08.31.10
New Products and Regulations Test the Testers
Changes in biocompatibility testing affect genotoxicity, bone replacement materials and other areas.
After remaining relatively stable for an extended period, the past year or two have witnessed major changes in the field of biocompatibility testing. Perhaps the greatest changes have been in genotoxicity, where the orthopedic industry has had to digest a guidance document from the U.S. Food and Drug Administration (FDA) that contradicts current standards set by the International Organization for Standardization (ISO), requiring different testing regimens for the United States and Europe.
Another area in ferment is the testing of fillers used in bone replacement therapy. The FDA now requires a higher level of proof from orthopedic device manufacturers claiming osteoconductivity or osteoinductivity. The more frequent use of antimicrobial coatings on implants also has affected biocompatibility testing by requiring additional testing to demonstrate the efficacy of the coatings.
Genotoxicity Testing in Flux
“The last year in biocompatibility testing has been the most dynamic since I entered the business,” said Thor Rollins, section leader for Nelson Labs, a Salt Lake City, Utah-based provider of life-cycle microbiology testing services to manufacturers in the medical device, pharmaceutical and nutraceutical industries. “The genotoxicity area has seen the most changes, many of them driven by the guidance document issued by the FDA in December 2008 that significantly departs from the ISO 10993-3 genotoxicity standard.”
The ISO standard offers two options. The first ISO option is to perform the Ames test, mouse lymphoma and a third chromosome aberration test that eliminates the need for colony sizing. The second option is the Ames test and mouse lymphoma plus colony sizing. The chromosome aberration test is quite expensive so the majority of companies have been sizing colonies instead when testing to the ISO standards. The Ames test is a biological assay to assess the mutagenic potential of chemical compounds. A positive test indicates that the chemical might act as a carcinogen.
“The FDA guidance reshuffled the deck by saying that colony sizing is no longer sufficient in the U.S. market,” Rollins said. “Instead the FDA wants the Ames test plus either mouse lymphoma or chromosome aberration plus a further test, mouse micronucleus. The mouse micronucleus test is a significant addition because it is an animal test while up to now only in vitro testing was required. This guidance covers Class II and III devices, so it applies to all orthopedic implants.”
In the mouse micronucleus test, mice are exposed to either the test article or an extract. The animals are harvested for bone marrow which is evaluated for the presence of micronuclei. Micronuclei are composed of chromosomes or fragments of chromosomes that provide an indicator of chromosomal damage.
“Meanwhile, Europe has been moving in a different direction,” Rollins added. “I recently returned from the June 2010 ISO meetings in Berlin where research was presented by European regulatory agencies to the effect that an in vitro mouse micronucleus test is essentially equivalent to the in vivo version of the test while being less expensive and faster to perform. It appears that European and possibly Japanese regulatory authorities are ready to accept the in vitro mouse nucleus test or at least should be accepting the in vitro test in the near future. Once enough information is presented to the FDA they might change but I don’t think it will happen soon.”
The research recently presented by the European regulatory authorities confirms earlier work, such as a study by David Kirkland, Marilyn Aardema, Leigh Henderson and Lutz Müller.1 Their research showed that a battery of three in vitro genotoxity tests— Ames plus mouse lymphoma assay plus in vitro micronucleus or chromosome aberrations test was able to discriminate rodent carcinogens and non-carcinogens. Only 19 carcinogens out of 206 tested gave consistently negative tests in the full three-test battery. Most of these 19 are either carcinogens via a non-genotoxic mechanism considered not necessarily relevant for humans or are extremely weak carcinogens.
The chromosome aberration test is another hot topic in the genotoxicity field.
“Here, Japan has taken the lead by requiring an exaggerated extraction process that uses organic solvents such as methanol or acetone to generate a residue,” Rollins explained. “The basic idea is to pull more leachables from the implant or other medical device so a test lasting a few weeks becomes more representative of 30 years of presence in the body. This approach will not have a significant impact on titanium and stainless steel but may generate significantly different results on polymers and biodegradable materials. It’s important to note that the FDA is taking a close look at this method so don’t be surprised to see it become mandatory in the future.”
