MPI Research has more than 15 years of experience in auditory safety testing, and continues to perform these studies on a regular basis. Our industry recognized team of experts conducts all procedures in accordance with the Food and Drug Administration’s guidance document for nonclinical safety evaluation of reformulated drug products and products intended for administration by an alternate route.

Over the last five years, MPI Research scientists have integrated all of these specialized functional and histopathological evaluations (ABR electrophysiology, middle ear assessment, screening approach to middle ear assessment and ototoxicity, definitive histopathological evaluations of the cochlea, cytocochleogram analyses, as well as evaluations to characterize vestibular functions) within one state-of-the-art facility. There is no commercial laboratory presently demonstrating a comparable breadth and depth of experience, including continuous Study Director engagement, in this unique area of drug safety research.

  • When developing a drug that is administered into the middle ear, is expected to come in contact with the middle or inner ear tissues, or new drug entities from a pharmacological class that has demonstrated ototoxicity following systemic administration (e.g., aminoglycoside antibiotics, sildenafil), functional auditory testing and relevant histopathology should be evaluated to assess possible ototoxicity.
  • Our industry recognized team of experts conducts all procedures in accordance with the Food and Drug Administration’s guidance document for nonclinical safety evaluation of reformulated drug products and products intended for administration by an alternate route.
  • These assessments provide data to support the safety of drugs intended for otic use. MPI Research collaborates with Sponsors and regulatory experts in developing the best program to determine if the drug is ready for further clinical evaluation and otic use in humans.
  • Guinea Pigs
  • Rats
  • Cats
  • Mice

The primary animal model for auditory drug safety characterization is the guinea pig.  Drug sensitivity and anatomical similarity to humans make the guinea pig most suitable for this type of specialized testing.  The use of alternative small animal species may be justified in  some cases.  There is a large body of literature supporting the use of mice and rats in auditory research.  When a large animal model is needed to support drug safety characterization, felines are the best option due to their less restrictive anatomical features and low incidence of middle and inner ear dysfunction.

Options for Study Design

The definitive data in an ototoxicity study is generated by correlating the auditory brainstem response with otic microscopy, including specific cytocochleogram analysis. As the assessments for middle ear histology and cytocochleogram each require one intact set of auditory structures, integrating these assessments into the study design can be problematic. To enable assessment of both of these post-life endpoints, and to maintain statistical power, there are two approaches to designing an ototoxicity study which requires direct middle ear exposure.

The first option is to use direct bilateral administration to the middle ears of each animal. This can be accomplished via transtympanic administration or via middle ear catheters implanted into both bullae. Using this technique, one ear may be used for middle ear histopathology and the other for the cytocochleogram analysis. This approach reduces the number of animals required for study and may allow for better correlation of middle ear histopathological and inner ear cytocochleogram analysis by reducing the inter-animal differences in drug distribution and metabolism.

The second option is to use direct administration to one ear (e.g., unilaterally) of each animal. This can also be accomplished via transtympanic administration or via middle ear catheters implanted into one bulla of each animal. To allow evaluation of both the cytocochleogram and the histopathology of relevant otic structures using this study design, twice as many animals are required for the assessment. Although most studies use one of the above alternative procedures for direct administration to the middle ear, in addition to this specialized route, systemic administration is a less complicated option relevant to some drug classes.

Endpoints

Auditory Brainstem Response Evaluation

Hearing thresholds are determined by repetitive recordings of the auditory brain stem responses, (ABR). The ABR procedure is conducted by evaluating the global neuronal activity within the auditory pathway from cochlea to cortex following stimulus presentations at specified frequencies and amplitudes. Determination of the response engendered in the auditory pathway at each frequency is used to interpolate the stimulus characteristics evoking the minimum hearing threshold. As this evaluation employs a quantitative EEG waveform to determine auditory thresholds, it is an ideal assessment which reduces behavioral variability and potential interpretational issues. Our test-retest data and interrater reliability with expert electrophysiologists in the field demonstrate our high level of expertise in auditory brainstem response data interpretation.

MPI Research has conducted multiple studies to validate our ABR methods for sensitivity, reliability, and predictive validity in several relevant species. The following graphs are representative data from these studies.

Cytocochleogram

In addition to otic micrsocopy, cytocochleograms are required on auditory safety studies to support or refute any potential test article related ABR changes. Cytocochleograms are graphical illustrations of the number of missing auditory hair cells over the length of the cochlea. Drug-induced hair cell toxicity produces permanent hearing loss making the cytocochleogram a critical endpoint on auditory safety studies.

Otic Microscopy

MPI Research characterizes middle ear toxicity by targeting those major structures associated with amplification/conduction of the sound pressure waves representing auditory stimuli. The structures evaluated are as follows: tympanum, middle ear ossicles (malleus, incus, stapes), tensor tympani, round window, cochlea, and the middle ear mucosa. If required, longitudinal sections of the Eustachian tube may also be included when possible. These procedures may be modified to include additional structures, as necessary.

Drug and Noise Induced Hearing Loss Models

Our test-retest data and interrater reliability, with expert electrophysiologists, in the field demonstrate our high level of expertise in auditory brainstem response data interpretation.

Establishing drug efficacy can only be achieved once you have a validated experimental model.  MPI Research offers several systemic and intratympanic drug-induced hearing loss models using aminoglycosides, diuretics, and platinum based chemotherapeutics.

32kHz-auditory-threshold

ABR threshold data in guinea pigs following intratympanic administration of gentamicin and saline.

auditory-threshold-line-graph

Guinea pig ABR data presented over time following a single 122 dB exposure to 8 kHz octave band noise for 2 hours.

Noise-induced hearing loss models require specialized noise chambers, speakers, and stimulus generators.  MPI Research offers a qualified method for noise induced hearing loss in both mice and guinea pigs.

auditory-threshold-line-graph2

auditory-missing-hair-cells

Average ABR and paired cytocochleogram data from mice following a single 118 dB exposure to an 8–16 kHz bandwidth noise for two hours.

Inner Ear Immunohistochemistry (IHC)

MPI Research offers multiple IHC markers for hair cell and synapse identification.  Our standard technique for inner and outer hair cell identification uses a phalloidin based marker which stains all F-actin in cells allowing for visualization of stereocilia and hair cell body both within the organ of Corti and vestibular epithelium of the utricle and saccule (available in all species).  Additionally, we offer a myosin Viia antibody marker which allows for the visualization of both inner and outer hair cell bodies (available in mouse only).

Characterizing the number of neuron synaptic connections to a regenerated or traumatized hair cell can elucidate and support functional audiometry data. Using antibody markers for presynaptic and postsynaptic markers, MPI Research offers a quantitative approach to synaptic contacts on inner hair cells.