Abstract
Background/Aim: Inflammatory bowel disease (IBD) is characterized by dysregulated immune responses and a multifactorial etiology. While imatinib has demonstrated efficacy in the treatment of immune-related diseases, its potential effects in IBD treatment remain underexplored. Materials and Methods: This study aimed to investigate the therapeutic effects of imatinib in colitis treatment. A dextran sulfate sodium (DSS)-induced colitis model was used to mimic IBD in mice. Imatinib was administered orally to mice simultaneously with DSS treatment. The effects of imatinib on DSS-induced colitis were evaluated by analyzing colitis-related pathology, including the disease activity index (DAI), histological lesions, inflammatory markers, and tight junction integrity. Additionally, western blot analysis and quantitative real-time polymerase chain reaction were used to assess inflammatory markers, tight-junction proteins, and cell death. Results: In the DSS-induced colitis model, imatinib treatment exerted protective effects by attenuating weight loss, restoring colon length, reducing spleen weight, and improving the DAI score and histological lesions. Additionally, imatinib reduced the level of proinflammatory cytokines, including TNF-α, IL-6, and IL-1β. Furthermore, imatinib treatment restored tight-junction integrity and decreased the expression of apoptosis marker proteins. Conclusion: Overall, imatinib treatment significantly alleviated the symptoms of DSS-induced colitis by influencing the expression of proinflammatory cytokines, tight junction proteins, and apoptotic markers in mice. These findings highlight imatinib as a potential therapeutic candidate for IBD.
Inflammatory bowel disease (IBD), which includes ulcerative colitis (UC) and Crohn’s disease, is characterized by dysregulated immune responses and a multifactorial etiology (1, 2). In recent years, there has been a consistent rise in the worldwide incidence of IBD (3). The development of this disease is influenced by a complex interplay of genetic and environmental factors, leading to abnormal immune responses (4). UC, one type of IBD, is characterized by chronic inflammation primarily affecting the colon and rectum, particularly within the mucosal and submucosal layers (5, 6).
Proinflammatory cytokines such as TNF-α and IL-6 play pivotal role in IBD progression (7, 8). In addition, inflammasome signaling has emerged as another crucial component of the innate immune response and plays a central role in regulating GI health and disease. Inflammasome activation not only leads to the release of pro-inflammatory cytokines, including IL-1β and IL-18, but also triggers a form of inflammatory cell death (8).
Despite the increasing prevalence of IBD, available therapies are primarily focused on symptom management rather than providing a cure. Non-steroidal anti-inflammatory drugs (NSAIDs) are commonly employed in treating patients with IBD (9). However, long-term use of NSAIDs is often associated with several adverse side effects, including fatigue, nausea, abdominal pain, and diarrhea (10). Despite the progress made in understanding IBD pathogenesis and the available therapeutic options, a significant gap persists in the quest for treatments that are effective with fewer adverse effects.
Imatinib is a tyrosine kinase inhibitor, which was initially developed for the treatment of gastrointestinal stromal tumors. It has shown anti-proliferative activity and immunomodulatory effects (11, 12). Additionally, imatinib modulates various immune cells, including lymphocytes, macrophages, and dendritic cells, by inhibiting inflammatory signal transduction pathways involved in the pathogenesis of autoimmune diseases.
Additionally, imatinib has been shown to suppress the secretion of pro-inflammatory cytokines, such as TNF-α, IL-1β, and matrix metalloproteinases (MMPs) (13). Consequently, it has shown efficacy in treating autoimmune conditions, such as rheumatoid arthritis, multiple sclerosis, and glomerulonephritis. However, the effectiveness of imatinib on colitis has not been adequately demonstrated in in vivo studies.
Our study aimed to investigate the potential therapeutic effect of imatinib in IBD using a dextran sulfate sodium (DSS)-induced colitis model. The efficacy of imatinib in ameliorating ulcerative colitis (UC) was evaluated through various parameters, including inflammatory markers, histological changes, cytokine levels, and cell death markers.
Materials and Methods
Animals. Male C57BL/6J mice (6 weeks old, 20-22 g) were obtained from Samtako (Gyeonggi-do, Republic of Korea) and housed in plastic cages under specific pathogen-free conditions (24±2°C, 12 h light/dark cycle). The mice were allowed to adapt to the experimental environment for one week. The animals had free access to sterile water and food throughout the entire experimental period. The animal experiments and protocols were reviewed and approved by the Institutional Animal Care and Use Committee at Kyungpook National University, following standard operating procedures (2022-0206).
