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PH-Assisted multichannel heat shock monitoring in the endoplasmic reticulum with a pyridinium fluorophore.

ABSTRACT: Heat shock is a global health concern as it causes permanent damage to living cells and has a relatively high mortality rate. Therefore, diagnostic tools that facilitate a better understanding of heat shock damage and the defense mechanism at the sub-cellular level are of great importance. In this report, we have demonstrated the use of a pyridinium-based fluorescent molecule, PM-ER-OH, as a 'multichannel' imaging probe to monitor the pH change associated with a heat shock in the endoplasmic reticulum. Among the three pyridinium derivatives synthesized, PM-ER-OH was chosen for study due to its excellent biocompatibility, good localization in the endoplasmic reticulum, and intracellular pH response signaled by a yellow fluorescence (λ _max = 556 nm) at acidic pH and a far red fluorescence (λ _max = 660 nm) at basic pH. By changing the excitation wavelength, we could modulate the fluorescence signal in 'turn-ON', single excitation ratiometric and 'turn-OFF' modes, making the fluorophore a 'multichannel' probe for both ex vitro and in vitro pH monitoring in the endoplasmic reticulum. The probe could efficiently monitor the pH change when heat shock was applied to cells either directly or in a pre-heated manner, which gives insight on cellular acidification caused by heat stress.

SUBMITTER: Chakraborty S

PROVIDER: S-EPMC11253182 | biostudies-literature | 2024 Jul

REPOSITORIES: biostudies-literature

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pH-Assisted multichannel heat shock monitoring in the endoplasmic reticulum with a pyridinium fluorophore.

Chakraborty Sandip S Bindra Anivind Kaur AK Thomas Anagha A Zhao Yanli Y Ajayaghosh Ayyappanpillai A

Chemical science 20240604 28

Heat shock is a global health concern as it causes permanent damage to living cells and has a relatively high mortality rate. Therefore, diagnostic tools that facilitate a better understanding of heat shock damage and the defense mechanism at the sub-cellular level are of great importance. In this report, we have demonstrated the use of a pyridinium-based fluorescent molecule, PM-ER-OH, as a 'multichannel' imaging probe to monitor the pH change associated with a heat shock in the endoplasmic ret ...[more]

PMID: 39027278

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Project description:BackgroundAlthough prostate cancer responds initially to androgen ablation therapies, progression to castration-resistant prostate cancer (CRPC) frequently occurs. Heat shock protein (Hsp) 90 inhibition is a rational therapeutic strategy for CRPC that targets key proteins such as androgen receptor (AR) and protein kinase B (Akt); however, most Hsp90 inhibitors trigger elevation of stress proteins like Hsp27 that confer tumor cell survival and treatment resistance.ObjectiveWe hypothesized that cotargeting the cytoprotective chaperone Hsp27 and Hsp90 would amplify endoplasmic reticulum (ER) stress and treatment-induced cell death in cancer.Design, setting, and participantsInducible and constitutive Hsp27 and other HSPs were measured by real-time reverse transcription-polymerase chain reaction and immunoblot assays. The combinations of OGX-427 with Hsp90 inhibitors were evaluated in vitro for LNCaP cell growth and apoptosis and in vivo in CRPC LNCaP xenograft models.Outcome measurements and statistical analysisTumor volumes were compared using the Kruskal-Wallis test. Overall survival was analyzed using Kaplan-Meier curves, and statistical significance was assessed with the log-rank test.Results and limitationsHsp90 inhibitors induced expression of HSPs in tumor cells and tissues in a dose- and time-dependent manner; in particular, Hsp27 mRNA and protein levels increased threefold. In vitro, OGX-427 synergistically enhanced Hsp90 inhibitor-induced suppression of cell growth and induced apoptosis by 60% as measured by increased sub-G1 fraction and poly(ADP-ribose) polymerase cleavage. These biologic events were accompanied by decreased expression of HSPs, Akt, AR, and prostate-specific antigen, and induction of ER stress markers (cleaved activating transcription factor 6, glucose-regulated protein 78, and DNA-damage-inducible transcript 3). In vivo, OGX-427 potentiated the anticancer effects of Hsp90 inhibitor PF-04929113 (orally, 25mg/kg) to inhibit tumor growth and prolong survival in CRPC LNCaP xenografts.ConclusionsHSP90 inhibitor-mediated induction of Hsp27 expression can be attenuated by OGX-427, resulting in increased ER stress and apoptosis, and synergistic inhibition of CRPC tumor growth.Patient summaryThis study supports the development of targeted strategies using OGX-427 in combination with Hsp90 inhibitors to improve patient outcome in CRPC.

Dataset Information

PH-Assisted multichannel heat shock monitoring in the endoplasmic reticulum with a pyridinium fluorophore.

Publications

pH-Assisted multichannel heat shock monitoring in the endoplasmic reticulum with a pyridinium fluorophore.

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