In this systematic review, we followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines [20]. The PICO criteria were used to develop the research questions: population—cancer cell lines derived from malignant tumors; intervention—treatment with Lanatoside C; comparison—not applicable; outcomes—both mechanistic and phenotypic outcomes were considered. Mechanistic outcomes included modulation of molecular pathways involved in cancer progression (e.g., PI3K/AKT, MAPK, Wnt, JAK/STAT, etc.). Phenotypic outcomes included changes in cellular behavior such as reduced proliferation, cell cycle arrest, inhibition of invasion and migration, and induction of cell death through apoptosis or other mechanisms. Our research questions were: “Is Lanatoside C a potential anticancer agent?”, “What are the molecular mechanisms regulated by Lanatoside C in malignant cell lines?” and “Does Lanatoside C exert a modulatory effect and act as a potential antineoplastic agent?”.

Search strategy

We searched the PubMed, EMBASE, Scopus, and Google Scholar databases to identify primary preclinical research that evaluated the in vitro and in vivo effects of Lanatoside C on cancer cells. Additionally, the references of pre-selected articles were hand-searched. The search was conducted from database inception to date. Several Medical Subject Headings (MeSH) terms and keywords were combined. Some of the important search terms used include Lanatoside C (Lan C), cardiac glycosides, anticancer effects, in vivo/in vitro studies, preclinical study. We also made use of boolean operators (AND, OR) to refine our search.

Study selection and selection criteria

All retrieved articles were screened independently by two independent reviewers (O.O.O, E.O.O) based on predefined eligibility criteria. Titles and abstracts were initially assessed for relevance, followed by a full-text review of potentially eligible studies. This was done with the Rayyan software (Rayyan Systems Inc, USA). Any discrepancies were managed by discussion with a third reviewer (T.J.O). Studies were included if they met the following criteria: The studies had to be preclinical studies (in vitro and in vivo studies) that evaluated the anticancer effects of Lanatoside C and highlighted the molecular mechanisms of Lanatoside C. Additionally, only full-text papers published in English were included. Studies were excluded from this review if they used only in silico analysis and if the methodology of the study was not properly reported.

Data extraction and analysis

Data from each eligible study were independently extracted by two reviewers (O.O.O, B.T.O) using a pilot-tested data extraction sheet which was custom-developed by the research team. The extracted information included: authors, year of publication, country, tissue origin of cell lines, dosage of Lan C, treatment duration, effects on biological pathways, effects on cell sensitization to chemotherapy and radiotherapy, and main findings of study (cell proliferation, cell viability, cell cycle arrest, apoptosis, Half-maximal inhibitory concentration (IC₅₀). Any discrepancies between the reviewers were resolved through discussion or, when necessary, adjudication by a third reviewer (E.O.O). The results of the individual studies were then summarized.

Methodological quality assessment

Due to lack of universally accepted tools for evaluating the quality of pre-clinical studies, the Toxicological Data Reliability Assessment Tool (ToxRTool)—a validated instrument specifically designed for assessing the reliability of toxicological data—was utilized [21, 22]. ToxRTool is divided into two assessment modules: one for in vitro studies, containing 18 criteria, and another for in vivo studies, comprising 21 criteria (supplementary material 2). Each criterion is scored as either 1 (yes) or 0 (no). Based on total scores, studies are categorized as follows: “reliable without restrictions” (scores of 15–18 for in vitro and 18–21 for in vivo), “reliable with restrictions” (scores of 11–14 for in vitro and 13–17 for in vivo), and “not reliable” (scores below 11 for in vitro and below 13 for in vivo) [23]. Additionally, if a study scored 0 on any of the critical items—six for in vitro and seven for in vivo—it was automatically classified as not reliable.

Results

Study characteristics

Out of a total of 1,195 records screened, 18 studies were incorporated into this systematic review, having met the inclusion criteria [24–41]. Figure 1 illustrates the study selection process in accordance with the PRISMA guidelines. All included studies examined the effects of Lanatoside C—an FDA-approved cardiac glycoside—across a range of cancer types, including breast, lung, colorectal, prostate, liver, and hematologic malignancies. All studies evaluated the anticancer potential of Lanatoside C, focusing on its effects on cellular proliferation, apoptosis, cell cycle regulation, migration, and invasion. All included studies employed in vitro experimental models, with ten of the eighteen also reporting in vivo findings; however, no human clinical studies had been published at the time of this review. These studies were published between 2001 and 2025 and were written in English. The majority were conducted in Asia (China: 6; Korea: 3; Japan: 1; Taiwan: 1), while four studies were conducted in the United States. Study designs, cancer types, dosage regimens, model systems, and outcome assessments varied widely across the studies. The general characteristics and findings are presented in the supplementary material 1.

Fig. 1.