Even the venerable cytoxicity test, which has been a staple for at least 20 years, is undergoing changes. The traditional approach is to soak the sample in media to extract material and put the media on L929 mouse fiberblast cells to see how they react. The test is fast, cheap and accurate. The big concern with this test always has been that it requires the technician to make a judgment call on a 0-to-4 scale, with 0 being nontoxic and 4 being highly toxic. The concern is that different labs and different technicians will tend to give different scores to the same sample.
In 2009, the ISO adopted a new version of the test that uses either the MTT or the XTT assay to provide machine-readable results. A stain is applied to the sample that healthy cells bring inside the cell walls, while sick cells do not. The cells are washed so that the stain remaining outside the cell walls is removed. The number of healthy cells then can be measured using a spectrophotometer that provides an objective standard that is not subject to human interpretation.
“Since both machine-readable and the technician-scored test are now being used, the question arises of how to correlate results from the two different testing methods,” Rollins said. “In theory, the FDA accepts the new test. But a lot of our clients are still using the old test because the FDA reviewers are more comfortable with it. In some cases I have seen FDA reviewers ask that the old test and new test be performed side by side so they could see the results of both. This is not a major burden because the old test is quite inexpensive. Germany, on the other hand, has been accepting the new test for years and reviewers are comfortable with the results. In most cases, European and Asian regulators will accept either the old or new test but I have seen cases where the old test has been turned down.”
Bone Replacement Therapy Testing Trends
Another area of biocompatibility testing that recently has seen major changes involves testing of materials used in bone replacement therapy. These materials are designed to temporarily fill gaps in bone and support the formation of permanent, new bone growth. Bone replacement materials have to comply with the same testing guidelines as permanent implants. What’s new in testing these materials is that many companies have entered the market with materials that support or stimulate new bone growth.
Specifically, osteoconductivity refers to material in which the graft supports the attachment of new osteoblasts and osteoprogenitor cells that form an interconnected structure that supports the growth of new bone. On the other hand, osteoinductivity refers to materials that go one step further by inducing non-differentiated stem cells or osteoprogenitor cells to differentiate into osteoblasts. A very common use of these materials is in combination with hardware in spinal fusion operations.
“The FDA is requiring a higher level of proof from companies claiming osteoconductivity or osteoinductivity,” said Dr. Joseph Carraway, director of toxicology for NAMSA, a medical device testing laboratory and contract research organization based in Northwood, Ohio, that specializes in the safety evaluation of medical devices. “Osteoconductivity or osteoinductivity must be demonstrated in an animal model and the FDA is looking for more quantitative end points. In the past, plain 2-D film radiographs were sufficient but the new gold standard is microcomputed tomography (micro-CT) that provides a 3-D image analysis of the site. Micro-CT delivers a more precise assessment, for example, that 60 percent of the volume in the site is new bone, 40 percent of which was due to natural growth and 20 percent of which was stimulated by the bone replacement material.”
The FDA also is placing greater emphasis on histomorphometry, which provides a microscopic view of the implant site and distinguishes between different types of cells as an endpoint in evaluating bone replacement materials. Staining and labeling techniques are used to make new bone show up as a different color from old bone. Administering short cycles of tetracycline to the animal allows multiple bone formation fronts to fluoresce under the right conditions. This is due to the ability of tetracyclines to incorporate at the border of the osteoid, which is actively mineralized. Histomorphometry can be used to assess a large number of parameters including total bone volume and bone produced in a given time period. The FDA reshuffled the deck in late July in animal studies by issuing “Guidance for Industry and FDA Staff: General Considerations for Animal Studies for Cardiovascular Devices.” While the guidance document does not apply strictly to orthopedic devices, the recommendations from the document can be expected to start being applied to animal studies used in orthopedic devices
almost immediately.
Carraway summarizes the impact of the document on biocompatibility testing for orthopedic devices as follows: “The FDA is asking for more robust studies with more quantitative data. Of course, this guidance document is just another milestone in a long trend of reviewers asking for more scientific information to make their decisions. We are seeing questions coming back from reviewers on almost a weekly basis. The recent guidance document recommends that studies include a negative control, such as an animal without any bone replacement material. The FDA is also asking device companies when they submit a 510(k) study to include a predicate in their comparison in order to prove the equivalence of their device to the device that has already been approved.”
