Nintedanib in TGF-β1-stimulated primary human lung fibroblasts
Cell-based study · Primary human lung fibroblasts from healthy donors (Lonza CC-2512); 12-day ficoll-crowded Scar-in-a-Jar culture in low-serum DMEM + GlutaMax (0.4% FBS), stimulated with 1 ng/mL TGF-β1. Methods describe passage 5 and 9 cells; nintedanib experiments used Lonza cells. Repeated experiments are not equated with independent donors. [1]
One coherent concentration/time experiment combines matrix/activation biomarker readouts with metabolic-activity and LDH controls. Biomarker reductions occur alongside reduced metabolic activity at higher concentrations; 10 μM was excluded from biomarker analysis. PRO-C3/α-SMA had no day-4 response. Healthy-donor, single-stimulus culture does not establish tissue fibrolysis, a direct kinase mechanism or animal/clinical benefit.
| Endpoint | Reported result | Reference |
|---|---|---|
| Supernatant PRO-C1, PRO-C3, PRO-C6, FBN-C and α-SMA | Qualitative concentration/time response PRO-C1, PRO-C6 and FBN-C decreased at days 4, 8 and 12; PRO-C3 and α-SMA decreased at days 8 and 12 but were not altered at day 4. Highest concentration (10 μM) omitted from biomarker analyses. | [1] |
| Cell metabolic activity at day 12 | Alamar Blue relative to TGF-β1 control 75%, 63% and 0% at 100 nM, 1 μM and 10 μM, respectively; all P < 0.0001 versus vehicle as described in Results. | [1] |
| Cytotoxicity marker LDH release | Statistical comparison with vehicle No significant change; small trend toward elevated LDH at 10 μM. | [1] |
Literature-Reported Treatment Conditions
- Treatment concentration
- Nintedanib: 1 nM, 10 nM, 100 nM, 1 μM and 10 μM; 10 μM excluded from biomarker analysis after complete Alamar Blue signal loss. TGF-β1: 1 ng/mL.
- Exposure duration
- 12 days; fresh stimulation medium and treatments exchanged on days 0, 4 and 8. Supernatant biomarkers collected on days 4, 8 and 12; displayed Alamar Blue day 12 and LDH day 4.
- Method
- Culture exposure in ficoll-containing low-serum medium. Nintedanib versus corresponding vehicle, with TGF-β1 control used for normalization. Each drug tested separately; other comparator compounds are not combined with nintedanib.
- Vehicle
- 0.1% DMSO kept constant for nintedanib and corresponding vehicle control.
- Co-treatment
- TGF-β1 serves as the profibrotic stimulus; no therapeutic drug combination adopted.
- Reported limits
- Exact donor count and per-experiment passage assignment are not established by the read nintedanib design. No Adooq material, lot, independent purity or stereochemical analysis is established. Detailed referenced ELISA validation papers were not retrieved.
Methods and source locations
Compound reported in the paper: Nintedanib (BIBF1120); Compound label in the paper: Kemprotec nintedanib, reported cat. no. 656247–17-5
Literature findings for the reported CAS-matched Kemprotec material in healthy-donor cultured fibroblasts. No Adooq-lot equivalence, IPF-patient-cell result, direct kinase IC50 or in-vivo efficacy claim.
Methods: Fibroblast cell culture, Metabolic activity assessed by Alamar blue, Cytotoxicity assessed by LDH release, Biomarker measurements, Statistical analysis; Results nintedanib subsection; Figure 5A–G and caption; Discussion model limitations
Supernatant PRO-C1, PRO-C3, PRO-C6, FBN-C and α-SMA: Competitive ELISAs; data expressed as % of TGF-β1 control, with corresponding 0.1% DMSO vehicle comparisons. Figure caption: mean ± SD of 3 separate experiments, each with 4 replicates/treatment; two-way ANOVA with Sidak’s or Dunnett’s multiple comparisons. No donor n or exact graph values inferred. — Methods Biomarker measurements and Statistical analysis; Results nintedanib subsection; Figure 5C–G/caption
Cell metabolic activity at day 12: Alamar Blue. Printed Results percentages, not estimates from the curve. Figure 5 caption gives mean ± SD of 3 separate experiments, each with 4 replicates/treatment. Loss of signal is not a separately measured cell-death percentage. — Methods Metabolic activity assessed by Alamar blue; Results nintedanib subsection; Figure 5A/caption
Cytotoxicity marker LDH release: LDH relative to maximum release from lysis control. Methods describe days 4, 8 and 12; Figure 5B displays day 4 only. No quantitative claim for undisplayed later LDH timepoints. Figure caption: mean ± SD of 3 separate experiments each with 4 replicates/treatment. — Methods Cytotoxicity assessed by LDH release; Results nintedanib subsection; Figure 5B/caption