Utility of Liver Biopsy in the Diagnosis and Management of Possible Drug-Induced Liver Injury in Patients Receiving Antituberculosis Therapy: A Retrospective Study

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

In the article "Role of liver biopsy in Drug-Induced Liver Injury from Anti- Tuberculosis Treatment: a retrospective cohort study" Gualano an co-workers describe 10 cases of putative tuberculostatic drug-induced liver injury who underwent liver biopsy and discuss the utility of liver biopsy in the setting of antituberculotic therapy (ATT) and liver injury.

While the findings are of interest, there are some issues that should be addressed:

1) page 2 (71-74): "In case of abnormal liver biochemistry, risk factors investigated included age, sex, baseline LTs, HIV, hepatitis B and C, alcohol intake, chronic liver disease. ANA, ASMA and LKM-1, AMA anti-neutrophil cytoplasmic (ANCA) and anti-double-strand DNA, were performed in all TB-DILI patients"

-> It is not mentioned that testing for Hepatitis A, Hepatitis E, CMV and other viruses have been performed/ no results are given. These investigations are prerequisites for RUCAM/CIOMS calculation. Please provide minimal necessary information for DILI causality assessment.

2) page 2 (95-96): "Severe and persistent elevations of liver transaminases and or hepatic cholestasis despite discontinuation of therapy, along with suspected other liver-related conditions were the indications of Liver biopsy (LB)."

-> Please define "severe and persistent" in terms of ULN and duration (time)

3) Table 3:

-> PLease provide results of clinical causality assessment (i.e. CIOMS/RUCAM)

 

4)  Page 9 (210-212): "In front of a clinical suspect of DILI, possible reasons to perform a biopsy could be: multiple candidates as etiologic agents; gain insight into potential mechanism of injury, assessment of the severity of injury to enable clinical decision; known underline clinical disease; exclusion of autoimmune hepatitis.

-> The usefulness of liver biopsy to differentiate between DILI and AIH needs to be substantiated: There are also studies that suggest that liver biopsy is not very helpful in this setting (Dig Dis . 2021;39(3):275-282. doi:10.1159/000511635. Epub 2020 Sep 17, Liver Int. 2019 Oct;39(10):1906-1917. doi:10.1111/liv.14195. Epub 2019 Aug 5.). The same might be true regarding the causality assessment when several drugs are involved. Please substantiate these claims

Regarding the differentiation between AIH and DILI, please use this reference: https://pubmed.ncbi.nlm.nih.gov/21674554/

 

5) The small number of study subjects and the retrospective setting complicate the interpretation of the results.

-> Please provide more detailled information on the DILI and perhaps nonDILI cases (i.e. liver biochemistries and where necessary time courses) 

 

Author Response

In the article "Role of liver biopsy in Drug-Induced Liver Injury from Anti- Tuberculosis Treatment: a retrospective cohort study" Gualano an co-workers describe 10 cases of putative tuberculostatic drug-induced liver injury who underwent liver biopsy and discuss the utility of liver biopsy in the setting of antituberculotic therapy (ATT) and liver injury.

While the findings are of interest, there are some issues that should be addressed:

1) page 2 (71-74): "In case of abnormal liver biochemistry, risk factors investigated included age, sex, baseline LTs, HIV, hepatitis B and C, alcohol intake, chronic liver disease. ANA, ASMA and LKM-1, AMA anti-neutrophil cytoplasmic (ANCA) and anti-double-strand DNA, were performed in all TB-DILI patients"

-> It is not mentioned that testing for Hepatitis A, Hepatitis E, CMV and other viruses have been performed/ no results are given. These investigations are prerequisites for RUCAM/CIOMS calculation. Please provide minimal necessary information for DILI causality assessment.

Thank you for your kind suggestion, In clinical medicine, DILI remains a diagnosis of exclusion. Liver test abnormalities may result from various liver diseases, including non-hepatic disorders. For diseases such as viral hepatitis, a single diagnostic test may confirm or exclude the diagnosis with high sensitivity and specificity. In contrast, DILI is a diagnosis based on suspicion of an adverse drug reaction and the exclusion of competing causes of liver injury. We agree that the aforementioned tests are imperative for DILI causality assessment and we confirm that testing for Hepatitis A, Hepatitis E, CMV and other viruses have been performed.

We completed the sentence at page 2, line 72

2) page 2 (95-96): "Severe and persistent elevations of liver transaminases and or hepatic cholestasis despite discontinuation of therapy, along with suspected other liver-related conditions were the indications of Liver biopsy (LB)."