New Tech Impacts Testing Methods
Anja Friedrich, head of marketing and responsible for quality control and biocompatibility at BSL Bioservice Scientific Laboratories GmbH, said that increased use of antimicrobial coatings on medical devices is one of the main trends that she is seeing in the European market. BSL Bioservice is an international contract research organization located in Munich, Germany. These products require a special panel of biological safety tests.
“It’s no secret that about 1.5 percent of implant patients suffer complications due to infections.” Friedrich said. “The aim is to reduce those complications by coating the implants with an antibiotic, often gentamycin. Gentamycin is usually given orally to patients after an operation but the coating provides a much higher concentration at the site of the implant. So the best way to prevent infections is to combine treatment with an oral and a local antibiotic.”
In testing coated implants, it’s necessary to perform the usual array of biocompatibility testing plus an assay to determine the antimicrobial effect such as the minimum inhibitory concentration tests to demonstrate the effectiveness of the coating in preventing infections. These tests either can be performed on agar plates showing an inhibition zone to the bacterial growth or as a broth dilution test. It’s important to note that the antimicrobial substance causes growth inhibition effects in biocompatibility testing in the test for cytotoxicity. Therefore, all the results of biocompatibility have to be reviewed thoroughly and evaluated by an expert.
Biodegradable implants also present special biocompatibility testing requirements.
“The advantage of a biodegradable implant is that the degradation of the material in some cases can eliminate the need for a subsequent procedure such as removing metal nails used to set a bone fracture,” Friedrich said.“What’s different here is that in addition to the usual array of biocompatibility tests it may be necessary to perform a toxicokinetic study in an in vivo test model in combination with physico-chemical tests. This requires a chemical study of the degradation products and testing of a series of frequently taken blood samples as well as macroscopical and histopathological examination of the target tissues. The aim of these studies is to achieve information about what happens with the degradation products themselves as well as their possible effects on the body.”
Friedrich added that verification of cleaning and sterilization procedures applied on the surgical instruments used for implantation is another area where the company is seeing increased demand and further development for testing services.
“Several hospitals in Germany had to temporarily stop doing implantation due to faults in their cleaning and sterilization procedure, which led to instruments contaminated with tissue and blood residuals,” Friedrich said. “We have been working with the manufacturers of the instruments and with the hospitals and performed validation testing to determine if the reprocessing procedure is sufficiently effective and suitable for the device tested. Standards such as AAMI TIR 31 describe methods to validate the cleaning step by contaminating instruments and evaluating cleanliness with biochemical tests after the cleaning step. In addition, prior to sterilization with moist heat, instruments are inoculated with specially resistant bacterial spores at the critical sites by representing the worst-case conditions for the sterilizing agent to penetrate. A bacterial growth test after the sterilization detects if devices are really sterile.”
Following the validated reprocessing procedures will avoid these severe hygienic problems in future.
In-House or Outsource Testing?
A question faced by most orthopedic device manufacturers is whether to perform testing in-house or to outsource to an independent laboratory.
“We are seeing a trend towards outsourcing testing to contract research organizations,” said James Rancourt, Ph.D., the founder and CEO of Polymer Solutions Inc. (PSI) in Blacksburg, Va. PSI offers cGMP testing services on large or small projects. “Testing requires highly skilled people and expensive equipment. The testing workload of a typical orthopedic device manufacturer alternates between short periods with very high levels of activity and long periods with not much happening. Many device manufacturers find that it’s too expensive to maintain a top-flight testing capability over these peaks and valleys. On the other hand, it’s much easier for an independent testing laboratory to maintain a high level of biocompatibility testing capabilities because its capabilities are utilized and paid for by multiple device manufacturers.”
Rancort added that demands placed on orthopedic device manufacturers by the FDA greatly have increased in the last five years.
“You need to know everything about everything and anything that you aren’t sure about is a big negative,” he said. “A third-party laboratory is typically better equipped to stay up with the developments in methods and technology that are continually affecting biocompatibility testing. Working for a multiplicity of companies on many different products provides a cross-pollination effect that leads to the proliferation of best practices.”
Athena Spencer, gas chromatography aboratory manager for Polymer Solutions said that “it goes without saying” that a trusted third-party laboratory often has more credibility than an in-house testing organization.
