11. Laboratory and Specimen Management Procedures 11.1
Contents
1. 03 395 454 and box inserts Fisher Scientific Cat No 03 395 462 Calibrated analytical balance capable of measuring to 0 0001g the same balance must be used throughout the procedure Leak proof transport container if required Preparation of the Dacron swabs Prepare prior to the procedure in a clean environment Tare the weighing balance and ensure balance has been calibrated Remove Dacron swabs from the box wear gloves at all times when handling swabs 4 The swabs are 6 inches long Using scissors cut off the top of the shafts to reduce the length of the swabs to approximately 4 5 inches so that each will fit inside the 15mL conical tubes 5 Place each Dacron swab into its own appropriately labeled 15mL conical tube labeled with a unique patient identifier 6 Label conical tubes so that they are identifiable for PTID and also for sample type eg label one conical tube Cervicovaginal label the other tube Rectal 7 Weigh the labelled conical tubes containing swabs and document the weight An example worksheet is shown in Figure 11 4 Record the weighing time 24hour clock 9 The Dacron swabs are now ready to be used for collection of fluid wN 9o See SSP section 9 for fluid collection instructions Storage of specimens Transport the conical tubes at ambient temperature to the location where post weight will be documented Following the documentation of the post collection sample wei2. documentation of training should be available for inspection at any time As transmission of HIV and other infectious agents can occur through contact with contaminated needles blood blood products and vaginal secretions all study staff must take appropriate precautions when collecting and handling biological specimens Guidance on universal precautions is available from the US Centers for Disease Control and Prevention at http www cdc gov ncidod dhqp bp universal precautions html HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 1 of 30 Additional reference information can be requested from the HPTN LC The information provided below is intended to standardize laboratory procedures for HPTN 076 across the study sites Adherence to the specifications detailed in this section is essential to ensure that primary secondary and exploratory endpoint data derived from laboratory testing will be considered acceptable to regulatory authorities 11 2 Specimen Labeling All containers into which specimens are initially collected e g blood collection tubes will be appropriately labeled according to local practices Participant Identification PTID labels will be provided by the HPTN Statistical Data and Management Center SDMC SCHARP if required for this function LDMS Tracking Forms will also be provided for use if required although sites may use their own specimen transport do
3. provide guidance on how to respond to the problem In addition to following this guidance designated site and lab staff will work together to document the problem take appropriate corrective and preventive action and document all action taken Reconciliation must be performed for all specimen types that are received by the laboratory and stored in the LDMS The lab staff should provide the HPTN LC with a detailed accounting of these issues in a regulatory Note to File or a specific agreed upon format HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 5 of 30 11 3 Protocol Related Testing and Sample Collection Samples will be collected and processed at the screening enrollment and follow up visits as indicated in tables 11 1 and 11 2 Collect specimens and label tubes according to local regulations and the specimen collection SOPs Blood collection tubes must be filled to the appropriate fill level as indicated by the tube manufacturer and must be collected according to the order of draw dictated by your local institution Please contact the HPTN LC for advice if your tube type is not listed below or if the order of draw differs from the following e Citrate Tube coagulation e Serum Tubes e EDTA Ethylenediaminetetraacetic acid Tubes e Fluoride glucose Tube After collection e EDTA tubes Lavender top and fluoride tubes grey top should be gently inverted at le
4. the appropriate number of LDMS cryovial labels The lab should store plasma in labeled cryovials Cryovial size may vary but 2 0 mL is recommended Reminder these vials hold 1 8 mL of liquid Do not add more than 1 8 mL due to expansion issues when freezing Blood processing and plasma storage should be performed within 6 hours of sample collection Centrifuge tube at 800 1000 x g for 10 minutes to separate cells and plasma Carefully remove plasma and avoid disturbing the cell layer Transfer the plasma to an appropriately labelled sterile centrifuge tube Centrifuge plasma again at 800 1000 x g for 10 minutes to remove any contaminating debris cells or platelets Log samples into LDMS and generate LDMS labels PL2 Each aliquot will have its own individual identification number Global Specimen ID Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 17 of 30 Store plasma in aliquot number order For example if there is only 3 mL of plasma for archive store 1 8 mL in aliquot 1 Store the remaining 1 2 mL in aliquot 2 and adjust the aliquot volume in LDMS to indicate 1 2 mL Store the aliquots in the freezer locations assigned in LDMS in a minus 70 to minus 90 freezer Plasma for storage will be stored on site until all protocol related testing is complete Note that some testing will be performed after study visits have been completed Study sites should plan to store sp