-> Please define "severe and persistent" in terms of ULN and duration (time):

Thank you for this pertinent observation.

 We have considered as “persistent” the cases in which liver enzymes remained elevated for more than four weeks despite discontinuation of therapy. For TB patients this interval is crucial, because it corresponds to the time after which we must re -initiate the TB treatment in case of suspension (low forgiveness or no forgiveness of TB treatment).

In our context we consider at risk of severe toxic hepatitis patiens with:

- Symptomatic acute hepatitis, whatever the ALT and bilirubin levels found,

- Asymptomatic hepatitis and transaminases values higher than 3 times the normal values in subjects with risk factors associated with the onset of toxic hepatitis

- Asymptomatic hepatitis with transaminases greater than 5 times or bilirubinemia greater than 2 times the normal values in subjects without risk factors.

In severe forms of hepatitis (transaminases above 10 times normal values), we also monitor glycemia and PT.

 

In these cases, hepatotoxic drugs (H, R and Z associated in drug sensitive TB; Pto/Eto, PAS and Bdq in MDR/XDR TB) must be suspended. In general, drug-induced hepatitis resolves with discontinuation of the suspected drugs.

• Palmieri F. per Gruppo di Lavoro Tubercolosi - INMI “L. Spallanzani”. Percorso Diagnostico Terapeutico Assistenziale (PDTA) sulla gestione del paziente con infezione / malattia tubercolare. Revisione n. 9, 19 / 01/ 2023. https://www.inmi.it/wp-content/uploads/2023/02/PDTA-Tubercolosi-Rev.-9_2023_DS_PW-1.pdf

 

3) Table 2:

-> Please provide results of clinical causality assessment (i.e. CIOMS/RUCAM)

Thank you for your suggestion. We accordingly modified the information in Table 2.

 4)  Page 9 (210-212): "In front of a clinical suspect of DILI, possible reasons to perform a biopsy could be: multiple candidates as etiologic agents; gain insight into potential mechanism of injury, assessment of the severity of injury to enable clinical decision; known underline clinical disease; exclusion of autoimmune hepatitis.

-> The usefulness of liver biopsy to differentiate between DILI and AIH needs to be substantiated: There are also studies that suggest that liver biopsy is not very helpful in this setting (Dig Dis . 2021;39(3):275-282. doi:10.1159/000511635. Epub 2020 Sep 17, Liver Int. 2019 Oct;39(10):1906-1917. doi:10.1111/liv.14195. Epub 2019 Aug 5.). The same might be true regarding the causality assessment when several drugs are involved. Please substantiate these claims.

.Regarding the differentiation between AIH and DILI, please use this reference: https://pubmed.ncbi.nlm.nih.gov/21674554/

Thank you for this suggestions. In Table 2 we researched the four anatomopathological criteria identified by the Authors as DILI defining (with sensitivity of 91%) in pathology description of our patient’s biopsies: portal inflammation; fibrosis, portal neutrophils and plasmacells, intracellular (hepatocellular) cholestasis. We found an association between the criteria and the diagnosis of DILI (mean value of five patients not DILI 1.6; mean value of five patients with DILI 3.6; P = 0.0077, comparison of two means – Med calc).

Traditionally, liver histology was considered the gold standard in diagnosis of chronic liver diseases.

Later on, specific liver diseases, including DILI, were diagnosed with the help of laboratory testing and, possibly, liver histology data. Based solely on laboratory criteria such as ALT and ALP, DILI and other liver injuries, are now classified as hepatocellular, cholestatic or mixed type of liver damage. It is clear that liver histology is not long required to achieve this classification.

Liver histology commonly has little impact in establishing the diagnosis of DILI. So, to arrive at the correct diagnosis, clinical and structured causality assessments are better approaches than liver histology results, obtained through an invasive procedure. Nevertheless, in rare instances of diagnostic uncertainty regarding alternative causes, liver biopsy should be considered as a final diagnostic approach, provided the patient will profit from this procedure.

• Teschke R, Frenzel C. Drug induced liver injury: do we still need a routine liver biopsy for diagnosis today? Ann Hepatol. 2013 Jan-2014 Feb;13(1):121-6. PMID: 24378275.

