Review Reports - Modes of (Inter)Actions of Polyvalent Immunoglobulins: Nonclinical and Clinical Research in Severe Bacterial Infections

Round 1

Reviewer 1 Report

Comments and Suggestions for Authors This review addresses an important clinical question: can polyvalent immunoglobulins help in severe bacterial infections by neutralizing toxins and affecting the body's response? The manuscript covers the topic well, but some sections need clearer definitions, better references, and a more careful link between how the treatment works and its actual clinical effects. 1. Can the authors clarify why pneumolysin (S. pneumoniae) is listed as a “type I exotoxin / superantigen” example? This does not match how pneumolysin is usually described and may weaken the toxin-mechanism argument. Please provide a reference for this classification or update the examples to match standard toxin categories. Adding a short explanation for this choice would also help readers. 2. Can the authors clarify the rationale for using the “type I / II / III exotoxin” framework? As it stands, “type II” includes several different types of toxins, and “type III” overlaps with secretion systems that many would not call “exotoxins.” It may be clearer to organize this section by mechanism, such as superantigens, A–B toxins, pore-forming toxins, and secretion systems, with clear definitions for each. This would make the section easier to follow and reduce confusion. 3. Can the authors give more details about the claim that in vitro studies show 95–100% inhibition at clinically relevant concentrations for different Ig preparations? Readers will want to know which assays were used, which toxins or pathogens were tested, how many studies there were, what concentration ranges were used, and how consistent the results were. A table or a short summary with this information would be helpful. It is also worth noting that in vitro results do not always predict clinical outcomes. 4. Can the authors expand the safety discussion to better represent practical ICU decisions in sepsis or septic shock? The authors mention immunological pathways, but it would help to clearly discuss known IVIg risks, such as blood clots, kidney problems, hemolysis, and infusion reactions, and how these might affect patients with sepsis. Summarizing the major trials' findings on side effects and whether risks varied by product or dose would make the paper more useful. Adding a short “Safety considerations” section would also help. 5. Can the authors clarify which patient-selection criteria (such as severity scores, biomarker patterns, or timing) are backed by prospective evidence, and which are based on post hoc or theoretical reasoning? Some parts suggest that certain groups “should” benefit, but the data may not be strong enough for that claim. It would be better to present these as ideas for future trials, such as testing low IgM, specific inflammation patterns, and early treatment. A simple table listing “proposed enrichment markers,” “supporting evidence type,” and “open questions” would make this section clearer.

Author Response

Title: Modes Of (Inter)actions of Polyvalent Immunoglobulins: Non-clinical and Clinical Research in Severe Bacterial Infections

Manuscript ID: biomedicines-4082743

The authors thank the Editor and the Reviewers for their time and effort in considering our manuscript. In response to their comments, we are submitting responses to the reviewers’ comments and a new revision of our manuscript. Our responses to their comments are included below in blue and the changes performed in the reviewed manuscript are highlighted using the track change function in the corresponding document (marked copy). The page numbers and lines correspond to those in the marked copy.

Reviewer 1

This review addresses an important clinical question: can polyvalent immunoglobulins help in severe bacterial infections by neutralizing toxins and affecting the body's response? The manuscript covers the topic well, but some sections need clearer definitions, better references, and a more careful link between how the treatment works and its actual clinical effects.

Can the authors clarify why pneumolysin (S. pneumoniae) is listed as a “type I exotoxin / superantigen” example? This does not match how pneumolysin is usually described and may weaken the toxin-mechanism argument. Please provide a reference for this classification or update the examples to match standard toxin categories. Adding a short explanation for this choice would also help readers.

ANSWER: Thank you for pointing this out. Pneumolysin was inadvertently listed in the text as an example of a Type I exotoxin during revision of this section. This was an editorial error rather than an intended classification. Pneumolysin is correctly described as a cholesterol‑dependent pore‑forming exotoxin, as reflected in Table 2. We have therefore removed pneumolysin as a superantigen exotoxin example from the text (page 7, line 218) to ensure consistency with standard toxin classifications and to avoid confusion.

