Reviewer's report
Title:Rapid Induction of Orthotopic Hepatocellular Carcinoma in
Immune-competent Rats by Ultrasound-guided Method and the Subsequent
Metronomic Chemotherapy
Version: 2Date:13 May 2010
Reviewer: Urban Emmenegger
Reviewer's report:
The manuscript by Chan et al describes a novel hepatocellular carcinoma (HCC)
model characterized by US-guided orthotopic implantation of syngeneic N1-S1
rat HCC cells into Sprague-Dawley rats, and a reasonable macroscopic tumor
take-rate of 60% after 10 days (similar to the take rate seen in rats undergoing
open implantation by laparotomy). The authors find an excellent correlation
between tumor size assessments using US, caliper measurements, functional
(perfusion) CT and blood biochemistry (GOT). Furthermore, the tumor model
appears to be susceptible to metronomic epirubicin therapy. Thus, this novel
orthotopic N1-S1 HCC model may be useful for the preclinical development of
novel treatment modalities for HCC, an important tumor entity from a global
perspective with limited treatment options especially for advanced disease.
The aforementioned positive aspects of the HCC model presented by Chan et al
by far outweigh certain limitations, such as the absence of cirrhotic changes
commonly seen in HCC patients, and the relatively short window of opportunity
for testing novel therapies given the rapid and aggressive tumor growth kinetics
observed.
Major Compulsory Revisions
Chan et al apply ‘metronomic epirubicin’ therapy. However, there is a lack of
rational given for the epirubicin dose chosen. By assuming a mean weight of 150
g per rat, the dose applied is 2 mg/kg/d, or 14 mg/kg given over 7 days (i.e.,
approximately 42 mg/sqm/week). This appears to be a relatively high dose of
epirubicin, which is also reflected by the degree of hematological toxicity
documented in Figure 4. Furthermore, one of the characteristics of metronomic
chemotherapy is the long-term administration of such treatment, which is clearly
not the case in the experiments by Chan et al. Admittedly, the definition of
metronomic chemotherapy remains vague.
Perpendicular tumor size measurements are commonly used to calculate tumor
volumes. Did the authors attempt such calculations by using the caliper/US/CT
data? And if yes: were the findings consistent with the data shown in this
manuscript using the mean of two perpendicular tumor diameters?
The authors apply functional (perfusion) CT scanning but do not show any
perfusion data – is there a special reason for this? Such data could confirm the
microvessel density changes presented in Figure 6c.
Minor Essential Revisions
Methods: The origin of the hepatocytes ‘clone 9’ used in Fig 5a should be
specified.
Methods: Under ‘Immunohistochemistry Analysis’ Chan et al list an
anti-PECAM-1 and an anti-CD31 antibody, yet CD31 and PECAM-1 are two
names for the same molecule. How many antibodies were used? The
terminology used in the Method/Results/Figure Legend sections as well as the
label of Figure 6c should be addressed to avert confusion.
Figure 1a: An arrow depicting the needle track and outlining the liver border
would be helpful.
Figure 1d: How many rats per group were used for this analysis?
Figure 3a: According to the text rats were sacrificed on day 17, not day 19 as
indicated in Figure 3a.
Discretionary Revisions
The future reader may be interested in whether rats without macroscopic tumor
growth 10 days after inoculation developed tumors at a later stage.
Abstract: ‘Microvessel density’ may be a more appropriate term to describe the
results presented in Figure 6c than ‘angiogenesis index’.
Background: Although Hanahan et al coined the term ‘metronomic’, the concept
of metronomic chemotherapy was described by others (Browder et al Cancer
Res. 2000;60(7):1878-86; Klement et al J Clin Invest. 2000;105(8):R15-24).
Level of interest: An article whose findings are important to those with closely
related research interests
Quality of written English: Needs some language corrections before being
published
Statistical review: No, the manuscript does not need to be seen by a
statistician.
Declaration of competing interests:
I declare that I have no competing interests.
