Impact of Histological Grade of Hepatocellular Carcinoma on the Outcome of Liver Transplantation FREE

HEPATOCELLULAR carcinoma (HCC) is a major cause of death worldwide, with a high incidence in Asia and South Africa.¹ More than 80% of patients with HCC have liver cirrhosis, particularly related to chronic viral hepatitis types B and C.² Although surgical resection of the tumor is considered the best option,³ it is possible in only a minority of patients owing to impaired functional reserve of the liver and multicentricity. To overcome the limited application of resection, other treatment modalities have been investigated.^4- 5

Use of orthotopic liver transplantation (OLT) for the treatment of HCC has been controversial. Although the results to date are comparable to those for conventional partial hepatectomy,^6- 7 the application of transplantation for treating cancer remains a difficult decision given the shortage of donor organs. Earlier studies^6,8 observed that patients with HCC did poorly compared with patients with benign disease. A recent experience,⁹ however, reported that patients with HCC, who were carefully selected on the basis of the size and number of lesions, did as well after OLT as patients without cancer.

In addition, data from a large international registry¹⁰ suggested that histological differentiation of the tumor was an important prognostic factor in the outcome of OLT for HCC. We describe our experience evaluating multiple factors, including clinicopathologic features, in the outcome of HCC after OLT.

Between June 1991 and January 1999, 952 liver transplantations were performed at Jackson Memorial Hospital/University of Miami Medical Center, Miami, Fla. A retrospective study revealed 56 OLT recipients with histologically proven HCC in the explant liver. Patients with so-called incidental tumor, ie, tumor not identified during pretransplant workup but found in explant liver, were also included in the study. Histological slides from 53 of the 56 patients were available for analysis, and these patients formed the study population. Data on demographics; pretransplant α-fetoprotein level; Child-Pugh class; presence of viral hepatitis types B and C; chemotherapy before and after OLT; size, number, and distribution of tumors; pTNM stage according to Union Internationale Contre le Cancer (UICC) criteria¹¹; histological differentiation of the tumor; presence of microscopic vascular invasion and lymph node metastasis; and donor bone marrow infusions were collected and analyzed.

Histological data on the tumor lesions of all patients were retrieved and reviewed by a single expert pathologist (M.B.) masked to the data on the outcome of patients after undergoing OLT. Histological differentiation was defined by the criteria of Kondo¹² and Sugihara et al,¹³ which classify HCC into 3 categories: well, moderately, and poorly differentiated. These classifications have overlapping and similar definitions to the Edmondson-Steiner classification.¹⁴

Overall survival and disease-free survival rates were calculated using the Kaplan-Meier method,¹⁵ and statistical significance was defined using the log-rank test.¹⁶ Factors related to survival were analyzed using the Cox proportional hazards regression model¹⁷ and SAS statistical software (SAS Institute Inc, Cary, NC). Statistical significance was defined as P<.05, corresponding to the test of null hypothesis that all coefficients associated with the risk factor in the regression are zero. Comparison of continuous ordinal data between 2 groups was performed using the Mann-Whitney U test.

Of 53 patients in the study, 36 (68%) were men and 17 (32%) were women. Mean age was 57 years (range, 20-77 years). Most patients (n = 44 [83%]) had viral hepatitis, 37 (70%) of whom were hepatitis C virus antibody positive. Twenty patients (38%) had Child-Pugh class B and 20 (38%) had Child-Pugh class C cirrhosis. Mean tumor size was 4.4 cm (range, 0.5-18 cm). After OLT, HCC (incidental tumor) was found in the explant livers of 13 patients (25%). Mean tumor size in incidental cases was 2.5 cm (range, 0.5-6.0 cm). Twenty-four patients (45%) received donor bone marrow infusions for induction of immunologic tolerance after OLT. Other demographic features are summarized in Table 1.

Median follow-up was 709 days and mean follow-up was 887 days, with the longest survival exceeding 8 years (2976 days to date). Of 53 patients, 17 (32%) died and 36 (68%) are currently alive. Four patients died of systemic infection, 1 of chronic rejection, and 1 of an acute cardiac event. None died of recurrent hepatitis. Overall survival at 1, 3, and 5 years was 79%, 65%, and 61%, respectively, whereas survival for patients with tumor size of 5 cm or less was 87%, 77%, and 71%, respectively. One-, 3-, and 5-year disease-free survival was 77%, 60%, and 60%, respectively (Figure 1). As of January 1999, there was no evidence of recurrence of HCC in 15 patients who had survived longer than 3 years. Of 14 patients (26%) in whom HCC recurred, 11 died of metastatic disease. One patient with recurrence died of systemic infection. Two patients who experienced recurrence were alive and well as of January 1999. Liver (n = 4 [29%]), lung (n = 3 [21%]), and bone (n = 2 [14%]) were the common sites of recurrent or metastatic disease. At the end of the study period, 34 patients (64%) were alive and free of disease.