A proportion of patients with drug-induced liver injury (DILI) present with autoantibodies, leading to the current concept of autoimmune-like DILI. The aetiology of autoimmune hepatitis (AIH) often remains uncertain but the disease can be triggered in susceptible patients by external factors such as viruses but also drugs. There are no specific histological features of either process and the pathological features may show only subtle differences making it challenging to distinguish between immune-mediated liver disease and hepatic toxicity. It has been reported that the presence of ANA titers is specific for DILI induced by certain medications. AMA positivity rather than ANA, has been associated with more pronounced liver injury. While there is an overlap of histologic findings between AIH and DILI, there are sufficient differences that allow pathologists to use the pattern of injury to suggest the correct diagnosis.

Notably, all patients in this series were tested for Antinuclear antibodies (ANA), and the results fell within the specified range limits. This suggests that ATT-drugs effectively plaied a role in causing liver damage in our patients.

References:

• Suzuki A, Brunt EM, Kleiner DE, Miquel R, Smyrk TC, Andrade RJ, Lucena MI, Castiella A, Lindor K, Björnsson E. The use of liver biopsy evaluation in discrimination of idiopathic autoimmune hepatitis versus drug-induced liver injury. Hepatology. 2011 Sep 2;54(3):931-9. doi: 10.1002/hep.24481. Epub 2011 Aug 8. PMID: 21674554; PMCID: PMC3192933.
• Weber S, Benesic A, Buchholtz ML, Rotter I, Gerbes AL. Antimitochondrial Rather than Antinuclear Antibodies Correlate with Severe Drug-Induced Liver Injury. Dig Dis. 2021;39(3):275-282. doi: 10.1159/000511635. Epub 2020 Sep 17. PMID: 32942273.
• Weber S, Benesic A, Rotter I, Gerbes AL. Early ALT response to corticosteroid treatment distinguishes idiosyncratic drug-induced liver injury from autoimmune hepatitis. Liver Int. 2019 Oct;39(10):1906-1917. doi: 10.1111/liv.14195. Epub 2019 Aug 5. PMID: 31319011.

5) The small number of study subjects and the retrospective setting complicate the interpretation of the results.

-> Please provide more detailled information on the DILI and perhaps non DILI cases (i.e. liver biochemistries and where necessary time courses) 

Thank you, we introduced the information required in table 1.

Reviewer 2 Report

Comments and Suggestions for Authors

DILI is an emerging liver disease often difficult to diagnose when other acute and chronic hepatitis risk factors may be involved. The paper by Gulano et al, has investigated the diagnostic process for the detection of DILI in their Tb patients treated with several anti Tb agents, all potentially hepatotoxic. Since this population has often comorbidities and other possible etiologies for hepatitis, including chronic HCV or HBV, it is important to follow specific algorithms for a correct diagnosis and an  appropriate treatment. I’d therefore accept the manuscript for publication provided that the authors organize a mother tongue English expert review to revise some long sentences (i.e: Materials and methods section, line 55-57) and use of preposition (i.e.: “for” instead of “of”, line 97).

Comments on the Quality of English Language

 I’d therefore accept the manuscript for publication provided that the authors organize a mother tongue English expert review to revise some long sentences (i.e: Materials and methods section, line 55-57) and use of preposition (i.e.: “for” instead of “of”, line 97).

Author Response

DILI is an emerging liver disease often difficult to diagnose when other acute and chronic hepatitis risk factors may be involved. The paper by Gualano et al, has investigated the diagnostic process for the detection of DILI in their Tb patients treated with several anti Tb agents, all potentially hepatotoxic. Since this population has often comorbidities and other possible etiologies for hepatitis, including chronic HCV or HBV, it is important to follow specific algorithms for a correct diagnosis and an appropriate treatment. I’d therefore accept the manuscript for publication provided that the authors organize a mother tongue English expert review to revise some long sentences (i.e: Materials and methods section, line 55-57) and use of preposition (i.e.: “for” instead of “of”, line 97).

 

Comments on the Quality of English Language

 I’d therefore accept the manuscript for publication provided that the authors organize a mother tongue English expert review to revise some long sentences (i.e: Materials and methods section, line 55-57) and use of preposition (i.e.: “for” instead of “of”, line 97).

