Liver Transplantation for Hepatocellular Carcinoma: Validation of the UCSF-Expanded Criteria Based on Preoperative Imaging

American Journal of Transplantation 2007; 7: 2587 2596 Blackwell Munksgaard C 2007 The Authors Journal compilation C 2007 The American Society of Transplantation and the American Society of Transplant Surgeons doi: 10.1111/j.1600-6143.2007.01965.x Liver Transplantation for Hepatocellular Carcinoma: Validation of the UCSF-Expanded Criteria Based on Preoperative Imaging F. Y. Yao a,b,, L. Xiao a, N. M. Bass a, R. Kerlan c, N. L. Ascher b and J. P. Roberts b Departments of a Medicine, b Surgery and c Radiology, University of California, San Francisco, San Francisco, CA Corresponding author: Francis Y. Yao, yaofyk@itsa.ucsf.edu Part of this work was presented in a plenary session of the 57th annual AASLD meeting, Boston, MA, October 29, 2006 (Hepatology 2006;44 (Suppl 1):191A). We previously suggested that in patients with heptocellular carcinoma (HCC), the conventional Milan criteria (T1/T2) for orthotopic liver transplantation (OLT) could be modestly expanded based on pathology (UCSF criteria). The present study was undertaken to prospectively validate the UCSF criteria based on pretransplant imaging. Over a 5-year period, the UCSF criteria were used as selection guidelines for OLT in 168 patients, including 38 patients exceeding Milan but meeting UCSF criteria (T3A). The 1- and 5-year recurrence-free probabilities were 95.9% and 90.9%, and the respective survivals without recurrence were 92.1% and 80.7%. Patients with preoperative T1/T2 HCC had 1- and 5-year recurrence-free probabilities of 95.7% and 90.1%, respectively, versus 96.9% and 93.6%, respectively, for preoperative T3A stage (p = 0.58). Under-staging was observed in 20% of T2 and 29% of T3A HCC (p = 0.26). When explant tumor exceeded UCSF criteria (15%), the 1- and 5-year recurrence-free probabilities were 80.4% and 59.5%, versus 98.6% and 96.7%, respectively, for those within UCSF criteria (p < 0.0001). In conclusion, our results validated the ability of the UCSF criteria to discriminate prognosis after OLT and to serve as selection criteria for OLT, with a similar risk of tumor recurrence and under-staging when compared to the Milan criteria. Key words: Hepatocellular carcinoma, liver transplantation, UCSF criteria Received 30 April 2007, revised 29 June 2007 and accepted for publication 25 July 2007 Introduction The field of orthotopic liver transplantation (OLT) for hepatocellular carcinoma (HCC) has evolved rapidly in the past decade (1). In 1996, the seminal publication by Mazzaferro et al. from Milan, Italy, set the stage for OLT to be widely accepted as the best treatment for early-stage HCC (2). In this prospective study of 48 patients whose preoperative imaging studies identified either a single lesion 5 cm or 2 3 lesions 3 cm (Milan criteria), the 4-year actuarial survival rate after OLT was 75% and the recurrencefree probability was 83%. Among the 13 patients (27%) with explant pathologic tumor stage under-estimated by preoperative imaging, their 4-year recurrence-free probability was only 59%, compared to 92% for the other 35 patients with pathologic tumor stage not exceeding the pretransplant criteria (2). Since that time, a number of other studies have reported 5-year posttransplant survival rates to be consistently 70% or better among patients with HCC meeting the Milan criteria (1). The United Network for Organ Sharing (UNOS) has incorporated the Milan criteria into T1 and T2 in a modified staging system for HCC (Table 1) (3). HCC has also evolved from being a contraindication to OLT in many transplant centers in the early to mid-1990s, to a prioritized condition for deceased donor allocation under the model for end-stage liver disease (MELD) system implemented in 2002 (3,4). After several subsequent modifications of the HCC-adjusted MELD organ allocation scheme, only the subgroup of patients meeting T2 criteria (Table 1) by radiologic staging are currently eligible to receive listing priority (3,4). This latest modification was largely driven by the observation that patients with a T1 lesion (solitary tumor <2cm) had a significantly lower risk of dropout from the waiting list due to tumor progression compared to T2 HCC (4,5). Furthermore, a high proportion of patients transplanted for a T1 lesion were found to have no HCC in the liver explant (1,4). In 2001, our group from the University of California, San Francisco (UCSF) reported that modest expansion of the tumor size limits beyond the Milan criteria did not appear to adversely impact survival after OLT (6). The proposed UCSF criteria (a single lesion 6.5 cm in diameter or 2 lesions 4.5 cm with total tumor diameter 8 cm) were derived from explant tumor pathology in 70 patients with 2587