Experimental design. To investigate the effects of imatinib on DSS-induced colitis (DSS; 160110; MP Biomedicals, Santa Ana, CA, USA), we randomly assigned 24 mice into four groups: the control group (Control), DSS-induced colitis group (DSS), DSS-induced colitis group treated with 10 mg/kg imatinib (Im10), and the DSS-induced colitis group treated with 20 mg/kg imatinib (Im20).
Imatinib (HY-15463, MedChemExpress, Monmouth Junction, NJ, USA) was dissolved in saline and administered orally for seven days, spanning from day 0 to the seventh day (Figure 1). Acute colitis was induced by providing the mice with 3% DSS (w/v) in their drinking water for five consecutive days, starting from day 0 and continuing until the fifth day. On the 10th day, the physical phenotypes of the colon tissues of mice were examined and the lengths of the large intestines were measured. Additionally, spleen weight was measured, and its percentage of the total body weight was calculated. Colon tissue samples were collected and stored at −150°C until further experimentation.
Overview of the study design. Mice were acclimatized for seven days before initiating group-specific treatment. Colitis was induced by administering 3% dextran sulfate sodium (DSS) in drinking water from day 0 to the fifth day. Concurrently, imatinib (at doses of 10 mg/kg or 20 mg/kg) was orally administered once daily from day 0 to the seventh day using oral gavage. All mice were euthanized on the 10th day.
Disease activity index. Disease Activity Index (DAI) scores were assessed daily during the 10-day period of DSS treatment. The following data were recorded: changes in body weight, stool consistency, and the presence of fecal occult blood. Body weight loss was categorized as follows: 0 (no loss), 1 (1-5% loss), 2 (5-10% loss), 3 (10-20% loss), or 4 (over 20% loss). Stool consistency was assessed as follows: 0 (normally formed), 2 (loose stools), and 4 (diarrhea). The presence of blood in the stool was scored as follows: 0 (negative), 2 (positive), and 4 (gross bleeding). The mean of all the scores obtained for each parameter was calculated to determine the DAI.
Hematoxylin and eosin (H&E) staining. Colon tissue samples were fixed in 4% formaldehyde in 1× phosphate-buffered saline, followed by embedding in paraffin. Afterward, the paraffin-embedded tissue was cut into 5-μm sections. Hematoxylin and eosin staining was conducted to visualize the morphological changes. The stained tissue sections were then examined using a MoticEasyScan One (Motic, Hong Kong, SAR) to check for cellular infiltration, mucosa length and epithelial damage caused by inflammatory cells. Scoring was conducted for all tissue sections.
Histological score. Colonic histological tissues were observed using a MoticEasyScan One (Motic) to assess histological damage. The severity of colonic inflammation, colonic epithelial cell infiltration, crypt destruction, and the extent of cell infiltration were calculated. Each subscore ranged from 0 to 4, representing the absence of changes to maximum tissue damage. Specifically, the scoring criteria were as follows: crypts: 0 (intact crypts), 1 (intact crypts), 2 (variable crypt diameter), 3 (atrophied crypts), and 4 (mucosa devoid of crypts); loss of surface epithelium: 0 (intact surface epithelium), 1 (sloughing off epithelial surface), 2 (patchy loss of surface epithelium), 3 (moderate loss of surface epithelium), and 4 (severe loss/erosion of surface epithelium); cell infiltration: 0 (clear mucosa/submucosa), 1 (mucosal/lamina propria infiltration), 2 (mucosal and submucosal infiltration), 3 (moderate cryptitis/infiltration into crypt epithelial cells), 4 (severe cryptitis). These scores were used to quantify the degree of histological damage.