Reviewer's report
Title:Rapid Induction of Orthotopic Hepatocellular Carcinoma in
Immune-competent Rats by Ultrasound-guided Method and the Subsequent
Metronomic Chemotherapy
Version: 2Date:13 May 2010
Reviewer: Urban Emmenegger
Reviewer's report:
The manuscript by Chan et al describes a novel hepatocellular carcinoma (HCC)
model characterized by US-guided orthotopic implantation of syngeneic N1-S1
rat HCC cells into Sprague-Dawley rats, and a reasonable macroscopic tumor
take-rate of 60% after 10 days (similar to the take rate seen in rats undergoing
open implantation by laparotomy). The authors find an excellent correlation
between tumor size assessments using US, caliper measurements, functional
(perfusion) CT and blood biochemistry (GOT). Furthermore, the tumor model
appears to be susceptible to metronomic epirubicin therapy. Thus, this novel
orthotopic N1-S1 HCC model may be useful for the preclinical development of
novel treatment modalities for HCC, an important tumor entity from a global
perspective with limited treatment options especially for advanced disease.
The aforementioned positive aspects of the HCC model presented by Chan et al
by far outweigh certain limitations, such as the absence of cirrhotic changes
commonly seen in HCC patients, and the relatively short window of opportunity
for testing novel therapies given the rapid and aggressive tumor growth kinetics
observed.
Major Compulsory Revisions
Chan et al apply ‘metronomic epirubicin’ therapy. However, there is a lack of
rational given for the epirubicin dose chosen. By assuming a mean weight of 150
g per rat, the dose applied is 2 mg/kg/d, or 14 mg/kg given over 7 days (i.e.,
approximately 42 mg/sqm/week). This appears to be a relatively high dose of
epirubicin, which is also reflected by the degree of hematological toxicity
documented in Figure 4. Furthermore, one of the characteristics of metronomic
chemotherapy is the long-term administration of such treatment, which is clearly
not the case in the experiments by Chan et al. Admittedly, the definition of
metronomic chemotherapy remains vague.
Perpendicular tumor size measurements are commonly used to calculate tumor
volumes. Did the authors attempt such calculations by using the caliper/US/CT
data? And if yes: were the findings consistent with the data shown in this
manuscript using the mean of two perpendicular tumor diameters?
The authors apply functional (perfusion) CT scanning but do not show any
perfusion data – is there a special reason for this? Such data could confirm the
microvessel density changes presented in Figure 6c.
Minor Essential Revisions
Methods: The origin of the hepatocytes ‘clone 9’ used in Fig 5a should be
specified.
Methods: Under ‘Immunohistochemistry Analysis’ Chan et al list an
anti-PECAM-1 and an anti-CD31 antibody, yet CD31 and PECAM-1 are two
names for the same molecule. How many antibodies were used? The
terminology used in the Method/Results/Figure Legend sections as well as the
label of Figure 6c should be addressed to avert confusion.
Figure 1a: An arrow depicting the needle track and outlining the liver border
would be helpful.
Figure 1d: How many rats per group were used for this analysis?
Figure 3a: According to the text rats were sacrificed on day 17, not day 19 as
indicated in Figure 3a.
Discretionary Revisions
The future reader may be interested in whether rats without macroscopic tumor
growth 10 days after inoculation developed tumors at a later stage.
Abstract: ‘Microvessel density’ may be a more appropriate term to describe the
results presented in Figure 6c than ‘angiogenesis index’.
Background: Although Hanahan et al coined the term ‘metronomic’, the concept
of metronomic chemotherapy was described by others (Browder et al Cancer
Res. 2000;60(7):1878-86; Klement et al J Clin Invest. 2000;105(8):R15-24).
Level of interest: An article whose findings are important to those with closely
related research interests
Quality of written English: Needs some language corrections before being
published
Statistical review: No, the manuscript does not need to be seen by a
statistician.
Declaration of competing interests:
I declare that I have no competing interests.
