Differences between radiology and histopathology: Are we judging correctly? Simona Onali 1, Emmanuel Tsochatzis 1, James O’Beirne 1, Aileen Marshall 1, TuVihn Luong 2, Massimo Pinzani 1, Pinelopi Manousou 1 1 The Royal Free Sheila Sherlock Liver Unit, Royal Free Hospital and UCL Institute for Liver and Digestive Health, London, UK 2 Department of Cellular Pathology, UCL Medical School, Royal Free Campus, London, UK

Background Selection criteria for OLT in patients with HCC are based on radiological assessment of the number and size of tumours  Retrospective studies about HCC patients undergoing LT reported different grade of discordance between radiological and histological findings 1-4. Factors other than tumour size and number may affect post-LT outcomes :  Pre-transplant aFP level >1000 ng/ml associated with higher risk of HCC recurrence even among patients transplanted within Milan criteria 5 1 Sotiropoulos 2005 Transplantation 2 Grasso 2006 Transplantation 3 Shah 2006 Transplantation 4 Chen 2009 HPB 5 Hameed 2014 Liver transplantation

Background Factors other than tumour size and number may affect post-LT outcomes : Pre-transplant aFP level >1000 ng/ml associated with higher risk of HCC recurrence even among patients transplanted within Milan criteria 1 strongest pre-transplant predictor of microvascular invasion New selection criteria based on the combination of size/number of HCC and aFP level have been proposed the French model (aFP model) 2 1Duvoux 2012 Gastroenterology

Aims of the study 1.To analyse any discrepancies between radiological reports and histopathological findings in the explant of patients undergoing LT for HCC 2.To identify potential factors that could predict HCC recurrence and survival post-LT between the parameters available during the pre-operative assessment period.

Patients and Methods All consecutive patients who underwent LT for HCC between January 1997 and February 2014 at the Royal Free Hospital Retrospectively: -Demographic and clinical data (sex, age, aetiology of liver disease, date of LT, pre-LT HCC treatment, aFP levels) -Pre-LT radiological findings (HCC number and size, macrovascular invasion, lymph-nodes invasion) -Histopathological findings on explant (HCC number and size, differentiation, micro/macrovascular invasion) -Donor characteristics -Immunosuppression type and levels -HCC recurrence and HCC-related death

Patients and Methods Discrepancy between radiological and histopathological findings was assessed comparing:  number of nodules  size of biggest nodule  fulfilment of Milan/UCSF criteria Patients were considered underestimated if: -nodule number and/or size was bigger on explant compared to imaging -they did not fulfil selection criteria on explant (in discordance to radiology) They were considered overestimated when the opposite occurred.

Results: baseline patients’ characheristics (n=185) Age median, range55 (27-68) Gender male150 (81%) Aetiology -HCV -HBV -ALD -Autoimmune -NAFLD -Other 101 (54%) 35 (19%) 28 (15%) 9 (5%) 7 (4%) 5 (3%) LT period (25%) 38 (20%) 59 (32%) 42 (23%) Pre-LT HCC treatment -Total -TAE/TACE -RFA -Ethanol injection -Resection -Combination 126 (68%) Pre-LT a-FP level (IU/L) median, range - > < (2-7936) 5 (3%) 25 (14%) 152 (83%)

Results: pre-LT imaging

Results: histopathological findings

RadiologyHistology O inside Milan O outside Milan O inside Milan O outside Milan

Discrepancy in selection criteria Classification according to Milan criteria Pathology Within Outside Radiologywithin 132/185 (71%)40/185(22%) outside4/185 (2%)9/185 (5%) Classification according to UCSF criteria Pathology Within Outside Radiologywithin 150/185 (81%)28/185(15%) outside2/185 (1%)5/185 (3%) 40/172 (23%) patients fell outside MC due to : number of HCC 10/40 size of HCC 20/40 both 10/40 28/178 (16%) patients fell outside UCSF criteria due to: number of HCC 19/28 size of HCC 5/28 both 4/28

Discrepancy between radiological and histological findings Number of nodules: 104/185 (56%) 78 underestimated - 60 pts -> 1-2 nodules - 18 pts -> 3-4 nodules 26 overestimated by 1-2 nodules -10 treated between imaging and LT Size of biggest nodule: 158/182 (87%) 93 underestimated - 44 pts -> difference ≥ 10 mm 65 overestimated - 25 pts -> difference ≥ 10mm

Results: HCC recurrence post-LT 29/185 (15.6%) developed HCC recurrence after a median of 37m post LT (4-157) 16 (55%) -> liver 13 (45%) -> metastatic disease involving lungs, bones or lymph nodes 20/29 died after a median of 48 m post-LT (7-157) Imaging Outside Milan criteria: 4/29 (14%) Outside UCSF criteria: 3/29 (10%) Histopathology Outside Milan criteria: 16/29 (55%) Outside UCSF criteria: 10/29 (35%)

Results: HCC recurrence post-LT 29/185 (15.6%) developed HCC recurrence after a median of 37m post LT (4-157) 16 (55%) -> liver 13 (45%) -> metastatic disease involving lungs, bones or lymph nodes 20/29 died after a median of 48 m post-LT (7-157) 16/29 (55%) outside Milan criteria according to explant 14/29 (48%) underestimated by imaging - size of biggest HCC (n=10) - n. of nodules and size of biggest (n=4) Vs 13/29 (45%) inside Milan criteria according to explant 3/29 (10%) had aFP >1000 IU/l -> vs 2/153 (1.3%) in non recurrence patients 13/29 (45%) had aFP >100 IU/l -> vs 17/153(11%) in non recurrence patients

