A systematic review reveals conflicting evidence for the prevalence of antibodies against the sialic acid ‘xenoautoantigen’ Neu5Gc in humans and the need for a standardised approach to quantification

Despite an array of hypothesised implications for health, disease, and therapeutic development, antibodies against the non-human sialic acid N-glycolylneuraminic acid (Neu5Gc) remain a subject of much debate. This systematic review of 114 publications aimed to generate a comprehensive overview of published studies in this field, addressing both the reported prevalence of anti-Neu5Gc antibodies in the human population and whether experimental variation accounts for the conflicting reports about the extent of this response. Absolute titres of anti-Neu5Gc antibodies, the reported prevalence of these antibodies, and the individual variation observed within experiments were analysed and grouped according to biological context (‘inflammation’, ‘xenotransplantation’, ‘biotherapeutic use’, ‘cancer’, and ‘healthy populations’), detection method, target epitope selection, and choice of blocking agent. These analyses revealed that the experimental method had a notable impact on both the reported prevalence and absolute titres of anti-Neu5Gc antibodies in the general population, thereby limiting the ability to ascribe reported trends to genuine biological differences or the consequence of experimental design. Overall, this review highlights important knowledge gaps in the study of antibodies against this important xenoautoantigen and the need to establish a standardised method for their quantification if the extent of the importance of Neu5Gc in human health is to be fully understood.


Eligibility criteria
5 Specify the inclusion and exclusion criteria for the review and how studies were grouped for the syntheses.Methods

Information sources
6 Specify all databases, registers, websites, organisations, reference lists and other sources searched or consulted to identify studies.Specify the date when each source was last searched or consulted.
Table 1 Search strategy 7 Present the full search strategies for all databases, registers and websites, including any filters and limits used.Table 1 Selection process 8 Specify the methods used to decide whether a study met the inclusion criteria of the review, including how many reviewers screened each record and each report retrieved, whether they worked independently, and if applicable, details of automation tools used in the process.

Data collection process
9 Specify the methods used to collect data from reports, including how many reviewers collected data from each report, whether they worked independently, any processes for obtaining or confirming data from study investigators, and if applicable, details of automation tools used in the process.

Methods
Data items 10a List and define all outcomes for which data were sought.Specify whether all results that were compatible with each outcome domain in each study were sought (e.g. for all measures, time points, analyses), and if not, the methods used to decide which results to collect.
Table 2 10b List and define all other variables for which data were sought (e.g.participant and intervention characteristics, funding sources).Describe any assumptions made about any missing or unclear information.
Table 2 Study risk of bias assessment 11 Specify the methods used to assess risk of bias in the included studies, including details of the tool(s) used, how many reviewers assessed each study and whether they worked independently, and if applicable, details of automation tools used in the process.

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Effect measures 12 Specify for each outcome the effect measure(s) (e.g.risk ratio, mean difference) used in the synthesis or presentation of results.Methods

Synthesis methods
13a Describe the processes used to decide which studies were eligible for each synthesis (e.g.tabulating the study intervention characteristics and comparing against the planned groups for each synthesis (item #5)).
Figure 2 13b Describe any methods required to prepare the data for presentation or synthesis, such as handling of missing summary statistics, or data conversions.

Methods
13c Describe any methods used to tabulate or visually display results of individual studies and syntheses.Methods

Section and Topic
Item # Checklist item Location where item is reported 13d Describe any methods used to synthesize results and provide a rationale for the choice(s).If meta-analysis was performed, describe the model(s), method(s) to identify the presence and extent of statistical heterogeneity, and software package(s) used.

Methods
13e Describe any methods used to explore possible causes of heterogeneity among study results (e.g.subgroup analysis, meta-regression).Methods 13f Describe any sensitivity analyses conducted to assess robustness of the synthesized results.N/A

Reporting bias assessment
14 Describe any methods used to assess risk of bias due to missing results in a synthesis (arising from reporting biases).N/A Certainty assessment 15 Describe any methods used to assess certainty (or confidence) in the body of evidence for an outcome.N/A

Study selection 16a
Describe the results of the search and selection process, from the number of records identified in the search to the number of studies included in the review, ideally using a flow diagram.
Figure 2 16b Cite studies that might appear to meet the inclusion criteria, but which were excluded, and explain why they were excluded.N/A

Study characteristics
17 Cite each included study and present its characteristics.Supplementary Materials

Risk of bias in studies
18 Present assessments of risk of bias for each included study.N/A

Results of individual studies
19 For all outcomes, present, for each study: (a) summary statistics for each group (where appropriate) and (b) an effect estimate and its precision (e.g.confidence/credible interval), ideally using structured tables or plots.

Supplementary Materials
Results of syntheses 20a For each synthesis, briefly summarise the characteristics and risk of bias among contributing studies.N/A 20b Present results of all statistical syntheses conducted.If meta-analysis was done, present for each the summary estimate and its precision (e.g.confidence/credible interval) and measures of statistical heterogeneity.If comparing groups, describe the direction of the effect.For more information, visit: http://www.prisma-statement.org/

Figures
Figures 3, 4, 5, 6 and 7 20c Present results of all investigations of possible causes of heterogeneity among study results.Results 20d Present results of all sensitivity analyses conducted to assess the robustness of the synthesized results.N/A Reporting biases 21 Present assessments of risk of bias due to missing results (arising from reporting biases) for each synthesis assessed.N/A Certainty of evidence