Yao et al. Table 1: UNOS staging classification for hepatocellular carcinoma T1: 1 nodule <1.9 cm T2: 1 nodule 2.0 5.0 cm; 2 or 3 nodules, all <3.0 cm T3: 1 nodule >5.0 cm; 2 or 3 nodules, at least one >3.0 cm T4a: 4 or more nodules, any size T4b: T2,T3 or T4a plus gross intrahepatic portal or hepatic vein involvement as indicated by CT, MRI or ultrasound N1: Regional (porta hepatis) nodes, involved M1: Metastatic disease, including extrahepatic portal or hepatic vein involvement Stage I: T1 Stage II: T2 Stage III: T3 Stage IVA1: T4a Stage IVA2: T4b Stage IVB: Any N1, any M1 For the purpose of this study, patients with T3 lesion meeting the proposed expanded criteria from UCSF single lesion 6.5 cm or 2 3 lesions none >3 cm with a total tumor diameter of 8 cm were classified as T3A. The other patients with T3 HCC exceeding these criteria were classified as T3B. HCC who underwent OLT. The 60 patients with pathologic tumor characteristics meeting the UCSF criteria had 1- and 5-year survival rates of 90% and 75%, respectively, which were significantly better than the 50% 1-year survival rate for the other 10 patients with pathologic tumor stage exceeding these limits. When the same criteria were retrospectively applied to preoperative radiologic staging, they still provided statistically significant prognostic power with respect to survival after OLT (6). The UCSF criteria also appeared to be better than the Milan criteria in predicting survival after OLT (7). Given that the proposed UCSF criteria were derived retrospectively from explant tumor pathology (6,7), several groups have advised caution in liberalizing the existing criteria for OLT (8 11). The limitations of imaging studies, exemplified by tumor stage under estimation in 20 30% of patients undergoing OLT (2,6,8), have been repeatedly cited as a major concern for pushing the envelope even to a modest degree (8 11). Proponents of maintaining the status quo might argue that the Milan criteria have so far been able to maintain acceptable outcome after OLT (12), and offer a wider safety margin for tumor under-staging by preoperative imaging (13). To address these concerns and the inherent limitations of our initial published data (6,7), we have applied the proposed UCSF criteria for OLT based on pretransplant imaging since 2001. The primary objective of the present study was to evaluate the impact of expansion of selection criteria for OLT on posttransplant tumor recurrence and survival. We also further examined the problem of tumor understaging by preoperative imaging studies and factors associated with HCC recurrence after OLT. Patients and Methods Tumor staging and baseline characteristics Since January 2001, we have applied the UCSF criteria for OLT based on pretransplant imaging. The study cohort included 168 patients who underwent OLT with known HCC between January 2001 and August 2006 and had pretransplant tumor staging that met UCSF criteria. There was no overlap with patients from the previous cohort of 70 patients (6). Patients with incidental HCC in the liver explant during this period were not included. During this period, an additional 28 patients transplanted under a downstaging protocol with further expansion of the UCSF criteria (14) were also excluded. Tumor staging was based on either contrast-enhanced computed tomography (CT) or magnetic resonance imaging (MRI) of the abdomen. Histologic confirmation of HCC was not routinely performed at our center. The diagnosis of HCC was supported by CT or MRI showing tumor enhancement during the arterial phase and the wash-out effect during delayed images. For small tumors under 2 cm that did not exhibit the typical radiographic characteristics of HCC, the diagnosis of HCC was supported by interval growth of the lesion on follow-up imaging. All patients underwent serial CT or MRI in combination with alpha-fetoprotein (AFP) every 3 months while on the waiting list for OLT. Chest CT was performed at baseline and at 3-month intervals to exclude metastatic disease. Bone scan was performed at baseline and if bone metastasis was suspected during follow-up. Staging by imaging according to the UNOS classification (Table 1) within 3 months prior to OLT revealed T1 HCC in 8 patients, T2 in 122 patients and T3A (exceeding T2 criteria but meeting UCSF criteria) in 38 patients. Under the MELD system of organ allocation implemented in February 2002, all patients with the diagnosis of HCC required approval by the regional UNOS review board to receive priority listing for OLT. During the study period, 23 patients were removed from the waiting list due to tumor progression (n = 19) or death from liver failure or other causes without OLT (n = 4). An additional 8 patients were removed from the waiting list due to other reasons, including psychosocial or substance abuse issues in 5 patients, decision not to undergo OLT in 3 patients, and OLT at another liver transplant center in 2 patients. The baseline characteristics of the entire cohort and according to pretransplant tumor stage are summarized in Table 2. The baseline characteristics of the 38 patients with pretransplant T3A tumor stage were not significantly different compared to the other 130 patients with T1/T2 tumor stage before OLT. According to the Child