Chilli extract shows preclinical activity against lung adenocarcinoma in rats

An ethanolic extract of Capsicum annuum , commonly known as chilli pepper, reduced lung-cancer-associated markers and inflammation in rats with experimentally induced lung adenocarcinoma, according to a study that combined chemical profiling, computational analyses, cell assays and animal experiments. The extract also lowered proliferation and nuclear factor kappa B staining while increasing the apoptosis marker caspase-3 in lung tissue. These findings remain preclinical: they show activity in cells and rats, not a proven treatment or preventive agent for lung cancer in people.

The researchers prepared the extract from fruits collected in North Sumatra, Indonesia, and used the same extract batch throughout the investigation. High-resolution liquid chromatography coupled to mass spectrometry tentatively identified 30 compounds across capsaicinoids and capsinoids, phenolic acids, flavonoids, carotenoids and organic acids. The profile included capsaicin, dihydrocapsaicin, homocapsaicin, rutin, quercetin, ferulic acid, chlorogenic acid, several carotenoids and ascorbic acid.

These annotations were obtained in positive electrospray-ionization mode and were semi-quantitative, while the plant’s cultivar was not characterized. The varied chemical composition means that the experimental effects cannot be attributed to capsaicin alone. The extract was also standardized by high-performance liquid chromatography with photodiode-array detection for capsaicin, dihydrocapsaicin, quercetin and ferulic acid, while the chemical profile was based on a batch from a single geographic source and an unspecified cultivar.

To identify potential molecular targets, the investigators combined predicted compound targets with genes associated with lung cancer. They found 503 shared targets and constructed a high-confidence human protein–protein interaction network, which identified AKT1, SRC, TNF, EGFR, IL6, CTNNB1, STAT3, HSP90AA1, ESR1 and HIF1A as highly connected hubs. Enrichment analysis highlighted EGFR tyrosine kinase inhibitor resistance, PI3K–Akt signalling, HIF-1 signalling and broader cancer pathways.

Molecular docking then predicted that rutin had the broadest interaction profile, ranking highest for six of the ten hubs, including SRC, AKT1, TNF, IL-6, CTNNB1 and STAT3. Rutin produced the strongest score in the screen for SRC, at −12.3 kcal/mol, and also ranked highly for AKT1 and EGFR. Other compounds had prominent predicted interactions: beta-carotene scored −11.7 kcal/mol for EGFR, zeaxanthin −9.2 kcal/mol for ESR1, and capsorubin −8.8 kcal/mol for HIF1A.

These docking scores estimate binding compatibility; they do not demonstrate that a compound inhibits a protein in cells or that a predicted target mediates an observed effect. Key predicted pathways, including AKT, EGFR, STAT3 and HIF-1α signalling, were not directly validated by immunoblotting or quantitative gene-expression analysis. In human A549 lung adenocarcinoma cells, the chilli extract reduced viability in a concentration- and exposure-dependent manner.

Its half-maximal inhibitory concentration was 148.6 ± 6.4 micrograms per millilitre after 24 hours and 86.3 ± 4.9 micrograms per millilitre after 48 hours. Normal human MRC-5 lung fibroblasts were less susceptible, producing selectivity indices of 2.26 and 2.75 at the respective exposure times. Those values indicate modest rather than wide selectivity between cancer and normal cells, and the assay did not include a reference chemotherapeutic agent for comparison.

The study also showed that the extract increased intracellular reactive oxygen species in A549 cells, with DCF fluorescence reaching 268.5 ± 11.4% of control after treatment with 200 micrograms per millilitre for 24 hours. The highest concentration also produced morphological signs of oxidative injury. Cell-based biomarker measurements revealed a more complex redox response.

Treatment increased superoxide dismutase, glutathione, catalase and glutathione peroxidase, consistent with activation of cellular antioxidant defenses, while reducing TNF-alpha, IL-6, NF-kappa B, cyclooxygenase-2 and vascular endothelial growth factor. At higher concentrations, however, malondialdehyde also increased, indicating greater lipid peroxidation. Together, the results are consistent with an adaptive antioxidant response that did not fully offset the extract-associated oxidative burden in A549 cells.

Separate cell-free DPPH, ABTS and ferric-reducing antioxidant assays showed moderate activity rather than strong radical-scavenging effects. The ethanol extract had the highest DPPH inhibition, at a maximum of 34.1% at 80 micrograms per millilitre, and the highest mass-normalized reducing activity alongside the methanol extract. The investigators therefore used the ethanol extract for the principal cell and animal experiments.