Quantitative real-time polymerase chain reaction (qRT-PCR) assay. Total RNA was isolated from mouse colon tissues using TRI Solution (TS200-001; Bio Science Technology, Daejeon, Republic of Korea). Next, total RNA was synthesized to cDNA using a PrimeScript™ 1st Strand cDNA Synthesis Kit (6110; Takara Biotechnology Co., Ltd., Shiga, Japan) with oligo-dT primers. The qRT-PCR mixture contained 8 μl of cDNA, 10 μl of Power SYBR® Green PCR Master Mix (4367659; Thermo Fisher Scientific, Waltham, MA, USA), 1 μl (0.2 pmol) of forward primer, and 1 μl (0.2 pmol) of reverse primer. The qPCR was run on the Applied Biosystems RT-PCR program (StepOnePlus™ Real-Time PCR System, Thermo Fisher Scientific) at 95°C for 10 min, followed by 40 cycles at 95°C for 15 s and 60°C for 1 min, and a melt curve stage at 95°C for 15 s. The primer sequences used were as follows: mTNF-α (F: 5′-ATGGGCTCCCTCTCATCAGT-3′, R: 5′-TGGTTTGCTACGACGTGGG-3′); mIL-1β (F: 5′-TGGTTTGCTACGACGTGGG-3′, R: 5′-CCCCAGGGCAT GTTAAGGA-3′); mIL-6 (F: 5′-TGACCCTGAGCGACCTG TCT-3′, R: 5′-RGTTGTGCAATGGCAATTCTGA-3′); and mβ-actin (F: 5′-GGCTCTTTTCCAGCCTTCCT-3′, R: 5′-GT CTTTACGGATGTCAACGTCACA-3′). All data were analyzed using the 2-ΔΔCq method and expressed as the fold change relative to the controls. The relative expression levels of TNF-α, IL-1β, and IL-6 were normalized to those of β-actin.
Western blot analysis. Colon samples were processed using the PRO-PREP for Cell/Tissue Protein Extraction Solution kit (17081; Intron Biotechnology, Kirkland, WA, USA) along with the Protease Inhibitor Cocktail (P3100-001; GenDEPOT, Katy, TX, USA) for lysis. The protein concentrations in the tissue lysates were determined using BCA Protein Assay Reagent A (WF322788; Thermo Fisher Scientific) and Reagent B (23224; PIERCE, Kyiv, Ukraine). Subsequently, 30 μg of protein from each sample was separated on a 10-12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gel and transferred onto polyvinylidene difluoride (PVDF) membranes. The membranes were then blocked with 5% skim milk in 1×Tris-buffered saline (TBS)-0.05% Tween-20 (TBST) at room temperature for 1 h. They were then incubated with primary antibodies against IL-1β (sc-52012; Santa Cruz Biotechnology, Dallas, TX, USA), TNF-α (3707; Cell Signaling Technology, Danvers, MA, USA), IL-6 (sc-57315; Santa Cruz Biotechnology), cleaved caspase-3 (9661; Cell Signaling Technology), p-p53 (377567; Santa Cruz Biotechnology), p53 (sc-126; Santa Cruz Biotechnology), cleaved caspase-8 (D35G2; Cell Signaling Technology), caspase-8 (D35G2; Cell Signaling Technology), ZO-1 (sc-33725; Santa Cruz Biotechnology) and claudin-2 (E1H90; Cell Signaling Technology). β-actin (sc-47778; Santa Cruz Biotechnology) was used as a loading control. After three washes with TBST, the membranes were incubated at room temperature for 1 h with the secondary antibodies normal rat IgG-HRP (sc-2750; Santa Cruz Biotechnology), goat anti-mouse IgG H&L horseradish peroxidase (HRP) (ab6789; Abcam, Cambridge, UK), and anti-goat IgG-HRP (sc-2354; Santa Cruz Biotechnology). Finally, protein bands were visualized using an Image Quant LAS 500 (Amersham™ ImageQuant™ 500; Cytiva, Uppsala, Sweden), with β-actin as an internal control. Data were quantified using ImageJ (United States National Institutes of Health, Bethesda, MD, USA).
Immunohistochemical analysis. The colon collected from each mouse was directly fixed in 4% buffered paraformaldehyde, dehydrated in 100% ethanol, and embedded in paraffin. Next, 5-μm sections were prepared from the embedded tissue. The colon sections were stained and ZO-1 and occludin were used as tight-junction markers. For ZO-1 and occludin staining, the sections were rehydrated and treated with 3% H2O2, soaked and boiled in citrate buffer for 15 min (three times each for 5 min), stored in a dark room with 10% normal serum for 1 h, and incubated with the primary antibody at 4°C overnight. The slides were then incubated overnight with ZO-1 (sc-33725; Santa Cruz Biotechnology) and occludin (E6B4R; Cell Signaling Technology), followed by the secondary antibody for 30 min. Finally, ZO-1– and occludin-positive cells were detected using the VECTASTAIN ABC streptavidin-HRP system (ZG0608; Vector Laboratories, Newark, CA, USA). The sections were then counterstained with hematoxylin before being dehydrated and mounted. All tissue sections were observed under a microscope (MoticEasyScan One; Motic).
