Critical GMP Validations for Drugs/Biologics Ⅰ Chung Keel Lee, Ph.D. Special Advisor to the Minister, MFDS Advisor, WHO Adjunct / Invited Professor, KU / EWU President, ISPE Korea July 19, 2013

Validation Definition: Establishing documented evidence which provides a high degree of assurance that a specific process will consistently produce a product meeting its pre-determined specifications and quality attributes. Method: Prospective Validation Retrospective Validation Concurrent Validation

Validation Elements of Validation Validation Report Revalidation Equipment validation Design Qualification Installation Qualification Operational Qualification Process validation Performance Qualification Validation Report Revalidation

Revalidation When there are changes in : Periodic revalidation Critical processes Equipment Materials Product characteristics Periodic revalidation Gradual process changes over a period of time Wear of equipment

Validation Validation Steps Validation Master Plan Validation Protocol Study Validation Report Writing SOP Step by At least 3 times consecutively Documented plan Easy to follow Conclusion

Validation Personnel qualification Analyist performance in laboratories Personnel following critical procedures Personnel doing data entry in computerized systems

Validation Steam-sterilization process using autoclave Calibration/certification Empty chamber heat distribution studies Heat penetration studies on various loading configurations Challenge studies with live bacterial organisms Depyrogenation process using hot-air oven General validation study procedures are similar to the steam-sterilization process validation except the challenge study. Endotoxin is used for the challenge study. Depyrogenation process via washing/rinsing All aseptic processes

Sterilization : Introduction 5 major methods for sterilization Moist heat Dry heat Ethylene oxide gas Ionizing radiation Filtration

Sterilization : Introduction Validation of sterilization – Biological Indicators Sterilization method Steam heat Dry heat Ethylene oxide Ionizing radiation Filtration Biological indicator Bacillus stearothermophilus B. subtilis var. niger B. subtilis var. globigii B. pumilus Brevundimonas diminuta

Sterilization : Introduction Validation of sterilization - Quantitation of indicator removal Heat or ETO Radiation 10 2 10 3 10 4 10 5 10 6 NO. Survivors (log) Time(min) 10 2 10 3 10 4 10 5 10 6 NO. Survivors (log) rads - D value = the amount of time (for heat or ETO) or dose (for radiation) to reduce the organisms by one order of magnitude

Sterilization : Introduction D Values Biological indicators B. Stearothermaphilus B. subtilis var.niger B. subtilis var.globigii B. pumilus Sterilization Process Saturated steam at 121 ℃ Dry heat at 170 ℃ Ethylene oxide at 54 ℃ 600mg/l, 50% RH Gamma radiation : wet Gamma radiation : dry D value 1.5 min 1.0 min 3.0 min 0.2 Mrad 0.15 Mrad

Validation of Autoclave Steam Sterilization : General Assurance of at least 10-6 probability of survival 2 basic approaches The probability of survival approach Heat labile products Overkill approach Heat stable products

Validation of Autoclave Steam Sterilization : Probability of Survival Approach Bioburden & Heat Resistance studies Calibration of biological indicators Validation Heat distribution Heat penetration Biological challenge Reproducibility of the sterilization cycle

Validation of Autoclave Steam Sterilization : Overkill Approach No need for bioburden & resistance studies At least a 12-log reduction of microorganisms having a D value of at least 1 min The most resistant isolates from production environment have D121 values ranging from 0.5 to 1.0 min. Calibration of biological indicators Validation

Validation of Autoclave Steam Sterilization : D value Survivor curve (microbial death rate curve) NO. Survivors (log) 10 2 10 3 10 4 10 5 10 6 1 2 3 4 5 6 7 8 9 D value at constant temp = 1 time (min) Dt = time at a given temperature(t) required to reduce the No.of microorganisms by 90%(1 logarithmic reduction)

Validation of Autoclave Steam Sterilization : Z value Thermal Resistance Curve 100 10 1.0 0.1 110 115 120 125 130 135 Z = 10℃ D value (log) Temperature - Z = the No. of degrees temperature change to change the D value by a factor of ten

Validation of Autoclave Steam Sterilization : Z value Steam sterilization : B. stearothermophilus - Base temperature : 121 ℃ Z = 10 ℃ in the range of 110~135 ℃ Dry heat sterilization : B. subtilis - Base temperature : 170 ℃ Z = 20 ℃ in the range of 150~190 ℃ Dry heat depyrogenation : Endotoxin - Base temperature : 250 ℃ - Z = 50 ℃ in the range of 225~300 ℃