“An independent testing lab is less likely to be questioned as to whether undue pressure might have influenced the results,” Spencer noted. “Even device manufacturers that perform testing in-house frequently transfer their methods to an independent lab to make sure that they come up with the same results.”
Cynthia Rancourt, PSI’s director of business operations and quality manager, told Orthopedic Design & Technology that her firm is seeing a “big demand” from clients for assistance in validating test methods as part of their efforts to comply with the FDA’s “Quality by Design” initiative.
“Quality by Design requires thinking about biocompatibility from the very beginning of the product development process,” she explained. “For example, suppose you are machining an implant and have three different choices of machining oils to use. From a traditional point of view, biocompatibility should not play a role in the decision because the oil will be cleaned off the part later. Still, it would be wise to use a biocompatible fluid from the beginning because it will increase the robustness of your manufacturing process. The FDA is now requiring that device manufacturers use risk management throughout the entire lifecycle of the product. These guidelines don’t specifically state, but do imply, that testing needs to be increased at the early stages of the design process. Design decisions throughout the product development process need to be documented and testing is often the best way to document these decisions.”
Manufacturer & Supplier Cooperation
Meredith May, vice president of engineering for Atlas Spine, pointed out that one way that orthopedic device manufacturers can reduce their testing burden is to work closely with materials suppliers. Jupiter, Fla.-based Atlas Spine’s mission is to create spinal implant and instrument systems that combine the highest level of performance with simplicity of use to enhance patient outcomes.
“If you are utilizing a new material in your device then you need to perform all of the tests specified in ISO 10993,” May said. “On the other hand, if you use an existing material that the manufacturer has already submitted to the FDA then the process can be streamlined. Instead of the completed 10993 biocompatibility panel, we submit cytotoxicity results and a reference to the material master file.”
Atlas Spine frequently uses polyether-etherketone (PEEK), a hard radiolucent plastic from Invibio that is used in conjunction with carbon fiber reinforcement or as pure PEEK. Laboratory studies during the 1990s confirmed that PEEK implants had the requisite combination of strength, wear, creep and fatigue resistance to replace metallic biomaterials for spine implants. The PEEK master file contains a range of studies, such as one performed by C. H. Rivard, S. Rhalmi and C. Coillard.2 These researchers studied a spinal implant system comprised of metal and PEEK polymer to correct scoliosis deformity without spine fusion. The in vivo study was designed to duplicate a worst case wear scenario and evaluated the biological response of the spinal chord and nerve roots to PEEK polymer particles. The study concluded that PEEK polymer was found to be harmless to the spinal cord and could safely be used as a component in spinal implant systems.
“If we get questions about the material, we contact Invibio [a leading manufacturer of PEEK] and they help us answer them,” May added. “We frequently use third-party laboratories such as Nelson Labs for biocompatibility and Empirical Testing Corporation for biomechanical testing. The ISO 10993-1 standard, ‘Guidance on the Selection of Tests,’ considers cytotoxicity tests to be so important that they should be performed on every orthopedic device regardless of the previous testing that may have been performed on the material used in the device. Besides their critical importance in submissions to regulatory agencies, we also find that cytotoxicity test methods are quite beneficial for troubleshooting and evaluating potential improvements in manufacturing processes.”
***
The field of biocompatibility testing is undergoing a period of substantial change. It is highly likely that the current dynamic atmosphere will persist into the foreseeable future as regulatory authorities continue to require ever-expanding data submissions and new device technologies demand new testing methods.
References:
1. David Kirkland, Marilyn Aardema, Leigh Henderson and Lutz Müller. “Evaluaton of the Ability of a battery of three in vitro genotoxicity tests to discriminate rodent carcinogens and non-carcinogens.” Mutation
Research 584 (2005) 1-256.
2. C. H. Rivard, S. Rhalmi and C. Coillard. “In vivo biocompatibility testing of PEEK polymer for a spinal implant system: a study in rabbits.” Journal of Biomedical Material
Research. 2002;62(4):488-498.
Jerry Fireman is based in Lexington, Mass., and is president of Structured Information, a company that produces articles and other technical documents for publications and companies in the medical device field as well as a wide range of other technologies. Hehas been writing since 1984 and has published more than 9,000 articles.
Changes in biocompatibility testing affect genotoxicity, bone replacement materials and other areas.