s classification of cirrhosis (15), almost 40% of the cohort had Child s class A cirrhosis or Child-Turcotte-Pugh (CTP) score of 5 6, with HCC as the sole indication for OLT. Chronic hepatitis C was the most common etiology of liver disease, accounting for more than 60% of cases, followed by chronic hepatitis B and alcoholic liver disease. The diagnosis in the remaining 15 patients included nonalcoholic fatty liver disease in 5 patients, cryptogenic cirrhosis in 3, hepatitis B and C coinfection in 2, hemochromatosis in 2, primary biliary cirrhosis in 2 and autoimmune hepatitis in 1 patient. The majority of the patients (88%) underwent OLT under the MELD system of organ allocation implemented in February of 2002. Eleven patients (7%) underwent right-lobe living donor liver transplantation (LDLT). The median and mean waiting time from listing to OLT or LDLT was 6.1 months and 6.7 months (range 0.7 15.1 months), respectively. Preoperative loco-regional therapies The general guidelines for pretransplant loco-regional therapies at our center are presented in Figure 1. Patients with decompensated liver disease with a CTP score of 10 (Child s C cirrhosis) generally did not receive elective loco-regional treatments. Among patients with well-compensated cirrhosis 2588 American Journal of Transplantation 2007; 7: 2587 2596

Table 2: Baseline clinical and demographic characteristics UCSF Criteria and Liver Transplantion for HCC Pretransplant HCC stage 1 All patients T1/T2 (N = 130) T3A (N = 38) (N = 168) Median age (years, range) 57.7 (35 74) 59.5 (27 73) 57.9 (27 74) Male gender (number/%) 101 (77.7%) 33 (86.8%) 134 (79.8%) Asian (number/%) 33 (25.4%) 15 (39.5%) 48 (28.6%) Child s class (CTP Score) A(5 6) 53 (40.8%) 11 (28.9%) 64 (38.1%) B(7 9) 46 (35.4%) 16 (42.1%) 62 (36.9%) C (10 15) 31 (23.8%) 11 (28.9%) 42 (25.0%) Median CTP (range) 7 (5 14) 7 (5 12) 7 (5 14) Etiology of liver disease HCV 84 (64.6%) 22 (57.9%) 106 (63.1%) HBV 30 (23.1%) 9 (23.7%) 39 (23.2%) Alcohol 4 (3.1%) 4 (10.5%) 8 (4.8%) Others 12 (9.2%) 3 (7.9%) 15 (8.9%) Median alpha-fetoprotein 13.2 19.7 14.5 and range (ng/ml) (0.7 6992) (2.1 1132) (0.7 6992) Transplanted under MELD 115 (88.5%) 33 (86.8%) 148 (88.1%) Living-donor liver transplant 9 (6.9%) 2 (5.3%) 11 (6.5%) 1 The two groups are not significantly different in all baseline characteristics (p > 0.05). HCC = hepatocellular carcinoma; CTP = Child-Turcotte-Pugh score; HCV = hepatitis C virus; HBV = hepatitis B virus; MELD = model for end-stage liver disease. with a CTP score <7 (Child s A cirrhosis), laparoscopic radiofrequency ablation (RFA) was the preferred treatment modality for the subgroup with multi-focal HCC, taking advantage of the fact that laparoscopic RFA could effectively treat all the lesions in a single session. For patients with wellcompensated or mildly decompensated liver disease (CTP <10) and one or more lesions 4 cm, selective trans-arterial chemoembolization (TACE) and laparoscopic RFA, either alone or in combination, were the mainstays of treatment. Percutaneous RFA or ethanol injection (PEI) were used mainly for patients with one or more lesions <4 cm, since the efficacy of these treatments is lower for tumors beyond 4 cm in maximal diameter. In this subgroup, PEI was considered a safer and preferred treatment modality versus percutaneous RFA in patients with a CTP score of 7 9 (Child s B cirrhosis). The choice of treatment was ultimately determined based on tumor size, number, location and hypervascularity according to CT or MRI, as well as the hepatic reserve of the individual patient, following thorough review of the clinical and radiologic information in a multi-disciplinary tumor conference. One hundred and twenty-five patients (73% for T1/T2 and 79% for T3A) received elective loco-regional therapies (Table 3). Thirty-one patients received elective TACE alone prior to OLT for a total of 50 treatment sessions. Fiftyfive patients received ablation only before OLT, including 7 treated with 18 sessions of PEI, and 48 patients receiving 49 session of RFA (22 laparoscopic, 6 open and 20 by percutaneous approach). Thirty-nine patients received a combination of TACE and ablation treatments, including 10 with PEI and TACE, 28 with TACE and RFA (12 laparosocpic, 2 open and 14 percutaneous) and 1 with PEI and laparoscopic RFA. A total of 101 treatments were delivered to these 39 patients. A significantly higher proportion of patients in the T3A group received combination therapy when compared with the T1/T2 group (p = 0.004). In the T3A group, tumor size at the time of last CTP < 7 CTP 7-9 < 4 cm Correct INR to < 1.5 and platelets to > 50K, if needed Single 4 cm 2 or 3 lesions Ascites better controlled Able to correct INR to < 1.5 and platelets > 50K < 4cm Single 4 cm 2 or 3 lesions Severe Ascites No treatment Figure 1: Protocol for elective loco-regional therapy for HCC before liver transplantation. TACE Percutaneous RFA PEI Laparoscopic RFA PEI Sequential PEI Highly selective TACE American Journal of Transplantation 2007; 7: 2587 2596 2589