Validation of Autoclave Steam Sterilization : Typical Cycle Produced by a Sterilizer

Validation of Autoclave Steam Sterilization : Lethal Rate where = temperature within the container(item) being heate = process temperature(reference temperature) Lethal rate is the equivalent time for any specific temperature relative to another temperature. ex) minutes at 121.1℃( ) per minute at 119℃( )

Validation of Autoclave Steam Sterilization : Lethal Rate Example The lethal rate for 117.0 ℃ relative to 121.1 ℃ (assuming a Z value of 10) : This means that for every full minute of the process at a temperature of 117 ℃, the process is credited with the equivalent of 0.389 min at 121.1 ℃.

(minutes at 121.1℃ per minute at T℃) Validation of Autoclave Steam Sterilization : Lethal Rate Lethal Rate Table (minutes at 121.1℃ per minute at T℃) tempera -ture (℃) Z value(℃) 7 8 9 10 11 12 100 0.001 0.002 0.005 0.008 0.012 0.017 101 0.003 0.006 0.010 0.015 0.021 102 0.004 0.018 0.026 103 0.023 0.031 104 0.007 0.013 0.019 0.028 0.038 105 0.016 0.025 0.034 0.046 106 0.042 0.055 107 0.027 0.039 0.052 0.067 108 0.035 0.049 0.064 0.081 109 0.045 0.062 0.079 0.098 110 0.041 0.058 0.078 0.119 111 0.036 0.075 0.121 0.144 112 0.050 0.073 0.097 0.123 0.149 0.174 113 0.070 0.126 0.155 0.184 0.211 114 0.130 0.163 0.195 0.226 0.256 114.5 0.114 0.150 0.185 0.219 0.251 0.282 115 0.134 0.173 0.210 0.245 0.279 0.310 115.5 0.158 0.200 0.239 0.275 0.341 116 0.187 0.230 0.271 0.309 0.344 0.376 116.5 0.220 0.266 0.308 0.347 0.382 0.414 117 0.260 0.307 0.350 0.389 0.424 0.455 117.5 0.306 0.355 0.398 0.437 0.471 0.501 tempera -ture (℃) Z value(℃) 7 8 9 10 11 12 118 0.361 0.410 0.452 0.490 0.523 0.552 118.5 0.425 0.473 0.514 0.550 0.580 0.607 119 0.501 0.546 0.584 0.617 0.644 0.668 119.5 0.591 0.631 0.664 0.692 0.715 0.736 120 0.696 0.729 0.755 0.776 0.794 0.810 120.5 0.821 0.841 0.858 0.871 0.882 0.891 121 0.968 0.972 0.975 0.977 0.979 0.981 121.5 1.141 1.122 1.108 1.096 1.087 1.080 122 1.345 1.296 1.259 1.230 1.207 1.189 122.5 1.585 1.496 1.431 1.380 1.341 1.308 123 1.868 1.728 1.626 1.549 1.488 1.440 123.5 2.202 1.995 1.848 1.738 1.653 124 2.596 2.304 2.100 1.950 1.835 1.744 125 3.607 3.073 2.712 2.455 2.262 2.113 126 5.012 4.097 3.503 3.090 2.789 2.561 127 6.964 5.464 4.524 3.890 3.438 3.102 128 9.676 7.286 5.843 4.898 4.239 3.758 129 13.445 9.716 7.547 6.166 5.226 4.553 130 18.682 12.957 9.747 7.762 6.443 5.517