Results: HCC recurrence post-LT 29/185 (15.6%) recurrence 20/29 (69 %) died aFP pre-LT ? aFP >1000 IU/l -> n=3/29 (10%) aFP > 100 IU/l -> n= 13/29 (45%) Versus aFP pre-LT in the non-recurrence ? aFP >1000 IU/l -> n=2/153 (1.3%) aFP >100 IU/l -> n= 17/153 (11%)

Results: discrepancy between radiology and histopathology in patients with HCC recurrence N. of nodules: 11/29 (38%) Size of biggest HCC: 22/ 29 (76%) Milan criteria : 14/29 (48%) - size of biggest HCC (n=10) - n. of nodules and size of biggest (n=4) UCSF criteria: 8/29 (28%) - size of biggest HCC or total size (n=3) - n. of nodules (n=2) - both size & number (n=3)

Log rank p<0.001 underestimated no discrepancy or overestimated Kaplan Meier curve for HCC recurrence according to Milan Criteria discrepancy

Results: predictors of HCC recurrence post- LT In Cox regression f actors significantly associated were: 1.aFP levels p=0.001, OR=4.1, 95% CI= Radiological size of biggest nodule p=0.001,OR=1.04, 95% CI= Number of nodules p=0.6 What about the number of nodules? Cut-off ?

Results: predictors of HCC recurrence post- LT In Cox regression f actors significantly associated were: 1.aFP levels p=0.001, OR=4.1, 95% CI= Radiological size of biggest nodule p=0.001,OR=1.04, 95% CI= Number of nodules p=0.6 What about the number of nodules? Cut-off ? aFP cut-off= 100 IU/l Sensitivity 41% Specificity 89% Cut-off of biggest nodule size = 3 cm Sensivity 62% Specificity 74%

Results: predictors of HCC recurrence post- LT aFP cut-off= 100 IU/l Sensitivity 41% Specificity 89% AUROC = p= % CI=0.6 – 0.8

Results: predictors of HCC recurrence post- LT Cut-off of biggest nodule diameter = 3 cm Sensivity 62% Specificity 74% AUROC =0.707 p=0.0001, 95% CI=0.6 – 0.8

…what if we combine aFP <100 IU/l and size of biggest nodule <3 cm ? AUROC=0.72, p=0.0001, 95% CI= Sensitivity=77% Specificity= 70%

We then compared our cut-offs with those proposed in the literature Total number of patientsRecurrence HCC aFP (IU/l) < (83%)16 (55%) (14%)10 (35%) >10005 (3%)3 (10%) Size of biggest nodule (cm) <3117 (76%)11 (38%) >336 (24%)18 (62%)

Kaplan Meier for HCC recurrence according to aFP and size of biggest HCC cut-offs aFP<100 and diam<3cm aFP>1000 and diam>3cm aFP 3cm aFP and diam<3cm aFP and diam>3cm Log rank p<0.001

Results: HCC recurrence post-LT 29/185 (15.6%) recurrence How many outside selection criteria on imaging? Outside Milan criteria: 4/29 (14%) Outside UCSF criteria: 3/29 (10%) Outside“aFP<100 IU/L + biggest nodule<3 cm”: 22/29 (76%)

Results: Survival post-LT 49/185 (27%) patients died after a mean of 42 months (6-160) post-LT HCC related 1-year survival 98%, 5-year survival 91% Overall related 1-year survival 95%, 5-year survival 80% Cause of death: HCC recurrence (41%) - median follow-up 40 months (7-157) - 1-year survival 89% - 5-year survival 53% de-novo malignancy (15%) decompensated cirrhosis (14%) sepsis (10%) chronic rejection (4%) Unknown, but not HCC-related (16%)

Results: predictors of HCC-related mortality post-LT Cox regression: pre-LT factors significantly associated: 1. aFP levels >100kU/l p<0.001, OR=5.1, 95% CI= Radiological size of biggest HCC p=0.021,OR=1.04, 95% CI=

aFP>1000 and diam>3cm aFP<100 and diam<3cm aFP 3cm aFP and diam<3cm aFP and diam>3cm Kaplan Meier for HCC-related survival according to aFP and size of biggest HCC cut-offs

Conclusions 1.Discordance between radiological and histological findings.  tumour progression between imaging and LT (tumour biology, waiting time)  innacurate imaging staging ? Does it really matter?  Recurrence occured in 35% of underestimated patients (vs 10% of the rest) according to Milan Criteria  4 (2%) overestimated according Milan criteria: how many patients we are overestimating and not listing ?

Conclusions 2. aFP and radiological diameter of the biggest HCC were the only pre-LT factors significantly associated with HCC recurrence and HCC related survival. A lower aFP cut-off of 100 IU/l showed higher sensitivity than the current in identifying HCC recurrence post LT Combination of aFP <100 Ku/l and size of biggest HCC <30 mm seems to perform better than the actual selection criteria. We do propose to consider patients with diameter>30mm for LT when aFP <100 IU/L.

Acknowledgements

Median TAC levels up to 30 days post LT was not associated with recurrence (p=0.6, OR=0.97, 95% CI )

aFP 400 IU/l Only 7 patients had aFP greater than 400 IU/l Cox regression: p=0.12, OR=2.4, 95% CI=

Underestimation over time In Milan criteria : 12/46 (26%) : 10/38 (26%) : 8/52 (15%) : 10/49 (20%) In number of nodules : 24/46 (52%) : 17/38 (45%) : 18/52 (34%) : 19/49 (38%) In size of biggest nodule : 29/46 (63%) : 21/38 (55%) : 25/52 (48%) : 18/49 (37%)