Can the authors clarify the rationale for using the “type I / II / III exotoxin” framework? As it stands, “type II” includes several different types of toxins, and “type III” overlaps with secretion systems that many would not call “exotoxins.” It may be clearer to organize this section by mechanism, such as superantigens, A–B toxins, pore-forming toxins, and secretion systems, with clear definitions for each. This would make the section easier to follow and reduce confusion.

ANSWER: Thank you for this helpful suggestion. We agree that organizing exotoxins by mechanism of action improves clarity. In the revised manuscript, we have removed the Type I/II/III terminology and now structure Section 2.2.1 and Table 2 exclusively by mechanistic classes—superantigens, membrane‑damaging (pore‑forming) toxins, and A–B toxins. This change simplifies the presentation, avoids overlapping classifications, and aligns the discussion more directly with immunoglobulin‑mediated toxin neutralization.

Can the authors give more details about the claim that in vitro studies show 95–100% inhibition at clinically relevant concentrations for different Ig preparations? Readers will want to know which assays were used, which toxins or pathogens were tested, how many studies there were, what concentration ranges were used, and how consistent the results were. A table or a short summary with this information would be helpful. It is also worth noting that in vitro results do not always predict clinical outcomes.

ANSWER: Thank you for this thoughtful suggestion. We agree that a quantitative statement such as “95–100% inhibition” requires detailed methodological context (assay formats, toxin/panel selection, concentration ranges, and number of studies). To avoid over‑generalization on the one hand and additional complexity in this already extensive review on the other hand, we have removed that sentence. We now emphasize that exotoxin‑neutralizing activity varies by toxin, assay, and immunoglobulin preparation, and that these observations derive from experimental (in vitro/ex vivo or animal) settings; comparative clinical evidence demonstrating superior toxin‑neutralizing activity of one preparation over another in a specific toxin‑related indication remains limited (page 8, lines 248-253). This revision preserves accuracy, improves balance, and avoids implying a level of consistency that the current literature does not uniformly support.

Can the authors expand the safety discussion to better represent practical ICU decisions in sepsis or septic shock? The authors mention immunological pathways, but it would help to clearly discuss known IVIg risks, such as blood clots, kidney problems, hemolysis, and infusion reactions, and how these might affect patients with sepsis. Summarizing the major trials' findings on side effects and whether risks varied by product or dose would make the paper more useful. Adding a short “Safety considerations” section would also help.

ANSWER: Thank you for this important comment. We agree that safety considerations are highly relevant when considering the use of immunoglobulins in critically ill patients with sepsis or septic shock. The special warnings and precautions for use of Ig are to be followed (according to the leaflet/core SmpCs) and patients must be monitored and observed for any symptoms of adverse reactions (like hypersensitivity reactions, thromboembolic events, ARF, and haemolytic anemia) throughout the infusion period.

The primary aim of this review, however, is to synthesize mechanistic insights into how polyvalent immunoglobulins interact with pathogens and the host immune response, and to evaluate the extent to which these mechanisms are supported by clinical data. For this reason, a detailed product‑ and dose‑specific safety analysis was not undertaken, as this would require a different type of review (e.g. systematic review) and a different review focus and would substantially expand the scope of this manuscript.

Nevertheless, we acknowledge that immunoglobulin safety is influenced by several factors, including formulation‑specific characteristics (e.g., excipients, IgA or anti‑A/B content), manufacturing processes, dosing, and patient‑related risk factors typical of severe sepsis. In addition, interpreting adverse events in this population is inherently complex, as complications such as acute kidney injury or thromboembolic events are also frequent consequences of sepsis itself. To address this aspect, we have added a concise paragraph on safety considerations in the new Section 4.3 (Limitations and Safety Considerations), highlighting IVIg‑associated risks and the need for standardized safety reporting in future clinical studies (page 24, lines 831-847).