Statistical analysis. Data were analyzed using SPSS Statistics for Windows, version 25.0 (IBM, Armonk, NY, USA). One-way analysis of variance (ANOVA) was used to compare the control and DSS groups, as well as the DSS group and each treatment group (n=6). All data are presented as mean±standard error of the mean (SEM). A p-value <0.05 was considered statistically significant.
Results
Effects of imatinib on symptoms in mice with DSS-induced colitis. To evaluate the therapeutic potential of imatinib in treating IBD, we employed a DSS-induced colitis mouse model that closely resembles human IBD in phenotypic features (Figure 1A). Mice with DSS-induced colitis exhibited a noticeable decline in body weight after seven days. Interestingly, treatment with imatinib at doses of 10 mg/kg and 20 mg/kg significantly attenuated the weight loss from day 6 (Figure 2A).
Imatinib alleviates disease severity in DSS-induced colitis. (A) Body weight changes following dextran sulfate sodium (DSS) administration in the presence or absence of imatinib treatment. (B) A representative photograph of colon in each group. (C) The colon lengths of each mouse group were measured. (D) Splenomegaly was accessed through the ratio between the spleen and body weight in each group. (E) The disease activity index (DAI) score includes scales for body weight, stool consistency, and fecal occult blood at D10. Data are presented as mean±standard error of the mean (SEM). p<0.05 indicates statistical significance (n=6). *p<0.05 represents control vs. DSS; #p<0.05 represents DSS vs. 10 mg/kg imatinib (Im10) and 20 mg/kg imatinib (Im20).
Furthermore, we evaluated the colon lengths of each experimental group. The DSS-induced mice displayed a significant reduction in colon length compared to that of the other groups. However, treatment with both 10 mg/kg and 20 mg/kg imatinib led to a significant restoration of colon length compared to DSS group (Figure 2B and C).
Splenomegaly is a common symptom that is observed in colitis models (14). Consistent with the prior results, the colitis mouse model exhibited a significant increase in spleen weight. Notably, treatment with imatinib at 10 mg/kg and 20 mg/kg resulted in significant reductions in spleen weight compared to DSS group (Figure 2D).
Moreover, DAI scores were measured to comprehensively assess disease severity. The group treated solely with DSS exhibited the highest DAI score, reflecting severe disease activity at D10. However, treatment with imatinib resulted in a significant dose-dependent reduction in the DAI score compared to the DSS group (Figure 2E). Taken together, imatinib treatment exerts protective effects against DSS-induced colitis by attenuating weight loss, restoring colon length, reducing spleen weight, and improving the DAI score.
Effects of imatinib on histological damage in colon tissues. Histological examination of the colon tissue was conducted through H&E staining. In the DSS group, there was a noticeable shortening of the mucosa, along with extensive damage to the goblet cells and colon tissue (Figure 3A). However, treatment with imatinib alleviated damage to colon tissue in a dose-dependent manner.
Protective effect of imatinib on colonic structure. (A) Hematoxylin & eosin (H&E) staining was performed to elucidate the efficacy of imatinib. (B) Mucosa length was measured in each group (magnification ×200 μm). (C) Histologic scores based on H&E staining. The data are presented as mean±standard error of the mean (SEM). p<0.05 indicates statistical significance (n=6). *p<0.05 represents control vs. dextran sulfate sodium (DSS); #p<0.05 represents DSS vs. 10 mg/kg imatinib (Im10) or 20 mg/kg imatinib (Im20).
Treatment with imatinib preserved mucosal length in a dose-dependent manner compared to the DSS group (Figure 3B). Additionally, the reduction in goblet cells caused by DSS treatment was relatively decreased by imatinib treatment (Figure 3A).
Histological scores accessing the severity of colonic inflammation, colonic epithelial cell infiltration, crypt destruction, and the extent of cell infiltration showed a significant decrease in the Im20 group compared to those of the DSS group (Figure 3C). Overall, imatinib treatment relieved the histological damage caused by DSS-induced colitis.
Anti-inflammatory effects of imatinib in colon tissues. The improvement in histological scores after imatinib treatment led us to investigate the inflammatory cytokines. mRNA expression levels of inflammatory cytokines, including TNF-α, IL-1β, and IL-6, were quantified using qRT-PCR (Figure 4A-C). In comparison to that of the control group, the DSS group exhibited a significant increase in TNF-α and IL-1β mRNA expression. However, the imatinib treatment group demonstrated a dose-dependent reduction in the mRNA expression of pro-inflammatory cytokines, such as TNF-α and IL-1β (Figure 4A and B).
