Pancreatic Circulating Tumor Cells: An Update

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors

This article provides a comprehensive overview of the latest research on circulating tumor cells in pancreatic ductal adenocarcinoma, highlighting recent advances, ongoing challenges, and future prospects.  It reveals insights into the disease and how these findings can inform future treatment decisions.  However, the article also has some limitations:

1. In the methods section of the abstract, "focusing on original studies and reviews published within the past 15 years " and " Attention was paid to studies published since 2018 and landmark earlier works", does this include articles published before 2018?
2. In " MDPI review of 17 PDAC studies found CTCs in anywhere from 11% to 92% of cases" under section 3.1, the sources of the literature were not cited;
3. It appears inappropriate for the "Combinatorial Multi-Analyte Analytical Method" in Table 1 to include "100% sensitivity for PDAC" in the detection rate;
4. In the "Clinical Applications in Selected Cancer Types" section of Table 2, the "Relevance to Treatment Decisions" for PDAC only describes "investigational applications" without specifying the "current phase of clinical trials.";
5. The layout of the discussion requires further optimization;
6. Some abbreviations in the article are not listed in the abbreviation table;
7. The reference format should be consistent, and there are relatively few references.

Author Response

We thank the reviewer for his constructive comments

1. In the methods section of the abstract "focusing on original studies and reviews published within the past 15 years "and"Attention was paid to studies published since 2018 and landmark earlier works". Does this include articles published before 2018?

Articles before 2018 are scarce. However, we decided to start our review from 2018 since previous results are included in the more recent studies.

2. In "MDPI review of 17 PDAC studies found CTCs in anywhere from 11% to 92% of cases" under section 3.1. the sources of the literature were not cited.

This is included in ref. 7. This has been added in the text.

3. It appears inappropriate for the "Combinatorial Multi-Analyte Analytical Method" in Table 1 to include "100% sensitivity for PDAC" in the detection rate:

We agree with the reviewer comments, but this has been found and published in a peer-reviewed journal (Cancer).

4. In the "Clinical Applications in Selected Cancer Types" section of Table 2. The "Relevance to Treatment Decisions" for PDAC only describes "investigational applications" without specifying the "current phase of clinical trials".

Considering the reviewers comments we were able to write what has been published. However, there are not sufficient results to make a standard practice yet.

5. The layout of the discussion requires further optimization

The layout of the discussion has been optimized.

6. Some abbreviations in the article are not listed in the abbreviation table.

This has been completed.

7. The reference format should be consistent, and there are relatively few references

The reference format is yet consistent

Reviewer 2 Report

Comments and Suggestions for Authors

The manuscript entitled “Pancreatic Circulating Tumor Cells: An Update” was well-designed, and the context and subtitles included are interesting and attractive to any readers. However, it needs minor revision for some typos and the following comments.

 

• The manuscript should be thoroughly revised for some typos.
• The authors frequently used the colon punctuation mark in the middle of the text, which can be considered the poor English-language writing. Some examples are as follows.
• In the introduction section (3^rd paragraph). It will be clearer if the context for unique challenges of PDAC CTC should be highlighted and listed by number or the letters of the alphabet instead of the colon punctuation mark.
• Page 10, the sentence started with “Similarly, serial CTC monitoring may predict outcomes “.
• Page 10, the sentence started with “Overall, the evidence strongly supports that CTCs in PDAC are prognostic”.
• The sentence “Indeed, both enrichment strategy and disease stage influence yield, localized early tumors often release CTCs intermittently if at all, whereas metastatic disease sheds more cells.” is not clear and probably not correct.
• Table-1, especially column 3, and Tble-2 are poorly organized and appear cluttered. It needs to be revised, properly formatted, and presented in a clearer and more visually appealing manner (e.g., using an appropriate font and size for the text).
• On page 8, the context “(No such clusters were found in 295 healthy donor controls [26])” was placed in the wrong place. There might have been a typo.
• The sentence “This phenomenon of tumor cell macrophage clusters has shown that in their presence, the advance of PDAC disease correlates significantly [26,29].” Does not express a clear sense. It should be revised.

 

Author Response

We thank the reviewer for his constructive comments

The authors frequently used the colon punctuation mark in the middle of the text, which can be considered the poor-English-language writing. Some examples are as follows: In the introduction section (3^rd paragraph). It will be clearer if the context for unique challenges of PDAC CTC should be highlighted and listed by number or the letters of the alphabet instead of the colon punctuation mark.