The animal experiment used 25 male Wistar rats divided among five groups, with five animals per group. Lung adenocarcinoma was induced with diethylnitrosamine, or DEN, a carcinogen that is bioactivated in lung tissue and produces lesions with features resembling human lung adenocarcinoma. One group served as an untreated normal control, another received DEN without treatment, and a third received the chemotherapy docetaxel at 10 milligrams per kilogram intraperitoneally once a week.

The remaining groups received the chilli extract orally at 100 or 200 milligrams per kilogram for 14 consecutive days. Investigators responsible for biochemical measurements, histopathological evaluation and data analysis were blinded to treatment allocation, and no unexpected mortality was reported during the treatment period. The researchers selected male rats to reduce hormonal fluctuations associated with the estrous cycle, a choice that also limits how directly the findings can be generalized across sexes.

Compared with normal animals, DEN-induced rats had higher serum carcinoembryonic antigen, neuron-specific enolase, malondialdehyde, IL-6 and TNF-alpha, together with lower superoxide dismutase and glutathione. The extract shifted these measurements toward the normal range in a dose-dependent fashion. At 200 milligrams per kilogram, it reduced carcinoembryonic antigen by 53% and neuron-specific enolase by 37% relative to DEN-only animals.

At the same dose, it reduced malondialdehyde by 48%, IL-6 by 55% and TNF-alpha by 52%, while increasing superoxide dismutase by 52% and glutathione by 107%. The 200-milligram effects approached those measured in the docetaxel group, but the comparison came from a preclinical experiment with five animals per group. It should not be interpreted as evidence that the extract is equivalent to chemotherapy, and serum marker changes do not by themselves establish reduced tumor burden.

Immunohistochemical observations were consistent with reduced proliferation and inflammation and increased apoptosis. DEN-treated lungs showed strong Ki-67 staining, weak caspase-3 staining and intense NF-kappa B immunoreactivity. Chilli extract reduced Ki-67 and NF-kappa B staining, with clearer effects at 200 milligrams per kilogram, while increasing caspase-3 staining and improving the appearance of alveolar architecture.

Docetaxel produced similar directional changes. However, these findings came from descriptive assessment of representative tissue fields rather than digital image analysis, and the study did not include hematoxylin-and-eosin histopathological scoring. The staining trends align with the biochemical results but cannot independently establish a reduction in tumor burden or confirm the proposed mechanisms.

In addition, the animal experiment assessed outcomes only at the endpoint because DEN produces multifocal intrapulmonary lesions that could not be followed with caliper measurements, and longitudinal imaging was unavailable. The contrast between the cell and animal results may reflect different exposure conditions and tissue contexts. Direct exposure of cultured cancer cells to relatively high extract concentrations increased ROS and coincided with reduced viability, whereas oral administration in rats was associated with lower serum malondialdehyde and higher antioxidant levels.

The authors noted that systemic exposure after oral dosing would be lower and metabolically transformed, while A549 cells have different basal redox conditions from normal tissue. These considerations may help explain why direct cellular exposure was associated with oxidative-stress markers while the treated rats showed restoration of serum antioxidant measures. They do not establish the mechanism underlying the observations.

Other limitations include the modest in vitro selectivity index, the absence of ROS and cell-death measurements in normal cells, use of only male animals, a 14-day treatment period, endpoint-only measurements, no pharmacokinetic analysis, and the lack of direct validation of predicted signalling pathways. The extract also requires standardization, quantitative marker assays, formal histopathological analysis, pharmacokinetic evaluation and repeated-dose toxicity testing before it could be considered for clinical study.

Subject of Research: Preclinical evaluation of Capsicum annuum extract against chemically induced lung adenocarcinoma

Article Title: Capsicum annuum suppresses lung adenocarcinoma through oxidative, inflammatory, and survival pathway modulation

Article References: Syahputra, R. A., Nugraha, S. E., Rimayani, L., Rohani, A. S., Juwita, N. A., Halim, P., Choi, J., Choi, M., Kim, H. J., Park, S., Akter, S., Rani, A., Kim, B., & Kim, S.-H. (2026). Capsicum annuum suppresses lung adenocarcinoma through oxidative, inflammatory, and survival pathway modulation. Current Research in Biotechnology, Article 100429. https://doi.org/10.1016/j.crbiot.2026.100429

Image Credits: AI Generated

DOI: 10.1016/j.crbiot.2026.100429

Keywords: Capsicum annuum, chilli pepper extract, lung adenocarcinoma, A549 cells, DEN-induced rat model, oxidative stress, inflammation, network pharmacology, molecular docking, apoptosis, Capsicum, annuum