Thanks for your suggestion. We have revised the document for the English language

Reviewer 3 Report

Comments and Suggestions for Authors

The authors need to revise the title and direction of the manuscript to reflect the utility of liver biopsy histology in the diagnosis and management of possible anti-TB DILI. As currently presented, the authors state at various times in the results and discussion that all 10 biopsies represent "TB DILI" , while at other times only 5 of 10 are DILI or 7 of 10 are DILI. Based on the biochemical and pathology findings and time to onset, only 5 of the 10 cases appear to actually represent anti-TB DILI with 2 cases being chronic hepatitis C, one case being hepatic tuberculosis , one case being NAFLD and 1 case having nonspecific findings. While the authors call for a causality assessment to be applied to possible DILI cases, I note that they fail to calculate a RUCAM score for any of the cases - but this should be done to help confirm the true etiology of the liver injury.  Their study should then be focused on the differential diagnosis of possible DILI in patients receiving anti-TB therapy, showing that the other causes mentioned above  can confound the diagnosis , and offers the best support for performing a liver biopsy in cases where the  clinical signature of anti-TB DILI  is atypical. 

I would title the paper: "The utility of liver biopsy in the diagnosis and management of possible DILI in patients receiving anti-TB therapy" or something similar.  They then need to be much more accurate in reporting the actual number of patients where DILI from anti-TB drugs was present (5 of 10). 

The paper would be strengthened if the total number of patients treated for active TB over the 5 year period was reported  and how many patients were thought to have anti-TB DILI who did not undergo liver biopsy. How did those patients compare to the 5 patients with anti-TB DILI in the biopsy series in terms of the clinical injury pattern, time to onset, recovery, etc. Was RUCAM used to diagnose any of these patients?  Also,  did  any additional TB patients undergo liver biopsy during this time period - and for what  indications ?  This would be in line with the new title reflecting the usefulness of liver histology in this patient population. 

Many of the patients resumed anti-TB treatment - a more complete description of their outcomes should be included in the results. Was any attempt made to desensitize patients to Isoniazid etc? 

Comments on the Quality of English Language

no comment

Author Response

The authors need to revise the title and direction of the manuscript to reflect the utility of liver biopsy histology in the diagnosis and management of possible anti-TB DILI. As currently presented, the authors state at various times in the results and discussion that all 10 biopsies represent "TB DILI" , while at other times only 5 of 10 are DILI or 7 of 10 are DILI. Based on the biochemical and pathology findings and time to onset, only 5 of the 10 cases appear to actually represent anti-TB DILI with 2 cases being chronic hepatitis C, one case being hepatic tuberculosis , one case being NAFLD and 1 case having nonspecific findings. While the authors call for a causality assessment to be applied to possible DILI cases, I note that they fail to calculate a RUCAM score for any of the cases - but this should be done to help confirm the true etiology of the liver injury. 

Thank you for the suggestion. We calculated RUCAM for each patient, as follow. We inserted the results in e table 2

	R value	pattern	diagnosis
Pat 1	12,5	hepatocellular	Chronic viral hepatitis
Pat 2	8,4	hepatocellular	Chronic viral hepatitis, HCV correlated
Pat 3	12,5	hepatocellular	Non-alcoholic steatohepatitis (nash). Secondary iron overload
Pat 4	8,4	hepatocellular	TB-DILI (acute)
Pat 5	11,6	hepatocellular	TB DILI (acute)
Pat 6	8,2	hepatocellular	TB-DILI ( partial resolution)
Pat 7	14,1	hepatocellular	TB-DILI (partial resolution)
Pat 8	4,6	mixed	minimal non-specific hepatitis
Pat 9	1,2	colestatic	TB granulomatous hepatitis
Pat 10	16,9	hepatocellular	TB-DILI

 

Their study should then be focused on the differential diagnosis of possible DILI in patients receiving anti-TB therapy, showing that the other causes mentioned above  can confound the diagnosis , and offers the best support for performing a liver biopsy in cases where the  clinical signature of anti-TB DILI  is atypical. 

I would title the paper: "The utility of liver biopsy in the diagnosis and management of possible DILI in patients receiving anti-TB therapy" or something similar.  They then need to be much more accurate in reporting the actual number of patients where DILI from anti-TB drugs was present (5 of 10). 

Thank you for your suggestion, we modified the title of the paper as follow:

Utility of liver biopsy in the diagnosis and management of possible drug-induced liver injury in patients receiving anti-TB therapy

The paper would be strengthened if the total number of patients treated for active TB over the 5 year period was reported  and how many patients were thought to have anti-TB DILI who did not undergo liver biopsy.

Thank you for the question and for the suggestion.

In the past 5 years we followed  1900, patients, with an incidence of liver injury 16%. As previously mentioned, we opted to perform liver biopsy in cases of prolonged and severe symptoms or when causes other than DILI were suspected.