5. to enrollment 11 4 Specimen Processing for Sample Storage 11 4 1 Plasma Processing for Storage Five aliquots of plasma will be prepared from the 20 ml blood tube at each study visit as indicated below HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 16 of 30 20 mL of EDTA whole blood will be drawn for plasma storage at each time point at which HIV testing is performed as indicated in Table 11 1 Sites are requested to store 5 x 1 8 mL aliquots of plasma if possible HPTN LC should be informed at any time that three or less aliquots of 1 8mL are stored This plasma will also be used for PK testing at the appropriate visits An additional 20 mL of EDTA whole blood will be drawn for plasma storage for participants with a reactive or positive HIV test at any time after enrollment as indicated in Table 11 3 This additional plasma will be stored in the same way Sites will follow site specific SOPs for plasma processing which will include the following HPTN 076 SSP Manual Collect blood into lavender top blood collection tubes EDTA labeled with a SCHARP provided PTID label Size and number of collection tubes may vary depending on local lab requirements Deliver this to the local LDMS laboratory along with the LDMS Specimen Tracking Sheet or site specific requisition Using the LDMS Specimen Tracking Sheet log the sample into LDMS specimen type BLD and generate
6. to the LDMS storage laboratory should be as soon as possible so that biopsies can be placed in the freezer within one hour of collection Specimens should be accompanied by a completed LDMS tracking sheet Procedure Label a cryovial with an LDMS generated label containing the appropriate sample study identification information Weigh the labeled cryovial using an analytical balance use the same analytical balance throughout the procedure Document the weight of the labeled cryovial on the appropriate sample study form See Figure 11 5 for an example of study form Receive biopsy in transport medium Biopsy should be delivered to the lab to allow freezing within one hour of collection Transfer biopsy to petri dish Work under sterile conditions Using pointed forceps pick up the biopsy and drain off excess medium by touching biopsy to side of petri dish Transfer biopsy to a pre weighed cryovial Ensure biopsy sits at bottom of cryovial Weigh the cryovial containing the biopsy Document the weight of the cryovial containing the vaginal biopsy on the appropriate sample study form HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 25 of 30 e Freeze the cryovial containing the biopsy in Liquid Nitrogen or a dry ice alcohol bath e Store the labeled cryovial containing the biopsy in a 80 C freezer Document the date time the cryovial containing the bio
7. 11 Laboratory and Specimen Management Procedures 11 1 Overview of Section 11 This section contains information on the laboratory procedures performed in HPTN 076 Laboratory procedures will be performed in a variety of settings including e Clinics e Local laboratories e The HPTN Laboratory Center LC Baltimore MD USA e Other laboratories designated by the HPTN LC Tables in this document list the time points testing location s and specimen requirements for each test In all settings laboratory procedures will be performed according to the guidelines included in this section of the SSP and in addition study site Standard Operating Procedures SOPs that have been reviewed and approved by the HPTN LC In addition package insert instructions must be followed Ideally one method test kit and or combination of test kits will be used for each test throughout the duration of the study If for any reason a new or alternative method kit or test must be used after study initiation site laboratory staff must inform the HPTN LC to determine if any test kit validation is required For international sites an updated PAL protocol analyte list will need to be submitted to the LC for approval by DAIDS Regardless of whether tests are performed in clinic or laboratory settings study staff that perform the tests must be trained in proper testing and associated quality control QC procedures before performing the tests for study purposes
8. 6 Safety Week 8 Safety Week 14 Primary Outcome Visit Week 64 Tail Phase Week 76 Tail Phase Eue Q RE El E S g Week 4 First Injection Week 12 Second Injection Week 20 Third Injection Week 28 Fourth Injection Week 36 Fifth Injection Week 44 Sixth Injection Week 52 wer FPCPEEPEEEEE e TT MTT ELELELLELL Coagulation testing PT INR x aPTT Vaginal tissue X processing storage 1 To be offered if a satisfactory Pap smear is not documented within the last 12 calendar months and is indicated See Protocol Section 3 1 1 2 Coagulation and Pap testing can also be done at later visits but results must be available for enrollment into the Tissue Subset 3 Vaginal tissue will be used for pharmacologic assessments Vaginal tissue will be collected only at US sites that have this capacity Collection at Week 36 is preferred but tissue may be collected Week 44 Vaginal tissue must be collected at the same visit as cervicovaginal and rectal fluid collection All samples for PK plasma vaginal tissue cervicovaginal fluid and rectal fluid should be collected within a 4 hour window HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 9 of 30 11 3 1 HIV Testing HIV testing will be performed using blood collected by phlebotomy no finger stick or oral fluid testing at participant visits in accordance with the testing algorithms de
9. HPTN SDMC will provide each site with a list of participants who did not consent to indefinite storage and possible future research testing Sites should follow local regulations for sample destruction and follow the information provided in the LC section of the HPTN Manual of Operations Documentation of the destruction should be provided to the HPTN LC 11 4 2 Whole Blood Storage for Pharmacogenomic Testing Specimen Type Whole blood collected in EDTA anticoagulant purple top tubes Specimen volume Minimum 4 mL whole blood Handling Instructions Whole blood is transferred to 5 mL cryovials and frozen at minus 70 C or colder Materials Materials Thermo Scientific Nalgene Cryovials 5 mL Nalgene Cat 5000 0050 Different cryovials may be used but the HPTN LC must be consulted before use Disposables Standard Disposable Transfer Pipets example Fisher Cat 137117M Procedure Stepwise 1 Anappropriately labeled and filled EDTA whole blood tube will be received 2 Log specimens into LDMS upon receipt using the following LDMS codes a PRE BLD b ADD EDT c DER BLD d Sub Add Der N A e Other Spec ID PGEN HPTNO76SSPManual VesoniO0 Aprii7 2015 Section 11 Laboratory and Specimen Management Procedures Page 19 of 30 3 Transfer the whole blood to a labeled Nalgene cryovial using a transfer pipet 4 Do not fill cryovials to more than 74 capacity 5 Use Parafilm to seal caps of the cryovials to pre