Flowchart of literature search and selection criteria

In vitro anticancer effect of lanatoside C on different cancer types

Breast cancer

A total of three studies examined the effects of Lanatoside C on breast cancer using cell lines such as MCF-7, a model for Estrogen Receptor (ER)-positive breast cancer, used to study hormone-driven growth and antiestrogen responses, and MDA-MB-231, a model for aggressive, ER-negative, Triple-Negative Breast Cancer (TNBC) [26, 36, 41]. Findings from these studies indicated that Lanatoside C inhibited cell proliferation in these cell lines in a dose-dependent manner. It also disrupted cell cycles at critical checkpoints (G2/M phase), halting cell division and ultimately inducing apoptosis in breast cancer cells. Hu et al. reported a number of the pathways through which Lanatoside C exerts its anticancer effects including inhibition of the Wnt/β-catenin signaling pathway via GSK-3β activation, c-Myc degradation via the proteasome-ubiquitin pathway, caspase-9 cleavage with upregulated cleaved PARP and mitochondrial-mediated apoptosis (evidenced by loss of mitochondrial membrane potential and increased reactive oxygen species generation) [26]. Similarly, Reddy et al. identified the Wnt/β-catenin, MAPK, JAK/STAT, PI3K/AKT/mTOR signaling pathways as key targets of Lanatoside C in breast cancer cells. Apoptosis was also mediated by decreased antiapoptotic BCL-2 and increased proapoptotic BAX expression [36]. Both studies also reported Lanatoside C-induced G2/M cell cycle arrest. Summaries of these pathways are presented in Table 1 and Fig. 2.

Table 1.