Can the authors clarify which patient-selection criteria (such as severity scores, biomarker patterns, or timing) are backed by prospective evidence, and which are based on post hoc or theoretical reasoning? Some parts suggest that certain groups “should” benefit, but the data may not be strong enough for that claim. It would be better to present these as ideas for future trials, such as testing low IgM, specific inflammation patterns, and early treatment. A simple table listing “proposed enrichment markers,” “supporting evidence type,” and “open questions” would make this section clearer.

ANSWER: We thank the reviewer for this important and constructive suggestion. We fully agree that the strength of evidence supporting different patient‑selection criteria varies substantially, and that this needs to be clearly distinguished to avoid over‑interpretation.

In the revised manuscript, we have therefore:

Explicitly separated patient‑selection concepts supported by prospective or randomized evidence from those drawn from post hoc analyses, observational studies, or mechanistic reasoning (Sections 4 and 5).
Softened prescriptive language (e.g., “should benefit”) where robust prospective evidence is lacking, reframing these statements as hypotheses or rationales for future trials.
Added a new summary table (Table 3) that lists proposed enrichment markers, the type of supporting evidence (prospective, retrospective, post hoc, or meta-analysis), and key open questions. This table is explicitly framed as a forward‑looking research tool rather than a clinical recommendation.

We believe these revisions improve transparency, align the manuscript with the current strength of evidence, and directly address the reviewer’s concern.

Reviewer 2 Report

Comments and Suggestions for Authors

The current submitted review manuscript presents an interesting and clinically important review for potential using polyvalent immunoglobulins, in particular IgM-IgA-enriched ones, in sepsis patients. The manuscript is well-structured with illustrative diagrams and written in perfect English.

However, there are some points that need to be addressed and rectified before this manuscript may be considered for publication. These are explained below:

Although this review manuscript presented and covered most reported data on polyvalent Igs in infection and sepsis the critical review of the reported findings and the authors own comments are not frequent and often leaving narrated text as story without critical explanation or comments. Comment no. 2 is a clear example of my point. I highly recommend doing more critical reading of some of the cited references and incorporating further explanation and/or possible justification of certain reported finings.
In page 15, line 495, 496: pasting from the text “IgM/IgA-enriched preparations also reduced IL-1 β plasma levels and tissue injury in rat CLP models.

Comment: put reference. This reference is evidently Ates I. et al. 2015, Pharmaceutical Biology, 2015; 53(1): 78–84. DOI: 10.3109/13880209.2014.910535.

Upon reading this paper, one can comment that despite the lower IL-1β in blood of IgM-enriched immunoglobulin in that study, there was no redundant confirmatory method for neutrophils in intestine tissue homogenate by myeloperoxidase activity assay, lung or blood neutrophils, lung and liver wet/dry ratio to assess if edema a hallmark of sepsis/acute lung injury was improved.

Page 16 line 504, pasting from the text “differences in treatment timing” Reference is missing. A proper reference to be cited here is Berlot G. et al. 2018. Effects of the timing of administration of IgM- and IgA-enriched intravenous polyclonal immunoglobulins on the outcome of septic shock patients. Ann Intensive Care. 2018 Dec 10;8:122. doi: 10.1186/s13613-018-0466-7.

It is also more informative to add right after “treating time” that : early administration of ivIgGAM was associated with lower mortality rate in septic shock patients (Berlot G. et al. 2018).

Page 16 line 504-506: pasting from the text “ Also, different animal models

of sepsis have shown differences in cytokine responses (e.g., LPS or CLP models) [191].

This paper, referenced herein (191, Remick D.G. et al. Shock 2000, 13, 110-116) compare cytokines in LPS vs. CLP without any IVIg or IgM-IgA treatment. This is a different context and from polyvalent Igs intervention, the axis point of this current review and may be removed. More priority of focus should be tailored to the primary clinical and mortality findings such as Berlot G. et al. 2018, including animal models then comparison of cytokine profile come after from the same papers that revealed amelioration in injury and multi-organ failure (hallmark of sepsis and septic shock).