10. anagement Procedures Page 22 of 30 Figure 11 4 Example of a Dacron Swab Collection and Storage Worksheet u s JI OOJS poo q jo BdUaSaJd 3a szu wwo 14519m 1sod pue aJd JO pasn aouejeq jo JaquINN elas 1eu4104 4H tc A4101e40qe Suisso204d ay ui paAra284 3J9M sueuumads OWI Tewo 4H vz eun uonejoo pinjj je29g 3eu04 JH pz eun uonoo oo pinj4 jeur8e o21 8 jyeqoay jeu ISeAO2IAJ9 2uDBI2M 24d qoms qDMS 1004404 1uBI2M 10U404 q07 2001 SJnoH JH pZ 1S0d JH bZ 4 aqn aqn pue 4U31IOM SW Ul SW ul u0onpo o2 u auSioM 1uSloM uaaMeq 01399 09 uo u Jaquinn Aq p JeAsaquUl au 3 Sui Ul ysod 13 9a 09 on epo5 yonbijy JWUOLdd aw uizaaJj 1uSi9M 39N Jo awiy 3SOd ad SING adAL pinj4 April 17 2015 Page 23 of 30 Version 1 0 HPTN 076 SSP Manual Section 11 Laboratory and Specimen Management Procedures 11 4 4 Vaginal Tissue for Processing and Storage Tissue Subset Participants Only Note It is important that cervicovaginal fluid be collected before the collection of vaginal tissue The following samples will be collected from participants in the tissue subset Select participants US sites only and will be processed and stored using LDMS Specimens should be collected as described in section 9 of the SSP Vaginal Biopsies Based on a 15 mg biopsy the following numbers of biopsies are suggested for each testing procedure e Vaginal t
11. asma samples to the LC upon receiving shipping request lists The site will batch the shipment export the LDMS data and notify the LC about the shipment Any additional samples may be specifically requested by the HPTN LC e g archive back up samples Contact the HPTN LC at Johns Hopkins University Estelle Piwowar Manning epiwowa jhmi edu 410 614 6736 and Paul Richardson prichal 8 jhmi edu to coordinate the timing and logistics of each shipment Sites will ship samples to the LC using the LDMS following the LC approved Shipping SOP indicating Lab 300 as the ship to lab ID number Personnel involved in the shipping process must be IATA trained and certified for the shipping of Category B Biological specimens UN 3373 Diagnostic Packing Instructions 650 Include a copy of the shipping manifest and box map For dry ice shipments use diagnostics packing code 650 UN 3373 and address the shipment to Estelle Piwowar Manning Johns Hopkins University Hospital Department of Pathology Pathology Building Room 311 600 North Wolfe Street Baltimore MD 21287 USA Notify the HPTN LC via email epiwowa jhmi edu when the shipment has been picked up from the site by the courier shipping company Attach an electronic copy of the shipping manifest and LDMS batch to the email notification and include the following information in the notification name of courier shipping company shipment tracking number number of boxes shipped date
12. ast 8 times or as specified by manufacturer after specimen collection to prevent clotting e Citrate tubes Light blue top should be gently inverted at least 4 times or as specified by manufacturer after specimen collection to prevent clotting e For plasma storage 20 ml of whole blood should be collected into spray dried EDTA tubes eg BD 366643 or other to yield 5 x 1 8mL plasma HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 6 of 30 Table 11 1 Schedule of Study Visits and Specimen Collection All Participants Mis Oral Phase E Injection and Tail Phase Follow Up Weeks Screening Day 0 Enrollment DOT Visits 1 2 DOT Visit 3 DOT Visit 4 Week 4 First Injection Week 6 Safety Week 8 Safety Week 12 Second Injection Week 20 Third Injection Week 28 Fourth Injection Week 36 Fifth Injection Week 44 Sixth Injection Week 64 Tail Phase Week 76 Tail Phase Week 14 HIV testing Bi NI Syphilis testing pem Hematology CBC with Differential and platelets mm Urine or vaginal swab GC CT testing Cervicovaginal and rectal fluid storage Plasma Storage Plasma for PK testing Whole blood storage for Pharmacogenomic testing HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 7 of 30 1 Following the HIV algorithms described in SSP section 11 3 1 HIV testing does not need to be p
13. cumentation The staff member who collects the samples will ensure that the visit code specimen collection date and time as well as their initials or code is documented More detailed information about the labeling procedures must be provided in the site s Chain of Custody SOP When specimens are tested at the laboratories any additional labeling required for in country specimen management or chain of custody will be performed in accordance with site specific SOPs Stored specimens will be entered into the LDMS and labeled with LDMS generated labels 11 2 1 Local Specimen Processing and Storage For samples that are processed and stored locally each sample will be labeled and entered into the LDMS 11 2 2 Local Specimen Testing Sites will follow local testing arrangements for the collection and testing of samples this will be described in the site SOPs AII lab results must be recorded following local guidelines 11 2 8 Remote Specimen Testing Samples that will be sent to the HPTN LC will be labeled and entered into the LDMS 11 2 4 Use of the LDMS LDMS must be used at all sites to track specimens that will be tested stored or shipped off site for testing Detailed instructions for use of LDMS are available in the LDMS User Manual HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 2 of 30 https www fstrf org apps cfmx apps ldms ldmsManual webhelp in
