TITLE: Mild Hypothermia to prevent Acute kidney injury in Liver Transplantation (MHALT) Trial - Statistical Analysis Plan for the Interim Analysis, v1.0 
AUTHORS: Michael Bokoch, Claus Niemann, Dieter Adelmann, Rishi Kothari

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block">Acute kidney injury (AKI), or worsening kidney function, is a common complication after liver transplantation (20-90% in published studies). Patients who experience AKI after liver transplantation have higher mortality, increased graft loss, longer hospital and intensive care unit stays, and more progression to chronic kidney disease compared with those who do not. In this study, half of the participants will have their body temperature cooled to slightly lower than normal (mild hypothermia) for a portion of the liver transplant operation, while the other half will have their body temperature maintained at normal. The study will evaluate if mild hypothermia protects from AKI during liver transplantation.</div><div class = "text-block">This study is a single-blinded, randomized controlled trial of mild hypothermia during liver transplantation to provide protection from AKI. Participants will be randomized to normothermia (36.5-37.5 °C) versus mild hypothermia (34-35 °C) during a portion of the liver transplant operation. The protocol is based on preliminary data from rodent models showing that hypothermia protects the kidneys from ischemia-reperfusion injury, as well as studies in deceased organ donors showing that cooling improves post-transplant organ function. Temperature will be maintained with standard techniques plus a minimally-invasive esophageal cooling device that is approved by the U.S. Food and Drug Administration. The investigators hypothesize that mild hypothermia will reduce the incidence and severity of AKI after LTx. Standard surrogates (e.g., change in serum creatinine, need for initiation of dialysis) and biomarkers will be used to assess the severity of kidney injury.</div></div>

[STEPS]
?. [STATISTICAL CONSIDERATIONS]
Statistical Analysis Plan for the Interim AnalysisNCT03534141UCSF IRB # 17-22384Version 1.0May 7, 2020
?. Analysis PopulationsThe following populations will be used for the summaries and analyses of the study data. These populations are defined as follows:Randomized:The Randomized population will consist of all subjects who are randomized to a treatment groupIntention-to-treat (ITT):The ITT population will consist of all randomized subjects not withdrawn from the study. The interim efficacy analysis will be based on the ITT population.Per-Protocol (PP):The PP population will consist of all ITT subjects who complete liver transplant, received the full randomized study treatment, and complete 72 hours of study follow up. Analyses of the PP population will be performed in the final statistical analysis.Intention-to-treat (ITT):The ITT population will consist of all randomized subjects not withdrawn from the study. The interim efficacy analysis will be based on the ITT population.Per-Protocol (PP):The PP population will consist of all ITT subjects who complete liver transplant, received the full randomized study treatment, and complete 72 hours of study follow up. Analyses of the PP population will be performed in the final statistical analysis.Adjustments may be made to refine the definition of the above populations prior to database lock for the final statistical analysis. The final definition of these populations will be documented in a prospectively developed final statistical analysis plan (final SAP) published on Protocols.io. The final SAP must be finalized and approved prior to database lock and unblinding.
?. Statistical HypothesesThe null (H0) hypothesis and the alternative (H1) hypothesis to be tested for the primary endpoint are that:H0: p1 = p2 versus H1: p1 ≠ p2Where p1 is the proportion of the ITT population meeting the primary outcome in the Control Arm (Normothermia), and p2 is the proportion of the ITT population meeting the primary outcome in the Experimental Treatment Arm (Mild Hypothermia).
?. Demographic and Baseline CharacteristicsFor the final statistical analysis, demographic and relevant baseline characteristics will be presented and summarized descriptively by treatment for the Randomized, ITT, and the PP populations. These characteristics will not be included for the interim statistical analysis for efficacy, as they have been summarized and reported elsewhere to the Data Safety Monitoring Board (DSMB). Additional summary of demographic and baseline characteristics may be specified in the final SAP, if deemed necessary.
?. Sample Size Justification and Power AnalysisConsidering one interim analysis for efficacy, and given the incidence of acute kidney injury (AKI) in liver transplantation (LTx) patients at UCSF (68%), we estimate a sample size of 101 subjects per arm (n = 202 total) is needed to detect a 30% reduction in AKI with 80% power at overall alpha = 0.05 (two-tailed).We will aim to enroll 230 total subjects to allow for just over a 10% drop-out rate, since some subjects may not be able to complete the protocol and be withdrawn from the study.
