1、Designation: F 1439 03Standard Guide forPerformance of Lifetime Bioassay for the TumorigenicPotential of Implant Materials1This standard is issued under the fixed designation F 1439; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revision,
2、the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon (e) indicates an editorial change since the last revision or reapproval.1. Scope1.1 This guide is intended to assist the biomaterials testinglaboratory in the conduct and evaluation of tumo
3、rigenicity teststo evaluate the potential for new materials to evoke a neoplasticresponse. The procedure is generally reserved only for thosematerials which have not previously been used for humanimplantation for a significant period of time.1.2 Assessment of tumorigenicity is one of several proce-d
4、ures employed in determining the biological response to amaterial as recommended in Practice F 748. It is assumed thatthe investigator has already determined that this type of testingis necessary for a particular material before consulting thisguide. The recommendations of Practice F 748 should beco
5、nsidered before a study is commenced.1.3 Whenever possible, it is recommended that a battery ofgenotoxicity procedures be initiated and proposed as an alter-native to an in-vivo tumorigenicity bioassay. Genotoxicityassays may also be considered as initial screening proceduresdue to the sensitivity o
6、f the assays, the significant reduction intime to gain valuable data, and the desire to reduce the use ofanimals for testing. Genotoxicity assays that may be consid-ered are outlined in Guides E 1262, E 1263, E 1280, andE 2186, and Practices E 1397 and E 1398. Additionally, othergenotoxicity testing
7、 which might be considered (but which donot yet have ASTM test methods) include Salmonella/Mammalian-Microsomal Plate Incorporation Mutagenicity As-say, In Vivo Cytogenetics Bone Marrow Chromosomal Dam-age Assay, BALB/3T3 Morphological Transformation ofMouse Embryo Cells, and the Mouse Micronucleus
8、Assay. Theinvestigator is advised to consider carefully the appropriatenessof a particular method for his application after a review of thepublished literature.1.4 This standard does not purport to address all of thesafety concerns, if any, associated with its use. It is theresponsibility of the use
9、r of this standard to establish appro-priate safety and health practices and determine the applica-bility of regulatory limitations prior to use.2. Referenced Documents2.1 ASTM Standards:2E 1262 Guide for the Performance of the Chinese HamsterOvary Cell/Hypoxanthine Guanine Phosphoribosyl Trans-fera
10、se Gene Mutation AssayE 1263 Guide for Conduct of Micronucleus Assays inMammalian Bone Marrow ErythrocytesE 1280 Guide for Performing the Mouse Lymphoma Assayfor Mammalian Cell MutagenicityE 1397 Practices for the in vitro Rat Hepatocyte DNARepair AssayE 1398 Practices for the in vivo Rat Hepatocyte
11、 DNARepair AssayE 2186 Guide for Determining DNA Single-Strand Damagein Eukaryotic Cells Using the Comet AssayF 748 Practice for Selecting Generic Biological Test Meth-ods for Materials and Devices2.2 Other Documents:National Toxicology Program General Statement of Workfor the Conduct of Toxicity an
12、d Carcinogenicity Studies inLaboratory Animals3OECD Guidelines for Testing of Chemicals: Guideline 451,Carcinogenicity Studies4OECD Guidelines for Testing of Chemicals: Guideline 453,Combined Chronic Toxicity/Carcinogenicity Studies4Good Laboratory Practice for Nonclinical Laboratory Stud-ies53. Ter
13、minology3.1 Definitions of Terms Specific to this Standard:3.1.1 carcinogenica substance is considered to be carci-nogenic if it can be shown to be causally related to an increasedincidence of malignant neoplastic formation.1This guide is under the jurisdiction of ASTM Committee F04 on Medical andSu
14、rgical Materials and Devices and is the direct responsibility of SubcommitteeF04.16 on Biocompatibility Test Methods.Current edition approved Nov. 1, 2003. Published December 2003. Originallyapproved in 1992. Last previous edition approved in 2002 as F 1439 02.2For referenced ASTM standards, visit t
15、he ASTM website, www.astm.org, orcontact ASTM Customer Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.3Available from National Institute of Environmental Health Sciences, ResearchTriangle Park, NC, Augu
16、st 1988.4Available from Organization for Economic Cooperation and Development, 200L St., NW, Suite 650, Washington, DC 200364922.5Available from 21 CFR, Part 58, U.S. Government Printing Office, Superin-tendent of Documents, 732 N. Capitol St., NW, Mail Stop: SDE, Washington, DC20401.1Copyright ASTM
17、 International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.3.1.2 maximum implantable dosethe maximum weight orvolume of the test article which can be reasonably implantedinto the test site taking into account the gross distention oftissue which can occur and
