WVU IACUC - APPROVED SOP: BrdU and Other Thymidine Analogues Used in Animals

Transcription

1 Institutional Animal Care and Use Committee Effective: 1/1/11 Reviewed 2/5/14 Ver.2 SOP # WVU IACUC - APPROVED SOP: BrdU and Other Thymidine Analogues Used in Animals Guidance document for handling BrdU and other thymidine analogues (IdU and CldU) in the Lab and other areas. 5-bromo-2 -deoxyuridine (BrdU), 5-iodo-2 -deoxyuridine (IdU), and 5-chloro-2 -deoxyuridine (CldU) can substitute for the nucleoside deoxythymidine (T) and be incorporated into DNA during S-phase of cell division. Precautions must be taken when working with concentrated solutions and/or solid forms of BrdU and other thymidine analogues because these agents are toxic and may be teratogenic or mutagenic at high concentrations, but not carcinogenic. Anyone working with BrdU, IdU, or CldU should refer to the specific Material Safety Data Sheets (MSDS) for these compounds and information provided in the appendices below. General: Each employee working with these compounds is required to follow rules, recommendations and guidelines as outlined in their chemical hygiene plan, SOPs and supervisor s instructions. PI Responsibilities for use: The Principal Investigator is responsible for training employees using these compounds. The training should include: o Discussion of the known and potential hazards & review of the (M)SDS o Explanation of the relevant policies, SOPs and use procedures o Techniques and procedures including the proper use of personal protective equipment o Clean up and disposal procedures Each Principal Investigator is responsible for ensuring that the OLAR Director and husbandry staff members are informed in advance of the intended dates of use in the animal facility. Proper room and dated cage signage (including time of injection and date of safe cage change) is required. Lab Personal Protection: Wear gloves (preferably nitrile), lab coat, and safety glasses or goggles. When preparing stock solutions in the lab, work is done in a ventilated enclosure fume hood or a direct ducted Biosafety Cabinet Class II (preferably B2, but A2 is acceptable as the compound is aqueous soluble and non-volatile). The biosafety cabinet surface can be sprayed with 10% bleach to help inactivate these compounds. Small spills may be cleaned with soap and water using paper towels. No eating or drinking in the area of use. Minimize the possibility of inadvertent ingestion or inhalation and direct skin or eye contact with the drug. Wash hands after BrdU, IdU, or CldU work. Limit access to authorized users only. Page 1 of 5

2 Properly label containers of the chemical and any secondary containers. Develop a PI specific SOP for handling and transport (i.e., double containment during transport) of the chemical for animal use and storage of the chemical between uses. The SOP should have a contingency for spills, broken or leaking bottles and needle sticks. Hoods should be cleaned after handling animals/cages with BrdU; the spill formulation should be used. Transportation of the chemical within the facility should be performed using sealed, non-breakable secondary containment. Ensure that you provide appropriate biological and chemical waste containers for solid and liquid waste designated for chemical waste disposal. Elimination of small amounts of parent compounds in animals: 1. Follow the Standard OLAR hazardous chemical/drug use SOP. 2. Typical concentrations of BrdU, IdU, and CldU used in animal studies range from mg/kg (i.p.), which is well beneath toxic exposure levels and considered to be safe and effective. Nevertheless, low concentrations of the parent compounds and inactive metabolites are excreted through animal saliva and waste streams up to 7 days following injection. To minimize dangers posed by eliminated waste, pregnant females injected with BrdU should be singly housed and left undisturbed for 7-14 days. Though human exposures to trace amounts of BrdU on unbroken skin are unlikely to be harmful, care should be used when handling animals and animal bedding containing eliminated BrdU and/or other thymidine analogues. Wearing double gloves, lab coat, and face mask should eliminate any risk posed by contact with or inhalation of the eliminated parent compounds. Bedding should be disposed of by bagging and labeling each cage where BrdU was used with the PI and the compound. Special disposal involves minimal dust creation, dirty-side cage dumping into a HEPA filtered dump station, with staff wearing a fitted N-95 or PAPR. Cages are then washed twice as dedicated loads with no water recycling between loads. Bedding can be disposed of as normal waste. Appendices Hazard Identification: 5-BROMO-2'-DEOXYURIDINE Synonyms: BrdU; BDU; 5-BDU; bromouracil deoxyriboside; bioxuridine; NSC 38297; uridine, 5-bromo-2 - deoxy; bromodeoxyuridine Physical-Chemical Properties: CAS#: Molecular formula: C9H11BrN2O5 Molecular weight: g Form: Solid; White needles when crystallized from absolute ethanol; no distinctive odor Solubility: Some solubility in water or saline Volatility: Stable dark and heat/humidity for 3 mos. at temperatures below 60 C. Sunlight causes discoloration. Frozen aqueous solutions exposed to UV result in formation of debrominated dimmer. Photo-oxidation occurs in air/light. BrdU is hydrolyzed at the N-glycosyl bond, this rate increases at alkaline ph. Flammability restrictions do not apply. Other: Laboratory operations should be conducted in a fume hood, glove box, or ventilated cabinet. BrdU is a probable combustible compound. BrdU is cytotoxic, teratogenic, and mutagenic. It is not carcinogenic; it is a topical antiviral agent and is useful in the treatment of neoplasms and sensitizes tissue to X-rays. Page 2 of 5

