FL496, an FL118-derived small molecule, induces growth inhibition, senescence, and apoptosis of malignant pleural mesothelioma (MPM) cells, and exhibits anti-MPM tumor efficacy strikingly superior to the pemetrexed-cisplatin combination

Our plain-English summary of FL496, an FL118-derived small molecule, induces growth inhibition, senescence, and apoptosis of malignant pleural mesothelioma (MPM) cells, and exhibits anti-MPM tumor efficacy strikingly superior to the pemetrexed-cisplatin combination, by I.A.M. Aljahdali, X. Ling, W. Wu, et al (Journal of Experimental & Clinical Cancer Research, 2025).

The study

• Authors: I.A.M. Aljahdali, X. Ling, W. Wu, W. Wang, D. Li, R. Zhang, E. Zhang, A. Stablewski, R. Azrak, Q. Li, F. Li
• Journal: Journal of Experimental & Clinical Cancer Research
• Published: 2025;44(1):293
• Cited by: 1 article, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: Malignant pleural mesothelioma (MPM) responds poorly to chemotherapy and is a highly progressive malignancy with a median survival time of only 6-9 months. Therefore, the development of anti-MPM tumor agents with high efficacy and low toxicity is urgent and addresses an unmet need for MPM patients.

Methods: Medicinal chemistry synthesis of small molecules based on the FL118 drug platform were further comparatively investigated using multiple MPM and osteosarcoma cell/tumor in vitro and/or in vivo models. The method includes cell viability assay, Western blot analysis, colony formation assay, immunocytochemical staining, β-galactosidase senescence staining, flow cytometry, DNA fragmentation cell death detection, vector-free CRISPR-Cas9-mediated gene knockout, bioinformatic analysis, FL496 efficacy determination using severe combined immunodeficiency (SCID) mice with human MPM tumor, and immunohistochemistry (IHC) analysis of MPM tumors.

Results: Here, we report that we identified a novel FL118-derived small molecule (FL496). FL496 appears to be strikingly more effective in inhibiting MPM tumor growth in MPM tumor animal models than the currently most prevalent pemetrexed-cisplatin combination in the clinic. The treatment of MPM cells with FL496 rapidly induced p53 and p21 accumulation, and Rb and p-Rb inhibition, which were associated with MPM cell senescence and G1/G0 arrest and apoptosis. Knockout (KO) of the TP53/p53 gene decreased the ability of FL496 to inhibit MPM cell growth (i.e., increase FL496 IC50 values) and colony formation. FL496-treated MPM cells resulted in strong inhibition of the expression of survivin, Mcl-1, Bcl-2, Bcl-XL, and the induction of active caspase-3, cleaved PARP, and PUMA, which were further confirmed using MPM tumor tissues via IHC analysis. High survivin in MPM patients' tumors is associated with poor patient survival. Similar to FL118, FL496 treatment reduces DDX5 expression in MPM cells, but FL496 is more potent than FL118 in inhibiting MPM cell growth. Therefore, the mechanism of action (MOA) of FL496 overlaps with, but is likely beyond the scope of FL118 MOA, which needs further investigation.

Conclusions: Together, these results indicate that FL496 is a promising anti-MPM small molecule, and its high anti-MPM potential is worthy of being further explored as a monotherapeutic agent to treat MPM patients in clinical trials.

© 2025. The Author(s).

