Multidrug-Resistant Profiles in Non-Small Cell Lung Carcinoma Patient-Derived Cells: Implications for Personalized Approaches with Tyrosine Kinase Inhibitors.

The impact of tyrosine kinase inhibitors (TKIs) on multidrug resistance (MDR) in non-small cell lung carcinoma (NSCLC) is a critical aspect of cancer therapy. While TKIs effectively target specific signaling pathways of cancer cells, they can also act as substrates for ABC transporters, potentially triggering MDR. The aim of our study was to evaluate the response of 17 patient-derived NSCLC cultures to 10 commonly prescribed TKIs and to correlate these responses with patient mutational profiles. Using an ex vivo immunofluorescence assay, we analyzed the expression of the MDR markers ABCB1, ABCC1, and ABCG2, and correlated these data with the genetic profiles of patients for a functional diagnostic approach. NSCLC cultures responded differently to TKIs, with erlotinib showing good efficacy regardless of mutation burden or EGFR status. However, the modulation of MDR mechanisms by erlotinib, such as increased ABCG2 expression, highlights the challenges associated with erlotinib treatment. Other TKIs showed limited efficacy, highlighting the variability of response in NSCLC. Genetic alterations in signaling pathways associated with drug resistance and sensitivity, including TP53 mutations, likely contributed to the variable responses to TKIs. The relationships between ABC transporter expression, gene alterations, and response to TKIs did not show consistent patterns. Our results suggest that in addition to mutational status, performing functional sensitivity screening is critical for identifying appropriate treatment strategies with TKIs. These results underscore the importance of considering drug sensitivity, off-target effects, MDR risks, and patient-specific genetic profiles when optimizing NSCLC treatment and highlight the potential for personalized approaches, especially in early stages.

Immunofluorescence-Based Assay for High-Throughput Analysis of Multidrug Resistance Markers in Non-Small Cell Lung Carcinoma Patient-Derived Cells.

Lung cancer remains the leading cause of cancer death globally, with non-small cell lung cancer (NSCLC) accounting for the majority of cases. Multidrug resistance (MDR), often caused by ATP-binding cassette (ABC) transporters, represents a significant obstacle in the treatment of NSCLC. While genetic profiling has an important role in personalized therapy, functional assays that measure cellular responses to drugs are gaining in importance. We developed an automated microplate-based immunofluorescence assay for the evaluation of MDR markers ABCB1, ABCC1, and ABCG2 in cells obtained from NSCLC patients through high-content imaging and image analysis, as part of a functional diagnostic approach. This assay effectively discriminated cancer from non-cancer cells within mixed cultures, which is vital for accurate assessment of changes in MDR marker expression in different cell populations in response to anticancer drugs. Validation was performed using established drug-sensitive (NCI-H460) and drug-resistant (NCI-H460/R) NSCLC cell lines, demonstrating the assay's capacity to distinguish and evaluate different MDR profiles. The obtained results revealed wide-ranging effects of various chemotherapeutic agents on MDR marker expression in different patient-derived NSCLC cultures, emphasizing the need for MDR diagnostics in NSCLC. In addition to being a valuable tool for assessing drug effects on MDR markers in different cell populations, the assay can complement genetic profiling to optimize treatment. Further assay adaptations may extend its application to other cancer types, improving treatment efficacy while minimizing the development of resistance.

Coleon U, Isolated from Plectranthus mutabilis Codd., Decreases P-Glycoprotein Activity Due to Mitochondrial Inhibition.

Multidrug resistance in cancer is often mediated by P-glycoprotein. Natural compounds have been suggested as a fourth generation of P-glycoprotein inhibitors. Coleon U, isolated from Plectranthus mutabilis Codd., was reported to modulate P-glycoprotein activity but the underlying mechanism has not yet been revealed. Therefore, the effects of Coleon U on cell viability, proliferation, and cell death induction were studied in a non-small-cell lung carcinoma model comprising sensitive and multidrug-resistant cells with P-glycoprotein overexpression. P-glycoprotein activity and mitochondrial membrane potential were assessed by flow cytometry upon Coleon U, sodium-orthovanadate (an ATPase inhibitor), and verapamil (an ATPase stimulator) treatments. SwissADME was used to identify the pharmacokinetic properties of Coleon U, while P-glycoprotein expression was studied by immunofluorescence. Our results showed that Coleon U is not a P-glycoprotein substrate and is equally efficient in sensitive and multidrug-resistant cancer cells. A decrease in P-glycoprotein activity observed with Coleon U and verapamil after 72 h is antagonized in combination with sodium-orthovanadate. Coleon U induced a pronounced effect on mitochondrial membrane depolarization and showed a tendency to decrease P-glycoprotein expression. In conclusion, Coleon U-delayed effect on the decrease in P-glycoprotein activity is due to P-glycoprotein's functioning dependence on ATP production in mitochondria.