Reviewer's report 
 Hepatocellular carcinoma and the penetrance of HFE C282Y mutations: a cross sectional Title: 
study. 
 1  14 March 2005 Version: Date: 
 Ernest Beutler Reviewer: 
Reviewer's report: 
General 
This manuscript reports the frequency of HFE genotypes among the 144 cases of hepatocellular carcinoma (HCC) collected from two hospitals in East Anglia over a 30 year span. The frequency of C282Y homozygotes among these cases is then applied to data on annual incidence of HCC and 
frequency of C282Y homozygosity among the overall population to calculate the lifetime risk of HCC 
among C282Y homozygotes, which is estimated to be 1.31% (0.52%-3.32%). The calculations were 
limited to males as none of the 8 C282Y homozygotes among the 144 HCC cases were female. 
The manuscript addresses a very important question, ie. what is the penetrance of the C282Y 
homozygous genotype. The approach they have used is a more robust than most, given that the 
phenotype is well-defined, allowing a relatively unbiased estimate of penetrance. However, the low incidence of HCC makes collection of these data difficult. Because the numbers are by necessity small, the variance of their estimate is large. 
This is the third paper by this group reporting similar data on hepatocellular carcinoma in C282Y 
homozygotes. As noted by the authors, the current manuscript includes some cases included in the prior publications. In the first of these, published in Lancet in 1997, a penetrance of 0.4% based on 2/28 homozygotes found and extrapolated to the 49 HCC cases in the series was reported. In the 
second, published in Gut in 2000, the penetrance was estimated to be 1.1% based on 3 
homozygotes found among 34 HCC cases. 
Although the current paper reports on a larger number of cases, the numbers are still quite small and 
the confidence interval around the percentage of homozygotes (8/102 male HCC cases or 7.8%) is 
therefore of necessity still very large. Compounding the problem is that fact that the calculation of 
lifetime risk for homozygotes is dependent on several other estimates, the variances of which do not 
appear to be taken into account: the 1.26/100,000 annual incidence among males in the total 
population, the 0.6% frequency of C282Y homozygotes in the population, and the number of 
subjects in the total population reaching the age of 54 annually (1260/100,000). Taking the variance 
of all the estimates used in the final calculation would make the variance of the overall estimate even larger. 
------------------------------------------------------------------------------- 
Major Compulsory Revisions (that the author must respond to before a decision on publication can be reached) 
The authors cite two papers finding a similar percentage of C282Y homozygotes among HCC cases, 
one reporting 2 homozygotes/35 HCC cases in France (Blanc JF, et al. J Hepatol 2000) and the 
other reporting 5 homozygotes/162 HCC cases in Austria (Cauza E, et al. Am J Gastroenterol 2003), 
but do not cite another French study reporting 0 homozygotes/133 HCC cases (Boige V, et al. Lack 
of association between HFE gene mutations and hepatocellular carcinoma in patients with cirrhosis. 
Gut 2003;52:1178-1181). This report should be cited, and discussed. Three other Italian and 
Spanish studies reporting no homozygotes among HCC cases are dismissed as having small populations with low HFE C282Y prevalence. Given the major risk of HCC associated with viral hepatitis, it is surprising that the authors did not 
test for these factors to analyze their interaction with C282Y homozygosity in this population. 
------------------------------------------------------------------------------- 
Minor Essential Revisions (such as missing labels on figures, or the wrong use of a term, which the 
author can be trusted to correct) 
------------------------------------------------------------------------------- 
Discretionary Revisions (which the author can choose to ignore) 
none 
The review of this manuscript for was a joint effort of Jill Waalen MD MPH and Prof Ernest Beutler 
 Accept after minor essential revisions What next?: 
 An article whose findings are important to those with closely related research Level of interest: 
interests 
 Acceptable Quality of written English: 
 No Statistical review: 
Declaration of competing interests: 
We declare that we have no competing interests 
Reviewer's report 
 Hepatocellular carcinoma and the penetrance of HFE C282Y mutations: a cross sectional Title: 
study. 