?. [STATISTICAL ANALYSES]
1.5.1. Interim Analysis for EfficacyA single interim analysis for efficacy is planned for this study after 50% of the required patients (101 of the 202 estimated by the power calculation) are accrued after accounting for withdrawn subjects. Based on the O'Brien-Fleming approach to calculate alpha spending, during the interim analysis step, if the p value is p value after 100% enrollment is 1.5.2. Analyses of Primary Efficacy EndpointΔThe primary efficacy endpoint is the incidence of AKI within 72 hours after LTx. The International Club for Ascites (ICA) 2015 criteria, a revision of the Kidney Disease Improving Global Outcomes (KDIGO) criteria for patients with cirrhosis, will be used to define AKI (Angeli et al., 2015). The definitions are as follows:AKI: increase in serum creatinine (sCr) ≥ 0.3 mg/dL within a 48-hour time window, or a percentage increase ≥ 50% from baseline, or initiation of renal replacement therapy (RRT)Baseline sCr: the most recent value of sCr prior to LTxThe primary outcome (presence or absence AKI) will be compared between the two arms by a two-sample test of proportions (Fisher’s exact test or chi-square, as appropriate).To determine whether a subject met the primary outcome, the following procedure is followed in order:Step #1. Need for renal replacement therapy If the patient had RRT within 72 hrs from the End of Surgery (“postop_rrt_72h” = yes), the primary outcome is met (subject had AKI).Step #2. Calculation of the fold-increase in sCrThe interim analysis database will contain four sCr values:a. creat_0 = Baseline sCr (most recent prior to Surgery)b. creat_24h = Highest sCr, 0-24 hrs from End of Surgeryc. creat_48h = Highest sCr, 24-48 hrs from End of Surgeryd. creat_72h = Highest sCr, 48-72 hrs from End of SurgeryThe highest of these values (b-d) will be used to calculate the following: If the Fold increase in sCr is ≥ 1.5, the primary outcome is met (subject had AKI).Step #3.Calculation of the difference in sCrThe following calculation should be performed: If ΔsCrmax If ΔsCrmax ≥ 0.3 mg/dL, but the patient does not meet either of the criteria described in Step #1 (RRT) or Step #2 (Fold increase), additional timing information may be required to determine if the subject had AKI. According to KDIGO criteria, the sCr must increase by ≥ 0.3 mg/dL within a 48 hour time window to diagnose AKI (i.e., if the sCr increased by 0.3 mg/dL over 72 h, this is not necessarily AKI) (Kellum et al., 2012). In this situation, the following calculations should be performed: * indicates a time interval that is unambiguous (definitively less than 48 h)If either Δ3 or Δ5 is ≥ 0.3 mg/dL, the timing is unambiguous (If any of Δ1, Δ2, or Δ4 is ≥ 0.3 mg/dL, the timing is ambiguous and further information is required to determine AKI status. The Data Management Team will provide the Statistician with the exact times and dates of collection for the relevant sCr values. The time interval (Δt) between sCr collections should be calculated for each Δ1, 2, or 4 that is ≥ 0.3 mg/dL. If any Δt is ≤ 48 h, the primary outcome is met (subject has AKI).If all Δt > 48 h, the primary outcome is NOT met (subject DOES NOT have AKI).Baseline sCr: the most recent value of sCr prior to LTxThe primary outcome (presence or absence AKI) will be compared between the two arms by a two-sample test of proportions (Fisher’s exact test or chi-square, as appropriate).To determine whether a subject met the primary outcome, the following procedure is followed in order:Step #1. Need for renal replacement therapy If the patient had RRT within 72 hrs from the End of Surgery (“postop_rrt_72h” = yes), the primary outcome is met (subject had AKI).Step #2. Calculation of the fold-increase in sCrThe interim analysis database will contain four sCr values:a. creat_0 = Baseline sCr (most recent prior to Surgery)b. creat_24h = Highest sCr, 0-24 hrs from End of Surgeryc. creat_48h = Highest sCr, 24-48 hrs from End of Surgeryd. creat_72h = Highest sCr, 48-72 hrs from End of SurgeryThe highest of these values (b-d) will be used to calculate the following: If the Fold increase in sCr is ≥ 1.5, the primary outcome is met (subject had AKI).Step #3.Calculation of the difference in sCrThe following calculation should be performed: If ΔsCrmax If ΔsCrmax ≥ 0.3 mg/dL, but the patient does not meet either of the criteria described in Step #1 (RRT) or Step #2 (Fold increase), additional timing information may be required to determine if the subject had AKI. According to KDIGO criteria, the sCr must increase by ≥ 0.3 mg/dL within a 48 hour time window to diagnose AKI (i.e., if the sCr increased by 0.3 mg/dL over 72 h, this is not necessarily AKI) (Kellum et