Cytotoxicity testing is one of the standard biocompatibility tests for nearly all medical devices irrespective of their duration and nature of patient contact per ISO 10993-1:2018 and the U.S. FDA’s Guidance on the Use of ISO 10993-1 (2020). Although cytotoxicity testing is useful as a screening tool, it is not a definitive indicator of device safety since in vitro tests do not fully represent the complex in vivo environment of the human body.
Consequently, cytotoxicity failures (i.e., positive test results) should be evaluated with other relevant biocompatibility endpoints to determine device safety. In the following sections, we explore risk management strategies for medical device cytotoxicity failures, focusing on two case studies demonstrating the importance of a comprehensive approach to evaluating device safety. The approaches described below relied upon data from material reviews, chemical characterization and toxicological risk assessment, and other biocompatibility endpoint data per ISO 10993-1:2018.
Case Study 1: Limited Contact – Surface Device, Intact Skin
In the first case study, an intact skin-contacting medical device with limited duration (less than five minutes in clinical use) was tested for biocompatibility in compliance with ISO 10993-1:2018. This included the MEM elution cytotoxicity assay, the Guinea Pig Maximization assay and the Intracutaneous Irritation test.
The MEM elution cytotoxicity assay failed with grade “4” (severe) cytotoxicity at all three observed time points (24, 48 and 72 hours). The Guinea Pig Maximization test and the Intracutaneous Irritation test produced passing results with no sensitization or irritation noted in normal saline or sesame oil extracts. Chemical characterization and toxicological risk assessment were not conducted as they would not be standard recommendations for a limited-duration device with intact skin contact.
The materials used in the device’s construction were reviewed to identify potentially cytotoxic compounds, and polychloroprene was identified as a potential cause for cytotoxicity failure. Polychloroprene (i.e., a synthetic black rubber) was one of the materials used to construct the device. It is also used as a positive control for the MEM elution cytotoxicity assay and thus is expected to be associated with cytotoxicity failure. Because a likely cause for the cytotoxicity failure was identified—and the sensitization and irritation results were favorable—the toxicological risk was considered acceptable for the nature and duration of patient contact (i.e., < 5 min with intact skin).
Case Study 2: Limited Contact – Externally Communicating Device, Blood Path, Indirect
In the second case study, an externally communicating indirect blood path medical device with limited duration exposure (up to eight hours in clinical use) was tested using a battery of biocompatibility testing. This testing included the MEM elution cytotoxicity assay, the Guinea Pig Maximization assay, the Intracutaneous Irritation test, Acute Systemic Toxicity and Hemolysis – Extract Method.
This device showed mild but passing (grade 2/4) cytotoxicity at 24 hours, increasing to a failing level (moderate; grade 3/4) for cytotoxicity at 48 and 72 hours. The device passed the sensitization and irritation testing in normal saline and sesame oil extracts. The device also demonstrated no signs of acute systemic toxicity or hemolysis using the extract method.
Chemical characterization was conducted, and toxicological risk was evaluated based on the extractables results. Several citrates, acrylates, methacrylates and metals were identified in the extractables, all of which are known to have cytotoxic potential. However, the Margin of Safety (MOS) values were greater than 10 for all the potentially cytotoxic compounds identified above. Thus, the toxicological risk was considered acceptable.
Based on the information obtained from the chemical characterization data, toxicological risk assessment, and other relevant biocompatibility test results, the toxicological risk was considered acceptable for the nature and duration of patient contact (i.e., up to 8 hours with indirect blood contact).
When Cytotoxicity Failure Happens
Every medical device requires a biological evaluation, the specifics of which are outlined in ISO 10993-1:2018 and the U.S. FDA’s guidance on ISO 10993-1: 2020, to identify potential risks resulting from the product’s materials and manufacturing processes.
Cytotoxicity testing is a primary screening biocompatibility assay required for nearly all medical devices. According to Section 10 of ISO 10993-5:2009, “cytotoxicity data shall be assessed in relation to other biocompatibility data and the intended use of the product.” Hence, cytotoxicity failures (i.e., a positive test result) should be evaluated with all other relevant biological endpoints to determine device safety. The following flowchart outlines how WuXi AppTec toxicologists determine the most appropriate steps to take when the potential for cytotoxicity is identified.
A final word
A thorough understanding of the materials used in the device’s construction and the manufacturing processes are critical aspects of medical device development and safety evaluation, as these can significantly impact cytotoxicity test results. Certain materials, such as metals, alloys, rubber, adhesives, antibiotics, anticoagulants, ethylene oxide residuals, lubricants and plasticizers, are associated with potential cytotoxicity failures. Hence, understanding the properties and potential risks of the various materials and processes is essential for interpreting cytotoxicity assay results. Cytotoxicity results are part of the screening process for the evaluation of medical devices but should not be considered a definitive indicator of device safety. Instead, a comprehensive risk management approach that considers all relevant data, which may include chemical characterization, toxicological risk assessment, other biocompatibility testing, materials/process review and the intended clinical use of the device, should be employed to determine the overall safety of the device.