0 2 3 μ g / m l . A f t e r overnight at 4°C, the plate is washed and all wells were blocked with PBS and 1% bovine serum albumin (BSA) (Sigma Chemical Co.), 100 µl/well for 1 hour at room temperature. The protein- coated plate is incubated with 100 μl sera diluted 1:800, 1:1600, 1:3200 and 1:6400 tested in dupli cate for 2 hours at room temperature. One set of duplicate is treated with 8M Urea for 5 minutes. Goat anti -human IgG (gamma) horseradish peroxidase (HRP) (Zymed, So. San Francisco, CA) are added to the wells at a 1:5,000 dilution in PBS/1% BSA and incubated for 45 minutes at room temperature (Amersham Co., Arlington Heights, IL). Following the final wash, TMB (Kirkegaard and Perry Laboratories, Gaithersburg, MD) developing reagent is added. Color reaction is read at an optical density of 650nm until the standard curve well containing 0.3 µg/ml reaches 0.3 OD. More dilute sera will be used if values are at the top or off of the standard curve. The reaction is stopped with 1N HCl and the optical density read at 450nm. Values are calculated in µg/ml bas ed on the standard curve. The values are reported using the 1:800 dilution (no urea treatment) with the assay cut -off value of 200 μg/ml. Results between 100 to 200 μg/ml will need to be confirmed with western blot. To calculate the % avidity index, the absorbance reading after urea wash is divided with absorbance reading without urea wash and multiplied by 100. Random normal blood donors (n=10) are run every 100 plates, or quarterly, whichever comes first, and the standard curve and antibody values compared to mean and standard deviations of the experimental runs. Validation runs will be tabulated and discussed by the IMC and a qu ality assurance report filed with the data manager. 6304p Protocol 118 04/06/2016 1 Appendix C Data and Safety Monitoring Plan (DSMP) Phase I -II Study of Combination Immunotherapy for the Generation of HER -2/neu (HER2) Specific Cytotoxic T Cells (CTL) in vivo A. Purpose To ensur e that the Tumor Vaccine Group clinical research staff (Table 1) follow NIH/NCI/CTEP guidelines with respect to (1) accurate assessment and timely reporting of adverse drug reactions associated with investigational drugs (2) adherence to protocol and (3) a ccurate reporting of data. The DSMP will be attached to all protocols as an appendix and be approved by Human Subjects Division Institutional Review Board (IRB) at the University of Washington, Seattle, WA as part of the IRB protocol approval process. The Principal Investigator is responsible for every aspect of the design, conduct and final analysis of the protocol. Regulations defining the responsibilities for assessment and reporting of all adverse events (AE), serious AE and unexpected AE are defined by the Code of Federal Regulations: 21 CFR 312.32 and Common Terminology Criteria for Adverse Events (CTCAE) Version 3.0 published June 10, 2003 by the Cancer Therapy Evaluation Program (CTEP), a division of the NCI/NIH. A matrix of reporting requirement s and schedules is in the CTEP web -site at http://ctep.infi.nih/gov . This clinical study will rely upon the monitoring of the trial by the Principal investigator in conjunction with a Research Nurse, a Statistician, and an Independent Study Monitor assigned by the Research Trial Office of the University of Washington/Fred Hutchinson Cancer Research Center Cancer Consortium. If an adverse event occurs the Principal Investigator will be notified along with the IRB and appropriate federal agencies: FDA, NCI/CTEP and the Clinical Data Update System and/or the Adverse Event Expedited Reporting System, (AdEERS). B. Objectives 1. To ensure that the Principal Investigator and clinical research staff follow federal and ins titutional regulatory guidelines with respect to timely reporting of adverse reactions associated with investigational drugs. 2. To provide instructions for completion of adverse events reporting forms as follows: a) MED WATCH FDA form 3500A, or b) Adver se Reaction (ADR) Form for Investigational Agents, or c) the Adverse Event Expedited Report for 24 hour phone/fax reporting of adverse events. 3. To define classification of adverse drug reactions as expected or unexpected. 4. To define classification of adverse drug reactions as serious or non- serious. 