(minutes at 121℃ per minute at T℃) Validation of Autoclave Steam Sterilization : Lethal Rate Lethal Rate Table (minutes at 121℃ per minute at T℃) tempera -ture (℃) Z value(℃) 7 8 9 10 11 12 100 .001 .002 .005 .008 .012 .018 101 .003 .006 .010 .015 .022 102 .004 .013 .019 .026 103 .016 .023 .032 104 .007 .020 .028 .038 105 .017 .025 .035 .046 106 .043 .056 107 .040 .053 .068 108 .014 .024 .036 .050 .066 .083 109 .063 .081 .100 110 .042 .060 .079 .121 111 .037 .077 .123 .147 112 .052 .075 .126 .152 .178 113 .072 .129 .158 .187 .215 114 .133 .167 .200 .231 .261 114.5 .118 .154 .190 .224 .257 .287 115 .139 .251 .285 .316 115.5 .164 .205 .245 .282 .348 116 .193 .237 .278 .351 .383 116.5 .228 .274 .355 .390 .422 117 .268 .359 .398 .433 .464 117.5 .365 .408 .447 .481 .511 tempera -ture (℃) Z value(℃) 7 8 9 10 11 12 118 .373 .422 .464 .501 .534 .562 118.5 .439 .489 .527 .593 .619 119 .518 .599 .631 .658 .681 119.5 .611 .649 .708 .731 .750 120 .720 .774 .794 .811 .825 120.5 .848 .866 .880 .891 .901 .909 121 1.00 .100 121.5 1.18 1.16 1.14 1.12 1.11 1.10 122 1.39 1.33 1.29 1.23 1.21 122.5 1.64 1.54 1.47 1.41 1.37 123 1.93 1.78 1.67 1.59 1.52 123.5 2.28 2.05 1.90 1.69 1.62 124 2.68 2.37 2.15 2.00 1.87 125 4.39 3.16 2.78 2.82 2.31 126 5.18 4.22 3.59 2.85 2.61 127 7.20 5.62 4.64 3.98 3.51 128 10.0 7.50 6.00 5.01 4.33 3.83 129 13.9 7.74 6.31 5.34 130 19.3 13.3 7.94 6.58

Validation of Autoclave Steam Sterilization : F value The accumulated lethality over the duration of the sterilization process (thermal death time) where F = the accumulated lethality over the data collection period t0 = the start of the data collection period tE = the end of the data collection period T = the temperature at each measurement point Tb = the base temperature at which the Z was determined Z = the z value within the process temperature range dt = the time interval between temperature data ·

Validation of Autoclave Steam Sterilization : F0 value For steam sterilization where F0 = the accumulated lethality over the data collection period t0 = the start of the data collection period tE = the end of the data collection period T = the actual temperature 121 = the usual base temperature for steam sterilization 10 = the z value usual for steam sterilization t = the time interval between temperature data · It assumes that the temperature is measured every minute.

Accumulated Lethal Rates : F0 (in minute) : 14.22 Validation of Autoclave Steam Sterilization : F0 Calculation from Temperature Data (Temperature measured every minute) Temperature T-121 (T-121)/10 10(T-121)/10 ∑ 101 -20 -2 0.01 111 -10 -1 0.10 0.11 121 1.00 1.11 124 3 0.3 2.00 3.11 131 10 1 10.00 13.11 14.11 14.21 14.22 Accumulated Lethal Rates : F0 (in minute) : 14.22

Validation of Autoclave Steam Sterilization : F0 Calculation from Temperature Data (Temperature measured every minute)

Validation of Autoclave Steam Sterilization : Probability of Nonsterile Unit (PNSU) · where B = probability of nonsterile unit A = No. of presterilization bioburden(BB) organisms F0 = the minimul F0 observed in the sterilization cycle D121 = D value of the BB organism

Validation of Autoclave Steam Sterilization : Probability of Nonsterile Unit (PNSU) Example from the previous Temperature Data : If the container has maximum of 100 organisms with D value of 0.5 min. at 121 ℃, This means that the probability of a non-sterile unit is approximately 1 in 1026 units.

Validation of Autoclave Steam Sterilization : Spore Log Reduction(SLR) where A = initial No. of spores B = the No. of spores surviving the heat treatment ·

Validation of Autoclave Steam Sterilization : Minimum Required F0 Overkill approach requires the lethality sufficient to provide a 12-log reduction of microorganisms having a D value of 1.0 Minimum Required F0 should be 12 minutes or more ·

Validation of Autoclave Steam Sterilization Example of Minute-by-Minute Temperature Observations of a Sterilizer Cycle with a 5-min Hold Time Along with the Incremental and Cumulative F (Lethal Rate) Values Time (min) Temperature (℃) Lethal rate Total lethal rate 1 25.0 0.000 2 55.0 3 85.0 4 100.0 0.008 5 110.0 0.079 0.087 6 115.5 0.282 0.369 7 119.0 0.631 1.000 8 121.1 1.00 2.000 9 3.000 10 4.000 Time (min) Temperature (℃) Lethal rate Total lethal rate 11 121.1 1.00 5.000 12 6.000 13 116.0 0.316 6.316 14 108.0 0.200 6.516 15 96.0 0.000 16 80.0 17 65.0 18 42.0 19 30.0 20 25.0

Validation of Autoclave Steam Sterilization