The anti-inflammatory effect of imatinib in mice with dextran sulfate sodium (DSS)-induced colitis. (A) The mRNA levels of pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), (B) IL-1β, and (C) IL-6, in colon tissues were determined by quantitative real-time PCR. (D) The expression of pro-inflammatory cytokines, including TNF-α, IL-1β, and IL-6, was assessed using western blot analysis (n=6). The data are presented as mean±standard error of the mean (SEM). *p<0.05 represents control vs. DSS; #p<0.05 represents DSS vs. 10 mg/kg imatinib (Im10) and 20 mg/kg imatinib (Im20).
To further assess the impact of imatinib treatment, protein expression levels of TNF-α, IL-1β, and IL-6 were also evaluated (Figure 4D). DSS treatment increased the cytokine levels compared to those of the control groups. Notably, DSS-induced production of cytokines, including TNF-α, IL-1β, and IL-6, was dose-dependently reduced by imatinib treatment (Figure 4D).
Effect of imatinib on the epithelial barrier in mice with DSS-induced colitis. The protective effect of imatinib on intestinal barrier integrity was investigated by examining the expression of tight-junction (TJ) proteins, including ZO-1, occludin, and claudin-2.
Immunohistochemical (IHC) staining was performed to assess the distribution and intensity of ZO-1 and occludin. In the control group, ZO-1 and occludin were observed on both apical and basolateral surfaces of the crypts. In contrast, the DSS group exhibited a significantly weakened ZO-1 and occludin expression level (Figure 5A). However, protein levels of ZO-1 and occludin were increased in the imatinib-treated groups compared to those of the DSS group.
Protective effect of imatinib on tight junctions and potential anti-apoptotic ability in mice with dextran sulfate sodium (DSS)-induced colitis. (A) Immunohistochemical staining was conducted to assess the protein levels of ZO-1 and occludin. (B) The expression of ZO-1 and claudin-2 was measured using western blot analysis. (C) The expression of apoptotic markers, including cleaved-caspase-3, cleaved-caspase-8, caspase-8, P53, and p-P53, was assessed using western blot analysis (n=6).
To further validate the TJ protein expression, western blotting was conducted on ZO-1, and claudin-2. The expression of ZO-1 was found to be decreased in the DSS group and increased following imatinib treatment. Another TJ protein, claudin-2, was found to be elevated in the DSS group and decreased following imatinib treatment (Figure 5B).
These results highlight the restorative effect of imatinib on intestinal barrier integrity. Imatinib treatment successfully recovered the expression and localization of TJ proteins, such as ZO-1, occludin, and claudin-2. These results support the notion that imatinib exerts a beneficial effect on intestinal barrier function.
Anti-apoptotic effect of imatinib in mice with DSS-induced colitis. The potential anti-apoptotic ability of imatinib was investigated using western blot analysis. The expression of apoptosis markers was increased in the DSS group; however, in the imatinib treatment group, the expression levels of cleaved caspase-3, cleaved caspase-8, caspase-8, p53, and p-p53 were redueced (Figure 5C). These results suggest that imatinib has an anti-apoptotic effect on colonic epithelial cells in mice with DSS-induced colitis.
Discussion
Ulcerative colitis (UC) is a chronic inflammatory condition characterized by cycles of relapse and inflammatory remission, accompanied by increased morbidity and a heightened risk of malignant tumor development (15). With an annual incidence rate of 10 per 100,000 individuals, concerns about UC are also escalating annually.
The long-term effects of UC include irreversible damage to the intestinal tract, leading to toxic megacolon, intestinal perforation, hemorrhage, polyps, and an increased risk of colon cancer (16). Given the gravity and long-term consequences of UC, there is an increasing need for effective treatment options capable of providing symptom relief, reducing inflammation, and preventing disease progression.
Imatinib, a tyrosine kinase inhibitor, has demonstrated efficacy in treating cancer and immune-related diseases by inhibiting tyrosine kinase activity. Tyrosine kinases play pivotal roles in numerous biological processes, including growth, differentiation, metabolism, and apoptosis (17). These kinases operate as signaling molecules that mediate signal transduction from a variety of leukocyte antigen receptors, innate immune receptors, and cytokine receptors (18). By targeting these signaling pathways, imatinib may regulate immune responses and inflammatory processes associated with UC. However, this possibility has not been studied.