In plain English

This was a laboratory study of FL496, a new small molecule derived from the FL118 compound platform, tested against malignant pleural mesothelioma (MPM), a cancer the authors describe as causally associated with asbestos exposure. The authors first screened three FL118 analogues (FL7N1, FL7N2, FL7N3) in mice carrying human cervical cancer HT-3 tumours, then compared SN-38, FL7N1, FL118 and FL496 in two human MPM cell lines (MSTO-211H and NCI-H226); FL496 and FL7N1 were separately compared in four osteosarcoma PDX-derived cell lines. Among the small molecules tested in the two MPM cell lines, FL496 gave the highest cell-growth inhibition. FL496 treatment raised p53, p21 and the DNA-damage marker gamma-H2AX, produced senescence and G0/G1 arrest, lowered survivin, Mcl-1, Bcl-2, Bcl-XL and DDX5, and raised cleaved caspase-3, cleaved PARP and PUMA. Knocking out TP53 made MPM cells less sensitive. In 45 SCID mice bearing MSTO-211H tumours (nine groups of five), FL496 alone slowed tumour growth more than the pemetrexed-cisplatin combination. In TCGA mesothelioma data (84 primary tumours), high survivin expression was associated with shorter survival. The authors concluded FL496 merits further development toward clinical trials.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: preclinical laboratory study: chemical synthesis plus in vitro cell experiments, CRISPR-Cas9 gene knockout, bioinformatic analysis of a public patient dataset, and mouse xenograft tumour experiments
• Who was studied: Human MPM cell lines MSTO-211H and NCI-H226, osteosarcoma U2OS and four osteosarcoma PDX-derived cell lines, cervical cancer HT-3 cells; 45 female SCID mice bearing MSTO-211H xenograft tumours (5 per group, 9 groups); and 84 primary tumours from the TCGA-MESO dataset
• Exposure: Treatment with the synthesised compounds FL496, FL118, FL7N1, FL7N2, FL7N3 (and comparators SN-38, etoposide) in cell culture, and in mice FL496 at 1.25-7.5 mg/kg, pemetrexed 15-30 mg/kg and cisplatin 1.5-3 mg/kg, alone or combined
• Main results:
  • Among FL496, FL118, FL7N1 and SN-38 compared in the two MPM cell lines, FL496 gave the greatest inhibition of cell growth (IC50 values shown only in the figures)
  • In four osteosarcoma PDX-established cell lines, FL496 was compared with FL7N1 and inhibited cell growth/viability
  • FL496 raised p53 and p21 and lowered Rb and phosphorylated Rb in both MPM cell lines, with senescence and G0/G1 arrest
  • In MPM cells with TP53 knocked out, FL496 was less effective; in knockout NCI-H226 cells 500 nM FL496 did not reach the IC50
  • In U2OS osteosarcoma cells, FL496 raised gamma-H2AX earlier than p53 and p-p53(S15); etoposide was used as the comparison drug
  • FL496 lowered Mcl-1, Bcl-2 and Bcl-XL and raised cleaved caspase-3, cleaved PARP and PUMA in MSTO-211H and NCI-H226 cells; after p53 knockout, PUMA and cleaved caspase-3 were absent
  • FL496 lowered survivin and DDX5 expression in both MPM cell lines
  • In the TCGA-MESO dataset, survivin was higher in biphasic than epithelioid MPM, and high survivin went with shorter overall survival
  • In the mouse MPM xenograft experiment, FL496 alone slowed tumour growth more than pemetrexed plus cisplatin; adding FL496 to the combination was not better than FL496 alone
  • Tumour tissue from FL496-treated mice showed more active caspase-3 and gamma-H2AX and fewer Ki67-positive cells at 4 and 8 weeks
  • Mouse body weight change across treatment groups stayed within 10%
• The authors' conclusion: The authors concluded that FL496 is a promising anti-MPM small molecule worth exploring as a single-agent treatment for MPM patients in clinical trials
• Limitations the authors state:
  • The authors noted the limitation of using only two mesothelioma cell models, which they tried to offset with non-mesothelioma cell models
  • The mechanism of action of FL496 was not fully worked out and needs follow-up study
  • The relationship between DDX5 and p53 in MPM cells was unclear, so why p53 knockout reduced FL496 activity remains to be explained
  • The initial in vivo compound screen used only 1-2 mice per drug dose level

Topics

The National Library of Medicine indexes this study under subjects our site covers: Lung cancer, Mesothelioma.

Medical terms in this study, explained in our Lexicon: Apoptosis, Pleural Mesothelioma, Pemetrexed (Alimta), Cisplatin, Chemotherapy, Median Overall Survival, Immunohistochemistry, Clinical Trial.

Related studies

• In vitro and in vivo therapeutic efficacy of the PPAR-γ agonist troglitazone in combination with cisplatin against malignant pleural mesothelioma cell growth — N. Hamaguchi, H. Hamada, et al. (2010)
• Interferon-β produces synergistic combinatory anti-tumor effects with cisplatin or pemetrexed on mesothelioma cells — Q. Li, K. Kawamura, et al. (2013)
• Synergistic antitumor activity of recombinant human Apo2L/tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in combination with carboplatin and pemetrexed in malignant pleural mesothelioma — G. Pasello, L. Urso, et al. (2014)
• Dexamethasone alleviates pemetrexed-induced senescence in Non-Small-Cell Lung Cancer — H. Ge, J. Ke, et al. (2018)
• Inhibition of autophagy potentiates pemetrexed and simvastatin-induced apoptotic cell death in malignant mesothelioma and non-small cell lung cancer cells — K.E. Hwang, Y.S. Kim, et al. (2015)
• Combination of onconase and dihydroartemisinin synergistically suppresses growth and angiogenesis of non-small-cell lung carcinoma and malignant mesothelioma — R. Shen, J. Li, et al. (2016)

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