The introduction has been improved according to the reviewer's comments.

Page 10, the sentence started with "Similarly, serial CTC monitoring may predict outcomes"

Page 10, this has been corrected according to the reviewer's suggestion.

Page 10, the sentence started with "Overall, the evidence strongly supports that CTCs in PDAC are prognostic".

Page 10, this has been corrected according to the reviewer's suggestion

The sentence "Indeed, both enrichment strategy and disease stage influence yield, localized early tumors often release CTCs intermittently if all, whereas metastatic disease sheds more cells" is not clear and probably not correct.

The sentence has been changed and is now more comprehensive.

Table 1: especially column 3, and Table 2 are poorly organized and appear cluttered. It needs to be revised, properly formatted and presented in a clearer and more visually appealing manner (e.g. using an appropriate font and size for the text).

Table 1 has been reformatted. However, we believe that all these informations should remain in order not to decrease the impact of the table. We believe that many readers will not read the whole text, but will concentrate on the table. Table 2 has been deleted.

On page 8, the context "(No such clusters were found in 295 healthy donor controls [26])" was placed in the wrong place. There might have been a typo.

On page 8 the typo mistake has been corrected.

The sentence "This phenomenon of tumor cell macrophage clusters has shown that in their presence, the advance of PDAC disease correlates significantly [26,29]." Does not express a clear sense. It should be revised.

The sentence has been corrected.

Reviewer 3 Report

Comments and Suggestions for Authors

The review provides a broad survey of CTCs) in PDAC, including detection technologies, CTC biology/heterogeneity, prognostic associations, and emerging translational directions (clusters, organoids). The topic is timely and relevant. However, the manuscript is presently overly descriptive, repetitive, and insufficiently critical in judging evidentiary strength and clinical readiness. The result reads closer to an annotated bibliography than a synthesis that advances reader understanding or sets priorities for the field.

Major comments

1. Insufficient critical synthesis / prioritisation: The manuscript summarises many studies however rarely ranks them by quality, reproducibility, sample size, or clinical relevance. This makes it difficult to distinguish robust conclusions from exploratory signals. Please add explicit “What is solid vs uncertain” statements at the end of each major section (detection platforms, EMT/heterogeneity, clusters, prognostic value, translational models).

2. Redundancy and length dilute impact: Key limitations (EpCAM bias, EMT-related detection failure, low abundance, lack of standardisation) are repeated across sections without new insight. Condense overlapping passages and use internal cross-references rather than restating. Tightening will materially improve readability and perceived rigour.
3. Clinical utility is not assessed rigorously: Prognostic associations are frequently presented without answering the clinical question: Does CTC status add incremental value beyond existing markers (CA19-9, imaging, stage, performance status) or standard prognostic models? Where possible, summarise effect sizes (HR/OR), timing (baseline vs on-treatment), and whether multivariable models were used. If this information is missing in primary studies, state that explicitly and downgrade confidence.
4. Predictive vs prognostic claims need sharper separation: Several sections blur prognostic correlation (risk stratification) with predictive utility (treatment selection/response guidance). PDAC evidence for predictive use remains limited. Reframe organoids/drug testing as promising but early, with practical constraints (yield rates, time-to-result, representativeness, and scalability).
5. Early detection section should be shortened and made more cautious: Given PDAC biology and low CTC abundance in early disease, standalone CTC-based early detection is not currently supported by strong evidence. The manuscript should state clearly that early detection remains speculative/adjunctive, likely requiring multi-analyte approaches and rigorous prospective validation.
6. Missing integrative framework: A high-quality review should provide a conceptual model that links: (i) PDAC biology and EMT/partial EMT states, (ii) technical capture biases, (iii) which CTC subsets associate with outcome, and (iv) how CTCs might be combined with ctDNA/exosomes/clinical variables.

Add at least one schematic figure and consider a concluding section on a realistic “near-term clinical roadmap” (what could be tested in the next 2–3 years vs longer term).

Minor comments

1. Standardise terminology ( CTC positivity, CTC detection rate, CTC enumeration) and define thresholds when reported.
2. Where studies are heterogeneous, explicitly note differences in patient cohorts (resectable vs metastatic; neoadjuvant vs palliative), sampling timepoints, and assays used.
3. Ensure tables do not simply list studies; add columns for cohort size, stage, platform, timepoint(s), endpoint(s), multivariable adjustment, and key limitations.
4. Consider reducing comparisons to non-PDAC cancers unless they directly inform PDAC-specific feasibility or mechanisms.

