April 2002 : Cisplatin-Induced Ototoxicity: Cell-Biological Aspects of Hair Cell Degeneration

1.  Introduction

        Cisplatin (cis-diamminedichloroplatinum (II); PtCl2(NH3)2) is one of the most potent cytotoxic drugs currently available for cancer chemotherapy, and is especially effective in the treatment of advanced and metastatic forms of solid tumours such as testicular, ovarian and head-and-neck carcinomas. Its clinical efficacy, however, is limited by severe side effects, which include renal injury, peripheral neuropathies, hearing impairment, nausea and vomiting, visual impairment, and myelosuppression. Of these, nephrotoxicity, peripheral neurotoxicity and ototoxicity are potentially the major dose-limiting factors, in that they are cumulative and in general only partially reversible with discontinuation of therapy. Several attempts have been made to reduce the toxic side effects of cisplatin. Although vigorous pre- and post-hydration and mannitol-induced diuresis are now included routinely in cisplatin chemotherapy, these techniques have proven to be only partially successful, since renal failure still occurs, especially after repeated administration of cisplatin. It has also been attempted to reduce the toxic side effects of cisplatin by concomitant administration of so-called rescue agents (usually sulfur-containing ligands; cf., Reedijk and Teuben, 1999), but many of these compounds seem not only to reduce the nephrotoxicity and ototoxicity but the antitumour effect of cisplatin as well. Cisplatin-induced ototoxicity in humans is generally manifested as tinnitus and sensorineural hearing loss. This hearing impairment is dose-related, cumulative, bilateral, usually permanent and is initially characterized by a high-frequency deficit, but in patients receiving repeated doses of cisplatin progressively extends toward frequencies that are important for speech perception.

2. Histological changes

        Histologically, the most prominent change seen in the cochlea after chronic administration of cisplatin is degeneration of the organ of Corti, consisting of loss of the outer hair cells (OHCs) and the inner hair cells (IHCs) as well as a disturbance of the organ of Corti's typical microarchitecture. Hair cell loss follows a specific topographical pattern similar to that observed after chronic aminoglycoside administration or acoustic overstimulation. Initially, OHC loss is present predominantly in the basal cochlear turn, with the first row of OHCs being more vulnerable than the other OHC rows and the IHCs. In more severely damaged cochleas loss of both OHCs and IHCs is found along the entire basilar membrane. These findings correlate with the permanent, frequency-dependent elevation in the auditory thresholds and with the irreversible suppression of both the cochlear microphonics and compound action potential. Post-treatment degeneration of the organ of Corti, frequently observed during aminoglycoside-induced ototoxicity, has never been observed in cisplatin-intoxicated cochleas. In fact, recent studies indicate that functional and morphological recovery of the guinea-pig organ of Corti may be possible after cessation of cisplatin administration (Stengs et al., 1997; Cardinaal et al., 2000a), suggesting that an intrinsic regenerative mechanism is present in the organ of Corti. This finding may have clinical implications, especially since recovery of cisplatin-induced hearing loss has been reported to occur occasionally in patients. Equally important is the finding that concomitant administration of the neuroprotective ACTH(4-9) analogue (melanocortin) ORG 2766, which has been proven not to interfere with the antitumour effect of cisplatin, significantly reduces cisplatin-induced loss of OHCs (Smoorenburg et al., 1999; Cardinaal et al., 2000b). However, these phenomenological findings require more insight into the cellular mechanism(s) of cisplatin ototoxicity in order to further develop the protection and recovery potentials.

3. The Mechanisms of Ototoxicity

         Although the ototoxic effect of cisplatin has been extensively studied during the past decades, the cellular mechanism(s) by which cisplatin induces loss of OHCs and subsequent degeneration of the organ of Corti remain elusive. Furthermore, it is unclear if the degenerative changes in the organ of Corti and the functional and morphological changes in the stria vascularis in cisplatin-intoxicated cochleas occur by the same mechanism(s). This is mainly due to the fact that the cellular sites of cisplatin uptake and accumulation in the cochlea have not been properly identified. Although [195mPt]-labelled cisplatin could be detected in homogenated samples of the organ of Corti and the stria vascularis (Schweitzer, 1993), ultrastructural X-ray microanalysis studies have failed to localize cisplatin in OHCs (Maruyama et al., 1993; Saito and Aran, 1994; Welb, 1995). Electrophysiological studies have demonstrated that cisplatin interferes with the mechano-electric transduction process, either directly by blocking the transduction channels in the apical membranes of the OHCs or indirectly by a blockage of the voltage-sensitive calcium-channels in the basolateral membranes of the OHCs. However, one must bear in mind that most of these data have been obtained with in vitro models and may not resemble actual physiological alterations in the intact cochlea. In the absence of proof, we will assume that the cellular mechanism(s) underlying cisplatin-induced ototoxicity, which involves damage to terminally-differentiated, post-mitotic OHCs, are identical to the toxic action of cisplatin in tumour cells, which are more susceptible for damage by cytotoxic agents than non-proliferating cells.