 1  14 March 2005 Version: Date: 
 Ernest Beutler Reviewer: 
Reviewer's report: 
General 
This manuscript reports the frequency of HFE genotypes among the 144 cases of hepatocellular carcinoma (HCC) collected from two hospitals in East Anglia over a 30 year span. The frequency of C282Y homozygotes among these cases is then applied to data on annual incidence of HCC and 
frequency of C282Y homozygosity among the overall population to calculate the lifetime risk of HCC 
among C282Y homozygotes, which is estimated to be 1.31% (0.52%-3.32%). The calculations were 
limited to males as none of the 8 C282Y homozygotes among the 144 HCC cases were female. 
The manuscript addresses a very important question, ie. what is the penetrance of the C282Y 
homozygous genotype. The approach they have used is a more robust than most, given that the 
phenotype is well-defined, allowing a relatively unbiased estimate of penetrance. However, the low incidence of HCC makes collection of these data difficult. Because the numbers are by necessity small, the variance of their estimate is large. 
This is the third paper by this group reporting similar data on hepatocellular carcinoma in C282Y 
homozygotes. As noted by the authors, the current manuscript includes some cases included in the prior publications. In the first of these, published in Lancet in 1997, a penetrance of 0.4% based on 2/28 homozygotes found and extrapolated to the 49 HCC cases in the series was reported. In the 
second, published in Gut in 2000, the penetrance was estimated to be 1.1% based on 3 
homozygotes found among 34 HCC cases. 
Although the current paper reports on a larger number of cases, the numbers are still quite small and 
the confidence interval around the percentage of homozygotes (8/102 male HCC cases or 7.8%) is 
therefore of necessity still very large. Compounding the problem is that fact that the calculation of 
lifetime risk for homozygotes is dependent on several other estimates, the variances of which do not 
appear to be taken into account: the 1.26/100,000 annual incidence among males in the total 
population, the 0.6% frequency of C282Y homozygotes in the population, and the number of 
subjects in the total population reaching the age of 54 annually (1260/100,000). Taking the variance 
of all the estimates used in the final calculation would make the variance of the overall estimate even larger. 
------------------------------------------------------------------------------- 
Major Compulsory Revisions (that the author must respond to before a decision on publication can be reached) 
The authors cite two papers finding a similar percentage of C282Y homozygotes among HCC cases, 
one reporting 2 homozygotes/35 HCC cases in France (Blanc JF, et al. J Hepatol 2000) and the 
other reporting 5 homozygotes/162 HCC cases in Austria (Cauza E, et al. Am J Gastroenterol 2003), 
but do not cite another French study reporting 0 homozygotes/133 HCC cases (Boige V, et al. Lack 
of association between HFE gene mutations and hepatocellular carcinoma in patients with cirrhosis. 
Gut 2003;52:1178-1181). This report should be cited, and discussed. Three other Italian and 
Spanish studies reporting no homozygotes among HCC cases are dismissed as having small populations with low HFE C282Y prevalence. Given the major risk of HCC associated with viral hepatitis, it is surprising that the authors did not 
test for these factors to analyze their interaction with C282Y homozygosity in this population. 
------------------------------------------------------------------------------- 
Minor Essential Revisions (such as missing labels on figures, or the wrong use of a term, which the 
author can be trusted to correct) 
------------------------------------------------------------------------------- 
Discretionary Revisions (which the author can choose to ignore) 
none 
The review of this manuscript for was a joint effort of Jill Waalen MD MPH and Prof Ernest Beutler 
 Accept after minor essential revisions What next?: 
 An article whose findings are important to those with closely related research Level of interest: 
interests 
 Acceptable Quality of written English: 
 No Statistical review: 
Declaration of competing interests: 
We declare that we have no competing interests 
Please review this paper.
user
Reviewer's report 
 Hepatocellular carcinoma and the penetrance of HFE C282Y mutations: a cross sectional Title: 
study. 