Author Response

Title: Modes Of (Inter)actions of Polyvalent Immunoglobulins: Non-clinical and Clinical Research in Severe Bacterial Infections

Manuscript ID: biomedicines-4082743

Reviewer 2

However, there are some points that need to be addressed and rectified before this manuscript may be considered for publication. These are explained below:

Although this review manuscript presented and covered most reported data on polyvalent Igs in infection and sepsis the critical review of the reported findings and the authors’ own comments are not frequent and often leaving narrated text as story without critical explanation or comments. Comment no. 2 is a clear example of my point. I highly recommend doing more critical reading of some of the cited references and incorporating further explanation and/or possible justification of certain reported findings.

ANSWER: Thank you for this valuable observation. We agree that adding more critical analysis and author commentary will strengthen the manuscript. We have revised the text throughout the review to include:

Critical interpretation of key findings from cited studies, highlighting limitations, variability in results, and implications for clinical practice.
Justifications for observed differences between nonclinical and clinical outcomes, including potential reasons such as patient selection, timing of administration, and study design.
Discussion of controversies and gaps in the literature to provide a balanced perspective.
Future directions for the design of new trials.

Specific examples of these changes have been incorporated in Sections 2.1.3 “Clinical Studies on Endotoxin Opsonization” (page 7, lines 198-200), 2.2.3 “Clinical Studies on Exotoxin Neutralization” (page 10, lines 273-274, 276-278, and 284-289), 2.3.3 “Clinical Studies on Additive Effects of Polyvalent Immunoglobulins and Antibiotics” (page 12, lines 359-362; page 13, lines 374-377), 3.3.4 “Clinical Studies on Modulation of Complement Activity” (page 20, lines 729-735), 4 “Important Lessons Learned” (page 24, lines 803-847), and 5 “Conclusions and Future Directions” (page 25, lines 878-888).

In page 15, line 495, 496: pasting from the text “IgM/IgA-enriched preparations also reduced IL-1 β plasma levels and tissue injury in rat CLP models.

Comment: put reference. This reference is evidently Ates I. et al. 2015, Pharmaceutical Biology, 2015; 53(1): 78–84. DOI: 10.3109/13880209.2014.910535.

Upon reading this paper, one can comment that despite the lower IL-1β in blood of IgM-enriched immunoglobulin in that study, there was no redundant confirmatory method for neutrophils in intestine tissue homogenate by myeloperoxidase activity assay, lung or blood neutrophils, lung and liver wet/dry ratio to assess if edema a hallmark of sepsis/acute lung injury was improved.

ANSWER: Thank you for pointing this out. We have added the missing reference (Ates I. et al., Pharmaceutical Biology, 2015; 53(1): 78–84. DOI: 10.3109/13880209.2014.910535) to support the statement regarding IL-1β reduction and tissue injury in rat CLP models. After reviewing the study, we note that the authors performed histopathological scoring and quantified polymorphonuclear leukocyte infiltration in lung and intestinal tissues. However, as the authors themselves state, the study did not assess in situ cytokine release, apoptotic indices, or animal survival. To reflect these limitations more accurately, we have revised the manuscript to include a general statement noting that more comprehensive experimental studies are needed to confirm and extend these findings (Section 3.2.3, page 17, lines 550-556).

Page 16 line 504, pasting from the text “differences in treatment timing” Reference is missing. A proper reference to be cited here is Berlot G. et al. 2018. Effects of the timing of administration of IgM- and IgA-enriched intravenous polyclonal immunoglobulins on the outcome of septic shock patients. Ann Intensive Care. 2018 Dec 10;8:122. doi: 10.1186/s13613-018-0466-7.

It is also more informative to add right after “treating time” that: early administration of ivIgGAM was associated with lower mortality rate in septic shock patients (Berlot G. et al. 2018).