Univariate analysis regarding survival was performed with the following factors: age, sex, presence of viral hepatitis, incidental tumor, pre-OLT α-fetoprotein level, Child-Pugh class, size of the tumor, presence of multiple tumors, presence of bilobar tumor, microscopic vascular invasion, microscopic lymph node metastasis, histological differentiation, UICC pTNM stage based on microscopic findings, donor bone marrow infusion, and chemotherapy (transarterial chemoembolization and intravenous chemotherapy). Eight factors—incidental tumor, size, multiple tumor, bilobar tumor, vascular invasion, lymph node metastasis, histological differentiation, and pTNM stage—significantly affected survival (P<.05, log-rank test). Bone marrow infusion after OLT did not affect survival (Table 1).

To elucidate the independent factors affecting survival, multivariate analysis using the Cox proportional hazards model was performed with the following factors: age, sex, presence of viral hepatitis, pre-OLT α-fetoprotein level, Child-Pugh class, size of the tumor, presence of multiple tumors, presence of bilobar tumor, microscopic vascular invasion, microscopic lymph node metastasis, histological differentiation, UICC pTNM stage based on microscopic findings, donor bone marrow infusion, and chemotherapy (transarterial chemoembolization and intravenous chemotherapy). Multivariate analysis revealed histological differentiation and UICC pTNM stage to be the independent factors affecting survival (Table 2). Further analysis without UICC pTNM stage revealed histological differentiation and tumor size to be the independent factors affecting survival (Table 2).

The outcome of patients with tumors larger than 5 cm with poorly differentiated HCC (range, 7-18 cm; median, 9.25 cm) was extremely poor (0% survival within 1 year of OLT). Patients with well- to moderately differentiated HCC with tumors larger than 5 cm (range, 5.5-15 cm; median, 6.25 cm) presented with unexpectedly good 3-year survival of 62% (Figure 2). Six of 8 patients in this group (well- to moderately differentiated tumors >5 cm) are alive without recurrence of tumors at a median follow-up of 38 months (range, 8 months to 8 years). These 6 patients all had a single nodule measuring 5.5 to 8 cm (median, 6.25 cm).

Patients with small tumors did well, with 3-year survival of 82% among patients with well- to moderately differentiated HCC and 67% among patients with poorly differentiated HCC. This difference in survival within the small tumor group was not statistically significant (P = .26, log-rank test) (Figure 3).

Univariate analysis for disease-free survival was performed with the factors studied in the survival analysis. Seven factors—incidental tumor, size, bilobar tumor, vascular invasion, lymph node metastasis, histological differentiation, and pTNM stage—showed a statistically significant difference (P<.05, log-rank test). Bone marrow infusion did not affect disease-free survival (Table 3). To characterize the independent factors affecting disease-free survival, multivariate analysis with Cox proportional hazards model was performed similarly to the overall survival analysis. Histological differentiation and microscopic lymph node metastasis were noted as the independent factors (Table 4). Excluding lymph node metastasis, only histological differentiation remained a significant factor (Table 4). To further understand the effect of histological differentiation of HCC on prognosis, recurrence was studied according to differentiation of the tumor. Nine of 15 patients with poorly differentiated HCC died; 7 of these deaths were directly related to recurrent disease. On the other hand, 8 of 38 patients with well- to moderately differentiated HCC died; 4 of these deaths were directly related to recurrent disease (Table 5). When combined with size criteria (Table 5), 5 of 6 patients with large (>5 cm), poorly differentiated HCC died of recurrence, although only 2 of 8 patients with large but well- to moderately differentiated HCC died of recurrence. When disease-free interval was compared in patients with recurrence based on histological differentiation, it was significantly greater in patients with well- to moderately differentiated HCC. The median disease-free interval of patients with well- to moderately differentiated HCC was 430 days, whereas that of patients with poorly differentiated HCC was 175 days (P = .006, Mann-Whitney U test) (Figure 4).

Previously, significant prognostic factors after OLT for HCC have included features such as size, number, and distribution of the tumor; nodal metastasis; vascular invasion; and TNM stage based on microscopic findings.^7,9,18- 21 The role of histological differentiation of the tumor was not studied.^7,9,18- 21 To date, to our knowledge, only one published study¹⁰ from an international registry has suggested that histological differentiation is a significant and independent prognostic factor after OLT for patients with HCC. This study¹⁰ was viewed critically by other investigators mainly because it was based on observations of different pathologists from multiple transplant centers. Our study is from the patient records of a single center where all the histological slides were collected and evaluated by a single pathologist, thereby excluding major intraobserver variability in histological differentiation.