14. cy report for each site Sites are expected to resolve all discrepancies within one week of receipt of the report The HPTN LC is responsible for reminding sites to adhere to the one week timeframe and for following up with sites that do not resolve discrepancies within one week The HPTN SDMC reviews the discrepancy reports for critical samples e g plasma needed for confirmatory HIV testing that appear to be missing and works with the LC and site staff to undertake appropriate corrective action All corrective action should be documented in paper based clinic and or laboratory records as appropriate and entered in the details section of LDMS The LC and SDMC will discuss and document any items that although resolved appear unresolvable in LDMS Any corrections to the LDMS need to be made following guidelines provided by FSTRF The LDMS system will back up the LDMS data daily It is the site s responsibility to transfer the backup file to an external device following procedures outlined by LDMS user support 11 2 6 LDMS Reconciliation All sites must follow the HPTN LC approved site specific SOP for regular reconciliation and verification of specimens that are stored these SOPs must be followed throughout the study In the event that the required volume or number of sample aliquots is not obtained at any time point designated site clinic and lab staff must immediately inform the HPTN CORE HPTN SDMC and LC The HPTN CORE SDMC and LC will
15. dex html As of the date of this version of the SSP the current version of LDMS is Version 9 0 1 All sites should upgrade to this version as soon as possible All sites must use the HPTN barcode label format in order to ensure that both the specimen ID and the global specimen ID assigned to each specimen are printed on LDMS generated labels An example of a two dimensional LDMS generated barcode label is below 50008000009 FEQO043F 01 999515640 057 03 Jan 2005 08 00 BLD EDT PL2 N A 1 00 ML 0 Scr Row 1 LDMS Specimen ID Row 2 Global Specimen ID Row 3 Patient Identifier ID1 and Study Protocol Identifier ID2 Row 4 Specimen Date or Harvest Date and Specimen Collection Time Row 5 Primary Type Additive Type Derivative Type and Sub Additive Derivative Type Row 6 Volume Volume Unit and Visit Visit Unit VID Row 7 Other Specimen ID Questions related to use of LDMS for HPTN 076 should be directed to Paul Richardson prichal 8 jhmi edu Technical support for the general use of LDMS is available from Frontier Science LDMS User Support at Frontier Science Regular Hours Monday to Friday 12 00 AM to 6 00 PM Eastern Standard Time Off hours See below Email ldmshelp fstrf org Phone 1 716 834 0900 extension 7311 HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 3 of 30 Fax 1 716 832 8448 should be used to fax Installation Reports
16. ditional plasma storage HIV Viral Load 1 Following approved HIV testing algorithm shown in figure 11 3 HIV rapid testing may be performed in the clinic or the laboratory 2 Sites may collect specimens for resistance testing at a local laboratory to assist with clinical management results from resistance testing performed at local laboratories will not be reported to the SDMC Stored plasma may not be used real time local resistance testing 3 Obtained from 20mL of EDTA whole blood that has been drawn at the same time and date as the sample for HIV testing See SSP Section 11 4 1 for specimen processing details HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 14 of 30 11 3 2 Hepatitis Testing Testing for HBV and HCV will be performed at Screening Sites will follow local testing arrangements for the collection and testing of samples this will be described in the site SOPs Test results are required for the enrollment visit 11 3 3 Safety Testing CBC and Chemistry testing will be performed at various time points throughout the study Additional potassium and magnesium testing will be performed at the screening visit Sites will follow local testing arrangements for the collection and testing of samples this will be described in the site SOPs Test results from the screening visit are required prior to enrollment Same day test results are not required prior t
17. each test is read and verified at appropriate time points Documentation is required for the testing start and stop times as well as result verification times These must be recorded on testing log sheets If a participant has a reactive or positive HIV test at any time after enrollment additional blood draw and testing is required as detailed in Table 11 3 HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 10 of 30 Figure 11 1 HIV Testing Algorithm at the Screening Visit HIV TestingAlgorithm at Screening All Participants Non reactive Non reactive RNA Screen US FDA cleared 4th Generation HIV EIA for acute HIV HIV Rapid Test infection Reactive Reactive Reactive Non reactive This individual is not eligible for enrollment if any HIV test is reactive positive Follow This individual is eligible local testing guidelines to determine HIV to attend the Enrollment infection status visit based on HIV status NOTES All site specific HIV testing plans must be documented and approved by the HPTN LC before the study opens A single reactive positive result from any HIV test is sufficient to exclude a participant at Screening For example if an HIV rapid test in the clinic is reactive the 4 generation HIV EIA and RNA Screen do not need to be performed If a participant is not eligible for enrollment because of a reactive posi