al., 2012). In this situation, the following calculations should be performed: * indicates a time interval that is unambiguous (definitively less than 48 h)If either Δ3 or Δ5 is ≥ 0.3 mg/dL, the timing is unambiguous (If any of Δ1, Δ2, or Δ4 is ≥ 0.3 mg/dL, the timing is ambiguous and further information is required to determine AKI status. The Data Management Team will provide the Statistician with the exact times and dates of collection for the relevant sCr values. The time interval (Δt) between sCr collections should be calculated for each Δ1, 2, or 4 that is ≥ 0.3 mg/dL. If any Δt is ≤ 48 h, the primary outcome is met (subject has AKI).If all Δt > 48 h, the primary outcome is NOT met (subject DOES NOT have AKI).The primary outcome (presence or absence AKI) will be compared between the two arms by a two-sample test of proportions (Fisher’s exact test or chi-square, as appropriate).To determine whether a subject met the primary outcome, the following procedure is followed in order:Step #1. Need for renal replacement therapy If the patient had RRT within 72 hrs from the End of Surgery (“postop_rrt_72h” = yes), the primary outcome is met (subject had AKI).Step #2. Calculation of the fold-increase in sCrThe interim analysis database will contain four sCr values:a. creat_0 = Baseline sCr (most recent prior to Surgery)b. creat_24h = Highest sCr, 0-24 hrs from End of Surgeryc. creat_48h = Highest sCr, 24-48 hrs from End of Surgeryd. creat_72h = Highest sCr, 48-72 hrs from End of SurgeryThe highest of these values (b-d) will be used to calculate the following:Fold increase in sCr = Maximum of {creat_24h, creat_48h, creat_72h} / creat_0If the Fold increase in sCr is ≥ 1.5, the primary outcome is met (subject had AKI).Step #3.Calculation of the difference in sCrThe following calculation should be performed:ΔsCrmax = Maximum of {creat_24h, creat_48h, creat_72h} – Minimum of {creat_0, creat_24h, creat_48h}If ΔsCrmax If ΔsCrmax ≥ 0.3 mg/dL, but the patient does not meet either of the criteria described in Step #1 (RRT) or Step #2 (Fold increase), additional timing information may be required to determine if the subject had AKI. According to KDIGO criteria, the sCr must increase by ≥ 0.3 mg/dL within a 48 hour time window to diagnose AKI (i.e., if the sCr increased by 0.3 mg/dL over 72 h, this is not necessarily AKI) (Kellum et al., 2012). In this situation, the following calculations should be performed:Δ1 = creat_24h – creat_0Δ2 = creat_48h – creat_0*Δ3 = creat_48h – creat_24hΔ4 = creat_72h – creat_24h*Δ5 = creat_72h – creat_48h* indicates a time interval that is unambiguous (definitively less than 48 h)If either Δ3 or Δ5 is ≥ 0.3 mg/dL, the timing is unambiguous (If any of Δ1, Δ2, or Δ4 is ≥ 0.3 mg/dL, the timing is ambiguous and further information is required to determine AKI status. The Data Management Team will provide the Statistician with the exact times and dates of collection for the relevant sCr values. The time interval (Δt) between sCr collections should be calculated for each Δ1, 2, or 4 that is ≥ 0.3 mg/dL. If any Δt is ≤ 48 h, the primary outcome is met (subject has AKI).If all Δt > 48 h, the primary outcome is NOT met (subject DOES NOT have AKI).Step #4. Death of a subject prior to 72 hours from the End of SurgeryIf a subject died 1.5.3. Analyses of Secondary EndpointsAnalyses of secondary endpoints are not included in the interim analysis for this study. A detailed analysis of the frequencies and distribution of KDIGO stages of AKI is an important component of the final statistical analysis.1.5.4. Analyses of Safety EndpointsAnalyses of safety endpoints are not included in the interim analysis for this study.
?. Stopping RuleThis study does not have a formal stopping rule that is based on statistical testing.
?. Handling of Missing DataSubjects with missing primary outcome data will be excluded from the interim analysis.
?. References    Angeli, P., Ginès, P., Wong, F., Bernardi, M., Boyer, T.D., Gerbes, A., Moreau, R., Jalan, R., Sarin, S.K., Piano, S., et al. (2015). Diagnosis and management of acute kidney injury in patients with cirrhosis: revised consensus recommendations of the International Club of Ascites. Gut 64, 531–537.Kellum, J.A., Lameire, N., Aspelin, P., Barsoum, R.S., Burdmann, E.A., Goldstein, S.L., Herzog, C.A., Joannidis, M., Kribben, A., Levey, A.S., et al. (2012). Kidney disease: Improving global outcomes (KDIGO) acute kidney injury work group. KDIGO clinical practice guideline for acute kidney injury. Kidney International Supplements 2, 1–138.Kellum, J.A., Zarbock, A., and Nadim, M.K. (2017). What endpoints should be used for clinical studies in acute kidney injury? Intensive Care Medicine 43, 901–903.