Yao et al. Table 3: Elective pretransplant loco-regional treatments T1/T2 T3A All patients (N = 130) (N = 38) (N = 168) Loco-regional treatments TACE only 1 26 (20%) 5 (13.2%) 31 (18.5%) Ablation only 2 47 (36.2%) 8 (21.1%) 55 (32.7%) PEI 5 (3.8%) 2 (5.3%) 7 (4.2%) RFA 42 (32.3%) 6 (15.8%) 48 (28.6%) Combination 3 23 (17.7%) 16 (42.1%) 39 (23.2%) TACE + PEI 6 (4.6%) 4 (10.5%) 10 (6.0%) 0TACE + RFA 17 (13.1%) 11 (28.9%) 28 (16.7%) PEI + RFA 0 (0%) 1 (2.6%) 1 (0.6%) No elective treatment 4 35 (26.9%) 8 (21.1%) 43 (25.6%) TACE within 24 h 13 (10%) 7(20.1%) 20 (11.9%) None 22 (16.9%) 1 (2.6%) 23 (13.7%) 1 The 31 patients in this group underwent 50 elective TACE treatments, not including TACE performed within 24 h prior to OLT. 2 The 7 patients treated with PEI had a total of 18 treatment sessions. Forty-eight patients received 49 RFA treatments, including 22 laparoscopic, 6 open and 20 by the percutaneous approach. 3 The difference between T1/T2 versus T3A groups in the proportion of patients receiving combination treatment was statistically significant (p = 0.004). 4 The reason for no elective treatment included severely decompensated liver disease in 22 patients, short waiting time ( 3 months) to OLT or LDLT in 16 patients, small tumor size <2 cm per decision of the physician in 4 patients and technical considerations in 1 patient. TACE = trans-arterial chemoembolization; PEI = percutaneous ethanol injection; RFA = radiofrequency ablation. imaging within 3 months before OLT when compared to baseline showed stable size of the largest lesion (+/ 0.3 cm) with varying degree of tumor necrosis in 24 patients, decrease in size of the largest tumor nodule by > 0.3 cm but still >T2 criteria in 14 patients. Forty-three patients (25.6%) did not receive elective loco-regional therapies, including 20 patients who received only TACE within 24 h prior to OLT when a donor was available. The reason for no elective loco-regional treatment included decompensated liver disease with a CTP score 10 in 22 patients, short waiting time of 3 months to OLT in 16 patients, small tumor size <2 cm per decision of the physician in 4 patients and technical considerations in 1 patient. Histopathologic studies In the analysis of HCC characteristics in the liver explant, tumor size, number, total tumor diameter, pathologic tumor stage and histologic grade based on the Edmondson and Steiner criteria (grade 1: well-differentiated; grade 2: moderately differentiated; grade 3: poorly differentiated) (16), the degree of tumor necrosis and the presence or absence of micro- or macro-vascular invasion were recorded. In a patient with multiple lesions in the liver explant, the highest tumor histologic grade was reported. Statistical analysis Survival and recurrence-free probabilities were estimated by the Kaplan Meier method and compared using the log-rank test. Chi-squared, paired t-test or Mann-Whitney U-test was used to compare differences between subgroups. Univariate and multivariate Cox proportional hazards model was used to identify predictors for HCC recurrence. The value p < 0.05 was considered statistically significant in all analyses. Results Survival and recurrence after liver transplantation The median follow-up was 26.1 months for the entire cohort (range 0.2 72.9 months), with a minimum follow-up of 6 months among survivors. Three patients in this cohort underwent liver retransplantation at 7, 29 and 159 days after OLT, due to primary graft nonfunction, hepatic artery thrombosis after LDLT and ductopenic rejection, respectively. Follow-up was censored on the day of retransplantation. Two patients were alive without recurrence after 1.0 0.8 0.6 Survival without recurrence (N=168) Recurrence-free probabilities (N=168) 0.4 0.2 0.0 0 1 2 3 4 5 Years After Liver Transplantation 168 126 83 58 36 14 Figure 2: The Kaplan Meier survival function following liver transplantation. The number of patients at risk at each time point is shown below the horizontal axis. 2590 American Journal of Transplantation 2007; 7: 2587 2596

UCSF Criteria and Liver Transplantion for HCC 1.0 0.8 0.6 Radiologic T1/T2 Stage (N=130) Radiologic T3A Stage (N=38) 0.4 P=0.58 by log-rank test Figure 3: The Kaplan Meier recurrence-free probabilities following liver transplantation according to preoperative tumor stage. The difference in survival was compared by the log-rank test. The number of patients at risk at each time point is shown below the horizontal axis. 0.2 0.0 0 1 2 3 4 5 Years After Liver Transplantation T2 130 96 65 45 29 11 T3A 38 30 18 13 7 3 retransplantation, and the other patient died 24 days after retransplant due to multi-organ failure. The Kaplan Meier 1- and 5-year recurrence-free probabilities were 95.9% and 90.9%, respectively, and the 1- and 5- year probabilities of survival without recurrence were 94.7% and 80.9%, respectively (Figure 2). Among the 12 patients who suffered from HCC recurrence after OLT, the median time to recurrence was 11.9 months with a range of 3.1 23.3 months. Nine of the 12 patients