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Han et al. Cancer Drug Resist 2024;7:16 https://dx.doi.org/10.20517/cdr.2024.01 Page 3 of 25

activity. Among its eight components of cucurbitacins with notable anticancer activity are cucurbitacin B,
[11]
[12]
D, E, I, C, II, A, L-glucoside, S and R . Its enantiomeric isomer, Cucurbitacin B , has long been known
[15]
for its role in lung cancer [13,14] , colorectal cancer , colon cancer , and glioma . Melon pedicle was first
[16]
[17]
documented in Shennong’s Herbal Classic, which describes its function in eliminating dampness and
inducing vomiting. Furthermore, many ancient texts, such as “Shengji General Record”, “Qianjin Yaofang”,
and “Ancient and Modern Medical Systems” also documented the use of melon pedicels for treating
toothache, malaria, and hemorrhoids. Modern research has revealed that melon stalks have antitumor,
hepatoprotective, and other beneficial effects. Clinical reports also indicate the utilization of melon stalks in
the treatment of acute jaundice, infectious hepatitis, chronic rhinitis, chronic hepatitis, primary liver cancer,
and other conditions. However, the anti-glioma effects of isocuB have not been proven.

While much is known about the molecular, structural, energetic, and chemical aspects of drug-target
interactions, challenges remain in the selection and definition of targets, hindering the advancement of
pharmacology and pharmacotherapy to an exact science . The development of network pharmacology and
[18]
bioinformatics will not only reduce the cost of drug development, but also shorten the time required for
drug development. Furthermore, molecular docking techniques enable the prediction of the binding
affinities and conformations of receptors and ligands [19,20] . This has been extremely helpful in our research. A
deeper comprehension of the role of microRNAs (miRNAs) in development and disease, particularly in
cancer, renders them attractive tools and targets for innovative therapeutic approaches. Functional studies
have indicated the causal role of miRNA dysregulation in many cancers. MiRNAs show promise in
preclinical development as tumor suppressors or oncogenes miRNA mimics, and molecules targeting
miRNAs .
[21]

The aim of our study was to predict the target genes and pathways of isocuB against glioma using network
pharmacology and data analysis. Afterwards, we validated these predictions through Counting Kit-8
(CCK-8), wound healing assay, transwell invasion, TdT-mediated dUTP-biotin nick end labeling (TUNEL)
staining, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), WB, and tumor growth
experiments in nude mice. Then, we further investigated and predicted the relationship between miRNA
and resistance to the drug TMZ. Our study demonstrated that isocuB exerts its inhibitory effects on
glioma by regulating specific pathways and gene expression, suggesting a novel and effective treatment
approach for glioma.

METHODS
Network pharmacology and databases analysis
Predicting the target genes of isocuB
In this study, we used the Comparative Toxicogenomics database (http://ctdbase.org/), the PharmMapper
database (http://www.lilab-ecust.cn/pharmmapper/), and the SwissTargetPrediction database (https://www.
sib.swiss/). We set our filter to “number of interactions > 1” to identify isocuB targets. In total, 300 potential
targets predicted for isocuB were identified.

Search related therapeutic target genes of glioma
We simultaneously used “glioma” as a keyword in the Online Catalog of Human Genes and Disorders
(OMIM) database (https://www.omim.org/), the CTD database, and the GeneCards database (https://www.
genecards.org). We set the filter to “Relevance score ≥ 10” to search for therapeutic targets associated with
glioma. After collating the results, we identified 22,320 relevant therapeutic targets by removing duplicate
values.