18. ecimens until all of the protocol specified testing including assessments at the HPTN LC has been completed and the primary research paper has been published LDMS Entry LDMS Specimen Code for Plasma Storage Primary Additive Derivative Sub Add Deriv Test LDMS Code Plasma ii EDT PL2 N A Storage Codes used in table BLD Blood EDT EDTA PL2 Plasma Double Spun N A Not Applicable Other Spec ID Not Applicable HPTN 076 SSP Manual All plasma vials are stored in the LDMS and in a 70 C to 90 C freezer Specimens will be shipped to HPTN Laboratory Center LC when requested Version 1 0 Section 11 Laboratory and Specimen Management Procedures April 17 2015 Page 18 of 30 All enrolled study participants must consent to collection and storage of their plasma for the duration of their study participation and until all protocol specified testing has been completed Participants are asked to consent separately to indefinite storage and possible future research testing of their plasma after the study is completed Participants may refuse to consent to indefinite storage and possible future research testing and still enroll in the study After all protocol specified testing has been completed the stored plasma of participants who do not consent to indefinite storage and possible future research testing must be destroyed After all protocol specified testing has been completed the
19. ein and glucose this testing may be performed in the clinic or the laboratory 6 Urine or vaginal swab for GC CT will be performed at Screening The choice of sample type is site dependant 7 Cervicovaginal and rectal fluid collection can be done at week 36 preferred or week 44 For participants in the Tissue Subset fluid collection and storage must be performed at the same visit as the vaginal tissue biopsy week36 preferred or week 44 See Table 11 2 All samples for PK plasma vaginal tissue cervicovaginal fluid and rectal fluid should be collected within a 4 hour window 8 Plasma for storage and for PK can be obtained from 20mL of EDTA whole blood that has been drawn at the same time and date as the sample for HIV testing See SSP Section 11 4 1 for specimen processing details At Enrollment day 0 and visits DOT visit 3 Weeks 2 4 12 20 28 36 and 44 blood for PK must be drawn prior to the administration of study product 9 Store 4 mL of EDTA whole blood for pharmacogenomics analysis The whole blood specimen must be entered into LDMS and stored frozen See section 11 4 2 for specimen processing details HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 8 of 30 Table 11 2 Additional Procedures for Participants in the Tissue Subset US Sites Only Injection and Tail Phase Follow Up Weeks Screening DOT Visits 1 4 Week 2 Oral Run in Safety Visit Week
20. equent visits if a woman had a positive pregnancy test at a previous visit and is still pregnant Sites will follow local testing arrangements for the collection and testing of samples this will be described in the site SOPs The test result from the screening visit is required prior to enrollment Same day test results must be available and confirmed to be negative PRIOR to the provision or continuation of study product This is a requirement at all visits at which study product is to be administered or continued enrollment weeks 4 12 20 28 36 and 44 Study product must NOT be given if the pregnancy test obtained at this or the prior visit is positive Study product must NOT be administered if the participant is pregnant This must be based on pregnancy test results from previous visits and on the result of the pregnancy test performed at the current study visit 11 3 7 Urinalysis Testing Sites will follow local testing arrangements for the collection and testing of urine for urinalysis protein and glucose This will be described in the site SOPs Results from the enrollment visit are not required prior to enrollment 11 3 8 Urine or Vaginal Swab for GC CT Testing and Syphilis Testing Sites will follow local testing arrangements for the collection and testing of Urine or Vaginal swabs for GC CT and plasma for syphilis testing This will be described in the site SOPs Results from the screening visit are required prior
21. erformed after confirmation of HIV infection based on results from samples collected on two separate dates At the oral dosing visit Day 0 and injection visits Weeks 4 12 20 28 36 and 44 at least one same day negative or non reactive HIV test result must be obtained PRIOR to administering the study product 2 Hepatitis testing includes hepatitis B surface antigen HBsAg hepatitis B surface antibody HBsAb and hepatitis C antibody HCAb 3 Chemistry testing includes total bilirubin CPK alkaline phosphatase creatinine AST ALT total protein glucose calcium and phosphorous 4 Pregnancy testing may be performed in the clinic or the laboratory at all visits where this testing is indicated Testing may be performed using a urine plasma or serum sample The assay used for pregnancy testing must have a limit of detection of 25 mIU mL or lower Women not of childbearing potential are excluded from pregnancy testing throughout their participation in this study Pregnancy testing is not required at subsequent visits if a positive result is obtained At the first oral dosing visit Day 0 and injection visits Weeks 4 12 20 28 36 and 44 a pregnancy test must be performed and pregnancy must be ruled out PRIOR to administering the study product Urine pregnancy testing may be performed in the clinic or the laboratory Pregnancy testing 1s not required if a positive result was obtained at a prior visit 5 Urinalysis includes prot
22. erformed at the HPTN LC or a laboratory designated by the HPTN LC This testing will be performed retrospectively at the end of the study If real time resistance testing is needed for clinical management that testing should be arranged by the site outside of the study separate specimens should be collected for that testing For sites that do not have the capacity for local resistance testing for clinical care results from resistance testing may be provided at the end of the study at the request of the site IoR with approval of the HPTN LC and Protocol Chair Results from specialized resistance testing e g minority variants analysis if performed will not be returned to study sites 11 6 Laboratory Monitoring LC staff will conduct periodic site visits to review in clinic documentation LDMS reports specimen storage and other laboratory documentation relevant to this protocol HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 30 of 30