18、its possible effects on test results.3.1.3 mutagenica substance is said to be mutagenic if itinduces alterations in the genetic code of the cell.3.1.4 tumorigenica substance is said to be tumorigenic ifit can be shown to be causally related to an increased incidenceof neoplastic formation whether ma
19、lignant or benign.4. Significance and Use4.1 This guide is not intended to specify the exact method ofconducting a test for any particular material but only to presentsome of the criteria that should be considered in method designand possible problems that could lead to misleading results. Inthe dev
20、elopment of the actual test protocol, it is recommendedthat recognized tumorigenesis bioassay procedures be con-sulted.4.2 The recommendations given in this guide may not beappropriate for all applications or types of implant materials.These recommendations should be utilized by experiencedtesting p
21、ersonnel in conjunction with other pertinent informa-tion and the requirements of the specific material application.5. Choice of Animal Model5.1 These types of bioassays for chemical substances havetraditionally been performed in mice or rats, or both, becauseof their small size, relative cost facto
22、rs, and lifespan. For thetesting of biomaterials, mice are not recommended because thesmall animal size is not conducive to the placement of solidimplants. The investigator should seriously consider the use ofone of the traditional models in order to draw upon theextensive information available abou
23、t typical tumor formationrates and sites in control animals. The National ToxicologyProgram3recommends the use of Fischer 344 (F344/N) rats.However, other readily available species and strains may alsobe acceptable for the performance of these studies. Other ratspecies which have been recommended in
24、clude Sprague-Dawley, Long-Evans, and Wistar. Some investigators haverecommended the use of Long-Evans or Wistar Rats because ofthe difficulty of achieving a two-year lifespan for Fischer andSprague-Dawley rats.5.2 The currently accepted level of testing in a particularsite of implantation or medica
25、l specialty should be carefullyresearched and regulatory requirements determined before astudy design is finalized to ensure acceptability of the finalresults.5.3 The appropriate choice of male or female animals or acombination should be carefully considered in light of theparticular material and ap
26、plication being investigated. If thedevice will ultimately be used only in the male or female, onlyone sex may need to be evaluated. Otherwise, both sexesshould be used.5.4 The decision to use other species for study should becarefully documented in terms of a clear need. The use ofspecies which hav
27、e not previously been used may reduce theamount of comparative data available on control animals.Typical tumor rates for hamsters, rats, and mice have beentabulated and are available in Refs. (1, 2, 3).66. Selection of Size and Form of Implant6.1 Tumorigenicity bioassays have traditionally been per-
28、formed using chemical substances as the challenge. Theevaluation of implant materials requires that solid material beimplanted in some form. It is important to realize that thedown-sized implants necessary for use in animals will have agreater surface area to volume ratio, and this difference must b
29、econsidered in experimental design.6.2 It may be important to determine the site of administra-tion of the test material that is most appropriate to the end usebefore determining implant size. The site of implantationshould be the paravertebral muscle unless the size of theimplant causes this site t
30、o be unacceptable. Alternatively, thesite of implantation should mimic the anticipated end use, ifpossible. Where a specific material may be utilized in morethan one type of device, multiple sites of administration shouldbe considered if different types of tissue will be contacted. (Forinstance, mat
31、erials that may be in contact with bone orimplanted into internal organ tissue might be tested in bothtissues.)6.3 It should be recognized that the response of the testanimal to an extract of a material may not fully represent theresponse that might be seen if the material itself were to beimplanted
32、. In general, an extract should not be used as asubstitute for the actual material of interest.6.4 The physical form of the test material should be repre-sentative of that intended for use in human patients and shouldconsider potential material debris, if appropriate. The investi-gator should be awa
33、re that tests have shown (4) that powderedpolymeric materials may not elicit a tumorigenic responsesubcutaneously even when prepared from polymers that doinduce tumors when implanted in the form of a film. Theimpact of physical form and surface properties on tumorigen-esis must be carefully consider
34、ed, in making decisions aboutthe physical form of the implants (5, 6, 7, 8, 9, 10).6.5 Researchers have found that the aspect ratio (length/diameter) of fiber materials may play a role in the tumorigen-esis of a particular material (11, 12). When new fibrousmaterials are being tested, the actual fib