This study aimed to investigate the therapeutic potential of imatinib for the first time in UC by using a well-established DSS-induced colitis model to replicate UC in mice (19, 20). The symptoms of UC, including loose stools, diarrhea, and weight loss, appeared with DSS administration. However, the groups administered imatinib (Im10 and Im20) during DSS treatment showed significantly reduced disease severity compared to the DSS group (Figure 2).
The GI tract consists of a single layer of epithelial cells specialized in nutrient absorption and serves as a physical barrier against the external environment (21, 22). Intestinal epithelial cells (IECs), regulated by TJ proteins that prevent the passage of substances from the lumen, regulate the permeability of ions, nutrients, and water (23, 24). Thus, integrity of the TJ is important in colitis progression. In DSS-induced colitis, imatinib (Im10 and Im20) treatment prevented abnormal changes in TJ proteins, including a decrease in ZO-1 and occludin levels and an increase in claudin levels. This suggests that imatinib restores intestinal barrier integrity (Figure 5).
Another indicator of colitis severity is the presence of goblet cells, which secrete mucins to form a protective mucus layer, enhancing the barrier between luminal contents and the epithelial surface (25). Histological analysis using H&E staining revealed physical damage to the intestinal mucosa in the DSS group, including reduced mucosal length and disrupted lamina propria integrity. Conversely, imatinib treatment improved the condition of the intestinal mucosa and restored goblet cells, indicating that imatinib protects mucin production and maintains intestinal barrier function in colitis (Figure 3).
Aberrant innate immune activation accelerates intestinal pathology in IBD patients (26, 27). Dysregulation of pro-inflammatory cytokines, such as TNF-α, IL-1β, and IL-6 drives the inflammatory processes in IBD, highlighting them as potential therapeutic targets (28). Hence, we investigated cytokine level changes during DSS treatment with imatinib and found that imatinib reduces TNF-α, IL-1β, and IL-6 levels (Figure 4), potentially attenuating the inflammatory response and improving disease symptoms (29).
Additionally, an increase in apoptosis is observed in the pathogenesis of IBD (29). Apoptosis, the programmed cell death process, plays a crucial role in regulating the immune system and removing potentially harmful cells. Imatinib treatment reduced the expression levels of apoptosis markers, including p53, p-p53, cleaved-caspase-8, caspase-8, and cleaved-caspase-3, as identified by western blot analysis (Figure 5C).
Conclusion
To our knowledge, this is the first study to investigate the therapeutic effects of imatinib in colitis. Our results show that imatinib reduces pro-inflammatory cytokines, preserves colon structures, and protects colonic epithelial cells from apoptosis. These findings suggest imatinib as a promising candidate for treating IBD.
Footnotes
Authors’ Contributions
Conceptualization: Myoung Ok Kim. Methodology: Hyeonjin Kim, Chae Yeon Kim, Dongwook Kim, Zae-Young Ryoo, Jun Koo Yi, Soyoung Jang, Weihong Wen. Investigation: Hyeonjin Kim, Chae Yeon Kim, Dongwook Kim. Writing-Original Draft: Hyeonjin Kim, Chae Yeon Kim, Dongwook Kim, Soyoung Jang. Visualization: Ke Huang, Zhibin Liu, Jiwon Ko, Su-Geun Lim, Soyoung Jang. Validation: Eungyung Kim, Lei Ma, Soyoung Jang. Resources: Eungyung Kim, Lei Ma, Kanghyun Park, Jiwon Ko, Su-Geun Lim, Weihong Wen. Formal analysis: Eungyung Kim, Lei Ma, Soyoung Jang. Software: Kanghyun Park. Data Curation: Ke Huang, Zhibin Liu, Soyoung Jang. Writing-Review & Editing: Yonghun Sung, Soyoung Jang, Myoung Ok Kim. Funding acquisition: Zae-Young Ryoo, Jun Koo Yi, Myoung Ok Kim. Supervision: Myoung Ok Kim. Project administration: Myoung Ok Kim, Soyoung Jang.
Conflicts of Interest
The Authors declare no conflicts of interest in relation to this study.
- Received April 15, 2024.
- Revision received May 22, 2024.
- Accepted May 23, 2024.
- Copyright © 2024 The Author(s). Published by the International Institute of Anticancer Research.
This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY-NC-ND) 4.0 international license (https://creativecommons.org/licenses/by-nc-nd/4.0).