In vitro anticancer effect of lanatoside C

Author	Cancer Type	Cell Lines	Pathway Affected	Cell Proliferation In Vitro	Apoptosis/Cell Cycle Interruption
Ha et al. [24]	Pancreatic cancer	PANC-1, CFPAC-1	ER stress/GRP78 pathway	Suppressed under ER stress and hypoxia	Caspase-7 activation; apoptosis confirmed via FACS
Durmaz et al. [25]	Liver cancer	Huh7, HepG2, FOCUS, Mahlavu	MAPK/ERK, PI3K/AKT	Inhibited in a dose-dependent manner; especially sensitive in MKN-45	G2/M arrest; PARP & caspase-3 cleavage; ROS accumulation
Hu et al. [26]	Gastric cancer, breast cancer, tongue cancer, liver cancer	MKN-45, SGC-7901 (gastric); MCF-7 (breast); HN4 (tongue); HepG2 (liver)	Wnt/β-catenin signaling via GSK-3β activation; c-Myc degradation via proteasome-ubiquitin pathway; mitochondrial apoptosis (loss of MMP, ROS generation)	Inhibited colony formation dose-dependently	Induced G2/M cell cycle arrest; increased apoptosis (Annexin V+, cleaved caspase-9, cleaved PARP upregulated)
Badr et al. [27]	Glioblastoma	U87, Gli36, U251, primary GBM	DR5 (DEATH RECEPTORS) upregulation, TNF pathway genes, caspase-independent necrosis-like death, autophagy, mitochondrial membrane potential loss, ATP depletion	Significantly decreased (70–80% decrease in viability with lanatoside C + TRAIL)	Caspase-independent death with lanatoside C alone; caspase activation only when combined with TRAIL; DR5-dependent apoptosis pathway activated with combo
Zhang. et al. [28]	Cholangiocarcinoma	HuCCT-1, TFK-1, HIBEpiC	STAT3 signaling pathway, ROS (oxidative stress), mitochondrial membrane potential (MMP), apoptosis-related proteins (Bcl-2, Bax, caspase-9, caspase-3)	Inhibited (via CCK-8, wound healing, transwell, and live cell imaging assays)	Induced apoptosis and S/G2 phase cell cycle arrest
Kang et al. [29]	Colorectal cancer	HCT116, HT-29	MAPK pathway (Erk1/2, JNK1/2), autophagy (LC3, p62), mitochondrial dysfunction, DNA damage repair (53BP1, RNF168)	Reduced colony formation; replication-dependent death	G2/M cell cycle arrest; no significant apoptosis (no PARP-1 or caspase 3 cleavage)
Huang et al. [30]	Prostate cancer	PC-3, DU145, LNCAP, WPMY-1, HPRF	TNF, IL-17, P53, HIF-1 signaling	Inhibited in a dose-dependent manner; strong anti-proliferative effect observed in colony formation assays	Induced apoptosis; cell cycle arrest at G2/M (PC-3) and S + G2/M (DU145)
Duan et al. [31]	Cervical cancer	HeLa, BEAS-2B	JAK2/STAT6/SOCS2 signaling pathway; also affects apoptosis-related proteins (cleaved caspase-3, caspase-9, PARP), p21, cyclin B1	Colony formation suppressed. Inhibited in a dose-dependent manner	Induced apoptosis; cell cycle arrest at S and G2/M phases (PI staining, flow cytometry)
Xia et al., [32]	Lung cancer (NSCLC)	A549	Ferroptosis (GPX4, SLC7A11 downregulation)	Inhibited (reduced clone formation, EDU staining, Ki67 staining in vivo)	Apoptosis not implicated (Z-VAD-FMK had no effect); cell death via ferroptosis (mitochondrial membrane potential loss, SLC7A11/GPX4 suppression)
Johannson et al., [33]	Multiple cancers	CI-H69, CCRF-CEM, H69AR, CEM-VM1, ACHN, 8226-S, 8226-LR5, 8226-Dox40, U-937-GTB, GTB-Vcr	linked to Na⁺/K⁺-ATPase inhibition	Decrease observed (dose-dependent cytotoxicity reported)	Not specified for Lanatoside C
Nagao et al., [34]	Uterine leiomyosarcoma (ULMS)	SKN, SK-UT-1 (ULMS cell lines), HFF2T	EIF2 signaling, Autophagy, Sirtuin signaling pathway (notably downregulation of UCP2), NRF2-mediated oxidative stress response	Significantly inhibited in SKN and SK-UT-1 cells	Induced apoptosis
Rasheduzzaman et al., [35]	Liver Cancer/Hepatocellular carcinoma	Huh-7, HepG2	ROS, p38MAPK, TRAIL sensitization, Caspase-8 and − 3 activation, Mitochondrial apoptosis (Bcl-2 downregulation, Bax translocation, cytochrome c release), Autophagy via AMPK phosphorylation and LC3-II accumulation, DR5 (death receptor) activation under autophagy inhibition	Decreased (observed via MTT and crystal violet assays). TRAIL resistance reversed by LanC	Induced apoptosis
Reddy et al., [36]	Breast, Lung, Liver cancers	MCF-7 (breast), A549 (lung), HepG2 (liver); normal: L132, WRL68, PBMC	MAPK, Wnt/β-catenin, JAK/STAT, PI3K/AKT/mTOR, cell cycle (G2/M arrest), apoptosis (decrease BCL-2/increase BAX)	Strong inhibition of colony formation; dose-dependent cytotoxicity	Induced apoptosis; G2/M phase cell cycle arrest
Tesselar et al., [37]	Anaplastic thyroid cancer	8505 C, Cal-62	Autophagy activation, intracellular Ca²⁺ signaling, early response genes (ATF3, cFOS), reactivation of iodide metabolism (hNIS expression), potential indirect modulation of TSHR pathway	Inhibited completely in surviving cells after treatment; proliferation arrest observed	CDKN1A/p21 upregulation suggests cell cycle arrest; induced apoptosis
Xu et al., [38]	Multiple Myeloma (MM)	MM1.S, LP1, RPMI-8226, KMS11, U266, HEK293T	Otub1/c-Maf axis (LanC increases K48-linked polyubiquitination of c-Maf, disrupting Otub1–c-Maf interaction)	Decreased colony formation in primary MM cells (especially those with high c-Maf expression)	Induced apoptosis
Chao et al., [39]	Hepatocellular carcinoma (HCC)	Hep3B, HA22T	PKCδ (activation, translocation, cleavage), Mitochondrial (BID, Bcl-2, Mcl-1, AIF), Caspases (2, 3, 6, 7, 8, 9, PARP), Akt/mTOR, ERK1/2.	Inhibited in a concentration-dependent manner	Induced apoptosis; cell cycle arrest indicated by sub-G1 accumulation
Crommentuijn et al., [40]	Glioblastoma (GBM)	U87, GBM8	TRAIL-mediated apoptosis pathway	Decreased—U87 and GBM8 cells showed reduced proliferation when treated with lanatoside C and sTRAIL	Induced apoptosis; specific cell cycle effects not detailed
Vinod et al., [41]	Gastric cancer and breast cancer	NCI-N87 (HER2+), MDA-MB231 (HER2−)	Not specified in this excerpt; suggested mechanism through enhanced sensitivity to radioimmunotherapy	Strong decrease in cell proliferation for both cell lines	NA

NA Not applicable

Fig. 2.

Common pathways for anticancer effect of Lan C

Liver cancer

Five studies assessed the effect of Lanatoside C on hepatocellular carcinoma (HCC) using Huh7, HepG2, FOCUS, Mahlavu (a poorly differentiated HCC cell line), Hep3B, HA22T cell lines, with HepG2 (used in 4 of the 5 studies) being the most commonly employed model [25, 26, 35, 36, 39]. These studies demonstrated that Lanatoside C suppressed the growth of hepatocellular carcinoma cells in a concentration-dependent manner. This cardiac glycoside also interfered with critical phases of the cell cycle, effectively blocking cell division and triggering apoptosis to eliminate the cancer cells. Durmaz et al., Reddy et al., Chao et al., and Rasheduzzaman identified the PI3K/AKT/mTOR and MAPK/ERK pathways as central to the anticancer effects of Lanatoside C in HCC [25, 35, 36, 39]. Additionally, Hu et al. and Reddy et al. reported that Wnt/β-catenin signaling via GSK-3β activation also played a significant role [26, 36]. Other identified anticancer mechanisms of lanatoside C included enhanced apoptosis, characterized by increased levels of cleaved caspase 8 and caspase-9, upregulation of cleaved PARP, increased BAX expression and decreased BCL-2 expression. All studies also reported G2/M phase cell cycle arrest. Summaries of these pathways are presented in Table 1 and Fig. 2. 