with HCC recurrence had died by the end of follow-up. There were 11 deaths unrelated to HCC, including 3 in the early postoperative period (within 2 months) from sepsis and multi-organ failure, 6 from graft failure due to severe recurrent hepatitis C infection at 9 45 months after OLT, 1 death from graft failure and sepsis at 8 months in the setting of biliary strictures and possible ischemic injury and 1 death from sepsis at 30 months after OLT. The 130 patients with pretransplant T1 or T2 stage had 1- and 5-year recurrence-free probabilities of 95.7% and 90.1%, respectively, versus 96.9% and 93.6%, respectively, for the 38 patients with T3A HCC by preoperative staging (p = 0.58) (Figure 3). The 25 patients (14.9%) with explant pathologic HCC stage exceeding UCSF criteria had 1- and 5-year recurrence-free probabilities of 80.4% and 59.5%, respectively, versus 98.6% and 96.7%, 1.0 0.8 Pathologic Stage within UCSF Criteria (N=143) 0.6 Pathologic Stage Exceeding UCSF Criteria (N=25) 0.4 Figure 4: The Kaplan Meier recurrence-free probabilities following liver transplantation according to pathologic tumor stage meeting or exceeding UCSF criteria. The difference in survival was compared by the log-rank test. The number of patients at risk at each time point is shown below the horizontal axis. 0.2 0.0 P<0.0001 by log-rank test HR 13.3, p<0.0001 0 1 2 3 4 5 Years After Liver Transplantation Within T3A 143 110 73 51 31 11 >T3A 25 16 10 7 5 3 American Journal of Transplantation 2007; 7: 2587 2596 2591

Yao et al. respectively, for the 143 patients with pathologic HCC stage within UCSF criteria (p < 0.0001) (Figure 4). Histopathologic tumor charactersitics in the liver explant The explant histopathologic characteristics of HCC according to the UNOS staging classification are summarized in Table 4. One patient had HCC found in a 1.6 cm lymph node in association with a completely necrotic 2.3 cm tumor in the liver explant (T2N1). The histologic grade of tumor differentiation was unknown in 44 patients due to complete tumor necrosis from preoperative ablation or TACE treatments. In the other 124 patients, 14 (11.3%) had poorly differentiated tumor grade. Vascular invasion was found in 18 patients (10.7%), including 14 with micro-vascular invasion and 4 with macro-vascular invasion (T4B). All four patients with macro-vascular invasion had involvement of the portal vein. HCC recurrence was found in 7 of 18 patients (38.9%) with micro- or macro-vascular invasion, versus only 5 of 150 patients (3.3%) without vascular invasion (p < 0.0001) (Table 4). Among the 44 patients with complete tumor necrosis in the liver explant, 19 of 44 patients (43.2%) had biopsy proven HCC before OLT, either by percutanoeus needle biopsy (n = 5) or during laparoscopic or open RFA (n = 14). The diagnosis of HCC was supported by imaging characteristics Table 4: Histopathologic tumor characteristics in the liver explant No. of patients No. with Pathologic tumor stage (N = 168) recurrence T1 11 (6.5%) 0 T2 103 (61.3%) 3 (2.9%) T3A 29 (17.3%) 1 (3.4%) T3B 8 (4.8%) 4 (50%) T4A 12 (7.1%) 1 (8.3%) T4B 4 (2.4%) 2 (50%) N1 1 1 (0.6%) 1 (100%) Histologic grade 2 # of patients # with recurrence (N = 124) Well-differentiated (grade 1) 52 (41.9%) 0 Moderately differentiated 58 (46.8%) 8 (13.8%) (grade 2) Poorly differentiated (grade 3) 14 (11.3%) 3 (21.4%) Vascular invasion 3 # of patients # with recurrence (N = 168) Micro-vascular 14 (8.3%) 5 (35.7%) Macro-vascular 4 (2.4%) 2 (50%) No 150 (89.3%) 5 (3.3%) 1 One patient had a 1.6 cm metastatic lymph node in association with a completely necrotic 2.3 cm tumor (T2) in the liver. This patient suffered from HCC recurrence. 2 Information on the histologic grade could not be determined in 44 patients due to complete tumor necrosis from prior loco-regional therapies. One patient with lymph node invasion developed HCC recurrence but the histologic grade is unknown. 3 The presence of micro- or macro-vascular invasion was significantly associated with tumor recurrence (p < 0.0001). of the tumor in conjunction with angiographic findings during TACE and uptake of lipiodol by the tumor in 14 patients. One patient had a completely necrotic 2.3 cm necrotic tumor in the explant, but evidence of lymph node invasion. This patient suffered from HCC recurrence at 9 months after OLT. The remaining 10 patients were all treated with percutanoeus ablations, including 4 with multifocal tumors, 5 with a solitary T2 lesion ranging from 2.4 3.6 cm and 1 patient with a 1.7 cm T1 lesion. Correlation between tumor stage and histopathologic tumor charactersitics in the explant The correlation between pathologic tumor stage and unfavorable histologic features, namely micro-vascular invasion and poorly differentiated tumor grade, is presented in Figure 5. None of the 11 patients with pathologic T1 HCC had either micro-vascular invasion or poorly differentiated grade, and they were excluded from this analysis. The single patient with lymph node metastasis was also excluded. Micro-vascular invasion was