 1  14 March 2005 Version: Date: 
 Ernest Beutler Reviewer: 
Reviewer's report: 
General 
This manuscript reports the frequency of HFE genotypes among the 144 cases of hepatocellular carcinoma (HCC) collected from two hospitals in East Anglia over a 30 year span. The frequency of C282Y homozygotes among these cases is then applied to data on annual incidence of HCC and 
frequency of C282Y homozygosity among the overall population to calculate the lifetime risk of HCC 
among C282Y homozygotes, which is estimated to be 1.31% (0.52%-3.32%). The calculations were 
limited to males as none of the 8 C282Y homozygotes among the 144 HCC cases were female. 
The manuscript addresses a very important question, ie. what is the penetrance of the C282Y 
homozygous genotype. The approach they have used is a more robust than most, given that the 
phenotype is well-defined, allowing a relatively unbiased estimate of penetrance. However, the low incidence of HCC makes collection of these data difficult. Because the numbers are by necessity small, the variance of their estimate is large. 
This is the third paper by this group reporting similar data on hepatocellular carcinoma in C282Y 
homozygotes. As noted by the authors, the current manuscript includes some cases included in the prior publications. In the first of these, published in Lancet in 1997, a penetrance of 0.4% based on 2/28 homozygotes found and extrapolated to the 49 HCC cases in the series was reported. In the 
second, published in Gut in 2000, the penetrance was estimated to be 1.1% based on 3 
homozygotes found among 34 HCC cases. 
Although the current paper reports on a larger number of cases, the numbers are still quite small and 
the confidence interval around the percentage of homozygotes (8/102 male HCC cases or 7.8%) is 
therefore of necessity still very large. Compounding the problem is that fact that the calculation of 
lifetime risk for homozygotes is dependent on several other estimates, the variances of which do not 
appear to be taken into account: the 1.26/100,000 annual incidence among males in the total 
population, the 0.6% frequency of C282Y homozygotes in the population, and the number of 
subjects in the total population reaching the age of 54 annually (1260/100,000). Taking the variance 
of all the estimates used in the final calculation would make the variance of the overall estimate even larger. 
------------------------------------------------------------------------------- 
Major Compulsory Revisions (that the author must respond to before a decision on publication can be reached) 
The authors cite two papers finding a similar percentage of C282Y homozygotes among HCC cases, 
one reporting 2 homozygotes/35 HCC cases in France (Blanc JF, et al. J Hepatol 2000) and the 
other reporting 5 homozygotes/162 HCC cases in Austria (Cauza E, et al. Am J Gastroenterol 2003), 
but do not cite another French study reporting 0 homozygotes/133 HCC cases (Boige V, et al. Lack 
of association between HFE gene mutations and hepatocellular carcinoma in patients with cirrhosis. 
Gut 2003;52:1178-1181). This report should be cited, and discussed. Three other Italian and 
Spanish studies reporting no homozygotes among HCC cases are dismissed as having small populations with low HFE C282Y prevalence. Given the major risk of HCC associated with viral hepatitis, it is surprising that the authors did not 
test for these factors to analyze their interaction with C282Y homozygosity in this population. 
------------------------------------------------------------------------------- 
Minor Essential Revisions (such as missing labels on figures, or the wrong use of a term, which the 
author can be trusted to correct) 
------------------------------------------------------------------------------- 
Discretionary Revisions (which the author can choose to ignore) 
none 
The review of this manuscript for was a joint effort of Jill Waalen MD MPH and Prof Ernest Beutler 
 Accept after minor essential revisions What next?: 
 An article whose findings are important to those with closely related research Level of interest: 
interests 
 Acceptable Quality of written English: 
 No Statistical review: 
Declaration of competing interests: 
We declare that we have no competing interests 
assistant