35、er length to beanticipated in practice should be studied. If fragmentation canbe anticipated or is a worse case possibility, an attempt shouldbe made to document a clinically relevant fiber length.6.6 The material to be tested should originate fromsample(s) representative of all processing including
36、 surfacefinishing, passivation, and sterilization or other final processingthat will occur to a finished device.6.7 Dosage:6.7.1 In most materials, the ratio between the surface area ofthe implant and the body weight of the animal or person willhave an effect on the amount of extractable substances
37、(if any)which leach out of the material. The total weight or volume ofmaterial used in each animal should be in excess of the6The boldface numbers in parentheses refer to the list of references at the end ofthis guide.F1439032anticipated dosages to be seen in clinical practice whencalculated based u
38、pon the ratio of surface area of sample tobody weight of the animal. Consideration should be given tousing the maximum implantable dose as the dosage or as oneof multiple dosage levels. For the special case of degradablematerials, the sample size should be calculated based on theratio of sample weig
39、ht to animal body weight.6.7.2 Whenever possible, more than one exposure levelshould be considered to evaluate a dose-response effect.7. Choice of Control7.1 Control groups for this type of study will usually consistof identical animals that have not received an implant of thetest material but have
40、been subjected to the remainder of thesurgical procedures. Additional groups such as housing (ani-mals which receive no treatment but are housed with the testanimals) and reference control groups may be included in thestudy design.7.2 The investigator should consider a negative controlgroup in addit
41、ion to the sham or untreated controls. Theseanimals would receive an implant or treatment identical to thetest animals but the implant would be manufactured from aselected negative reference material. This group would thenserve to isolate any results due to the implant trauma ormechanically induced
42、changes.8. Size of Test Groups8.1 The test group and the control group should eachcontain enough animals which will be scheduled to survive tothe end of the study to allow statistically valid conclusions tobe drawn from the study. If both male and female animals arebeing used, each group should cont
43、ain an equal number ofanimals of each sex. The National Toxicology Program3requires 60 animals/sex/group for chemical studies with tenanimals being sacrificed earlier than two years. Other interna-tional organizations recommend 50 animals/sex/group.4Theinvestigator should ascertain that the number o
44、f animals ineach group is adequate for statistical and regulatory purposesbefore proceeding. In order to ensure valid data analysis, theanimals should be randomly assigned to control and experi-mental groups. Considerations specific to the particular implantapplication or medical specialty may manda
45、te a greater numberof animals in each group. Additional animals in interimsacrifice groups or satellite groups may be added.8.2 The number of test animals in each group shall bedetermined based upon a sound statistical analysis of thescientific questions to be addressed by the study. This analysissh
46、ould take into account predicted survival rates (if available)for the species being used as well as being consistent withresponsible use of experimental animals. If a statistically validexperiment can be performed with fewer than the usual numberof animals per group, that fact should be documented a
47、nd thestudy design should proceed accordingly.9. Duration of Study9.1 Recommended durations for evaluation of tumorigenic-ity in rats is two years.9.2 Depending upon the material being evaluated, the earlyresults may suggest that the study can be terminated earlierthan two years without compromising
48、 the validity of the study.Examples might include studies in which a significantlyincreased rate of tumor formation or toxicity is being seen inthe test animals or in one or more dosage groups.9.3 At the termination of the study, a majority of the animalsin each group should have survived for euthan
49、asia or beenterminated early for study-related reasons such as increasedtumor incidence, spontaneous tumors, or toxicity of the testarticle. It is expected that a minimum of 50 % of the animalsper sex and per group should survive until final study termi-nation barring the above reasons. Moreover, the number ofsurvivors or study-related terminations should be sufficient fordetection of effects at the p 0.05 level of significance. Ifattrition is occurring due to reasons which cannot be attributedto the test articles or spontaneous tumor formation, otherfactors should be considered