23. ght samples must be placed in a 80 C freezer or placed in dry ice within an hour of collection Samples placed in dry ice should be transported for final storage in a 80 C freezer as soon as is possible HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 21 of 30 Weigh the Dacron swabs and capped conical tubes again using the same analytical balance used for the pre weigh 10 Create an entry for each participant in LDMS print aliquot labels and label conical tubes The following LDMS codes must be used for cervicovaginal fluid Primary Code VAG Additive NON Derivative FLD Sub Add Deriv NON Enter the appropriate weight of the fluid in the Volume field and time of collection If a negative value for weight is obtained please add this information as a comment The following LDMS codes must be used for rectal fluid Primary Code REC Additive NON Derivative FLD Sub Add Deriv NON Enter the appropriate weight of the fluid in the Volume field and time of collection If a negative value for weight is obtained please add this information as a comment 11 Place the conical tubes in a 80 C freezer for storage until shipment is requested by the LC 12 Record the time that the sample is introduced to the freezer Time Interval between Collection and Freezing Time unit hours HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen M
24. he primary pharmacologic assessments will be performed using assays that have been validated and approved by the Clinical Pharmacology Quality Assurance CPQA Committee Results will not be returned to the sites or study participants Stored plasma may also be tested for the presence of other ARV drugs or other substances 11 5 3 Pharmacogenomic Testing A whole blood specimen for pharmacogenomic analysis will be collected at the enrollment visit Samples will be stored on site for shipment to the HPTN LC upon request Assays will be performed at the HPTN LC Results will not be returned to the sites or study participants HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 29 of 30 11 5 4 Other testing The HPTN LC will perform QA testing including testing to determine HIV infection status in selected cases Additional assays may be performed at the HPTN LC or a laboratory designated by the HPTN LC This testing may include the following tests for participants who acquire HIV infection HIV viral load HIV resistance testing HIV subtyping and other tests to characterize HIV viruses and or the host response to HIV infection Results will not be returned to the sites or study participants with the exception of HIV testing if results obtained at the HPTN LC do not agree with site results and the exception for resistance test results noted below Resistance testing will be p
25. issue for PK ideally 2 biopsies minimum 1 e Required Materials and Equipment 2 mL Cryovial Corning 430659 or equivalent e Medium Tischler forceps Gynex Order 1008 W or equivalent e Storage box Fisher Scientific Cat No 03 395 464 supplied with insert e Clean pointed forceps e Petridish Falcon 35 3803 or equivalent e Analytical Balance e Liquid Nitrogen or Dry Ice Alcohol bath Transport medium See below Vaginal biopsies should be collected using medium Tischler forceps 3 mm x 5 mm bite size Gynex Order 1008 W Vaginal biopsies should be placed into a suitably labeled 50mL conical tube specimen pot that contains 30mL of the following Transport Medium HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 24 of 30 Transport Medium Volume per 100mL RPMI Medium 1640 w L glutamine w HEPES 90 5mL Invitrogen Cat 22400 105 Fetal Bovine Serum Invitrogen Cat 10082 147 final 7 5mL 7 594 Antibiotic Antimycotic Invitrogen 100X Cat 4 15240 ImL 104 final 196 50mg mL Zosyn Wyeth NDC 0206 8852 16 ImL Prepare transport medium in a biological cabinet using sterile conditions and place in a suitable sterile container Store at 4 C for up to 14 days Discard any unused portion Transport the samples at ambient temperature from the biopsy suite to the location where the sample will be weighed Transport
26. may be enrolled and the oral drug may be given Non reactive HIV seronegative Before providing study drug the site must ensure that the HIV rapid test from the Enrollment visit and all HIV results from the Screening visit are available and are negative or non reactive If acute HIV infection is suspected the participant is not eligible for Enrollment at this time In this case the site should follow the algorithm but should also send sample for an RNA test that in the opinion of the site investigator is able to detect early HIV infection If possible the site should select an assay that is FDA cleared for early HIV diagnosis such as the APTIMA HIV 1 RNA Qualitative Assay Contact the HPTN LC Site PI and Protocol Chair for additional guidance HPTN 076 SSP Manual Section 11 Laboratory and Specimen Management Procedures Version 1 0 April 17 2015 Page 12 of 30 Figure 11 3 HIV Testing Algorithm at Follow Up Visits HIV TestingAlgorithm atFollow up Visits 4 generation HIV EIA Study drug may be provided before this result is available All Participants U S FDA cleared HIV Rapid Test Non Non reactive Reactive Reactive reactive 3 T a HIV HIV seronegative Possible HIV infection 3 Immediately consult the Protocol Chair seronegative This result and all HIV test results from prior visits must be non reactive negative before any further