Author Response

The review provides a broad survey of CTCs in PDAC, including detection technologies. CTC biology/heterogeneity, prognostic associations, and emerging translational directions (clusters, organoids). The topic is timely and relevant. However, the manuscript is presently overly descriptive, repetitive, and insufficiently critical in judging evidentiary strength and clinical readiness. The result reads closer to an annotated bibliography than a synthesis that advances reader understanding or sets priorities for the field.

We appreciate the pertinent comments by the reviewer. We agree that he raises the right issues. However, this topic is like a puzzle, we know some pieces, but we are fare away from knowing the whole picture. Therefore, some of the points mentioned by the reviewer cannot be either answered or commented. The aim of this review is to provide the current knowledge in this rapidly evolving area. We are only at the beginning of elucidating the impact of these researches. Therefore, criticisms on clinical recommendations are too early to be raised.

Major comments

1. Insufficient critical synthesis / prioritization: The manuscript summarizes many studies however rarely ranks them by quality, reproducibility, sample size or clinical relevance. This makes it difficult to distinguish robust conclusions from exploratory signals. Please add explicit "What is solid vs uncertain" statements at the end of each major section (detection platforms, EMT/heterogeneity, clusters, prognostic value, translational models).

Cf. previous comments. The mentioned studies are from high rated peer-review journals. We believe that the reviewers made their job and it is not our goal to check this. We decided not to use "solid vs uncertain", but we have made some comments with limitations.

2. Redundancy and length dilute impact: Key limitations (EpCAM bias. EMT-related detection failure, low abundance, lack of standardization) are repeated across sections without new insight. Condense overlapping passages and use internal cross-references rather than restating. Tightening will materially improve readability and perceived rigour

The manuscript has been shortened in order to avoid diluting the impact. Some repetitions are necessary as they are relevant to the different methodology.

3. Clinical utility is not assessed rigorously: Prognostic associations are frequently presented without answering the clinical question. Does CC status add incremental value beyond existing markers (CA 19-9, imaging, stage, performance status) or standard prognostic models? Where possible, summarize effect sizes (HR/OR), timing (baseline vs on-treatment), and whether multivariable models were used. If this information is missing in primary studies, state that explicitly and downgrade confidence.

The reviewer comments are appropriate, but the clinical questions cannot be answered at this time. However, there is emerging promising evidence that CTC can be used as an adjuvant in the diagnostic and prognostic in the management of pancreatic cancer, along with the CA 19-9, imaging, and the clinical findings. However, the clinical impact of the use of CTC still needs to be established. This is one of the messages of this review, which has been clearly emphasized in this work.

4. Predictive vs prognostic claims need sharper separation: Several sections blur prognostic correlation (risk stratification) with predictive utility (treatment selection/response guidance). PDAC evidence for predictive use remains limited. Reframe organoids/drug testing as promising but early, with practical constraints (yield rates, time-to-result, representativeness, and scalability).

 There is no evidence between what is predictive vs prognostic claims and therefore there is no sharp clear difference at this time. We thank the reviewer concerning the organoids. A sentence has been added in the manuscript.

5. Early detection section should be shortened and made more cautious: Given PDAC biology and low CTC abundance in early disease, standalone CTC-based early detection is not currently supported by strong evidence. The manuscript should state clearly that early detection remains speculative/adjunctive, likely requiring multi-analyte approaches and rigorous prospective validation.

We agree with the reviewer. Early detection is still speculative. This point has been emphasized in the manuscript.

6. Missing integrative framework: A high-quality review should provide a conceptual model that links (I) PDAC biology and EMT/partial EMT states, (II) technical capture biases, (III) which CTC subsets associate with outcome, and (IV) how CTCs might be combined with ctDNA/exosomes/clinical variables.

We agree with the reviewer, but we are waiting for more evidence to find out whether sub-groups would or would not benefit from these new approaches in this context. We are working actually on the PDAC biology and the results are not ready to be published. In the future we hope that we can get the answers to the queries.

Minor comments:

1. Standardize terminology (CTC positivity, CTC detection rate, CTC enumeration) and define thresholds when reported.

Terminology has been standardized. Thresholds have not been included since across the studies different methodologies have been used.

2. Where studies are heterogenous, explicitly note differences in patient cohorts (resectable vs metastatic; neoadjuvant vs palliative), sampling timepoints, and assays used.

Clinical studies are scarce and to date there is no valuable data dealing with the patient's subgroup. Therefore, this point goes beyond the scope of this review.

