PROTOCOL # 9777 EXPIRES:

Transcription

1 PROTOCOL FOR ANIMAL USE AND CARE Handwritten forms are not accepted CRPRC Last Name: First: Middle: Department: Phone / Fax: Investigator Last Name: First: Middle: Department: Phone: After hrs. #: After hrs. #: EH&S USE ONLY PROTOCOL # EXPIRES: Contact Species (common names): Number: Source: Cynomolgous macaque 54 CRPRC Rhesus macaque 54 CRPRC Project Title Recombinant Polyvalent Viral Vaccines for use in Pregnancy & infants Overnight housing location:: CRPRC Day use only : Animals will be maintained by: [ X ] Vivarium [ ] Investigator (If investigator maintained, attach husbandry SOP s.) Procedures: Provide a one or two sentence layman's description of the procedures employed on the animals in this project. This information will help the animal care staff understand any conditions they may encounter while caring for your animals. This proposal will use cynomolgus and rhesus macaques as experimental hosts to improve the immunogenicity of vaccine strategies based on genetically modified vaccine strains of poliovirus that express antigens from SIV. Animals will be immunized and blood, mucosal secretions, and fecal samples will be taken. Special Husbandry Requirements: Describe any special requirements your animals have with respect to food, water, temperature, humidity, light cycles, caging type, bedding, or any other conditions of husbandry. Follow established CRPRC BSL2 housing criteria and standard SAIDS protocols. Other instructions for animal care staff: (check applicable entries) Sick Animals Dead Animals Pest Control [ X ] Call Investigator [ X ] Call Investigator [ X ] Call Investigator [ X ] Clinician to treat [ X ] Save for Investigator [ X] OK to use pesticides [ ] Terminate [ ] Bag for disposal [ ] No Pesticides in animal area [ ] Necropsy [ ] Necropsy Hazardous Materials (only if in the animal room): Infectious Agents? [X ] Yes [ ] No Agent(s): Poliovirus, SIV Radioisotopes? [ ] Yes [ X] No Agent(s): Chemical Carcinogens? [ ] Yes [ X] No Agent(s): Toxic Chemicals? [ ] Yes [X ] No Agent(s): Version 7/21/ :39:13 AM Page 1

2 Funding source: NIH, NIAID Previously approved? [ ] Yes [ X ] No Is the project already funded? [ ] Yes [X ] No Previous protocol number (if any): What Veterinarian or veterinary clinic will provide care for your animals? (check one) [ ] Lab Animal Health Clinic ( ) [X ] California Primate Research Center ( ) [ ] VMTH Large Animal Field Service ( ) [ ] Another Veterinarian If you checked Another Veterinarian, please provide: Veterinarian: Address: Day phone: Emergency phone: If your veterinarian is not affiliated with one of the three service units listed above, please contact the campus veterinarian, ( pctillman@ucdavis.edu) for current information about training and record keeping requirements. Summary of Procedures: a) Briefly describe the overall intent of the study. Include in your description a statement of your hypothesis, the objectives and significance of the study. Your target audience is a faculty member from a discipline unrelated to yours. Do not use jargon. Collaborator (UCSF) has developed a novel method for the production of vaccines utilizing recombinant polioviruses. This method uses the well-characterized Sabin vaccine strains of poliovirus (Sabin isolates of type 1 and 2) as an expression vector for antigenic determinants derived from simian immunodeficiency virus (SIV). In recent studies at the CRPRC, Drs. and have shown that recombinant polioviruses expressing SIV antigens are capable of eliciting antibody responses in serum and secretions as well as cellular immune responses in cynomolgus macaques. Also, macaques immunized with a cocktail of recombinant Sabin-SIV viruses showed protection from challenge with virulent SIV. We hypothesize that Sabin-SIV RNA genome (instead of recombinant poliovirus itself) administered to monkeys can elicit SIV-specific immune responses that will protect macaques against SIV challenge; if successful, this approach could be used as a novel, efficient vaccine strategy for preventing HIV infection. In our proposed studies, we aim to determine if a cocktail of Sabin-SIV RNA can elicit long-lasting immune responses to SIV antigens 1) in adult female cynomolgus and rhesus monkeys before and during pregnancy and, 2) in infant macaques with SIV- and polio-specific antibodies acquired by transplacental transfer from their vaccinated mothers. Our ultimate goal is to develop safe and efficacious vaccines against HIV that can be used