Lung cancer

Two studies investigated the effects of Lanatoside C on lung cancer, both utilizing the A549 cell line [32, 36]. The findings revealed that Lanatoside C inhibited lung cancer cell proliferation in a dose-dependent manner. This cardiac glycoside also disrupted key stages of the cell cycle, halting cell division and initiating apoptosis to eliminate cancer cells. Xia et al. identified ferroptosis as the primary mechanism of Lanatoside C’s anticancer activity, marked by downregulation of Glutathione Peroxidase 4 (GPX4) and SLC7A11 (Fig. 3). This led to reduced antioxidant protection, accumulation of toxic lipid peroxides, leading to loss of antioxidant protection, and iron-dependent cell death in lung cancer cells [32]. In contrast, Reddy et al. reported that Lanatoside C targeted multiple pathways, including the MAPK, Wnt/β-catenin, JAK/STAT, PI3K/AKT/mTOR signaling pathways. They also observed G2/M phase cell cycle arrest and activation of apoptotic pathways mediated by decreased BCL-2 and increased BAX expression [36]. Summaries of these findings are presented in Table 1 and Fig. 2.

Fig. 3.

Cell death mechanisms of Lan C

Glioblastoma

Only two studies assessed the effect of Lanatoside C on glioblastoma, using cell lines such as U87, Gli36, U251, GBM8 [27, 40]. Both studies demonstrated that Lanatoside C suppressed glioblastoma cell growth in a concentration-dependent manner. By interfering with essential phases of the cell cycle, this cardiac glycoside effectively blocked cell division and triggered apoptosis, leading to cancer cell destruction. Badr et al. reported that upregulation of DR5 (death receptors), activation of TNF pathway genes, caspase-independent necrosis-like cell death, autophagy, loss of mitochondrial membrane potential and ATP depletion were central to the anticancer effects of Lanatoside C [27]. They also noted the already established ability of cardiac glycosides including Lan C to cross the blood brain barrier, which is important to intracranial malignancies. Similarly, Crommentuijn et al., identified the TRAIL (TNF-Related Apoptosis-Inducing Ligand)-mediated apoptosis pathway as a key mechanism underlying Lanatoside C’s anticancer activity [40]. Summaries of these findings are presented in Table 1.

Gastrointestinal cancers

A total of four studies examined the effects of Lanatoside C on several gastrointestinal cancers including gastric, colorectal, and pancreatic cancers [24, 26, 29, 41]. The cell lines used included PANC-1 and CFPAC-1 for pancreatic cancer, MKN-45, NCI-N87 and SGC-7901 for gastric cancer, HCT116 and HT-29 for colorectal cancer. All four studies demonstrated that Lanatoside C reduced cancer cell growth in a dose-dependent manner. It also disrupted vital checkpoints in the cell cycle, halting proliferation and triggering programmed cell death, ultimately leading to cancer cell elimination. In pancreatic cancer, Ha et al. reported that Lanatoside C exerted its anticancer effects via the ER stress/GRP78 pathway, where its downregulation promoted apoptosis in cancer cells [24]. Additionally, Kang et al. identified involvement of the MAPK pathway (Erk1/2, JNK1/2), autophagy (LC3, p62), mitochondrial dysfunction, and DNA damage repair mechanisms (53BP1, RNF168) as contributors to Lanatoside C’s anticancer activity in colorectal cancer cells—although limited cytotoxicity was observed [29]. Furthermore, Hu et al., highlighted the Wnt/β-catenin signaling pathway via GSK-3β activation as the primary mechanism underlying Lanatoside C’s anticancer action in gastric cancer cells [26]. Kang et al. and Hu et al. also reported that Lanatoside C induced G2/M phase cell cycle arrest.