3 Toxicity (NIH; DOHS; UCLA) Toxic Effects: BrdU is a toxic substance. Toxicity of BrdU is low in rodents. Acute Effects: Wheeze and cough, shortness of breath, burning in the mouth, throat, or chest. Rodents: inflammatory skin lesions, decreased weight/hair growth, and hematological adversity (anemia, leukocytopenia, thrombocytopenia). Chronic Effects: Reproductive disorders and genetic alterations. BrdU is incorporated into tissue DNA in place of thymidine in animals. BrdU-substituted DNA replaces normal DNA and chromosomal proteins are altered through: chromosome lengthening, chromatid breakage, and changed sister chromatid exchange frequencies. Meiosis and mitosis are affected. Carcinogenic Effects: Brdu is suspected to be a carcinogen resulting in heritable genetic damage. It is harmful to the unborn child. Mutagenic & teratogenic Effects: BrdU is not mutagenic Ames test; but BrdU is mutagenic micronucleus and sperm abnormality assay. BrdU is a strong teratogen in rodents. Systemic effects: BrdU integrates into DNA and affects the colon, stomach, bone marrow, and spleen. Also including the liver in pregnant mice and embryos. Exposure Assessment Route: BrdU is absorbed via body tissues in the gastrointestinal tract after parenteral injection and transplacentally. Avoid aerosol exposure and inhalation. BrdU is metabolized via hydrolysis and debromination. Toxicity: Acute LD50s in mice and rats are in the range of g/kg (parenteral), and 8.4 g/kg (oral). Control Plan Storage: Solid BrdU is stored in dark-colored, tightly-closed containers (original container covered in foil and stored in a secondary container; double contained) in the -20 C freezer. Storage of liquid stock concentrations is performed at -20 C in dark-colored plastic tubes or light-tight outer containers, which is non-breakable. Exposures: Refer to specific (M)SDS (these are only general guidelines) Skin: Remove contaminated clothing and immediately flush with plenty of water. Wash skin with soap and water; avoid rubbing and increases in temperature. Seek medical attention immediately. Eye: Rinse immediately with copious amounts of running water for at least 15 minutes. Seek attention of a licensed Ophthalmologist promptly. Ingestion: Seek medical attention immediately. Inhalation: Remove rapidly to clean air. Administer rescue breathing if necessary and call emergency services. Seek medical attention if needed. Waste Management Procedures: BrdU becomes present in DNA/RNA adducts. Major routes of excretion: urine, feces, saliva. Minor route: skin desquamation. Excretion period lasts one day to one week. Metabolites are nonhazardous. Disposal: Disposal procedures will be determined by Environmental Health and Safety with OLAR personnel, and detailed in the specific project s SOP. Page 3 of 5

4 Potentially hazardous waste: Absorbent materials (e.g., associated with spill cleanup) grossly contaminated shall be handled in accordance with the chemical waste disposal system. For additional information see: Spill and Accident Procedures: (NIH; DOHS; UCLA) Decontamination: Turn off equipment that could be affected by BrdU or the materials used for cleanup. If there is any uncertainty regarding the procedures to be followed for decontamination, call the WVU HSC Dispatch office at and ask for the Safety Office or, on the main campus, contact EH & S at o Ensure that appropriate Personal Protective Equipment (PPE) is being worn and absorb liquid using paper or pads for a small spill. o Wipe surfaces and clean with large amounts of soapy water. o Wash glassware and animal cages with soapy water. o Treat waste as Special Waste and dispose of properly. o Complete the WVU HSC Spill Response Notification form and forward to HSC Safety Office. The form can be found at: Safety/Spill_Response_Notification_Form_1.pdf In the event of a large spill: o Evacuate the area o Call o Provide first responders with information regarding the spill o For additional information see: Handling instructions: Preparing stock BrdU solutions: o Always handle powdered BrdU inside a certified chemical fume hood o Wear proper PPE as stated above o Always wash hands after removing gloves following handling BrdU o Clean powdered and stock solution BrdU spills with absorbent towels, soap and water o The paper towels should be placed into a Ziploc bag with a label for BrdU o The bag should be placed in a designated area for Special Waste disposal Page 4 of 5

5 References: NIH: Division of Occupational Health and Safety. 5-Bromo-2 -deoxyuridine Safety Data Sheet. (February 2009). UCLA: EH&S Staff. 5-Bromo-2-Deoxyuridine FACT SHEET. (February 2009) Hormel Institute: University of Tennessee Health Science Center Page 5 of 5