How did those patients compare to the 5 patients with anti-TB DILI in the biopsy series in terms of the clinical injury pattern, time to onset, recovery, etc. Was RUCAM used to diagnose any of these patients? 

The vast majority of (DILI) cases resolved in the following 4 weeks after the ATT treatment was suspended . We have considered as “persistent” the cases in which liver enzymes remained elevated for more than four weeks despite discontinuation of therapy. For TB patients this interval is crucial, because it corresponds to the time after which we must re -initiate the TB treatment in case of suspension (low forgiveness or no forgiveness of TB treatment).

Also, did  any additional TB patients undergo liver biopsy during this time period - and for what  indications? This would be in line with the new title reflecting the usefulness of liver histology in this patient population. 

Thank you for the question. No other patients underwent liver biopsy in the period indicated. As stated main indication was prolonged and severe symptoms or when causes other than DILI were suspected.

 

Many of the patients resumed anti-TB treatment - a more complete description of their outcomes should be included in the results. Was any attempt made to desensitize patients to Isoniazid etc? 

Thank you for the question. In our institution, drug rechallenge is performed in accordance with a local protocol that aligns with the primary international guidelines. In case of failure at the rechallenge of single drugs, we must stop it definitely and find an alternative one. You can access the protocol at the following URL:

https://www.inmi.it/wp-content/uploads/2023/02/PDTA-Tubercolosi-Rev.-9_2023_DS_PW-1.pdf

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

Thank you for the clarification 

Author Response

Thank you for your valuable contribute

Reviewer 3 Report

Comments and Suggestions for Authors

The authors have revised the title to reflect their findings and the manuscript is improved overall. However, a few comments remain:

1. the English must be improved

2. since none of the patients who had biopsies had positive autoantibodies, the section on drug-induced  autoimmune hepatitis in the discussion should be deleted as it is not relevant to the patients under discussion

3. Since 16% of 1900 patients at their center  had elevated aminotransferases, the authors should provide further details on the level of these elevations, the time to onset, the specif anti-TB drugs involved,   and the outcomes. Since only 10 individuals underwent liver biopsy, the other 1890 serve as an excellent comparison group. The authors should state how many  others  had persistent unexplained elevations, but  that were followed without a liver biopsy . This would strengthen the indication for biopsy in this population. 

Comments on the Quality of English Language

the English  will require a native speaker to review and correct the usage in many sentences.

Author Response

1. the English must be improved

Thank you for your suggestion. We have corrected the entire manuscript

 

2. since none of the patients who had biopsies had positive autoantibodies, the section on drug-induced  autoimmune hepatitis in the discussion should be deleted as it is not relevant to the patients under discussion

 

Thanks again. We have corrected the text

 

3. Since 16% of 1900 patients at their center  had elevated aminotransferases, the authors should provide further details on the level of these elevations, the time to onset, the specif anti-TB drugs involved,   and the outcomes. Since only 10 individuals underwent liver biopsy, the other 1890 serve as an excellent comparison group. The authors should state how many  others  had persistent unexplained elevations, but  that were followed without a liver biopsy . This would strengthen the indication for biopsy in this population. 

Thanks very much for the comments. We conducted a retrospective analysis of a series of patients treated at a tertiary care tuberculosis hospital center in Italy, who underwent LB for suspected TB-DILI, with the aim of describing the utility of biopsy for the differential diagnosis of liver disease and its treatment during anti TB treatment.

Severe and persistent elevation of liver transaminases and or hepatic cholestasis despite discontinuation of therapy, along with suspected other liver-related conditions, were the indications for Liver biopsy (LB) in ten patients. We have considered as “persistent” the cases in which liver enzymes remained elevated for more than four weeks despite discontinuation of therapy. For TB patients this interval is crucial, because it corresponds to the time after which we must re-initiate the TB treatment in case of suspension.

In the whole cohort of 1,900 TB patients treated from 2017 to 2022 we had 304 (16%) patients who developed hypertransaminasemia with TB treatment. Eighteen (6%) were severe; 216 (71%) moderate; 70 (23%) were mild forms. The most frequently encountered situation was pyrazinamide-related toxicity, observed in 45% of cases within the first three months of treatment. In 8% of cases, treatment was resumed after normalization of transaminases without further changes. Other patients stopped isoniazid, or isoniazid and pyrazinamide.

These data appear of interest and may be the subject of further publication, for which we thank you for the suggestion.