ANSWER: Thank you for your suggestion. The sentence in question (currently page 17, lines 563-565: “These differences may reflect species-specific responses, immune status, inflammation severity, or differences in treatment timing and dosage”) is an authors’ interpretation referring to differences in the inflammatory stage of sepsis and other biological factors that influence IL‑10 modulation across experimental models and clinical settings. It is not intended to refer to the timing of immunoglobulin administration in clinical studies; therefore, the Berlot et al. (2018) paper is not directly applicable in this context.

To avoid ambiguity, we have revised the sentence to clarify that we are referring to differences in inflammatory stage, immune status, and experimental conditions.

We would also like to note that the study by Berlot et al. (2018) is already appropriately cited and discussed in Section 4.1 (page 21, lines 761-766), where timing of Ig administration is explicitly addressed in the context of clinical outcomes.

Page 16 line 504-506: pasting from the text “ Also, different animal models of sepsis have shown differences in cytokine responses (e.g., LPS or CLP models) [191].

ANSWER: Thank you for this observation. We agree that the cited reference (Remick et al., 2000) does not directly relate to the intervention with polyvalent immunoglobulins, which is the main focus of this review. To maintain relevance and clarity, we have removed this sentence and its associated reference (Section 3.2.3, page 17, lines 565-567). We have also ensured that the discussion prioritizes findings from studies evaluating the effects of Ig interventions on pathogens and the host.

Reviewer 3 Report

Comments and Suggestions for Authors

The manuscript addresses the role of polyvalent immunoglobulin preparations as adjunctive therapy in severe bacterial infections, including sepsis and septic shock. The review is comprehensive in scope and provides an extensive synthesis of nonclinical and clinical evidence regarding immunoglobulin–pathogen and immunoglobulin–host interactions. However, several issues should be addressed before the manuscript can be considered for publication.

Abstract
While the potential benefits of immunoglobulin therapy are emphasized in the abstract, the authors are encouraged to briefly mention the heterogeneity of the clinical evidence and the lack of definitive benefit observed in trials with low risk of bias in order to provide a more balanced summary.

Methods and Review Scope
Although this is a narrative review, the authors may should clarify whether any structured literature search strategy was used, including the databases consulted and the time frame covered. The process by which clinical studies were selected should be described. Providing a brief description of the literature identification approach would improve the transparency, reproducibility, and credibility of the review.

Results and Discussion
Several immunoglobulin products discussed in the manuscript are manufactured by the authors’ affiliated institution. While disclosures may be provided elsewhere, the narrative occasionally lacks critical distance, particularly in comparisons between IgM and IgA enriched preparations and standard intravenous immunoglobulin. The authors should explicitly acknowledge this potential limitation and ensure that alternative interpretations, as well as neutral or negative findings, are discussed with equal emphasis.

Several key limitations remain under-addressed, including variability in dosing regimens, heterogeneity of study designs, and the predominance of small or retrospective studies within the clinical evidence base. These issues should be more explicitly acknowledged in a dedicated subsection to avoid an overly optimistic interpretation of the available data.

Author Response

Title: Modes Of (Inter)actions of Polyvalent Immunoglobulins: Non-clinical and Clinical Research in Severe Bacterial Infections

Manuscript ID: biomedicines-4082743

Reviewer 3

Abstract
While the potential benefits of immunoglobulin therapy are emphasized in the abstract, the authors are encouraged to briefly mention the heterogeneity of the clinical evidence and the lack of definitive benefit observed in trials with low risk of bias in order to provide a more balanced summary.

ANSWER: Thank you for this important suggestion. We agree that the abstract should reflect both the potential benefits and the limitations of the current evidence. We have revised the abstract to include a statement acknowledging the heterogeneity of clinical findings and the lack of definitive benefit in trials with low risk of bias, ensuring a more balanced summary (page 1, lines 23-25).

Methods and Review Scope
Although this is a narrative review, the authors may should clarify whether any structured literature search strategy was used, including the databases consulted and the time frame covered. The process by which clinical studies were selected should be described. Providing a brief description of the literature identification approach would improve the transparency, reproducibility, and credibility of the review.