present in 6 of 103 patients (5.8%) with pathologic T2 HCC, 3 of 29 patients (10.3%) with pathologic T3A tumors (p = 0.41) and 5 of 20 patients (25%) with pathologic T3B or T4A HCC (p = 0.017 vs. T2). Poorly differentiated tumor grade was observed in 6 of 92 patients (6.5%) with pathologic T2 tumors, versus 1 of 27 patients (3.7%) with T3A (p = 1.0), and 5 of 20 patients (25%) with T3B or T4A HCC (p = 0.14). Two of the four patients with T4B HCC had poorly differentiated histologic grade. When tumor stage according to pretransplant imaging data were used in this comparative analysis (Figure 6), 12 of 122 patients (9.8%) with T2 lesions had micro- or macrovascular tumor invasion in the liver explant, versus 6 of 38 patients (15.8%) with T3A tumors by pretransplant staging (p = 0.38). Poorly differentiated tumor grade was found in 10 of 111 patients (9.0%) with HCC classified as T2 before % 50 40 30 20 10 0 P = 0.41 5.8% 6.5% pt2 (N=103) P = 0.017 P = 1.0 10.3% 3.7% pt3a (N=29) Micro-vascular invasion Poorly differentiated grade P = 0.14 25% 25% pt3b/t4a (N=20) Figure 5: Correlation between pathologic tumor stage and histologic tumor characteristics micro-vascular invasion and poorly differentiated grade. The eight patients with pathologic T1 HCC and four patients with pathologic T4B (defined by macro-vascular invasion) were not included. 2592 American Journal of Transplantation 2007; 7: 2587 2596

UCSF Criteria and Liver Transplantion for HCC % 50 40 30 20 10 0 9.8% 9% T2 (N=122) P = 0.38 Micro- or macro-vascular invasion Poorly differentiated grade P = 0.76 15.8% T3A (N=38) 11.1% Figure 6: Correlation between pretransplant tumor stage and histologic tumor characteristics micro- or macro-vascular invasion and poorly differentiated grade. The eight patients with T1 HCC according to pretransplant staging were not included. Table 6: Tumor under-staging by preoperative imaging studies Pretransplant No. with under- No. with tumor stage staging 1 recurrence T2 (N = 122) 24 (19.7%) 6 (25%) T3A = 10 1 T3B = 4 2 T4A = 7 0 T4B = 2 2 N1 = 1 1 T3A (N = 38) 2 11 (28.9%) 2 (18.2%) T3B = 4 1 T4A = 5 1 T4B = 2 0 1 The difference in the incidence of under-staging between T1/T2 and T3A was not statistically significant (p = 0.26). 2 Among 38 patients with T3A by pretransplant staging, 8 (21%) had over-staging of HCC (T3A T2). CI 1.3 18.6, p = 0.019) remained significant predictors of HCC recurrence after OLT. OLT, versus 4 of 36 patients (11.1%) with T3A tumor stage (p = 0.76). Predictors of recurrence A total of 8 variables were included in univariate analysis of predictors of recurrence. The results, including hazard ratio with 95% confidence intervals, are summarized in Table 5. Pretransplant HCC stage T3A based on imaging was not associated with a greater risk for HCC recurrence compared to T1 or T2. Significant predictors of HCC recurrence included pathologic stage exceeding UCSF criteria, micro- or macro-vascular invasion, and AFP > 1000 ng/ml or > 500 ng/ml. Poorly differentiated histologic grade, loco-regional treatments versus no treatment and LDLT were not predictive of HCC recurrence. In bi-variate analysis after controlling for pathologic tumor stage exceeding UCSF criteria, micro- or macro-vascular invasion (hazard ratio 4.6, 95% CI 1.2 17.1, p = 0.023) and AFP > 1000 (hazard ratio 4.9, 95% Table 5: Predictors of tumor recurrence after liver transplantation in univariate analysis Variables Hazard ratio (95% CI) p-value Pretransplant T3A stage 0.65 (0.14 3.0) 0.58 by imaging (vs. T1 and T2 combined) Pathologic stage > T3A 13.3 (4.0 43.5) <0.0001 (Exceeds UCSF Criteria) Micro-or macro-vascular invasion 11.4 (3.6 35.9) <0.0001 AFP > 1000 6.3 (1.7 23.2) 0.006 AFP > 500 4.2 (1.3 13.9) 0.02 Loco-regional treatments 0.86 (0.19 3.9) 0.84 (vs. no treatment) Poorly differentiated grade 2.6 (0.69 9.8) 0.16 (grade 3) (vs. grade 1 or 2) LDLT 1.2 (0.15 9.0) 0.89 AFP = alpha-fetoprotein; LDLT = living donor liver transplantation. Accuracy of preoperative tumor staging Tumor under-staging was not found in any of the eight patients with T1 lesions before OLT. Among the 122 patients with tumors classified as T2 before OLT, tumor under-staging was observed in 24 patients (19.7%). Six of these patients (25%) suffered from HCC recurrence after OLT. Among the 38 patients with pretransplant T3A tumor stage, tumor under-staging was found in 11 patients (28.9%), of whom 2 (18.2%) had developed HCC recurrence (Table 6). The difference in the risk for understaging between tumors staged as T2 versus T3A before OLT did not achieve statistical significance (p = 0.26). Tumor stage was over-estimated in 8 of 38 patients (21%) classified as T3A before OLT. Discussion For the past decade, the Milan criteria for HCC have been the paradigm for the selection of the best candidates for OLT. Based on a combination of tumor size and number, the Milan criteria have served as a reliable surrogate for favorable tumor behavior, which translates into a low risk of recurrence following OLT. In the United States, the adoption of the Milan criteria for OLT was largely responsible for the steady improvement in posttransplant survival in recent years (17). However, as the experience with OLT for HCC continues to grow, there is also increasing evidence that the Milan criteria may be too restrictive (6,18 25). The proposed UCSF criteria (6), which represent a modest extension of the tumor size limits beyond the Milan criteria, have generated considerable interest but also attracted their full share of controversy (8,11,25). Based on mostly retrospective data, several other groups have independently tested the UCSF criteria (20,24,26 28). In a multi-center study by Leung et al. from UNOS region American Journal of Transplantation 2007; 7: 2587 2596 2593