After remaining relatively stable for an extended period, the past year or two have witnessed major changes in the field of biocompatibility testing. Perhaps the greatest changes have been in genotoxicity, where the orthopedic industry has had to digest a guidance document from the U.S. Food and Drug Administration (FDA) that contradicts current standards set by the International Organization for Standardization (ISO), requiring different testing regimens for the United States and Europe.
Compression testing biocompatible polymer components. Photo courtesy of Polymer Solutions. |
Another area in ferment is the testing of fillers used in bone replacement therapy. The FDA now requires a higher level of proof from orthopedic device manufacturers claiming osteoconductivity or osteoinductivity. The more frequent use of antimicrobial coatings on implants also has affected biocompatibility testing by requiring additional testing to demonstrate the efficacy of the coatings.
Genotoxicity Testing in Flux
“The last year in biocompatibility testing has been the most dynamic since I entered the business,” said Thor Rollins, section leader for Nelson Labs, a Salt Lake City, Utah-based provider of life-cycle microbiology testing services to manufacturers in the medical device, pharmaceutical and nutraceutical industries. “The genotoxicity area has seen the most changes, many of them driven by the guidance document issued by the FDA in December 2008 that significantly departs from the ISO 10993-3 genotoxicity standard.”
The ISO standard offers two options. The first ISO option is to perform the Ames test, mouse lymphoma and a third chromosome aberration test that eliminates the need for colony sizing. The second option is the Ames test and mouse lymphoma plus colony sizing. The chromosome aberration test is quite expensive so the majority of companies have been sizing colonies instead when testing to the ISO standards. The Ames test is a biological assay to assess the mutagenic potential of chemical compounds. A positive test indicates that the chemical might act as a carcinogen.
“The FDA guidance reshuffled the deck by saying that colony sizing is no longer sufficient in the U.S. market,” Rollins said. “Instead the FDA wants the Ames test plus either mouse lymphoma or chromosome aberration plus a further test, mouse micronucleus. The mouse micronucleus test is a significant addition because it is an animal test while up to now only in vitro testing was required. This guidance covers Class II and III devices, so it applies to all orthopedic implants.”
In the mouse micronucleus test, mice are exposed to either the test article or an extract. The animals are harvested for bone marrow which is evaluated for the presence of micronuclei. Micronuclei are composed of chromosomes or fragments of chromosomes that provide an indicator of chromosomal damage.
“Meanwhile, Europe has been moving in a different direction,” Rollins added. “I recently returned from the June 2010 ISO meetings in Berlin where research was presented by European regulatory agencies to the effect that an in vitro mouse micronucleus test is essentially equivalent to the in vivo version of the test while being less expensive and faster to perform. It appears that European and possibly Japanese regulatory authorities are ready to accept the in vitro mouse nucleus test or at least should be accepting the in vitro test in the near future. Once enough information is presented to the FDA they might change but I don’t think it will happen soon.”
The research recently presented by the European regulatory authorities confirms earlier work, such as a study by David Kirkland, Marilyn Aardema, Leigh Henderson and Lutz Müller.1 Their research showed that a battery of three in vitro genotoxity tests— Ames plus mouse lymphoma assay plus in vitro micronucleus or chromosome aberrations test was able to discriminate rodent carcinogens and non-carcinogens. Only 19 carcinogens out of 206 tested gave consistently negative tests in the full three-test battery. Most of these 19 are either carcinogens via a non-genotoxic mechanism considered not necessarily relevant for humans or are extremely weak carcinogens.
The chromosome aberration test is another hot topic in the genotoxicity field.
“Here, Japan has taken the lead by requiring an exaggerated extraction process that uses organic solvents such as methanol or acetone to generate a residue,” Rollins explained. “The basic idea is to pull more leachables from the implant or other medical device so a test lasting a few weeks becomes more representative of 30 years of presence in the body. This approach will not have a significant impact on titanium and stainless steel but may generate significantly different results on polymers and biodegradable materials. It’s important to note that the FDA is taking a close look at this method so don’t be surprised to see it become mandatory in the future.”