Please review this paper.
user
Reviewer's report
Title:Rapid Induction of Orthotopic Hepatocellular Carcinoma in
Immune-competent Rats by Ultrasound-guided Method and the Subsequent
Metronomic Chemotherapy
Version: 2Date:13 May 2010
Reviewer: Urban Emmenegger
Reviewer's report:
The manuscript by Chan et al describes a novel hepatocellular carcinoma (HCC)
model characterized by US-guided orthotopic implantation of syngeneic N1-S1
rat HCC cells into Sprague-Dawley rats, and a reasonable macroscopic tumor
take-rate of 60% after 10 days (similar to the take rate seen in rats undergoing
open implantation by laparotomy). The authors find an excellent correlation
between tumor size assessments using US, caliper measurements, functional
(perfusion) CT and blood biochemistry (GOT). Furthermore, the tumor model
appears to be susceptible to metronomic epirubicin therapy. Thus, this novel
orthotopic N1-S1 HCC model may be useful for the preclinical development of
novel treatment modalities for HCC, an important tumor entity from a global
perspective with limited treatment options especially for advanced disease.
The aforementioned positive aspects of the HCC model presented by Chan et al
by far outweigh certain limitations, such as the absence of cirrhotic changes
commonly seen in HCC patients, and the relatively short window of opportunity
for testing novel therapies given the rapid and aggressive tumor growth kinetics
observed.
Major Compulsory Revisions
Chan et al apply ‘metronomic epirubicin’ therapy. However, there is a lack of
rational given for the epirubicin dose chosen. By assuming a mean weight of 150
g per rat, the dose applied is 2 mg/kg/d, or 14 mg/kg given over 7 days (i.e.,
approximately 42 mg/sqm/week). This appears to be a relatively high dose of
epirubicin, which is also reflected by the degree of hematological toxicity
documented in Figure 4. Furthermore, one of the characteristics of metronomic
chemotherapy is the long-term administration of such treatment, which is clearly
not the case in the experiments by Chan et al. Admittedly, the definition of
metronomic chemotherapy remains vague.
Perpendicular tumor size measurements are commonly used to calculate tumor
volumes. Did the authors attempt such calculations by using the caliper/US/CT
data? And if yes: were the findings consistent with the data shown in this
manuscript using the mean of two perpendicular tumor diameters?
The authors apply functional (perfusion) CT scanning but do not show any
perfusion data – is there a special reason for this? Such data could confirm the
microvessel density changes presented in Figure 6c.
Minor Essential Revisions
Methods: The origin of the hepatocytes ‘clone 9’ used in Fig 5a should be
specified.
Methods: Under ‘Immunohistochemistry Analysis’ Chan et al list an
anti-PECAM-1 and an anti-CD31 antibody, yet CD31 and PECAM-1 are two
names for the same molecule. How many antibodies were used? The
terminology used in the Method/Results/Figure Legend sections as well as the
label of Figure 6c should be addressed to avert confusion.
Figure 1a: An arrow depicting the needle track and outlining the liver border
would be helpful.
Figure 1d: How many rats per group were used for this analysis?
Figure 3a: According to the text rats were sacrificed on day 17, not day 19 as
indicated in Figure 3a.
Discretionary Revisions
The future reader may be interested in whether rats without macroscopic tumor
growth 10 days after inoculation developed tumors at a later stage.
Abstract: ‘Microvessel density’ may be a more appropriate term to describe the
results presented in Figure 6c than ‘angiogenesis index’.
Background: Although Hanahan et al coined the term ‘metronomic’, the concept
of metronomic chemotherapy was described by others (Browder et al Cancer
Res. 2000;60(7):1878-86; Klement et al J Clin Invest. 2000;105(8):R15-24).
Level of interest: An article whose findings are important to those with closely
related research interests
Quality of written English: Needs some language corrections before being
published
Statistical review: No, the manuscript does not need to be seen by a
statistician.
Declaration of competing interests:
I declare that I have no competing interests.
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