The role of OLT in the management of HCC has been controversial. Orthotopic liver transplantation has an advantage over partial resection in that it can be applied regardless of the functional reserve of the liver. Also, it is now known that there is a group of patients with HCC that presents with multicentric, metachronous hepatocarcinogenesis.^22- 24 Total hepatectomy performed at the time of OLT allows removal of a potentially carcinogenic liver. Long-term results from earlier studies^6,8 were, nevertheless, disappointing. These series noted 20% to 30% overall 5-year survival, a high recurrence rate, and occasional long-term survival free of recurrent disease. Further studies^7,18- 20,25 revealed that successful transplantation for HCC yielding results comparable to conventional partial hepatectomy can be achieved in patients with early-stage disease. These results were encouraging and suggested that OLT should be added to the therapeutic spectrum for HCC in selected patients. The most recent studies,^9,26 based on strict criteria for the size and number of tumors, observed 4-year actual survival of 75% and recurrence-free survival of 83%⁹ or 5-year survival of 74%. Similarly, our overall survival of 78% and disease-free survival of 69% in 3 years for patients with tumor size of 5 cm or less reinforces the recommendation of OLT for such patients. Among patients with low-grade (well- to moderately differentiated) HCC, even better 3-year survival of 81% was observed. Furthermore, patients with large (>5 cm) but well-differentiated tumors demonstrated 3-year survival of 62%, suggesting that these patients might also be candidates for OLT in selected circumstances. On the contrary, patients with large (>5 cm) poorly differentiated tumors had a dismal prognosis. In our study, most of these patients presented with recurrence within a short period, and none survived more than a year.

Our single-center study reaffirms the findings of the multicenter study and suggests that, aside from size and multiplicity, histological differentiation of the tumor is a statistically significant factor affecting prognosis. In our multivariate analysis, histological differentiation of the tumor, an indicator of biological aggressiveness and progression,^13,27 was an independent, statistically significant factor affecting survival. Histological differentiation of the tumor was also a statistically significant factor in multivariate analysis for disease-free survival. From these results, it seems that indication for OLT should not be defined solely by size of the tumor but also with consideration of biological aggressiveness of the tumor cells as indicated by histological differentiation.

Orthotopic liver transplantation for HCC has several problems that must be considered. Waiting time for a graft is often 1 year or longer, during which HCC might grow and progress. To circumvent this problem, adjunctive measures might be necessary. Effective adjuvant therapy to prevent further progression of the tumor could be applied. Because most OLT candidates have poor functional reserve of the liver, nonsurgical local ablation modalities such as thermal ablation^28- 29 or percutaneous ethanol injection therapy³⁰ might be useful. In our series, a small number of patients received transarterial chemoembolization before OLT and chemotherapy after OLT. Although a previous study³¹ suggested the beneficial effect of chemotherapy or adjuvant therapy combined with OLT for HCC, this was not clearly demonstrated in our series, probably because of the small number of patients. Further prospective studies are required. More attractive options, such as living-related liver transplantation,³² split-liver transplantation,³³ or domino transplantation,^34- 35 should be serious considerations because these measures can effectively reduce waiting time.

In our experience, histological differentiation had a statistically significant effect on prognosis. Patients with small-sized HCC, regardless of histological grading, should be acceptable candidates for OLT. For relatively large tumors (slightly greater than 5 cm in diameter), OLT should not be denied solely on the basis of the size of the tumor. Because ultrasound-guided biopsy is a reliable and relatively safe method for obtaining histological information,³⁶ we suggest a preoperative biopsy to determine the histological differentiation should the HCC be large. Nonetheless, such an approach might have inherent, albeit small, risks of bleeding and tumor seeding. The reported rates of tumor implantation attributable to needle biopsies have varied from 0.005% to 5.1%,^37- 38 with most of the implanted tumors occurring in the chest or the abdominal wall. Furthermore, tumor differentiation obtained through needle biopsy may not be well appreciated because of the small sample size. An alternative, therefore, may be an open or laparoscopic biopsy that can only be applied for superficial lesions. For deeper intraparenchymal lesions, an intraoperative ultrasound-guided approach seems appropriate.

In conclusion, treatment protocols for HCC with OLT with or without nonsurgical modalities should consider histological grade of the tumor as a possible factor affecting survival. In addition, before we alter patient selection criteria, prospective trials are needed to confirm the impact of histological grade on the outcome of OLT for HCC.

Corresponding author and reprints: Tomoaki Kato, MD, Division of Transplantation, Department of Surgery, University of Miami School of Medicine, 1801 NW Ninth Ave, Highland Professional Building, Suite 511, Miami, FL 33136.