Even the venerable cytoxicity test, which has been a staple for at least 20 years, is undergoing changes. The traditional approach is to soak the sample in media to extract material and put the media on L929 mouse fiberblast cells to see how they react. The test is fast, cheap and accurate. The big concern with this test always has been that it requires the technician to make a judgment call on a 0-to-4 scale, with 0 being nontoxic and 4 being highly toxic. The concern is that different labs and different technicians will tend to give different scores to the same sample.
In 2009, the ISO adopted a new version of the test that uses either the MTT or the XTT assay to provide machine-readable results. A stain is applied to the sample that healthy cells bring inside the cell walls, while sick cells do not. The cells are washed so that the stain remaining outside the cell walls is removed. The number of healthy cells then can be measured using a spectrophotometer that provides an objective standard that is not subject to human interpretation.
“Since both machine-readable and the technician-scored test are now being used, the question arises of how to correlate results from the two different testing methods,” Rollins said. “In theory, the FDA accepts the new test. But a lot of our clients are still using the old test because the FDA reviewers are more comfortable with it. In some cases I have seen FDA reviewers ask that the old test and new test be performed side by side so they could see the results of both. This is not a major burden because the old test is quite inexpensive. Germany, on the other hand, has been accepting the new test for years and reviewers are comfortable with the results. In most cases, European and Asian regulators will accept either the old or new test but I have seen cases where the old test has been turned down.”
Bone Replacement Therapy Testing Trends
Another area of biocompatibility testing that recently has seen major changes involves testing of materials used in bone replacement therapy. These materials are designed to temporarily fill gaps in bone and support the formation of permanent, new bone growth. Bone replacement materials have to comply with the same testing guidelines as permanent implants. What’s new in testing these materials is that many companies have entered the market with materials that support or stimulate new bone growth.
Specifically, osteoconductivity refers to material in which the graft supports the attachment of new osteoblasts and osteoprogenitor cells that form an interconnected structure that supports the growth of new bone. On the other hand, osteoinductivity refers to materials that go one step further by inducing non-differentiated stem cells or osteoprogenitor cells to differentiate into osteoblasts. A very common use of these materials is in combination with hardware in spinal fusion operations.
“The FDA is requiring a higher level of proof from companies claiming osteoconductivity or osteoinductivity,” said Dr. Joseph Carraway, director of toxicology for NAMSA, a medical device testing laboratory and contract research organization based in Northwood, Ohio, that specializes in the safety evaluation of medical devices. “Osteoconductivity or osteoinductivity must be demonstrated in an animal model and the FDA is looking for more quantitative end points. In the past, plain 2-D film radiographs were sufficient but the new gold standard is microcomputed tomography (micro-CT) that provides a 3-D image analysis of the site. Micro-CT delivers a more precise assessment, for example, that 60 percent of the volume in the site is new bone, 40 percent of which was due to natural growth and 20 percent of which was stimulated by the bone replacement material.”
The FDA also is placing greater emphasis on histomorphometry, which provides a microscopic view of the implant site and distinguishes between different types of cells as an endpoint in evaluating bone replacement materials. Staining and labeling techniques are used to make new bone show up as a different color from old bone. Administering short cycles of tetracycline to the animal allows multiple bone formation fronts to fluoresce under the right conditions. This is due to the ability of tetracyclines to incorporate at the border of the osteoid, which is actively mineralized. Histomorphometry can be used to assess a large number of parameters including total bone volume and bone produced in a given time period. The FDA reshuffled the deck in late July in animal studies by issuing “Guidance for Industry and FDA Staff: General Considerations for Animal Studies for Cardiovascular Devices.” While the guidance document does not apply strictly to orthopedic devices, the recommendations from the document can be expected to start being applied to animal studies used in orthopedic devices
almost immediately.
Carraway summarizes the impact of the document on biocompatibility testing for orthopedic devices as follows: “The FDA is asking for more robust studies with more quantitative data. Of course, this guidance document is just another milestone in a long trend of reviewers asking for more scientific information to make their decisions. We are seeing questions coming back from reviewers on almost a weekly basis. The recent guidance document recommends that studies include a negative control, such as an animal without any bone replacement material. The FDA is also asking device companies when they submit a 510(k) study to include a predicate in their comparison in order to prove the equivalence of their device to the device that has already been approved.”
