Is Photodynamic Therapy Resistance a Special Case of Photobiomodulation?

Abstract

Red-to-near infrared photobiomodulation (R-NIR-PBM) is a growing field in photobiology, with (as of the date of writing) 606 citations in PubMed, and a total of 123 in the first 5 months of 2018 alone. Many biological processes are said to be affected by PBM, from improved cellular metabolism, antiapoptotic effects, and improvements in disease states.^1,2 While many mechanisms have been proposed, the greatest consensus seems to revolve around improved cellular energetics caused by an improvement in the kinetics of cytochrome c oxidase (CCO), the terminal enzyme in the electron transfer chain of the mitochondria.³ In particular, relief of CCO inhibition by NO is seen as a likely candidate.⁴

However, appropriate basic science studies, particularly involving isolated, purified CCO, have been sorely lacking, a deficiency we pointed out in an earlier PMLS editorial in 2011.⁵ In that editorial, we laid out a reasonable path forward and called for an increased focus on basic science.

Having worked some length along this path, current results now appear to contradict this hypothesis, as R-NIR, in our hands, does not show effects on cytochrome c oxidation,⁶ oxygen consumption, or NO inhibition of CCO (unpublished results) using isolated CCO. It seems that R-NIR, while undoubtedly showing observable biological effects in complex systems, must be employing other means than simple kinetic enhancement of CCO. These means may or may not ultimately affect CCO activity or expression, but a simple model where the metal centers in CCO act as photoreceptors, with enhancement of kinetic activity and relief of NO-induced CCO inhibition, appears to be simplistic. The basic molecular mechanism (or mechanisms) still remains unknown.

Despite our inability so far to demonstrate a PBM effect on isolated CCO, PBM stills appears to be a real phenomenon, as too many studies show positive results for this therapeutic modality to be a phantom. Something is happening, and it may take physiological conditions for it to happen. In a study on ischemia-reperfusion cardioprotection, R-NIR had no effect on infarct size in isolated Langendorff hearts using artificial blood substitute, but significantly reduced infarct size in an in vivo rat model.⁷ Similarly, CCO activity enhancement has been reported in cell and mitochondrial concentrates,^8,9 but not in isolated CCO.⁶ PBM therefore may be a more complicated phenomenon than our original hypothesis posits, illustrating the need for a broader framework in which to understand PBM.

Photodynamic therapy (PDT) resistance may offer us this broader framework. PDT requires a photosensitizer (PS) and, typically, light of similar wavelengths to many PBM studies. The consequences of R-NIR irradiation are in opposition to PBM, as cell destruction in PDT results from PS activation. However, PDT resistance involving cell survival develops upon exposure to NIR. This is usually ascribed to signaling effects of NO due to upregulation of inducible nitric oxide synthase (iNOS).^10
–12 Endogenous low flux NO can significantly reduce PDT effectiveness in mouse tumor models. Use of iNOS inhibitors and NO scavengers showed that iNOS and NO played a key role in cellular resistance to apoptosis. The results are similar to those ascribed to PBM: cell survival, antiapoptosis, and antioxidant activity. These effects are counter-productive for killing tumor cells in PDT, but would prove beneficial in PBM.

This leads to the following questions: do the observed antiapoptotic effects of PBM work through these same signaling pathways as seen in PDT resistance, and not through improved cellular energy metabolism? Is PDT resistance perhaps part of the same phenomenon as PBM? Is PDT resistance therefore a special case of PBM? Is the cellular target of R-NIR just another PS, but endogenous? Perhaps hemes or other metal centers, or other nitrosyl compounds, are endogenous “low-level photosensitizers” for PBM-induced NO release or generation, analogous to exogenous PSs in PDT.

In this scenario, the importance of antiapoptotic signaling in PBM would be in proportion to the significance of apoptosis in the disease state, thus more effect should be seen after cellular insults or in degenerative conditions. Consequently, there should be less chance of observable beneficial effects of PBM in “life-style enhancing” modes (such as cognitive enhancement in nondisease states).

We must acknowledge the differences in R-NIR dosages between PDT and PBM. Dosages in malignant brain tumor PDT clinical trials¹³ range from 27 to 250 mJ/cm², with the most promising results at the higher end.^14,15 PBM studies typically require lower fluences, from 45 mJ/cm² for an in vivo rat model to as low as 4 mJ/cm² for cellular work.¹⁶ Whether the higher fluences used in PDT allow the cells to access mechanisms not available under lower fluences is something that would need to be considered, and investigated. Still, the possibility that these two phenomena, PBM and PDT resistance, are connected, could provide us with a much-needed framework to potentially understand the mechanisms underlying PBM.

In addition to R-NIR-stimulated iNOS activity, other sources of NO could also be considered. In a study of hypoxia and reoxygenation injury in cardiomyocytes, it was found that R-NIR conferred protection in a manner dependent on NO derived from both NOS and non-NOS sources.¹⁷ R-NIR can also release NO from nitrosyl hemoglobin and myoglobin, and this released NO can enhance the cardioprotective effects of nitrite.¹⁸ Nitrite is also implicated as a source of NO in a novel nitric oxide synthase activity identified for CCO, with nitrite taking the place of oxygen as the electron acceptor, forming NO.^19,20 Perhaps we have come full circle, back to CCO as the cellular target for R-NIR energy in PBM! Another potential source of NO could be S-nitrosothiols and dinitrosyl iron complexes, which have been shown in solution to release NO upon irradiation with R-NIR.²¹

Whatever the ultimate source of NO, the NO signaling antiapoptotic pathways identified in PDT resistance mechanisms¹⁰ could also play a role in PBM. Perhaps we should consider them together under a more general heading such as “light activated cell survival strategies”?

Footnotes

Acknowledgments

This work was supported, in part, by The Bleser Endowed Chair of Neurology and The Chad Baumann Neurology Research Endowment, to Harry T. Whelan, MD.

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