Yao et al. I (24), the UCSF criteria, when applied to both pathologic and clinical (radiologic) staging in 144 patients, were better than the Milan criteria in discriminating prognosis after OLT. Marsh and Dvorchik from the University of Pittsburgh applied the UCSF criteria to their cohort of 393 patients who underwent OLT for HCC (20). Of the 265 patients who met UCSF criteria based on pathology, the 5-year recurrence-free survival was 94%. The Pittsburgh study, however, did not separately analyze recurrence-free survival in the subgroup of patients meeting the UCSF criteria but exceeding the Milan criteria. In a multi-center study from Korea involving 237 patients with HCC who underwent LDLT and 75 patients who received deceased donor livers (26), 7.3% of the patients met UCSF criteria but not Milan criteria. In the LDLT group, the 3-year survival rates for those either within or beyond the UCSF criteria were 90.6% and 58.5%, respectively (p < 0.001). In a National Institute of Health Single Topic Conference, Schwartz from Mount Sinai Hospital in New York tested the UCSF criteria according to explant tumor pathology in 353 patients (27). The 37 patients (10.5%) meeting UCSF criteria but exceeding Milan criteria had overall 5-year survival similar to patients meeting Milan criteria, but significantly better than those exceeding UCSF criteria. However, in terms of recurrence-free probability, the UCSF criteria fell short of the Milan criteria but were superior to those exceeding UCSF criteria. According to a recent report by Decaens et al. from 14 French transplant centers (28), 39 of the 461 patients (8.5%) had explant tumors beyond Milan but within UCSF criteria. Their 5-year survival rate was similar to the 184 patients meeting Milan criteria only (64% vs. 70%, p = 0.33), but significantly better than the 34% 5-year survival rate among 238 patients exceeding both criteria (p < 0.0001). However, based on an intention-to-treat principle and by pretransplant imaging data on 468 patients, the 44 patients (9.4%) meeting UCSF but exceeding Milan criteria had a 5-year intention-to-treat survival rate of 46%, compared to 60% for the 279 patients within Milan criteria (p = 0.1), and 35% for the 121 patients with pretransplant tumor stage beyond both criteria (p < 0.0001). Although the authors suggested that the UCSF criteria should not be used for patient selection based on a 5-year intentionto-treat survival of < 50% according to preoperative radiologic staging, the difference in survival between UCSF and Milan criteria did not achieve statistical significance (46% vs. 60%, p = 0.1). Furthermore, the 5-year cumulative risk of HCC recurrence was also not significantly different between the two groups (27% vs. 20%, p = 0.15) (28). Although the proposed UCSF criteria have been externally validated by pathologic analysis (20,24,26,27), our initial data need to be further validated by prospective application of the UCSF criteria to HCC staged by pretransplant imaging. In the present study using the UCSF criteria for patient selection for OLT, the 5-year patient survival without recurrence was 81% and the recurrence-free probability exceeded 90%. No significant difference was observed in the recurrence-free probabilities of the subgroup meeting UCSF criteria but exceeding Milan criteria versus those meeting Milan criteria. The ability of the UCSF criteria to discriminate prognosis after OLT was further supported by the observation that pathologic tumor stage exceeding the UCSF criteria, as a result of preoperative tumor under-staging, was associated with a significantly worse outcome compared to those with pathologic tumor stage within UCSF criteria. Thus the application of the UCSF criteria to preoperative staging still resulted in excellent posttransplant outcome with a similar risk for tumor recurrence and under-staging when compared to the Milan criteria. According to a study by Sotiropoulops et al., the accuracy of pretransplant staging based on the UCSF criteria appeared to be the same as the Milan criteria (29). Over two-thirds of our cohort received elective preoperative loco-regional therapies. Although a favorable impact of preoperative loco-regional therapy on outcome after OLT has not been proven (30 33), we cannot exclude in the present