New Tech Impacts Testing Methods
Anja Friedrich, head of marketing and responsible for quality control and biocompatibility at BSL Bioservice Scientific Laboratories GmbH, said that increased use of antimicrobial coatings on medical devices is one of the main trends that she is seeing in the European market. BSL Bioservice is an international contract research organization located in Munich, Germany. These products require a special panel of biological safety tests.
“It’s no secret that about 1.5 percent of implant patients suffer complications due to infections.” Friedrich said. “The aim is to reduce those complications by coating the implants with an antibiotic, often gentamycin. Gentamycin is usually given orally to patients after an operation but the coating provides a much higher concentration at the site of the implant. So the best way to prevent infections is to combine treatment with an oral and a local antibiotic.”
In testing coated implants, it’s necessary to perform the usual array of biocompatibility testing plus an assay to determine the antimicrobial effect such as the minimum inhibitory concentration tests to demonstrate the effectiveness of the coating in preventing infections. These tests either can be performed on agar plates showing an inhibition zone to the bacterial growth or as a broth dilution test. It’s important to note that the antimicrobial substance causes growth inhibition effects in biocompatibility testing in the test for cytotoxicity. Therefore, all the results of biocompatibility have to be reviewed thoroughly and evaluated by an expert.
Biodegradable implants also present special biocompatibility testing requirements.
“The advantage of a biodegradable implant is that the degradation of the material in some cases can eliminate the need for a subsequent procedure such as removing metal nails used to set a bone fracture,” Friedrich said.“What’s different here is that in addition to the usual array of biocompatibility tests it may be necessary to perform a toxicokinetic study in an in vivo test model in combination with physico-chemical tests. This requires a chemical study of the degradation products and testing of a series of frequently taken blood samples as well as macroscopical and histopathological examination of the target tissues. The aim of these studies is to achieve information about what happens with the degradation products themselves as well as their possible effects on the body.”
Friedrich added that verification of cleaning and sterilization procedures applied on the surgical instruments used for implantation is another area where the company is seeing increased demand and further development for testing services.
“Several hospitals in Germany had to temporarily stop doing implantation due to faults in their cleaning and sterilization procedure, which led to instruments contaminated with tissue and blood residuals,” Friedrich said. “We have been working with the manufacturers of the instruments and with the hospitals and performed validation testing to determine if the reprocessing procedure is sufficiently effective and suitable for the device tested. Standards such as AAMI TIR 31 describe methods to validate the cleaning step by contaminating instruments and evaluating cleanliness with biochemical tests after the cleaning step. In addition, prior to sterilization with moist heat, instruments are inoculated with specially resistant bacterial spores at the critical sites by representing the worst-case conditions for the sterilizing agent to penetrate. A bacterial growth test after the sterilization detects if devices are really sterile.”
Following the validated reprocessing procedures will avoid these severe hygienic problems in future.
In-House or Outsource Testing?
A question faced by most orthopedic device manufacturers is whether to perform testing in-house or to outsource to an independent laboratory.
“We are seeing a trend towards outsourcing testing to contract research organizations,” said James Rancourt, Ph.D., the founder and CEO of Polymer Solutions Inc. (PSI) in Blacksburg, Va. PSI offers cGMP testing services on large or small projects. “Testing requires highly skilled people and expensive equipment. The testing workload of a typical orthopedic device manufacturer alternates between short periods with very high levels of activity and long periods with not much happening. Many device manufacturers find that it’s too expensive to maintain a top-flight testing capability over these peaks and valleys. On the other hand, it’s much easier for an independent testing laboratory to maintain a high level of biocompatibility testing capabilities because its capabilities are utilized and paid for by multiple device manufacturers.”
Rancort added that demands placed on orthopedic device manufacturers by the FDA greatly have increased in the last five years.
“You need to know everything about everything and anything that you aren’t sure about is a big negative,” he said. “A third-party laboratory is typically better equipped to stay up with the developments in methods and technology that are continually affecting biocompatibility testing. Working for a multiplicity of companies on many different products provides a cross-pollination effect that leads to the proliferation of best practices.”
Nelson Labs scientist scores cells during a chromosomal aberration test. Photo courtesy of Nelson Labs. |
Athena Spencer, gas chromatography aboratory manager for Polymer Solutions said that “it goes without saying” that a trusted third-party laboratory often has more credibility than an in-house testing organization.