in pregnant women and infants. b) Procedures employed in this project: Please check the appropriate boxes if any of these procedures will be employed in your project: [ ] Monoclonal Antibody Production ** [ ] Food or water restriction [ ] Special diets; food or water treatment. [ ] Polyclonal Antibody Production ** [ ] Non-recovery surgical procedures [ X ] Induced illness, intoxication, or disease [ ] LD 50 or ID50 studies. [ ] Survival surgical procedures [ ] Death as an endpoint (see i below) [X ] catheters, blood collection, intubation [ ] Multiple survival surgery [ ] Trapping, banding or marking wild animals [ ] Prolonged restraint. (8 hrs+) [ ] Behavioral modification. [ ] [X ] Fasting prior to a procedure. [ ] Aversive conditioning. [ ] ** If this protocol only describes antibody production, you may use the attached antibody production page in lieu of completing section c below. Version 7/21/ :39:13 AM Page 2

3 c) Describe the use of animals in your project in detail, with special reference to any of procedures checked above. Include any physical, chemical or biological agents that may be administered. List each study group, and describe all the specific procedures that will be performed on each animal in each study group. Use terminology that will be understood by individuals outside your field of expertise. (Note: This cell will expand to whatever length you require. You may make this section as long as you wish, but try to be concise. Some projects may require one or two pages.) 1. Animals used for these studies: (1) mature female cynomolgous and rhesus macaques, and (2) offspring of the mature female cynomolgus and rhesus macaques. Adult, cycling female macaques will be inoculated 4 times intra-muscularly (IM) and intra-nasally (IN) over the course of two weeks with the designated cocktail of Sabin-SIV RNA (see #3 below). After the first in the four inoculations, blood, vaginal and rectal secretions, saliva and fecal samples will be collected weekly for 8 weeks, then every 2 weeks for the duration of the experiment to determine viral replication and immune response. Sample collection described in # 2 below. Pregnant female macaques: By 6 months after the first inoculation with Sabin-SIV RNA, all adult females will be bred and evaluated for pregnancy using standard CRPRC procedures. At gestational day (GD) 120 (±3 days), pregnant animals will be inoculated four times over two weeks with the designated cocktail of Sabin-SIV RNA (see # 3 below). Bi-weekly blood, secretions, saliva and fecal samples will be collected throughout pregnancy. Adult females after vaginal delivery: infants will remain with their mothers until weaning (as per CRPRC SOP) at 6 months of age. Bi-weekly blood, secretions and fecal samples will be continue to be collected after delivery until intravaginal challenge with SIV. At 6 months after delivery, adults will be inoculated intravaginally 4 times (twice a week for 2 weeks) with virulent SIV (100,000 infectious units in 1 ml per intravaginal inoculation, see #4 below). Bi-weekly blood, secretions, saliva and fecal samples will be collected weekly for 4 weeks after the first intravaginal challenge, then biweekly until the animals develop clinical signs of immunodeficiency disease or by 6 months after vaginal SIV challenge (whichever is first) when the animals will be euthanized and a complete necropsy will be done. Infants of vaccinated females: Within 3 days of birth infants will be inoculated 4 times intra-muscularly (IM) and intra-nasally (IN) over the course of two weeks with the designated cocktail of Sabin-SIV RNA (#3 below). After the first in the four inoculations, blood, rectal secretions, saliva and fecal samples will be collected biweekly until SIV challenge to determine viral replication and immune response. Sample collection described in # 2 below. At 6 months of age, infants will be inoculated orally 4 times (twice a week for 2 weeks) with virulent SIV (100,000 infectious units in 1 ml per oral inoculation, see #4 below). Bi-weekly blood, secretions, saliva and fecal samples will be collected weekly for 4 weeks after the first oral challenge, then biweekly until the animals develop clinical signs of immunodeficiency disease or by 6 months after oral SIV challenge (whichever is first) when the animals will be euthanized and a complete necropsy will be done. 2. Samples to be taken from animals: Blood will be collected by femoral venipuncture under ketamine (10 mg/kg, intramuscularly in hind leg) anesthesia; blood volume sample\d will not exceed 12 ml/kg/month). At the time of blood collection, vaginal and rectal secretions will be obtained by lavage with PBS, and swabs will be used to collect saliva. Collection of blood, secretions and saliva will be under ketamine anesthesia (10 mg/kg, intramuscularly in hind leg). All animals will be fasted 12 hours prior to anesthesia. 