Urogenital cancer

Three studies examined the effects of Lanatoside C on various urogenital cancers, including cervical cancer, prostate cancer, and uterine leiomyosarcoma (ULMS) [30, 31, 34]. The cell lines used included PC-3, DU145, LNCAP, WPMY-1 for prostate cancer; HeLa for cervical cancer; and SKN, SK-UT-1 for ULMS. These studies revealed that Lanatoside C reduced the proliferation of urogenital cancer cells in a dose-dependent manner. It also interfered with key checkpoints in the cell cycle, effectively halting cell division and, in some cases, triggering programmed cell death via apoptosis. Huang et al. in their study on prostate cancer cell lines found that upregulation of TNF, IL-17 and P53, along with downregulation of HIF-1 signaling, played important roles in Lanatoside C’s anticancer activity [30]. They emphasized that Lanatoside C’s dual targeting of inflammation and apoptosis offers a novel therapeutic approach for prostate cancer, particularly in metastatic cases. Targeting the TNF/TNFR2 signaling pathway could also enhance cancer immunotherapy. Furthermore, Duan et al. found that Lanatoside C targets the JAK2/STAT6/SOCS2 signaling pathway and modulates apoptosis-related proteins (cleaved caspase-3, caspase-9, PARP) in cervical cancer cells. They also observed cell cycle arrest at the S and G2/M phases and highlighted its potential role not only in inhibiting primary tumor growth but also in suppressing metastatic progression [31]. In their evaluation of Lanatoside C’s effects on ULMS, Nagao et al. found that the drug targets the EIF2 signaling, autophagy, sirtuin signaling pathways—most notably through downregulation of UCP2—and induces an NRF2-mediated oxidative stress response. The authors concluded that Lanatoside C holds promise for repurposing as an anti-cancer agent, particularly for ULMS, where therapeutic options remain limited [34].

Other cancers

Researchers have also evaluated the effects of Lanatoside C on other cancer types, including cholangiocarcinoma (HuCCT-1, TFK-1), anaplastic thyroid cancer (8505 C, Cal-62) and Multiple Myeloma (MM1.S, LP1, RPMI-8226, KMS11, U266, HEK293T) [28, 37, 38]. These studies showed that Lanatoside C can suppress the growth of cholangiocarcinoma, anaplastic thyroid cancer, and multiple myeloma cells in a concentration-dependent manner. Lanatoside C also disrupted key cell cycle checkpoints, thereby halting cellular division and, in some cases, initiating apoptosis to induce cancer cell death. Zhang et al., in their study on cholangiocarcinoma cell lines, reported that Lanatoside C targets the STAT3 signaling pathway and promotes apoptosis—evidenced by decreased Bcl-2, increased Bax, and activation of cleaved caspase-9 and caspase-3 [28] (Fig. 3). In contrast, Tesselar et al. reported upregulation of CDKN1A/p21, suggesting cell cycle arrest, along with autophagy activation in anaplastic thyroid cancer cells [37]. In a study by Xu et al. on multiple myeloma cell lines, Lanatoside C was found to increase K48-linked polyubiquitination of c-Maf, thereby disrupting the Otub1–c-Maf interaction. Based on these findings, the authors proposed that targeting the Otub1/c-Maf axis with compounds such as Lanatoside C could represent a novel and promising therapeutic strategy for multiple myeloma [38].

In-vivo anticancer effects of lanatoside C across cancer models

Ten in vivo studies investigated the anticancer efficacy of Lanatoside C (Lan C) across various cancer types, using different administration routes and tumor models. These studies consistently demonstrated the anticancer effects of Lan C, both as a monotherapy and in combination with other therapeutic agents. The findings are summarized in Table 2.

Table 2.

In vivo anticancer effect of Lan C

Author	Study Design	Dose given	Cancer Type	Route of Administration	In vivo antitumor activity
Durmaz et al.	In vivo	6 mg/kg/day	Liver cancer	Intraperitoneal injection (mice)	In nude mice xenografts, Lanatoside C significantly reduced tumor volume over 21 days compared to controls, indicating potential as a chemotherapeutic agent
Badr et al.	In vivo	10 mg/kg/day (Max tolerance dose)	Glioblastoma	In vivo	lanatoside C alone slowed tumor growth, and in combination with TRAIL led to > 85% tumor regression over 40 days
Zhang. et al.	In vivo	40 mg/kg/day	Cholangiocarcinoma	Gavage (oral administration in mice)	significant tumor reduction in HuCCT-1 xenografts in mice; reduced tumor volume and weight; increased caspase-9 and caspase-3, decreased STAT3
Kang et al.	In vivo	6 mg/kg/day	Colorectal cancer	subcutaneous (model; mouse xenograft models)	Alone: minimal effect; With radiation: significant tumor growth delay, especially in HCT116 xenograft (77.76% inhibition vs. 44.92% with radiation alone)
Xia et al.	In vivo	4-8 mg/kg/day	Lung cancer (NSCLC)	subcutaneous (xenograft model in nude mice)	suppressed tumor growth, reduced tumor size and weight, increased cell death (xenograft model)
Nagao et al.	In vivo	2.5 mg/kg/day	Uterine leiomyosarcoma (ULMS)	Intraperitoneal. xenograft (subcutaneous and orthotopic transplantation) mouse models	Significantly inhibited tumor growth in both subcutaneous and orthotopic ULMS xenograft mouse models
Xu et al.	In vivo	3 or 6 mg/kg/day	Multiple Myeloma (MM)	intraperitoneal (for mouse models)	Potent antimyeloma activity observed in two mouse models (subcutaneous and orthotopic)
Chao et al.	In vivo	2.5 mg/kg/day	Hepatocellular carcinoma (HCC)	subcutaneously(mouse xenograft models)	decreased tumor volume and delayed tumor growth without significant body weight loss
Crommentuijn et al.	In vivo	7.5 mg/kg/day	Glioblastoma (GBM)	intracranial injection (stereotactic), intraperitoneal (i.p.)	tumor size reduction, decreased Fluc signal, delayed progression, prolonged survival in combination treatment
Vinod et al.	In vivo	6 mg/kg/day	Gastric cancer and breast cancer	Intravenous (xenograft)	Significant tumor growth inhibition in NCI-N87 xenografts when treated with 131I-trastuzumab + lanatoside C compared to controls (p < 0.01)