3. Ensure tables do not simply list studies; add columns for cohort size, stage, platform, timepoint(s), endpoint(s), multivariable adjustment, and key limitations.

 Table 2 has been deleted. According to the available literature dealing with clinical studies, the vast majority includes observational trials, and no one goes over a phase 2 study. For these reasons we decided not to include details concerning patients, cohort data etc., since no valuable clinical informations can be extracted from those. These informations would be helpful in the future, when a correlation with clinical outcome can be achieved. This is the scope for future investigations.

4. Consider reducing comparisons to non-PDAC cancers unless they directly inform PDAC-specific feasibility or mechanisms.

We agree with the reviewer. Comparisons with non-PDAC cancers have been either reduced or deleted.

Round 2

Reviewer 1 Report

Comments and Suggestions for Authors

The author answered all my questions and made appropriate revisions in the revised manuscript. I am satisfied with most of the answers. However, as a review, 49 references are not enough. I suggest the author add references. For example, in the "Introduction" section, there are only 8 references, and this part does not cite a large number of literature to prove the author's viewpoint. The repeated appearance of references 3, 7, and 8 in the 'discussion' indicates a lack of relevant references by the author. These sections require the addition of new references.

Author Response

Answers to Reviewer 1

 

The author answered all my questions and made appropriate revisions in the revised manuscript. I am satisfied with most of the answers. However, as a review, 49 references are not enough. I suggest the author add references. For example, in the "introduction" section, there are only 8 references, and this part does not cite a large number of literatures to prove the author's viewpoint. The repeated appearance of references 3, 7, and 8 in the "discussion" indicates a lack of relevant references by the author. These sections require the addition of new references.

References have been added in the manuscript, there are now 91 references.

Reviewer 3 Report

Comments and Suggestions for Authors

Thank you for the revised submission. The revised manuscript is improved in structure and coverage and remains suitable for the journal’s readership. However, the review still reads predominantly as a catalogue of studies rather than a critical synthesis, and it would benefit from sharper framing around heterogeneity, evidence strength, and what is clinically actionable in PDAC.

Major points

1. Stronger critical synthesis is still needed. The manuscript summarises detection platforms and reported detection rates, but the discussion does not consistently explain why studies disagree (such cohort composition and stage mix, sampling site and blood volume, marker/assay bias, and inconsistent definitions of ‘CTC-positive’). A short, explicit subsection on “sources of heterogeneity and evidence quality” would materially strengthen the review.
2. Clinical utility needs clearer separation (prognostic vs predictive vs monitoring). The manuscript references associations with survival/recurrence and treatment response, but the narrative would benefit from clearer signposting of which claims are supported by stronger evidence (multi-centre / prospective cohorts) versus exploratory work. A concise “what is ready now vs what remains investigational” summary (even 6–8 sentences) would help readers.
3. Terminology and thresholds should be standardised. Where cut-offs/thresholds are reported, it should be made explicit that they are platform- and cohort-dependent and not currently standardised for PDAC clinical use. Please ensure consistent notation and definitions throughout (PD-L1 formatting, CTC positivity terminology, abbreviations).

Author Response

Major points

 

1. Stronger critical synthesis is still needed. The manuscript summarizes detection platforms and reported detection rates, but the discussion does not consistently explain why studies disagree (such cohort composition and stage mix, sampling site and blood volume, marker/assay bias, and inconsistent definitions of CTC-positive). A short, explicit subsection on "sources of heterogeneity and evidence quality" would materially strengthen the review.
2. Clinical utility needs clearer separation (prognostic vs predictive vs monitoring). The manuscript references associations with survival/recurrence and treatment response, but the narrative would benefit from clearer signposting of which claims are supported by stronger evidence (multi-center / prospective cohorts) versus exploratory work. A concise "what is ready now vs what remains investigational" summary (even 6-8 sentences) would help readers.
3. Terminology and thresholds should be standardized. Where cut-offs/thresholds are reported, it should be made explicit that they are platform- and cohort-dependent and not currently standardized for PDAC clinical use. Please ensure consistent notation and definitions throughout (PD-L1 formatting, CTC positivity terminology, abbreviations). 

To points 1 - 3           

The issues raised by the reviewer have been included in a paragraph at the end of the discussion to emphasize the methodological issues, the uncertainty between prognostic and prediction, the missing crucial clinical evidence, which leads to the fact that no conclusive information can be drawn at this time