ANSWER: Thank you for this suggestion. We agree that clarifying the literature identification process improves the transparency of the review. In response, we have modified the last paragraph of the Introduction to include a brief description of the approach used to identify and select relevant studies, including the databases consulted and the time frame covered (page 2, lines 64-81).

Results and Discussion
Several immunoglobulin products discussed in the manuscript are manufactured by the authors’ affiliated institution. While disclosures may be provided elsewhere, the narrative occasionally lacks critical distance, particularly in comparisons between IgM and IgA enriched preparations and standard intravenous immunoglobulin. The authors should explicitly acknowledge this potential limitation and ensure that alternative interpretations, as well as neutral or negative findings, are discussed with equal emphasis.

ANSWER: Thank you for this important comment. We agree that, given that our institution manufactures several of the immunoglobulin products discussed (both IVIg and IgM/IgA enriched preparations), it is essential to present the evidence with appropriate critical distance. While our conflicts of interest are clearly stated, we have carefully reviewed the manuscript to ensure that the narrative remains balanced and that mechanistic and clinical findings are presented objectively.

To address your concern, we have taken the following steps:

We ensured that, for each mechanism of action, results from both IVIg and IgM/IgA‑enriched preparations are reported whenever available, and that neutral or negative findings—including the absence of mortality benefit in low risk‑of‑bias randomized trials and instances of inconclusive mechanistic data—are discussed with equal emphasis to positive findings. These outcomes are also transparently summarized in the Supplementary Tables (where results are indicated as significant difference/marked change/no difference).
We rephrased selected mechanistic descriptions to avoid implying clinical superiority of IgM/IgA‑enriched preparations where this is not supported by comparative clinical evidence (new balancing sentences at page 8, lines 248–253; page 12, lines 359–362; page 13, lines 381–383; page 14, lines 434–435; and page 16, lines 538–541).
For scope clarity, we note that this review excludes (i) studies on viral infections and neonatal/pediatric populations and (ii) IVIg studies unrelated to severe bacterial infections, sepsis, or septic shock. Likewise, studies not providing evidence for at least one specific mode of action (i.e., reporting only efficacy/safety without mechanistic support) are not summarized in the Supplementary Tables, consistent with our mechanistic focus.
For readers seeking a broader discussion of immunoglobulin tissue‑protective functions in severe infections, we refer to a recent review by Schmidt et al. (2023), which complements our MOA‑centered approach (Schmidt et al., 2023, Multifaceted Tissue-Protective Functions of Polyvalent Immunoglobulin Preparations in Severe Infections—Interactions with Neutrophils, Complement, and Coagulation Pathways, Biomedicines 2023, 11, 3022. https://doi.org/10.3390/biomedicines11113022).

These revisions collectively reinforce balance and critical distance across the narrative, while maintaining the review’s mechanistic focus.

Several key limitations remain under-addressed, including variability in dosing regimens, heterogeneity of study designs, and the predominance of small or retrospective studies within the clinical evidence base. These issues should be more explicitly acknowledged in a dedicated subsection to avoid an overly optimistic interpretation of the available data.

ANSWER: Thank you for highlighting this. For most clinical studies mentioned in the respective sections, the type of study (retrospective, prospective, case study, meta etc) is already indicated. We agree that the major limitations of the evidence base should be made more explicit.

Accordingly, we have:

Revised several sections to foreground study limitations at the point of discussion: Section 2.1.3 (page 7, lines 198–200), Section 2.2.3 (page 10, lines 273–274 and 276-278), Section 2.3.3 (page 13, lines 374-377), and Section 3.3.4 (page 20, lines 729–735).
Added Section 4.3, “Limitations and Safety Considerations” (page 24, lines 803–847), which consolidates key constraints across the literature, including heterogeneity of study designs and patient selection, variability in dosing and timing, the predominance of small or retrospective studies, and the observation that trials with low risk of bias have not consistently demonstrated a definitive survival benefit.

These additions and revisions ensure that limitations are explicitly acknowledged and visible both locally (within each subsection) and globally (in Section 4.3), thereby preventing an overly optimistic interpretation of the available data.