Durmaz et al. found that intraperitoneal administration of Lan C significantly reduced tumor volume in nude mouse xenograft models of hepatocellular carcinoma (HCC) over a 21-day period, highlighting its potential as a chemotherapeutic agent. The study emphasized Lan C’s effectiveness regardless of PTEN status, which is clinically significant given that PTEN loss is associated with more aggressive, treatment-resistant liver cancers [25]. Additionally, Chao et al. demonstrated that Lan C suppressed HCC growth in subcutaneous xenografts without causing significant body weight loss [39].

Badr et al. found that Lan C intraperitoneal administration slowed the growth of glioblastoma (GBM) tumor cells as a monotherapy and achieved over 85% tumor regression when combined with TRAIL in nude mice over a 40-day period [27]. Furthermore, Crommentuijn et al. confirmed that Lan C enhanced the efficacy of AAV-mediated soluble TRAIL therapy in intracranial GBM models, resulting in delayed tumor progression and prolonged survival. While Lan C alone did not improve survival, the combination therapy yielded notable therapeutic benefits [40].

Zhang et al. demonstrated that oral administration of Lan C (via gavage) significantly reduced cholangiocarcinoma (CCA) tumor volume and weight in HuCCT-1 xenografts. The antitumor effect was associated with increased caspase-9 and caspase-3 activity, along with downregulation of STAT3 [28]. For colorectal cancer, Kang et al. observed minimal antitumor activity with Lan C alone; however, when combined with radiation, there was a marked delay in tumor growth in HCT116 xenografts (77.76% inhibition vs. 44.92% with radiation alone), implicating Lan C as a potential radiosensitizer [29]. Additionally, Xia et al. reported that subcutaneous lung cancer xenografts treated with Lan C exhibited significantly reduced tumor size and increased cancer cell death.

Nagao et al. demonstrated that intraperitoneal administration of Lan C suppressed tumor growth in both subcutaneous and orthotopic uterine leiomyosarcoma (ULMS) xenograft models, identifying UCP2 as a novel therapeutic target [34]. Vinod et al. evaluated Lan C in HER2-positive NCI-N87 gastric cancer xenografts and found that combining Lan C with 131I-trastuzumab (a radiolabeled antibody targeting HER2) resulted in significantly greater tumor growth inhibition compared to controls [41]. Majority of the included studies did not report the safety profile of Lan C. This is likely due to the fact that the drug is already FDA approved, with known pharmacokinetics, pharmacodynamics and safety profile.

The anticancer efficacy of Lan C demonstrated in vitro across a wide range of cancer types has been substantiated by in vivo studies. In vitro experiments consistently showed that Lan C exerts dose-dependent antiproliferative effects, induces cell cycle arrest (commonly at the G2/M phase), and triggers apoptosis through modulation of key oncogenic pathways including Wnt/β-catenin, PI3K/AKT/mTOR, MAPK, JAK/STAT, and ferroptosis-related mechanisms. These molecular alterations were further reflected in animal models, where Lan C significantly suppressed tumor growth across various xenograft models without evident toxicity. The in vivo outcomes not only validated the mechanistic insights obtained from cell-based assays but also highlighted Lanatoside C’s potential as a repurposed anticancer agent, given its established safety profile and multi-targeted action across diverse malignancies.

Quality assessment for included studies

Among 18 In vitro studies, the majority (> 95%) of the studies were regarded as reliable without restriction (supplementary material 3, Table S1) indicating high methodological quality. 70% of the in vivo studies were considered reliable without restriction, while the others were graded as not reliable (supplementary material 3, Table S2).