3. Vaccines, routes of immunization, and vaccine dose: (performed under ketamine anesthesia, 10 mg/kg injected intramuscularly in hind leg): Vaccines: Two cocktails of Sabin-SIV RNA will be used: Sabin-1-SIV RNA and Sabin-2-SIV RNA. [200 ug in Routes of Immunization: Each Sabin-SIV RNA cocktail contains recombinant Sabin RNA carrying different SIV antigens representing 90% of all SIV protein sequences. In addition, non-recombinant Sabin-1 RNA and Sabin-2 RNA carrying NO SIV antigens will be used to immunize control monkeys. Routes of immunization: Animals will be inoculated intramuscularly (IM) in 4 sites (each bicep & quadricep) and intranasally (IN, both nostrils). Vaccine dose-adults: The dose of vaccine administered into each IM site will consist of 200 ug of a cocktail of Sabin-SIV RNA in 0.5 mls of phosphate buffered saline (PBS). The dose of vaccine given IN will also consist Version 7/21/ :39:13 AM Page 3

4 of 200 ug of a cocktail of Sabin-SIV RNA in 0.5 mls PBS. The dose for adults is based on previous successful IM and IN immunization of rhesus juveniles and adults with other SIV vaccines. Vaccine dose-infants: The dose of vaccine administered into each IM site will consist of 200 ug of a cocktail of Sabin-SIV RNA in 0.25 mls of PBS. The dose of vaccine given IN will also consist of 200 ug of a cocktail of Sabin-SIV RNA in mls PBS. The dose for infants is based on previous successful IM and IN immunization of rhesus infants with other SIV vaccines. 4. SIV challenge: (performed under ketamine anesthesia, 10 mg/kg injected intramuscularly in hind leg). The SIV inoculum will consist of either tissue culture supernatant from virus-infected rhesus PBMC or plasma from virus-infected macaques and will contain 10 5 infectious units per ml of inoculum. All adult females will be inoculated intravaginally 4 times (twice a week for 2 weeks) with virulent SIV at 6 months after delivery of infants. All infants will be inoculated orally 4 times (twice a week for 2 weeks) with 1 ml of SIV at 6 months of age. 5. Experimental endpoint: All animals will be euthanatized by 24 weeks after SIV challenge to obtain tissues for virological, immunological, histochemical and molecular assays. A complete necropsy will be performed for each animal and peripheral and systemic lymphoid tissues will be prepared for histological, immunohistochemical and PCR analysis to detect virus-infected cells. 6. Based on the guidelines for categories for invasiveness, all SIV-infected animals may experience moderate to severe stress (category 3) due to immunosuppression as a results of their infection. Our goal is to circumvent or prevent any severe compromise or pain (category 4); for example, CRPRC Veterinary staff will have the option to treat any secondary infections or to administer analgesics. Thus, categories listed in the Table in section d below are designated as level 3. GROUP A- Adult females: Inoculation with Sabin-1 based recombinant RNA, boosting with Sabin-2 recombinant RNA will induce SIV-immunity. Group A will require a total of 18 adult female macaques: 9 cynomolgus macaques & 9 rhesus macaques. Monkeys will be inoculated IM + IN with Sabin-1-SIV RNA. Sixteen weeks later, animals will be inoculated IM + IN with Sabin-2-SIV RNA. At gestational day (GD) 120 (± 3 days), pregnant animals will be inoculated IM + IN with Sabin-1-SIV RNA l; two weeks later, animals will be inoculated IM + IN with Sabin-2-SIV RNA. Six months after delivery of infants, animals will be challenged intravaginally with SIV. Blood, vaginal and rectal secretions, saliva and fecal samples will be collected weekly for the first month and then every two weeks until the animals develop clinical signs of SIV or by 6 months post challenge (whichever is first) when the animals will be euthanized. GROUP B- Adult females: Inoculation with Sabin-2 based recombinant RNA, boosting with Sabin-1 recombinant RNA will induce anti-siv-immunity. Group B will require a total of 18 adult female macaques: 9 cynomolgus macaques & 9 rhesus macaques. Monkeys will be inoculated IM + IN with Sabin-2-SIV RNA. Sixteen weeks later, animals will be