Monitoring doxorubicin cellular uptake and trafficking using in vitro Raman microspectroscopy: short and long time exposure effects on lung cancer cell lines

Howe³

et al. 2017

Self Cite

Raman micro‐spectroscopy is a non‐invasive analytical tool, whose potential in cellular analysis and monitoring drug mechanisms of action has already been demonstrated, and which can potentially be used in pre‐clinical and clinical applications for the prediction of chemotherapeutic efficacy. To further investigate such potential clinical application, it is important to demonstrate its capability to differentiate drug mechanisms of action and cellular resistances. Using the example of Doxorubicin (DOX), in this study, it was used to probe the cellular uptake, signatures of chemical binding and subsequent cellular responses, of the chemotherapeutic drug in two lung cancer cell lines, A549 and Calu‐1. Multivariate statistical analysis was used to elucidate the spectroscopic signatures associated with DOX uptake and subcellular interaction. Biomarkers related to DNA damage and repair, and mechanisms leading to apoptosis were also measured and correlated to Raman spectral profiles. Results confirm the potential of Raman spectroscopic profiling to elucidate both drug kinetics and pharmacodynamics and differentiate cellular drug resistance associated with different subcellular accumulation rates and subsequent cellular response to DNA damage, pointing towards a better understanding of drug resistance for personalised targeted treatment.

“…The DOX band was determined, as Figure , by PCA control vs exposed cells. The evolution of similar bands in A549 cells has been demonstrated previously .…”

Section: Resultssupporting

confidence: 77%

Section: Resultssupporting

confidence: 76%

Section: Resultssupporting

confidence: 58%

Section: Resultsmentioning

confidence: 99%

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Doxorubicin kinetics and effects on lung cancer cell lines using in vitro Raman micro‐spectroscopy: binding signatures, drug resistance and DNA repair

Farhane¹,

Howe³

et al. 2017

Self Cite

“…Confocal laser scanning fluorescence images of (A) A549 cells and (B) Calu‐1 cells after DOX exposure and corresponding controls: 1.2‐hour DOX exposure, 2.6‐hour DOX exposure, 3.12‐hour DOX exposure, 4.24‐hour DOX exposure, 5.48‐hour DOX exposure and 6.72‐hour DOX exposure …”

Section: Raman Microspectroscopy For Chemotherapeutic Pre‐clinical Scmentioning

confidence: 99%

In vitro label‐free screening of chemotherapeutic drugs using Raman microspectroscopy: Towards a new paradigm of spectralomics

Farhane¹,

Nawaz

et al. 2018

Self Cite

This overview groups some of the recent studies highlighting the potential application of Raman microspectroscopy as an analytical technique in preclinical development to predict drug mechanism of action and in clinical application as a companion diagnostic and in personalised therapy due to its capacity to predict cellular resistance and therefore to optimise chemotherapeutic treatment efficacy. Notably, the anthracyclines, doxorubicin and actinomycin D, elicit similar spectroscopic signatures of subcellular interaction characteristic of the mode of action of intercalation. Although cisplatin and vincristine show markedly different signatures, at low exposure doses, their signatures at higher doses show marked similarities to those elicited by the intercalating anthracyclines, confirming that anticancer agents can have different modes of action with different spectroscopic signatures, depending on the dose. The study demonstrates that Raman microspectroscopy can elucidate subcellular transport and accumulation pathways of chemotherapeutic agents, characterise and fingerprint their mode of action, and potentially identify cell-resistant strains. The consistency of the spectroscopic signatures for drugs of similar modes of action, in different cell lines, suggests that this fingerprint can be considered a "spectralome" of the drug-cell interaction suggesting a new paradigm of representing spectroscopic responses.

An in vitro study of the interaction of the chemotherapeutic drug Actinomycin D with lung cancer cell lines using Raman micro‐spectroscopy

Farhane¹,

Byrne³

2017

Self Cite

The applications of Raman microspectroscopy have been extended in recent years into the field of clinical medicine, and specifically in cancer research, as a non-invasive diagnostic method in vivo and ex vivo, and the field of pharmaceutical development as a label-free predictive technique for new drug mechanisms of action in vitro. To further illustrate its potential for such applications, it is important to establish its capability to fingerprint drug mechanisms of action and different cellular reactions. In this study, cytotoxicity assays were employed to establish the toxicity profiles for 48 and 72 hours exposure of lung cancer cell lines, A549 and Calu-1, after exposure to Actinomycin D (ACT) and Raman micro-spectroscopy was used to track its mechanism of action at subcellular level and subsequent cellular responses. Multivariate data analysis was used to elucidate the spectroscopic signatures associated with ACT chemical binding and cellular resistances. Results show that the ACT uptake and mechanism of action are similar in the 2 cell lines, while A549 cells exhibits spectral signatures of resistance to apoptosis related to its higher chemoresistance to the anticancer drug ACT. The observations are discussed in comparison to previous studies of the similar anthracyclic chemotherapeutic agent Doxorubicin. A, Preprocessed Raman spectrum of ACT stock solution dissolved in sterile water and mean spectrum with SD of (B) nucleolus, (C) nucleus and (D) cytoplasm of A549 cell lines after 48 hours exposure to the corresponding IC 50 . K E Y W O R D SA549, Actinomycin D, binding signature, Calu-1, Raman micro-spectroscopy | INTRODUCTIONRaman micro-spectroscopy is a non-invasive analytical tool, whose potential in clinical applications to distinguish between normal and cancer cells and monitoring drug mechanisms of action has already been demonstrated [1][2][3][4][5]. Therefore, it can potentially be used in preclinical development for the prediction of chemotherapeutic efficacy and potentially ultimately as a companion diagnostic tool. In order to assure its application as a predictive tool, Raman spectroscopy should provide information about drug mechanisms of action and cellular reactions or resistance. Previous studies were made using Raman microspectroscopy and chemotherapeutic drugs such as Cisplatin [6], Vincristine [6], Erlotinib [7], Panitumumab [8] and Doxorubicin (DOX) [9] showing the potential of this