Discussion

Cancer encompasses multiple diseases affecting various organs, either primarily or secondarily, characterized by the gradual and uncontrolled growth of cells [42, 43]. While each cancer type has its distinct features, they all share common underlying molecular mechanisms that drive disease progression [44, 45]. Rising healthcare costs—particularly in drug development—pose challenges to system sustainability, prompting the need to repurpose and enhance existing drug therapies used for other diseases [46]. Lanatoside C, an FDA-approved cardiac glycoside, is one of the drugs under consideration for repurposing. This systematic review examined 18 preclinical studies investigating the anticancer properties of Lan C, across different cancer types. The findings provide substantial evidence for the broad-spectrum anticancer activity of Lan C through multiple molecular mechanisms, while also highlighting important considerations for its potential clinical translation.

Our study demonstrates that Lanatoside C exhibits anticancer effects across multiple cancer types, including breast, liver, lung, glioblastoma, gastrointestinal, and urogenital cancers. The consistency of these effects across various cancer models suggests that Lanatoside C has the potential for broad therapeutic application. In vitro studies revealed dose-dependent inhibition of cancer cell proliferation, with Lan C suppressing growth in most tested cancer cell lines. This broad-spectrum activity is noteworthy, as it indicates that Lan C may target fundamental vulnerabilities of cancer cells at the molecular level rather than cancer-specific alterations. The inhibitory effects on cell proliferation were associated with disruption of cell cycle progression, predominantly manifesting as G2/M phase arrest—a mechanism central to the anticancer properties of several established chemotherapeutic drugs [47]. This cell cycle arrest appears to be a common mechanism across the majority of cancer types, including breast cancer [36], hepatocellular carcinoma [24], and cervical cancer [31]. Furthermore, the anticancer effects extend beyond cytostatic activity to include cytotoxicity through various cell death mechanisms, most notably apoptosis and ferroptosis [32]. Such cytotoxic activity is characteristic of many established chemotherapeutic agents [48, 49].

A salient finding of this study is the convergence of Lan C’s effects on several key cancer-associated signaling pathways across different cancer types. The PI3K/AKT/mTOR pathway, which plays a crucial role in cancer cell survival and proliferation [50], was found to be downregulated by Lan C treatment in hepatocellular carcinoma [25, 39], lung cancer and breast cancer [36]. Similarly, MAPK pathway, a key signaling pathway for tumorigenesis [51], was observed to be inhibited by Lan C in multiple cancer models [25, 29, 35, 36], suggesting that MAPK pathway inhibition is a core mechanism of Lan C’s anticancer activity. The Wnt/β-catenin signaling pathway is also another common target, with studies in breast cancer and hepatocellular carcinoma [36], demonstrating Lan C-mediated disruption of this pathway through GSK-3β activation. This is particularly significant given the established role of Wnt signaling in cancer stemness, metastasis, and therapy resistance [52]. Also, the JAK/STAT pathway was identified as the target for Lan C in cervical cancer [31], cholangiocarcinoma [28], and breast cancer [36]. These findings not only point to Lanatoside C’s multifaceted anticancer mechanisms but also highlight its potential as a repurposed therapeutic agent that targets core oncogenic pathways shared across tumor types. Its ability to disrupt critical signaling networks and induce both cytostatic and cytotoxic effects makes Lanatoside C a promising candidate in the ongoing search for cost-effective, readily deployable cancer therapies through drug repurposing.

While several mechanisms appear to be shared across different studies and cancer types, our review also identified some cancer-specific pathways targeted by Lan C. In a study on lung cancer, Lan C induced ferroptosis through the downregulation of glutathione peroxidase 4 (GPX4) and SLC7A11, resulting in the loss of antioxidant protection and iron-dependent cell death. This represents an important alternative cell death pathway that may overcome apoptosis resistance in lung cancer. In prostate cancer cells, Lan C modulated the TNF/IL-17 signaling pathways, accompanied by p53 upregulation and HIF-1 downregulation [30]. The dual targeting of inflammatory and apoptotic pathways is particularly relevant for prostate cancer, especially in metastatic cases where immunomodulatory strategies may be beneficial. For pancreatic cancer, Lan C’s anticancer effects were mediated through the endoplasmic reticulum (ER) stress pathway via GRP78 downregulation [24]. These findings suggest that Lan C’s molecular mechanisms may be shaped by the specific oncogenic drivers and vulnerabilities of different cancer types.

The in vivo studies included in our review (10 out of 18 studies) provide supportive evidence for Lan C’s translational potential for possible clinical trial on human patients. These studies demonstrated significant tumor growth inhibition across multiple xenograft models, including hepatocellular carcinoma, glioblastoma, cholangiocarcinoma, colorectal cancer, lung cancer, and uterine leiomyosarcoma. Importantly, in some studies, these anticancer effects were achieved without significant body weight loss, suggesting a potentially manageable toxicity profile. The efficacy of Lan C as a monotherapy varied across studies and cancer types, with notable results observed in hepatocellular carcinoma [25] and cholangiocarcinoma [28]. Additionally, a number of studies reported enhanced efficacy when Lan C was combined with other therapeutic modalities. For instance, Lan C improved the efficacy of radiation therapy in colorectal cancer xenografts (77.76% inhibition versus 44.92% with radiation alone) [29], highlighting its potential as a radiosensitizer. Similarly, in glioblastoma models, combining Lan C with TRAIL therapy resulted in over 85% tumor regression, delayed tumor progression, and prolonged survival [27, 40]. These findings suggest that while Lan C may have therapeutic value as a monotherapy, its greatest potential could lie in combinatorial strategies, owing to its ability to simultaneously target multiple signaling pathways. This multifaceted action may help overcome treatment resistance—a common challenge when single pathways are selectively targeted. A key challenge to the translational potential of Lan C lies in the wide variation in dosages reported across the included studies, ranging from 2.5 mg/kg to 40 mg/kg. This variability limits the ability to draw reliable inferences regarding its safety thresholds as an anticancer agent.