inoculated IM + IN with Sabin-1-SIV RNA. At gestational day (GD) 120 (± 3 days), pregnant animals will be inoculated IM + IN with Sabin-1-SIV RNA 2 two weeks later, animals will be inoculated IM + IN with Sabin-1-SIV RNA. Six months after delivery of infants, animals will be challenged intravaginally with SIV. Blood, vaginal and rectal secretions, saliva and fecal samples will be collected weekly for the first month and then every two weeks until the animals develop clinical signs of SIV or by 6 months post challenge (whichever is first) when the animals will be euthanized. GROUP C- Adult females: Control group for groups A&B, inoculation with non-recombinant Sabin-1 and Sabin 2 RNA will not induce anti-siv-immunity. Group C will require a total of 18 adult female macaques: 9 cynomolgus macaques & 9 rhesus macaques Control monkeys will be inoculated IM + IN with non-recombinant Sabin-1RNA and non-recombinant Sabin-2 RNA following the same schedule as groups A&B. Six months after delivery of infants, animals will be challenged intravaginally with SIV. Blood, vaginal and rectal secretions, saliva and fecal samples will be Version 7/21/ :39:13 AM Page 4

5 collected weekly for the first month and then every two weeks until the animals develop clinical signs of SIV or by 6 months post challenge (whichever is first) when the animals will be euthanized. GROUP D- Infants of Group A vaccinated females that have passive immunity against recombinant poliovirus can still produce immunity to SIV antigens using a poliovirus recombinant. Group D will use a total of 18 infant macaques (born to Group A females): 9 cynomolgus & 9 rhesus macaques Monkeys will be inoculated IM + IN with Sabin-1-SIV RNA within 72 hours of birth. Sixteen weeks later, animals will be inoculated IM + IN with Sabin-2-SIV RNA. At 6 months of age animals will be challenged orally with SIV. Blood, vaginal and rectal secretions, saliva and fecal samples will be collected weekly for the first month and then every two weeks until the animals develop clinical signs of SIV or by 6 months post challenge (whichever is first) when the animals will be euthanized. GROUP E- Infants of Group B vaccinated females that have passive immunity against recombinant poliovirus can still produce immunity to SIV antigens using a poliovirus recombinant. Group E will use a total of 18 infant macaques (born to Group B females): 9 cynomolgus & 9 rhesus macaques Monkeys will be inoculated IM + IN with Sabin-2-SIV RNA within 72 hours of birth. Sixteen weeks later, animals will be inoculated IM + IN with Sabin-1-SIV RNA. At 6 months of age animals will be challenged orally with SIV. Blood, vaginal and rectal secretions, saliva and fecal samples will be collected weekly for the first month and then every two weeks until the animals develop clinical signs of SIV or by 6 months post challenge (whichever is first) when the animals will be euthanized. GROUP F- Control group for groups D&E, animals inoculated with Sabin-1 will not induce anti-sivimmunity. Group F will use a total of 18 infant macaques (born to Group C females): 9 cynomolgus & 9 rhesus macaques Control monkeys will be inoculated IM + IN with non-recombinant Sabin-1 RNA within 72 hours of birth. Sixteen weeks later, animals will be inoculated IM + IN with non-recombinant Sabin-2 RNA. At 6 months of age animals will be challenged orally with SIV. Blood, vaginal and rectal secretions, saliva and fecal samples will be collected weekly for the first month and then every two weeks until the animals develop clinical signs of SIV or by 6 months post challenge (whichever is first) when the animals will be euthanized. d) Study Groups and Numbers: Define, in the form of a table, the numbers of animals to be used in each experimental group described above. The table may be presented on a separate page as an attachment to this protocol if you prefer. The Normal format should be three columns: Study Group, Procedure, Number of animals. The number of rows should follow from the number of study groups; you may add as many rows as you require. The chart must fully account for the number of animals you intend to use under this protocol. Assign each group to an invasiveness category according to the chart below. Group Procedures / Drugs Number of Animals Category A Intra-muscular (IM) + intranasal (IN) Sabin-1-SIV RNA inoculation followed by IM + IN Sabin-2-SIV RNA inoculation; intravaginal SIV challenge 18 adult females 3 B C D E F IM+ IN Sabin-2-SIV RNA inoculation followed