5. To ensure compliance and accuracy of documentation of adverse drug reactions reportable to the FDA/NIH/CTEP. 6. To ensure compliance and accuracy of reporting and documentation of adverse drug reactions reportable to the NCI by the Clinical Data Update System (CDUS, for NCI -funded trials). C. Adverse Event Reporting Policy and Procedures 1. Evaluation of adverse events (Table 2): Patients are monitored for the development of end organ damage by assessing adverse events with serum chemistries, liver function studies, complete blood counts, urine analysis and physical exams performed every month until 30 days after the final vaccine administration. All adverse events for all systems are graded on a scale of 1 -5 and attribution is assigned, using the NCI Common Terminology Criteria. If an adverse event occurs during more than one cycle of treatment, then only the most severe adverse event is reported. All information pertaining to toxicity is recorded in th e source documents. 2. Definitions of adverse events: 6304p Protocol 118 04/06/2016 2 a. Adverse Event - any unfavorable and unintended sign (including abnormal laboratory finding), symptom or disease temporally associated with the use of a medical treatment or procedure regardless of whether it is considered related to the medical treatment or procedure. b. Expected Adverse Event - an event that may be reasonably anticipated to occur as a result of the study procedure. c. Unexpected Adverse Event - any adverse event that is unanticipat ed. An event that might have been anticipated but is more serious than expected or occurs more frequently than expected, would be considered an unexpected adverse event. d. Serious Adverse Event - grade 4 or 5 toxicity or any adverse event occurring that results in any of the following outcomes: death, a life threatening adverse event, inpatient hospitalization or prolongation of existing hospitalization due to the adverse event, a persistent or significant disability/incapacity or a congenital anomaly/bir th defect. e. Life-threatening Adverse Event - the patient was at substantial risk of dying at the time of the adverse event or it is suspected that the use or continued use of the product would have resulted in the patient’s death. 3. Scale of scoring adv erse events a. Grade 1 = mild adverse event b. Grade 2 = moderate adverse event c. Grade 3 = severe adverse event d. Grade 4 = life -threatening or disabling adverse event e. Grade 5 = death related to adverse event. 4. Attribution of adverse event a. 5 (definite): the adverse event is clearly related to the investigational agent b. 4 (probable): the adverse event is likely related to the investigational agent c. 3 (possible): the adverse event may be related to the investigational agent d. 2 (unlikely): the adverse event is doubtfully related to the investigational agent e. 1 (unrelated): the adverse event is clearly NOT related to the investigational agent. 5. Adverse event reporting is outlined below by regulatory agency (Table 1): University of Washington Human Subjects Division - expected adverse events are routinely reported as part of the annual renewal. Unexpected or more serious than expected adverse events are reportable by phone within 24 hours and followed in writing with a completed Human Sub jects Adverse Effect Report FDA (for trials using an Investigational New Drug (IND) - any unexpected, fatal or immediately life -threatening reaction must be reported to the FDA (Center for Biologics) by telephone or facsimile transmission (using Medwatch Form 3500A) as soon as possible but no later than seven calendar days after initial receipt of the information concerning the event. All unexpected, serious adverse events are reported in writing (using Medwatch Form 3500A) to the FDA (Center for Biologic s) within 15 calendar days after initial receipt of the information concerning the event. Yearly written progress reports to the FDA describe all expected or non- serious unexpected adverse events. NCI (for NCI- sponsored trials only) - adverse reactions t hat are expected and grade 1 -3 or unexpected and grade 1 are reportable in a routine report. Adverse reactions that are unexpected and grade 2 are reportable in writing, using the Adverse Event Expedited Report (AdEERS) within 10 days to the NCI for NCI -sponsored trials, via the Clinical Trials Evaluation Program (CTEP). Adverse reactions that are expected and grade 4 or 5, or unexpected and grade 3, 4 or 5 are reportable by phone/fax, using the Adverse Event Expedited Report (AdEERS) within 24 hours. Te mporary or permanent suspension of an NCI funded clinical trial is to be reported to the NCI grant program director responsible for the grant. 6.