study a potential role of preoperative loco-regional therapy in preserving excellent posttransplant outcome following modest expansion of tumor size limits and possible selection bias in transplanting those with a favorable response to these treatments. Consequently, the potential pre- and posttransplant effects of loco-regional therapy must be further evaluated and factored into future considerations for expanding the current HCC criteria for OLT. Further studies are currently in progress at our center to evaluate the impact of radiologic and pathologic response to preoperative loco-regional therapy, rather than treatment itself, on outcome after OLT for HCC. In this study, vascular invasion was found to be an independent predictor of HCC recurrence after OLT in accord with the results of several other studies (33 35). The overall prevalence of micro- or macro-vascular invasion in our series was only 11%. In the Milan series (2), none had microvascular invasion, in sharp contrast to the 40% prevalence of micro-vascular invasion in the study by Jonas et al. from Germany in which 120 patients with HCC within Milan criteria underwent OLT (34). In a large series of patients with HCC who underwent resection, tumor size >5 cm correlated with a higher incidence of micro-vascular invasion compared with smaller tumors, thus providing a cautionary note for expansion of the conventional tumor size limits for OLT (36). Nevertheless, resection data may not be equally applicable to patients undergoing OLT, since patients awaiting OLT frequently receive elective loco-regional therapy and go through a selection process during which the most aggressive tumors may progress rapidly beyond acceptable criteria for OLT. In the present study, we found micro-vascular invasion but not poorly differentiated tumor grade to be strongly associated with advanced pathologic tumor stage beyond UCSF criteria (Figure 5). Neither vascular invasion nor poorly differentiated tumor grade were significantly more prevalent in patients with radiologic or pathologic tumor stage meeting UCSF criteria but exceeding Milan criteria compared to those meeting Milan 2594 American Journal of Transplantation 2007; 7: 2587 2596

UCSF Criteria and Liver Transplantion for HCC criteria only (Figures 5 and 6). It is possible that some of the patients with vascular invasion or poorly differentiated tumor grade had been removed from the waiting list as a result of tumor progression to partly account for the relatively low prevalence of these unfavorable histologic tumor features in our cohort. In the current era of increasing demand and unrelenting organ shortage, the foundation of the debate regarding expansion of the conventional criteria for HCC may ultimately rest upon what the transplant community would consider to be an acceptable survival after OLT for HCC (4). Several groups have proposed a 50% 5-year patient survival to be the minimum acceptable cut-off (8,28). Our results have surpassed this mark by a considerable margin. However, we recognize that the number of patients in this study with HCC exceeding the Milan criteria but meeting the UCSF criteria is relatively small, and that the median posttransplant follow-up of 26 months is still too short to assess the full impact of posttransplant HCC recurrence. Independent validation of our results in a large-scale study based on pretransplant imaging may be difficult under the current UNOS policy for organ allocation. Adult-to-adult, right lobe LDLT has been advocated as an alternative way to investigate the feasibility of expanded HCC criteria (8). However, whether LDLT will achieve the same results as deceased donor OLT for HCC meeting Milan criteria is still not clear (1,37 40). Furthermore, existing data on LDLT for HCC using expanded indications are scant and limited by the small sample size (21,39,40). In conclusion, we report favorable results following prospective selection of OLT candidates with HCC meeting the UCSF criteria by radiologic staging. The UCSF criteria offer the potential benefit of OLT to an additional 5 20% of patients with HCC (26 28) who would have otherwise been excluded from OLT under the more restrictive Milan criteria. The safety margin for choosing the UCSF criteria over the Milan criteria in the selection of candidates for OLT appears to be sufficiently wide with respect to posttransplant survival and tumor recurrence. The risk of tumor under-staging also does not appear to be significantly higher when the preoperative staging criteria are modestly expanded. Acknowledgments We thank Peter Bacchetti and Chengshi Jin for providing statistical assistance in the preparation of this manuscript. This work was supported in part by a grant from the National Institute of Health to the University of California, San Francisco Liver Center (P30DK26743). References 1. Befeler AS, Hayashi PH, Di Bisceglie AM. Liver transplantation for hepatocellular carcinoma. Gastroenterology 2005; 128: 1752 1764. 2. 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