         It is generally thought that the free (dichloro) form of cisplatin is passively transported across the plasma membrane of both normal and tumour cells. The lower chloride concentration of the cytoplasm and the lower cytosolic pH favour aquation of the dichloro form and successive deprotonation, which yields positively charged aquated species of the drug. These reactive species are potent electrophilics that will react with the nucleophilic groups of nucleic acids (DNA and RNA) and the sulfhydryl moieties of peptides (e.g., glutathione), proteins (e.g., metallothionein) as well as other cellular macromolecules (for a review, see Reed et al., 1996). It is commonly accepted that binding of cisplatin to DNA results in the formation of cisplatin-DNA adducts, which produce severe local distortions in the DNA double helix, and that this so-called DNA-platination is an essential first step in the cytotoxic action of cisplatin. The cellular consequences of cisplatin-induced DNA damage and the mechanism by which these adducts cause cell death, however, are less well-understood.

         One potentially important way by which cisplatin-DNA adducts may kill cells is by the induction of programmed cell death or apoptosis. Despite a large volume of literature describing the ability of cisplatin to induce apoptosis in various tumour cells and the recent observation that the ototoxic aminoglycoside antibiotic amikacin induces apoptosis in OHCs in the neonatal rat cochlea (Vago et al., 1998), reports presenting evidence for cisplatin-induced apoptosis of OHCs in the adult mammalian cochlea remain sparse (Alam et al., 2000). Moreover, in most ultrastructural studies, morphological features typical of apoptosis, such as nuclear condensation and segmentation, cell fragmentation and apoptotic bodies, were never observed in the organ of Corti of cisplatin-intoxicated cochleas. However, in cisplatin-intoxicated kidneys, and also in dorsal root ganglia, high levels of cisplatin-DNA adducts and cisplatin-induced apoptosis have been demonstrated. Since most nephrotoxic drugs usually exhibit ototoxic activity and nephroprotective agents also ameliorate cisplatin ototoxicity, it cannot be excluded that cisplatin induces OHC loss by apoptotic cell death subsequently to the formation of cisplatin-DNA adducts.

         Although genomic DNA is generally accepted as the critical cellular target of cisplatin, there is evidence that other cellular targets may also be involved, such mitochondrial DNA, cytosolic proteins, cell membrane phospholipids and cytoskeletal proteins, and hence, alternative cellular mechanisms might be involved in cisplatin-induced OHC loss. X-ray microanalysis studies of cisplatin-intoxicated kidneys have demonstrated that cisplatin is internalized through an endocytotic process and is accumulated in lysosomes in the proximal tubular epithelial cells (Makita et al., 1982; Berry et al., 1985), and it has been suggested that subsequent release of the lysosomal contents into the cytoplasm provokes cellular necrosis. In addition, a role of glutathione has been implicated. Alterations in the activity of anti-oxidant enzymes and depletion of cochlear glutathione levels have been demonstrated after cisplatin administration and this suggests a role for reactive oxygen species in cisplatin-induced ototoxicity (Rybak and Somani, 1999). Depletion of glutathione levels are associated with the generation of reactive oxygen species, and these can cause not only DNA strand breaks but also lipid peroxidation and aberrant expression of membrane-bound enzymes, which can alter the cell's ability to maintain ionic gradients. Most experimental data indicate that the ototoxic effect of cisplatin is a multi-target phenomenon, involving not only the organ of Corti but also in the stria vascularis, both at the physiological and morphological level (Schweitzer, 1993). Cisplatin administration results in a marked decrease in the endochlear potential, which is generated in the stria vascularis (Klis et al., 2000; Tsukasaki et al., 2000), but this decrease is not permanent, contrary to the elevated auditory thresholds. Histologically, cisplatin-induced strial changes, which consist of blebbing of the marginal cells, intermediate cell atrophy and strial oedema, are dose-related and resemble those observed after administration of loop diuretics. More evidence for an effect on the stria vascularis derives specifically from the finding that cisplatin inhibits strial adenylate cyclase activity, which is present predominantly in the basolateral membranes of the marginal cells and seems to be involved in electrolyte and solute transport in the stria vascularis. This is supported by the observation that both sodium and potassium concentrations in the endolymph are increased after cisplatin administration, probably due to changed passive transport of solutes. Furthermore, recent studies suggest that cisplatin also may affect the auditory neurons (Zheng and Gao, 1996; Gabaizadeh et al., 1997) and the spiral ganglion cells (Cardinaal et al., 2000b). If, and how, OHC loss and subsequent degeneration of the organ of Corti are related to the strial changes and the changes in the spiral ganglion is as yet unknown. A precise localization of the cellular site(s) of uptake and accumulation of cisplatin in the cochlea as well as proper identification of the cellular mechanisms involved in drug-induced OHC degeneration may shed more light onto this issue and this will be helpful in the development and implementation of more sophisticated treatment protocols to prevent the ototoxic side effects of cisplatin.