by IM + IN Sabin-1-SIV RNA inoculation; intravaginal SIV challenge IM+ intranasal IN non-recombinant Sabin-1 RNA inoculation followed by IM + IN non-recombinant Sabin-2 RNA inoculation; intravaginal SIV challenge IM+ IN Sabin-1-SIV RNA inoculation followed by IM + IN Sabin-2-SIV RNA inoculation; oral SIV challenge IM + IN Sabin-2-SIV RNA inoculation followed by IM + IN Sabin-1-SIV RNA inoculation; oral SIV challenge IM + IN non-recombinant Sabin-1 RNA inoculation followed by IM + IN non-recombinant Sabin-2 RNA inoculation; oral SIV challenge 18 adult females 3 18 adult females 3 18 (infants of Group A females) 18 (infants of Group B females) 18 (infants of Group C females) 3 3 Version 7/21/ :39:13 AM Page 5

6 Category Description Little or no discomfort or stress Categories of invasiveness Examples: domestic flocks or herds being maintained in simulated or actual commercial production management systems; the short-term and skillful restraint of animals for purposes of observation or physical examination; blood sampling; injection of material in amounts that will not cause adverse reactions by the following routes: intravenous, subcutaneous, intramuscular, intraperitoneal, or oral. Minor stress or pain of short duration Examples:: cannulation or catheterization of blood vessels or body cavities under anesthesia; minor surgical procedures under anesthesia, such as biopsies or laparoscopy; short periods of restraint beyond that required for simple observation or examination, but consistent with minimal distress Moderate to severe distress Examples: major surgical procedures conducted under general anesthesia, with subsequent recovery; prolonged (several hours or more) periods of physical restraint; induction of behavioral stresses such as maternal deprivation Severe pain near, at or above the pain tolerance threshold Examples: exposure to noxious stimuli or agents whose effects are unknown; exposure to drugs, chemicals, or infectious agents at levels that markedly impair physiological systems and which cause death, severe pain, or extreme distress: Surgical experiments which have a high degree of invasiveness. Further descriptions of these categories are included in the instructions following this document. e) Rationale for species and numbers: How did you determine that 1) the species choice was appropriate and 2) the number of animals in each study groups was the minimum number necessary to achieve sound scientific results? Cynomolgus and rhesus macaques were chosen for these experiments because these species of monkeys are known to support poliovirus replication as well as being an excellent host for subsequent SIV challenge. Infection of rhesus macaques with SIV/SHIV is currently the best non-human primate model for human AIDS. The neonatal rhesus macaque is currently the most sensitive and biologically appropriate model in which to evaluate the transmission and efficacy of potential strategies to prevent HIV-infection of human newborns. We need to test the novel proposed recombinant poliovirus RNA-based immunization strategy in both cynomolgus and rhesus macaques because (1) this vaccine strategy has never been tested in any primate species and, (2) it is unknown which macaque species will produce immune responses that can protect against vaginal or oral challenge with SIV. Nine animals per group are required to ensure sufficient statistical power to determine vaccine failure (no difference in number of animals infected, virus load, or disease in vaccinated and unvaccinated controls) as well as vaccine efficacy (partial or complete protection). Nine animals per group are also needed to ensure that we can distinguish statistically significant as well as biologically meaningful differences among the vaccine regimens. The numbers of animals per group was calculated based on the need to detect with biologically acceptable statistical power (~80%) a minimum of 30-fold difference in measures of vaccine success between controls (AUC #8447) and vaccinated animals. In consultation with Dr. (CRPRC) who has expertise in biostatistics, we used estimated variances of parameters from our recent vaccine experiments in rhesus neonates conducted at the CRPRC and followed the methods outlined in, 1995, Intuitive Biostatistics. We anticipate that the results of our experiments will be used to plan clinical trials to vaccinate pregnant women and infants of HIV-infected women. Thus, we must obtain the most accurate information possible comparing vaccine immunogenicity and efficacy in pregnant and infant non-human primates. f) Surgery: If the project involves survival surgery, where will the surgery be conducted? Building: Who will be the surgeon? Room: Version 7/21/ :39:13 AM Page 6