“An independent testing lab is less likely to be questioned as to whether undue pressure might have influenced the results,” Spencer noted. “Even device manufacturers that perform testing in-house frequently transfer their methods to an independent lab to make sure that they come up with the same results.”
Cynthia Rancourt, PSI’s director of business operations and quality manager, told Orthopedic Design & Technology that her firm is seeing a “big demand” from clients for assistance in validating test methods as part of their efforts to comply with the FDA’s “Quality by Design” initiative.
“Quality by Design requires thinking about biocompatibility from the very beginning of the product development process,” she explained. “For example, suppose you are machining an implant and have three different choices of machining oils to use. From a traditional point of view, biocompatibility should not play a role in the decision because the oil will be cleaned off the part later. Still, it would be wise to use a biocompatible fluid from the beginning because it will increase the robustness of your manufacturing process. The FDA is now requiring that device manufacturers use risk management throughout the entire lifecycle of the product. These guidelines don’t specifically state, but do imply, that testing needs to be increased at the early stages of the design process. Design decisions throughout the product development process need to be documented and testing is often the best way to document these decisions.”
Manufacturer & Supplier Cooperation
Meredith May, vice president of engineering for Atlas Spine, pointed out that one way that orthopedic device manufacturers can reduce their testing burden is to work closely with materials suppliers. Jupiter, Fla.-based Atlas Spine’s mission is to create spinal implant and instrument systems that combine the highest level of performance with simplicity of use to enhance patient outcomes.
“If you are utilizing a new material in your device then you need to perform all of the tests specified in ISO 10993,” May said. “On the other hand, if you use an existing material that the manufacturer has already submitted to the FDA then the process can be streamlined. Instead of the completed 10993 biocompatibility panel, we submit cytotoxicity results and a reference to the material master file.”
Atlas Spine frequently uses polyether-etherketone (PEEK), a hard radiolucent plastic from Invibio that is used in conjunction with carbon fiber reinforcement or as pure PEEK. Laboratory studies during the 1990s confirmed that PEEK implants had the requisite combination of strength, wear, creep and fatigue resistance to replace metallic biomaterials for spine implants. The PEEK master file contains a range of studies, such as one performed by C. H. Rivard, S. Rhalmi and C. Coillard.2 These researchers studied a spinal implant system comprised of metal and PEEK polymer to correct scoliosis deformity without spine fusion. The in vivo study was designed to duplicate a worst case wear scenario and evaluated the biological response of the spinal chord and nerve roots to PEEK polymer particles. The study concluded that PEEK polymer was found to be harmless to the spinal cord and could safely be used as a component in spinal implant systems.
“If we get questions about the material, we contact Invibio [a leading manufacturer of PEEK] and they help us answer them,” May added. “We frequently use third-party laboratories such as Nelson Labs for biocompatibility and Empirical Testing Corporation for biomechanical testing. The ISO 10993-1 standard, ‘Guidance on the Selection of Tests,’ considers cytotoxicity tests to be so important that they should be performed on every orthopedic device regardless of the previous testing that may have been performed on the material used in the device. Besides their critical importance in submissions to regulatory agencies, we also find that cytotoxicity test methods are quite beneficial for troubleshooting and evaluating potential improvements in manufacturing processes.”
***
The field of biocompatibility testing is undergoing a period of substantial change. It is highly likely that the current dynamic atmosphere will persist into the foreseeable future as regulatory authorities continue to require ever-expanding data submissions and new device technologies demand new testing methods.
References:
1. David Kirkland, Marilyn Aardema, Leigh Henderson and Lutz Müller. “Evaluaton of the Ability of a battery of three in vitro genotoxicity tests to discriminate rodent carcinogens and non-carcinogens.” Mutation
Research 584 (2005) 1-256.
2. C. H. Rivard, S. Rhalmi and C. Coillard. “In vivo biocompatibility testing of PEEK polymer for a spinal implant system: a study in rabbits.” Journal of Biomedical Material
Research. 2002;62(4):488-498.
Jerry Fireman is based in Lexington, Mass., and is president of Structured Information, a company that produces articles and other technical documents for publications and companies in the medical device field as well as a wide range of other technologies. Hehas been writing since 1984 and has published more than 9,000 articles.