Page 161 - SRPSKO DRUŠTVO ISTRAŽIVAČA RAKA

SDIRSACR Oncology Insights

MtSR-positive subpopulation was dominant in all treated A549 cells. The true degree of ROS overburden in Mcf7 cells
was revealed by a massive increase in DCFDA-positive events. MTR staining of A549 cells showed the simultaneous
presence of hypo and hyperpolarized mitochondria within the same cell, indicating a variety in cellular mitochondrial
polarization, whereas integrity of mitochondrial network was severely affected in Mcf7 cells.
Conclusions: Increased survival of A549 and Mcf7 cells co-incubated with 1a-1d and NAC served as a validation that
both cell lines were affected by ROS generation upon treatment. Dysfunctional oxidative phosphorylation (OXOPHOS)
in KRASmut A549 cells indicated that ROS generation was not launched at the level of mitochondrial respiration
complexes I-V. Mcf7 cells were more susceptible to experimental treatments than A549 cells at both 24 h and 6 h of
investigation. These results imply that fully operative OXOPHOS in KRASwt cells exceedingly amplifies ROS production
in response to investigated compounds.

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Dual diagnostic and therapeutic use of photoactivable BODIPY dyes in cancer

Marija Mioč , Mladen Paradžik , Lidija Uzelac , Mladena Glavaš , Mihaela Matovina , Marina Oskomić , Lea Barbarić
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2 , Nikola Basarić , Marijeta Kralj 1
1Division of Molecular Medicine, Ruđer Bošković Institute, Zagreb, Croatia
2Division of Organic Chemistry and Biochemistry, Ruđer Bošković Institute, Zagreb, Croatia
Keywords: anticancer phototherapy, BODIPY, protein labelling, theranostics
Background: Quinone methides (QMs) are reactive intermediates derived from phenols, capable of alkylating and
cross-linking DNA, and are linked to the anticancer activity of drugs like mitomycin and anthracyclines. Their targeted
photochemical generation in tumors presents a promising strategy for novel phototherapeutics. We have developed
several QM precursors and explored their photocytotoxicity. Recently, we focused on QM precursors linked to BODIPY
chromophores, aiming to activate them using visible light. Notably, we observed unusual photoreactivity from higher
excited states, enabling fluorescent photolabeling of proteins. To advance potential theranostic applications, it is
essential to evaluate the subcellular localization, photoinduced cytotoxicity, and protein-labeling capabilities of these
compounds.
Materials and Methods: The antiproliferative activity of BODIPY–QM compounds was assessed using the MTT assay
on various human and mouse cancer and non-cancer cell lines. Intracellular localization was analyzed by confocal
microscopy. Photolabeling experiments were conducted by irradiating model proteins (e.g., bovine serum albumin,
human serum albumin, and dipeptidyl peptidase III – DPP III), followed by SDS-PAGE. The enzymatic activity of irradiated
DPP III was also evaluated.
Results: In the absence of light, the compounds exhibited little or no cytotoxicity. However, upon visible-light irradiation,
several compounds showed marked cytotoxicity, confirming photoactivation-dependent effects. Confocal imaging
revealed that BODIPY derivatives localize predominantly in the cytoplasm and membrane-rich organelles, as confirmed
using organelle-specific antibodies. Irradiation of model proteins led to covalent dye–protein binding, with efficiency
varying between proteins. Optimal concentrations and irradiation times were determined for effective photolabeling.
Conclusions: BODIPY-based QM precursors show strong potential for the development of photoactivatable anticancer
agents, effective under both normoxic and hypoxic conditions. Furthermore, their capacity for fluorescent protein
labeling supports their use in innovative diagnostic or theranostic approaches.

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