7 g) Anesthetics, Analgesics, Tranquilizers, Neuromuscular blocking agents: Post procedural analgesics should be given whenever there is possibility of pain or discomfort that is more than slight or momentary. If postoperative analgesics are not to be given, justify the practice under part (i) below. Provide the following information about any of these drugs that you intend to use in this project. Species Drug Dose (mg/kg) Route When and how often will it be given? Cynomolgus and Ketamine 10 mg/kg IM Before all procedures rhesus macaques Cynomolgus and rhesus macaques Oxymorphone 1 mg/kg IM As per judgement of CRPRC vets h) Neuromuscular blocking agents can conceal inadequate anesthesia and therefore require special justification. If you are using a neuromuscular blocking agent, please complete the following: Why do you need to use a neuromuscular blocking agent? What physiologic parameters are monitored during the procedure to assess adequacy of anesthesia? Under what circumstances will incremental doses of anesthetics-analgesics be administered? i) Adverse effects: Describe any potential adverse effects of the experiment on the animals (such as pain, discomfort; reduced growth, fever, anemia, neurological deficits; behavioral abnormalities or other clinical symptoms of acute or chronic distress or nutritional deficiency) All procedures will be performed under anesthesia administered by the CRPRC animal care or veterinary staff; to minimize pain or discomfort for the animals. We have observed 1/5 (20%) monkeys exposed to poliovirus via IN route developed neuropathology (paralysis). If paralysis develops in an animal, disposition of that animal will be left to the discretion of the veterinary staff. SIV-infected animals may display a spectrum of clinical disease that ranges from mild (such as skin rash, transient diarrhea) to moderate (e.g. lymphadenopathy, splenomegaly, chronic diarrhea, failure-to-thrive), to severe (multiple opportunisitic infections, anorexia, weight loss, pneumonia). Minimal discomfort is associated with blood collections, IN, or IM immunizations, and collection of vaginal rectal secretions and salia; animals will be anesthetized with ketamine for all of these procedures. All possible measures are taken to minimize discomfort from all experimental procedures. SIV infection of rhesus and cynomolgous macaques results in a fatal immunodeficiency and wasting syndrome. The animals will be euthanized when they experience 3 of the following: weight loss >15% in 2 weeks or >30% in 3 months; persistent hypothermia <96F even with heat supplementation; leukopenia (total WBC <3,000); lymphopenia (lymphocytes <800); anemia (hemoglobin <10); dehydration >10%; nonresponsive to therapy for opportunistic infections; persistent anorexia (> 3 days); animal significantly obtunded. These criteria are based on CRPRC guidelines. How will the signs listed above be ameliorated or alleviated? If signs are not to be alleviated or ameliorated by means of postoperative analgesics or other means, explain why this is necessary. Should any evidence of pain, discomfort, or neurologic deficit be seen in the experimental animals, analagesics and/or anesthetics will be administered promptly if considered appropriate after discussions with the CRPRC veterinary staff. Animals that fail to respond to supportive therapy will be euthanized. The SIV-infected animals will be euthanized prior to or at the time they develop clinical signs of AIDS. The decision to euthanize will be based on the judgment of the CRPRC veterinarians (as specified in the CRPRC guidelines, "Criteria for euthanasia of retrovirus-infected macaques"). Note: if any unanticipated adverse effects not described above do occur during the course of the study, a complete description of those effects and the steps taken to mitigate them must be submitted to the committee as an amendment to this protocol. Version 7/21/ :39:13 AM Page 7