This study has certain limitations. Despite the favorable findings, several methodological variations were observed across the included studies, which should be taken into account when interpreting the results. First, there was considerable heterogeneity in experimental design, including differences in Lan C dosage, treatment duration, and the specific cancer cell lines used. Also, in critically appraising the included studies, we found that while most were methodologically robust, the studies by Johannson et al. and Tesselar et al. demonstrated weaknesses, as they did not clearly report all methodological and experimental procedures [33, 37]. These variabilities complicate direct cross-study comparisons and underscore the need for greater standardization in preclinical cancer drug repurposing research. Additionally, most studies employed traditional 2D cell culture systems, which do not adequately replicate the complex tumor microenvironment [53]. This is a significant limitation, as new evidence has shown that the tumor microenvironment can influence drug response and contribute to therapy resistance [54]. Future studies would benefit from adopting more physiologically relevant models, such as 3D organoids or patient-derived xenografts [55]. Also, the in vitro studies on lung cancer were limited to a single cell line, which weakens the strength of the conclusions. Future studies should incorporate multiple cell lines to more accurately evaluate the anticancer effects of Lan C in lung cancer. Another important limitation relates to the potential toxicity of cardiac glycosides generally, including Lan C [56]. Although several studies in this review reported no apparent signs of toxicity in animal models, comprehensive toxicological evaluations were not performed, which will be key to translational research on human subjects. Given that Lan C is already FDA-approved for cardiac conditions, determining its therapeutic window in the context of cancer treatment will be crucial for its potential clinical translation. Another limitation of preclinical studies is the underreporting of negative findings, which can contribute to publication bias and may distort the overall understanding of a compound’s therapeutic potential. Our review was therefore limited to the available published data.

Future studies should focus on establishing optimal dosing regimens that maximize the anticancer efficacy of Lan C while minimizing toxicity and other adverse effects. This may involve exploring different administration routes, formulation strategies to enhance tumor targeting and reduce systemic exposure. Given the encouraging results seen with combination therapies, further research should investigate Lan C in conjunction with other therapeutic approaches, particularly in a setting where resistance to standard treatment has been established. Especially promising are combinations with radiotherapy, targeted therapies that act on complementary pathways, or immunotherapeutic strategies that could benefit from Lan C’s modulation of inflammatory signaling. Ultimately, well-designed clinical trials will be essential to validate these preclinical findings. Initial trials could prioritize cancer types that showed the most robust preclinical responses—such as hepatocellular carcinoma, cholangiocarcinoma, or colorectal cancer in combination with radiotherapy.

Conclusion

This systematic review provides substantial evidence for the broad-spectrum anticancer activity of Lanatoside C across various cancer types, mediated through both common and cancer-specific molecular mechanisms. The existing FDA approval of Lan C for cardiac indications presents an opportunity to facilitate its repurposing for oncology, offering a novel therapeutic approach, both as a monotherapy and in combination with other treatments. Although some methodological limitations and knowledge gaps remain, the consistency of findings across different experimental models—both In vitro and in vivo—supports further investigation of Lanatoside C as a potential anticancer agent. As research continues to elucidate the complex mechanisms underlying its anticancer effects, this cardiac glycoside may ultimately find a place in the expanding arsenal of repurposed drugs for cancer therapy, particularly for patients with limited treatment options or with therapy-resistant disease.

Abbreviations

Lan C

Lanatoside C

FDA

Food and Drug Administration

ToxRTool

Toxicological Data Reliability Assessment Tool

ULMS

uterine leiomyosarcoma

Authors’ contributions

O.O.O conceptualized the study idea and design. Systematic literature search and review, data extraction was carried out by O.O.O, B.T.O, E.O.O, and T.J.O. Qualitative synthesis of findings was carried out by O.O.O, and T.J.O. The first draft of the manuscript was written by O.O.O and B.T.O. A.I.A and T.J.O commented on previous versions of the manuscript. All authors read and approved the final manuscript. The entirety of the project was supervised by A.I.A.

Funding

No funding was received for conducting this study.

Data availability

All data generated during this study are included in this published article [and its supplementary material files].

Declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.