27. o the issue of study product 11 3 3 1 Creatinine Clearance Calculated creatinine clearance will be performed using the Cockcroft Gault formula for females eCcr female in mL min 140 age in years x actual body weight in kg x 85 72 x serum creatinine in mg dL 11 3 4 Coagulation Testing US sites only Coagulation testing PT INR and aPTT will be performed at the week 6 visit only for participants in the Tissue Subset This testing can also be done at later visits but results must be available for enrollment into the Tissue Subset Sites will follow local testing arrangements for the collection and testing of samples this will be described in the site SOPs 11 3 5 PapTest US sites only This will be offered at week 8 to participants who elect to enroll into the tissue subset if a satisfactory Pap smear is not documented within the last 12 calendar months See protocol section 3 1 1 for details This can also be done at later visits but results must be available for enrollment into the Tissue Subset Sites will follow local testing arrangements for the collection and testing of Pap smears HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 15 of 30 11 3 6 Pregnancy Testing All women of reproductive potential will have a BHCG test for pregnancy sensitivity of lt 25 mIU mL at each visit listed in table 11 1 Pregnancy testing is not required at subs
28. of shipment and expected date of arrival The following types of specimens will be shipped to the HPTN LC for testing HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 28 of 30 11 5 1 HIV QA testing Selected plasma aliquots will be shipped to the HPTN LC for HIV QA testing according to the HPTN Manual of Operations additional testing may be performed e g ABO typing When samples are received at the HPTN LC the LC will perform additional QA and HIV testing This will include e Quality assurance testing to confirm results of in country testing e Testing to confirm seroconversion events Data from the HPTN LC will be submitted to the SDMC 11 5 2 Pharmacology Testing Plasma samples for drug levels will be collected beginning at the enrollment visit Day 0 and at weeks 4 through week 76 These samples will be collected from all participants although PK testing may be limited to a subset of the samples At these visits the blood will be collected PRIOR to the administration of study product The actual date and time of each blood sample collection will be recorded as well as the time of each injection This information should be captured on the relevant CRF Specimens for pharmacology testing will be stored on site for shipment to the HPTN LC upon request Pharmacology testing will be performed at the HPTN LC or at an outside laboratory designated by the HPTN LC T
29. only LDMS User Support can be contacted during off hours as well as on weekends on U S holidays by completing the LDMS help form on the Frontier Science portal This form can be found on the portal by clicking the Contact LDMS User Support link You will need a portal account to access this form While it is preferred that users use the Contact LDMS User Support link on the portal there may be times when you need immediately assistance during off hours and cannot access the portal In these situations you can contact LDMS User Support by emailing the pager email addresses directly Pager 1 Idmspager 1 fstrf org Pager 2 Idmspager2 fstrf org Pager 3 Idmspager3 fstrf org Try pager 1 first If you do not receive a response within 15 minutes try pager 2 and then finally pager 3 When you contact LDMS user support there are certain pieces of information that you can provide to help them better respond to your question Please provide the following information in your email support 1 Your name 2 Your laboratory s LDMS ID number This is a 3 digit number assigned by Frontier Science to uniquely identify your laboratory It appears when you start LDMS and can also be found in the bottom right corner of the screen 3 A full explanation of the issue Your explanation should include any error messages or error numbers that appeared what you were doing in LDMS at the time the issue occurred and steps needed to reprod
30. psy was placed in the freezer e Biopsy should be frozen within one hour of collection Biopsy should be shipped upon request to the HPTN Laboratory Center at JHU Copies of the completed study forms should also be sent upon request LDMS Specimen details must be entered into LDMS Cryovials must be stored using labels produced from the LDMS The following LDMS codes must be used LDMS FIELD Primary code Additive Derivative Subderivative Volume Units Time Time unit Other Spec ID Comments Code Information to be entered VAG BTM VAG N A Weight of biopsy in mg mg Interval between Collection and Freezing hours PK Note any contamination or other details HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 26 of 30 Figure 11 5 Example of Study Form for Vaginal Biopsy Storage Storage of Vaginal Biopsy for PK Participant ID Number PTID Date of Biopsy Collection Visit Number Time actual 24 h BIOPSY ID Weight Milligrams format Time interval LDMS aliquot ID Cryovial Cryovial Biopsy Collection Freezing collection to freezing Biopsy hh mm hh mm Biopsy 1 Biopsy 2 NM Serial Number of Balance Used Weighing Performed By HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 27 of 30 11 5 Shipping of Samples to the HPTN LC Each site will ship pl
31. scribed in Figures 11 1 11 3 For further help on implementing the HIV testing algorithm seek guidance from the HPTN LC Whole blood will be collected according to site specific procedures Participants with one or more reactive HIV test results at either the screening or enrollment visit will not be eligible for enrollment regardless of subsequent test results RNA testing for acute HIV infection must be performed within 28 days prior to Enrollment The Protocol Chair Site PI CMC and HPTN LC must be notified immediately if one or more reactive HIV test results are obtained at any follow up visit after enrollment This includes the result of the 4 generation EIA test that is collected at the enrollment visit Additional HIV testing may be performed at any time at the discretion of the site investigator HIV infection must be confirmed using two independent samples collected on different days Plasma storage is required at every visit at which HIV testing is performed All tests and associated QC procedures must be documented on local laboratory log sheets or other laboratory source documents Kit lot numbers and expiry dates must also be documented All staff involved in HIV testing and verification of HIV test results should be aware of the testing time frame for the HIV test so that all tests are performed and verified within the specified time frame Place appropriate timekeeping devices in all test settings to ensure that