8 Is death an endpoint in your experimental procedure? [ ] Yes [ X ] No (Note: "Death as an endpoint" refers to acute toxicity testing, assessment of virulence of pathogens, neutralization tests for toxins, and other studies in which animals are not euthanized, but die as a direct result of the experimental manipulation). If death is an endpoint, explain why it is not possible to euthanize the animals at an earlier point in the study. If you can euthanize the animals at an earlier point, describe the clinical signs which will dictate that an animal will be euthanized. j) Literature search for alternatives and unnecessary duplication: This section is specifically required by Federal law. You are required to conduct a literature search to determine that either 1) there are no alternative methodologies by which to conduct this study, or 2) there are alternative methodologies, but these are not appropriate for your particular study. "Alternative methodologies" refers to reduction, replacement, and refinement (the three R's) of animal use, not just animal replacement. You must also show that the study is not unnecessarily duplicative of other studies. What was the date on which you conducted this search? 9/26/01 List the databases searched or other sources consulted (there should be more than one). Include the years covered by the search. Database Name Years Covered Keywords / Search Strategy Advanced Pub Med/Medline Current Contents Poliovirus, SIV, vaccines, protection, pregnancy Poliovirus, SIV, vaccines, protection, pregnancy What were your findings with respect to alternative methodologies? The poliovirus receptor transgenic mouse and the monkey are the only available animal model systems for assessing poliovirus vaccine efficacy. The SIV/monkey is the most satisfactory animal model system for the study of vaccine efficacy and the effects of vaccination during human pregnancy and infancy. Prior experiments have produced encouraging results, i.e. protection from vaginal challenge with SIV. Has this study been previously conducted? [ ] Yes [ X ] No If the study has been conducted previously, explain why it is scientifically necessary to replicate the experiment. k) Disposition of animals: At what point in the study, if any, will the animals be euthanized? Animals will be euthanized when they develop clinical signs of SIV, or at the end of the study period. l) Methods of euthanasia: Even if your study does not involve killing the animals, you should show a method that you would use in the event of unanticipated injury or illness. If anesthetic overdose is the method, show the agent, dose, and route. Species Method Drug Dose (mg/kg) route Macaca fasicularis IV overdose pentabarbitol 60 mg/kg IV Macaca mulatta IV overdose pentabarbitol 60 mg/kg IV m) Surplus animals: What will you do with any animals not euthanized at the conclusion of the project? All SIV-challenged animals will be euthanized as a part of this study. Version 7/21/ :39:13 AM Page 8

9 n) Project Roster: Please provide the names of all the individuals who will work with animals on this project. This page will not be made available to the public. Give either the University Employee ID # or a valid UC Davis address so that we can document training and occupational health compliance for regulatory agencies. Include all investigators, student employees, post-doctoral researchers, staff research associates, post-graduate researchers and laboratory assistants who will actually work with the animals. You don t need to include the staff of the vivarium in which your animals will be housed. The principal investigator is responsible for keeping this roster current. If any staff is added or subtracted from this project, you must amend the protocol by sending the campus veterinarian a memo describing any changes. Last Name First Name Middle Name UC ID Number or SSN Address Occupational Health Program: Supervisors must enroll their employees in the campus Occupational Health Program if the workers are at increased risk of illness or injury (such as allergy, physical injury, or infectious disease) because of their work. Enroll workers by having them complete an "Animal Contact History Form", available from Employee Health Services (phone ). For further information, visit our web site at or read the UC Davis Policy & Procedure Manual Training: Supervisors are responsible for insuring that their employees are adequate trained, both in the specifics of their job and in the requirements of the Federal Animal Welfare Act. EH&S offers free, basic wet labs in laboratory animal handling and techniques, and lecture format classes in the requirements of the Animal Welfare Act. To schedule a class for your unit, contact EH&S at Autotutorials are also available on the world wide web at Version 7/21/ :39:13 AM Page 9

