Nitro-substituted tert-butyl peresters induce apoptosis and exhibit multi-targeted anticancer activity in breast
Hedyeh Zafari1, Raheleh Shakeri1, Khadijeh Pouraghajan2
1Department of Biological Science, Faculty of Science, University of Kurdistan, Sanandaj, Iran.
Abstract:
The anticancer effects of six tert-butyl peresters derivatives were investigated against MDA‑MB‑231 (triple‑negative) and MCF‑7 (hormone receptor‑positive) breast cancer cells. Specifically, only the nitro‑substituted derivatives, including tert-butyl-4-nitroperbenzoate (1SS) and tert-butyl-3,5-dinitroperbenzoate (6SS), showed significant cytotoxicity against the cancer cells while displaying lower cytotoxicity against the non-cancerous L929 cell line. The triple‑negative cell line was consistently more sensitive. Compound 6SS was the more potent agent, with IC₅₀ values reaching 0.35 µg/mL in MDA‑MB‑231 monolayer cultures after 72 h (compared to 3.18 µg/mL for 1SS). Both compounds were cytotoxic in three‑dimensional (3D) spheroid models, although higher IC₅₀ values indicated greater resistance in the 3D environment. Caspase‑3/7 activity increased by 3.0‑fold (6SS) and 2.0‑fold (1SS) in MDA‑MB‑231 cells, and by 4.4‑fold (6SS) and 2.8‑fold (1SS) in MCF‑7 cells, suggesting that apoptosis is the primary cell death pathway. Neither compound inhibited the enzymatic activity of MMP‑9 or MMP‑2. Notably, while MMP‑9 appeared as a hub protein in the 1SS network, no direct enzymatic inhibition was detected, suggesting a non‑enzymatic or indirect regulatory mechanism. In silico predictions showed favorable drug‑likeness, high gastrointestinal absorption, absence of P‑glycoprotein substrate activity, and lower predicted acute toxicity (LD₅₀ of 914-1200 mg/kg) compared to doxorubicin (205 mg/kg), all of which await experimental confirmation. Network pharmacology predicted distinct target profiles: 1SS hubs include PTGS2, MMP9 and RELA (inflammation and tumor microenvironment), whereas 6SS targets EGFR, ERBB2, CASP3, PTGS2 and HSP90AA1 (proliferation, apoptosis and protein stability). Taken together, the dinitro derivative 6SS exhibits potent, multi‑targeted anti‑breast cancer activity through a proposed caspase‑dependent apoptotic mechanism, with a safety profile that supports further development.
Supplementary Information:
The online version contains supplementary material available at 10.1007/s13205-026-04893-x.
Insights
Nitro-substituted tert-butyl peresters show potent anticancer effects against triple-negative and hormone receptor-positive breast cancer cells. The dinitro derivative 6SS is particularly effective, inducing apoptosis with a favorable safety profile for further development.
Area of Science:
- Medicinal Chemistry
- Pharmacology
- Oncology
Background:
- Breast cancer remains a leading cause of cancer-related deaths globally.
- Development of novel therapeutics with improved efficacy and safety profiles is crucial.
- Tert-butyl peresters represent a class of compounds with potential anticancer properties.
Purpose of the Study:
- To investigate the anticancer effects of novel tert-butyl perester derivatives against human breast cancer cell lines.
- To identify specific derivatives with significant cytotoxicity and favorable safety profiles.
- To elucidate the mechanism of action, including cell death pathways and molecular targets.
Main Methods:
- Synthesis and evaluation of six tert-butyl perester derivatives.
- Cytotoxicity assays against MDA-MB-231 (triple-negative) and MCF-7 (hormone receptor-positive) breast cancer cells, and L929 non-cancerous cells.
- Caspase-3/7 activity assays to assess apoptosis.
- Inhibition assays for MMP-2 and MMP-9.
- In silico drug-likeness and toxicity predictions.
- Network pharmacology analysis to predict molecular targets.
Main Results:
- Nitro-substituted derivatives, particularly tert-butyl-3,5-dinitroperbenzoate (6SS), exhibited significant cytotoxicity against both breast cancer cell lines.
- Compound 6SS demonstrated higher potency (IC50 of 0.35 µg/mL in MDA-MB-231 cells) compared to tert-butyl-4-nitroperbenzoate (1SS).
- Apoptosis was identified as the primary cell death pathway, evidenced by increased caspase-3/7 activity.
- In silico studies predicted favorable drug-likeness and a lower toxicity profile compared to doxorubicin.
- Network pharmacology identified distinct target profiles for 1SS and 6SS, involving inflammation, tumor microenvironment, proliferation, and apoptosis pathways.
Conclusions:
- The dinitro derivative 6SS displays potent, multi-targeted anti-breast cancer activity.
- The mechanism of action involves caspase-dependent apoptosis.
- Compound 6SS possesses a promising safety profile, warranting further preclinical and clinical development.
- Tert-butyl peresters, especially nitro-substituted ones, represent a viable class of novel anticancer agents for breast cancer therapy.
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