32. study product is given Site PI and HPTN LC Follow local testing guidelines and consult the HPTN LC to determine HIV infection status Do not administer any further study product without approval from the Protocol Chair Site Pl and HPTN LC NOTES At any visit where study product will be given the site must ensure that the HIV rapid test from this visit and all HIV results from prior study visits are negative or non reactive If acute HIV infection is suspected do not administer any further study product Immediately consult the Protocol Chair Site Pl PSRT and HPTN LC In addition to following the algorithm above the site should send sample for an RNA test that in the opinion of the site investigator is able to detect early HIV infection If possible the site should select an assay that is FDA cleared for early HIV diagnosis such as the APTIMA HIV 1 RNA Qualitative Assay The site should contact the HPTN LC Site PI and Protocol Chair for additional guidance once all of the test results from this visit including the HIV RNA test are available HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 13 of 30 Table 11 3 Additional Procedures for Participants who have a Reactive or Positive HIV test at any Time after Enrollment HIV confirmation visit following a reactive or positive HIV result HIV testing CD4 cell count HIV resistance testing Ad
33. tive test result the site should follow local testing guidelines to determine HIV status Screening for acute infection should be performed using an RNA test that in the opinion of the site investigator is able to detect early HIV infection If possible the site should select an assay that is FDA cleared for early HIV diagnosis such as the APTIMA HIV 1 RNA Qualitative Assay RNA test results must be obtained from a specimen collected within 28 days prior to enrollment HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 11 of 30 Figure 11 2 HIV Testing Algorithm at the Enrollment Visit HIV Testing Algorithm at Enrollment 4th generation HIV EIA All Participants U S FDA cleared HIV Rapid Test before this result is available Reactive If result back before enrollment Reactive Non reactive Reactive If result back after enrolled Possible HIV HIV seronegative infection gossie HIV ee The individual is eligible for This individual is not enrolled immediately ole the enrollment only if this result eligible for Protocol Chair Site Pl and and all HIV test results from enrollment Follow RR bi ba N the Screening visit are local testing LC to determine HIV infection available and are non guidelines to status Donot administer an reactive negative determine HIV antl DUE Brecon Sialis Site PI and HPTNLC NOTES The participant
34. uce the issue The more details that you can provide the faster LDMS User Support can help you 4 How you want to be contacted If you want LDMS user support to call a specific telephone number please provide that number and extension 5 If applicable The license code or challenge code being generated by LDMS Note If you are contacting user support about a license or challenge code do not close the window with the code Doing so will cause LDMS to generate a new code Below are a few other details that can also be helpful to include in your email HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 4 of 30 1 Have there been any recent changes to the computer with LDMS such as new hardware installed a firewall upgrade a network name change or another change 2 Are you or another user able to repeat the issue 3 Ifyou have LDMS installed on multiple computers does the issue occur on all of them or does it only occur on a specific computer 11 2 5 LDMS exportation discrepancies and backup Each site must export its LDMS data to Frontier Science FSTRF at a minimum on a weekly basis or whenever changes or additions are made to the LDMS database Exported data are used by the HPTN SDMC to generate a discrepancy reports comparing the data from the LDMS with that entered onto the CRFs Any discrepancies identified during the reconciliation are included in a discrepan
35. vent leakage during shipping 6 Ensure PTID date visit number and laboratory identifier are on the LDMS label 7 Store whole blood ina freezer at minus 70 C or colder until requested for shipment 8 Ship when requested on dry ice for arrival on Monday through Friday only site must follow appropriate shipping regulations 9 Batch shipment to Estelle Piwowar Manning Johns Hopkins University Hospital Department of Pathology Pathology Building Room 313 600 North Wolfe Street Baltimore MD 21287 USA 11 4 3 Cervicovaginal and Rectal Fluid Processing and Storage Collection and Storage of Fluid Using a Dacron Swab This procedure outlines the preparation of Dacron swabs to be used for the collection of cervicovaginal fluid and rectal fluid Note It is important that the rectal fluid be collected before an enema is performed if performed It is important that cervicovaginal fluid be collected before the collection of vaginal tissue select participants at US sites only The following order of collection is recommended rectal fluid cervicovaginal fluid vaginal tissue Required Materials and Equipment e Two Dacron Swabs Fisher Scientific Cat No 22 029 574 HPTN 076 SSP Manual Version 1 0 April 17 2015 Section 11 Laboratory and Specimen Management Procedures Page 20 of 30 Two 15 ml Conical tubes with caps Fisher Scientific Cat No 14 959 49B or equivalent Storage box Fisher Scientific Cat No
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