10 Assurances for the Humane Care and Use of Vertebrate Animals: Principal Investigator's Statement: I have read and agree to abide by the UC Davis Policy and Procedure Manual section (Animal Use and Care). This project will be conducted in accordance with the ILAR Guide for the Care and Use of Laboratory Animals, and the UC Davis Animal Welfare Assurance on file with the US Public Health Service. (These documents are available from the Campus Veterinarian and at ). I will abide by all Federal, state and local laws and regulations dealing with the use of animals in research. I will advise the Animal Use and Care Administrative Advisory Committee in writing of any significant changes in the procedures or personnel involved in this project. Principal Investigator Rank / Title Date ** Conditions necessary for Committee Approval: Committee Use Only Below Final Disposition of this protocol: Approved Not Approved Withdrawn by Investigator Date of Action: / / I verify that the Institutional Animal Care and Use Committee of the, acted on this protocol as shown above. Version 7/21/ :39:13 AM Page 10

11 ANIMAL ROOM SAFETY INFORMATION Complete this form if you will be using biohazards, radioisotopes, carcinogens, or toxic chemicals in the animal room. PROTOCOL # EXPIRES: RUA#: BUA#: 0477 CCA#: Identity ofhazard: Poliovirus vaccine (types 1 & 2) Simian immunodeficiency virus (SIV) Investigator Last Name: Department: First Name: Phone: Fax: Provide a short description of the agent: SIV is a primate lentivirus that is genetically similar to HIV and causes fatal immunodeficiency (AIDS) in infected rhesus macaques. SIV can infect humans, but it is unknown whether SIV causes human disease. Poliovirus vaccine isolates are attenuated forms of poliovirus that infect humans, but do not cause any disease except very rarely in healthy people or in immunosuppressed people. This agent / material is hazardous for: [ ] Humans only [ ] Animals only [ X] Humans and Animals For which Animal Species? primates The agent can be spread by: [X] Blood [ X ] Feces/urine (polio virus) [ ] Saliva/nasal droplets [ ] Does not leave animal [x] Other: Mucosal (eye/mouth/nose/genital) Describe any human health risk associated with this agent: SIV can infect humans; thus, it is possible that SIV could cause a fatal, AIDS-like disease in humans. Infectious virus and SIV antibodies have been detected in SIV-infected humans but there have been no reports of disease in SIV-infected people. Poliovirus vaccines can infect humans and are used to prevent poliomyelitis (flaccid muscular paralysis); very rarely in healthy people or in some immunosuppressed people, poliovirus vaccines can induce muscle paralysis The precautions checked below apply to this experiment: [ ] The researcher or his/her technicians are responsible for the feeding and care of these animals. [ ] The following items must be assumed to be contaminated with hazardous material and must be handled only by the researcher or his/her technicians. [ ] Cage [ ] Stall [ ] Water Bottle [ ] Animal Carcasses [ ] Bedding [ ] Other: [ x] Cages must be autoclaved before cleaning. [ ] Label cages and remove label after decontamination. [ ] Animal carcasses must be labeled and disposed of as follows: [ ] Incineration [ ] Biohazardous Waste Container [ ] Bag and Autoclave [ ] EH&S will pick-up (2-1493). [x ] All contaminated waste (soiled bedding or other animal waste) must be properly labeled and disposed of as follows [ ] Incineration [ x] Biohazardous Waste Container [ ] Bag and Autoclave [ ] EH&S will pick-up (2-1493). Personal Protective Equipment Required: [ x] The following personal protective equipment must be worn/used in the room: [ ] Lab Coat/Coveralls [ x] Shoe Covers/Booties [ x] Disposable Gloves [x ] Head Cover [ ] NIOSH Certified Dust Mask [ ] Disinfectant footbath [ x] Eye Protection/Face Shield [ ] [ ] Fitted Respirator Type: [ x] Other: Describe: Plastic disposable gown/coveralls [ x] [ ] Personal protective equipment must be removed before leaving the room. Personal protective equipment must be discarded or decontaminated at the end of the project [ x] Hands, arms, and face must be thoroughly washed upon leaving the room [ ] Full shower, including washing of hair, must be taken upon leaving the room. [ ] Decontaminate Room (Inform ARS area supervisor when cage and/or room can be returned to general use). Provide any other information needed to safely work in this room: Biosafety Level 2+ (BSL2+) precautions must be used at all times Version 7/21/ :39:13 AM Page 11