4. References

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  • Berry, J.P., Brille, P., LeRoy, A.F., Gouveia, Y., Ribaud, P., Galle, P. and Mathé, G. (1985) Experimental ultrastructural and X-ray microanalysis study of cisplatin in the rat: Intracellular localization of platinum. Cancer Treatment Rep. 66: 1529-1533.

  • Cardinaal, R.M., De Groot, J.C.M.J., Huizing, E.H., Veldman, J.E. and Smoorenburg, G.F. (2000b) Dose-dependent effect of 8-day cisplatin administration upon the morphology of the albino guinea pig cochlea. Hearing Res. 144: 135-146.

  • Cardinaal, R.M., De Groot, J.C.M.J., Huizing, E.H., Veldman, J.E. and Smoorenburg, G.F. (2000a) Cisplatin-induced ototoxicity: Morphological evidence of spontaneous outer hair cell recovery in albino guinea pigs? Hearing Res. 144: 147-156

  • Gabazaideh, R., Staecker, H., Liu, W., Kopke, R., Malgrange, B., Lefebvre, P.P. and Van de Water, T.R. (19970 Protection of both auditory hair cells and auditory neurons from cisplatin induced damage. Acta Otolaryngol. (Stockh.) 117: 232-238.

  • Klis, S.F.L., O'Leary, S.O., Hamers, F.P.T., De Groot, J.C.M.J. and Smoorenburg, G.F. (2000) reversible cisplatin ototoxicity in the albino guinea pig. NeuroReport 11: 623-626.

  • Makita, T., Hakoi, K. and Ohokawa, T. (1982) X-ray microanalysis and elctron microscopy of platinum complex in the epithelium of proximal renal tubules of the cisplatin-administered rabbit. Cell Biol. Int. Rep. 10: 447-454.

  • Maruyama, T., Furuya, N. and Daimon, T. (1993) Distribution of platinum in the inner ear of guinea pigs treated with cisplatin. J. Otolaryngol. Jpn. 96: 1758-1759.

  • Reed, E., Dabholkar, M. and Chabner, B.A. (1996) Platinum analogues. In: Chabner, B.A. and Longo, D.L. (Eds.) Cancer Chemotherapy and Biotherapy, 2nd Edition. Lippincott-raven Publishers, Philadelphia, pp. 357-378.

  • Reedijk, J. and Teuben, J.M. (1999) Platinum-sulfur interactions involved in antitumour drugs, rescue agents and biomolecules. In: Lippert, B. (Ed.) Cisplatin. Chemistry and Biochemistry of A Leading Anticancer Drug. Wiley-VCH, Weinheim, pp. 339-362.

  • Rybak, L.P. and Somani, S. (1999) Ototoxicity. Amelioration by protective agents. Ann. NY Acad. Sci. 884: 143-151.

  • Saito, T. and Aran, J.-M. (1994) X-ray microanalysis and ion microscopy of guinea pig cochlea and kidney after cisplatin treatment. ORL 56: 310-314.

  • Schweitzer, V.G. (1993) Cisplatin-induced ototoxicity: Effect of pigmentation and inhibitory agents. Laryngoscope Suppl. 59.

  • Smoorenburg, G.F., De Groot, J.C.M.J., Hamers, F.P.T. and Klis, S.F.L. (1999) Protection and spontaneous recovery from cisplatin-induced hearing loss. Ann. NY Acad. Sci. 884:192-210

  • Stengs, C.H.M., Klis, S.F.L., Huizing, E.H. and Smoorenburg, G.F. (1997) Cisplatin-induced oto-oxicity: Electrophysiological evidence of spontaneous recovery in the albino guinea pig. Hearing Res. 111: 103-113.

  • Tsukasaki, N., Whitworth, C.A. and Rybak, L.P. (2000) Acute changes in cochlear potentials due to cisplatin. Hearing Res. 149: 189-198.

  • Vago, P., Humert, G. and Lenoir, M. (1998) Amikacin intoxication induces apoptosis and cell proliferation in rat organ of Corti. NeuroReport 9: 431-436.

  • Welb, R. (1995) Experimentelle Studie über die Wirkung von Cisplatin auf das Innenohr. Ph.D. Thesis, Heinrich-Heine-Universität, Düsseldorf, Germany.

  • Zheng, J.L. and Gao, W.Q. (1996) Differential damage to auditory neurons and hair cells by ototoxins and neuroprotection by specific neurotrophins in rat cochlear organotypic cultures.Eur. J. Neurosci. 8: 1897-1905.

5. Contributing Author

John De Groot, Ph.D.
Hearing Research Laboratories (Histology Unit), Department of Otorhinolaryngology,
University Medical Center Utrecht, Room G.02.531,
P.O. Box 85.500, NL-3508 GA Utrecht,
The Netherlands