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Efficacy and Safety of Pathogen-Reduced Platelets Compared with Standard Apheresis Platelets: A Systematic Review of RCTs

In this systematic review, we evaluate the efficacy and safety of blood components treated with pathogen reduction technologies (PRTs). We searched the Medline, Embase, Scopus, Ovid, and Cochrane Library to identify RCTs evaluating PRTs. Risk of bias assessment and the Mantel–Haenszel method for data synthesis were used. We included in this review 19 RCTs evaluating 4332 patients (mostly oncohematological patients) receiving blood components treated with three different PRTs. Compared with standard platelets (St-PLTs), the treatment with pathogen-reduced platelets (PR-PLTs) does not increase the occurrence of bleeding events, although a slight increase in the occurrence of severe bleeding events was observed in the overall comparison. No between-groups difference in the occurrence of serious adverse events was observed. PR-PLT recipients had a lower 1 and 24 h CI and CCI. The number of patients with platelet refractoriness and alloimmunization was significantly higher in PR-PLT recipients compared with St-PLT recipients. PR-PLT recipients had a higher number of platelet and RBC transfusions compared with St-PLT recipients, with a shorter transfusion time interval. The quality of evidence for these outcomes was from moderate to high. Blood components treated with PRTs are not implicated in serious adverse events, and PR-PLTs do not have a major effect on the increase in bleeding events. However, treatment with PRTs may require a greater number of transfusions in shorter time intervals and may be implicated in an increase in platelet refractoriness and alloimmunization.


Introduction
In recent years, much progress in ensuring the safety of blood and blood components has been observed, especially in order to reduce the risk of transmission of infections.
In Italy, donations are routinely screened for known virus infections (hepatitis C virus (HCV), human immunodeficiency virus (HIV), hepatitis B virus (HBV), and Treponema pallidum) [1]. However, there remains a residual risk of transfusion transmission both for known pathogens and for emerging pathogens for which blood donations are not routinely tested [2][3][4].
In addition, the safety of blood and blood components has also improved with the introduction of measures to reduce the risk of bacterial contamination (diversion of the first 10 mL of blood, accurate skin disinfection at the venipuncture site, and the adoption of standardized and validated operating procedures for blood processing) [1,5].
However, cases of transfusion-associated bacterial sepsis (TABS) from contaminated blood components have been described [6,7]. The transmission of the infection to the recipient is directly related to the amount of blood component transfused, the pathogen concentration, and the degree of immunocompetence of the transfused patient [8,9].

Results
The search identified a total of 776 potentially relevant records. After removal of duplicates, 523 records remained, of which 480 were excluded on the basis of the abstract and/or title. The search identified 43 records that appeared relevant on the basis of their full text or abstract using the original inclusion/exclusion criteria (Figure 1). and/or title. The search identified 43 records that appeared relevant on the b full text or abstract using the original inclusion/exclusion criteria (Figure 1). Twenty-four of them were excluded (reviews, protocols of RCTs, non studies, duplicates, studies containing no informative data). Nineteen RC cluded in the systematic review (see Table 1 for the main characteristics and included studies). Twenty-four of them were excluded (reviews, protocols of RCTs, nonrandomized studies, duplicates, studies containing no informative data). Nineteen RCTs were included in the systematic review (see Table 1 for the main characteristics and results of the included studies).  Overall, 4606 patients were enrolled in the 19 RCTs selected for the review. Of the 19 trials included in the systematic review, 10 compared Intercept ® PR-PLTs with St-PLTs [16,[18][19][20]22,23,25,30,32,34], 6 Mirasol ® PR-PLTs with St-PLTs [17,[26][27][28][29]31], 2 subgroups of patients receiving either Intercept ® or Mirasol ® PR-PLTs with St-PLTs [21,33], and 1 Theraflex ® PR-PLTs with St-PLTs [24] (Table 1).
In 17 studies, PR-PLTs was compared with St-PLTs. Two other studies considered the treatment of whole blood and RBCs with PRT, respectively.
Fifteen were parallel-group RCTs, and 4 were randomized crossover trials. Of the 4606 patients enrolled in the trials, 4332 received at least 1 platelet transfusion (2613 in Intercept ® platelet trials, 1299 in Mirasol ® platelet trials, and 171 Theraflex ® platelet trials).
One trial included children requiring cardiac surgery (16 participants) or adults requiring a liver transplant (28 participants). All of the other participants were thrombocytopenic patients who had a hematological or oncological diagnosis.
With the exception of one trial from Ghana, studies were conducted in industrialized countries, including the USA, Canada, and Europe (France, Germany, Belgium, the Netherlands, Sweden, Spain, and Italy).

Risk of Bias in Included Studies
Fourteen out of 19 reports were at high or unclear risk of bias for 1 or more domains; 6 were at high risk of bias in 1 domain, and 10 were at unclear risk of bias in 1 or more domains ( Figure 2).
Five reports [18,20,21,28] were at low risk of bias in all the domains. We assessed 5 studies as being at unclear risk of selection bias because they were unclear about the random sequence generation and the allocation concealment, while 14 studies were at low risk of selection biases. There were 4 open-label trials, and they were graded as high risk of performance bias (blinding of participants and personnel). Six studies were graded at unclear risk of detection bias due to the fact that they did not provide information to permit judgement about "high" or "low" risk of bias related to the blinding of participants and personnel. Nine studies were reported as double blind. Fourteen studies (73.6%) were graded at low risk of detection bias due to the fact that the assessor was blinded to treatment allocation, whereas 5 studies were graded at unclear risk of detection bias because they did not provide information to permit judgement about "high" or "low" risk of bias related to the blinding of outcome assessors. One trial [33] was judged at high risk of attrition bias because only 179 out of the 237 patients enrolled in the Intercept ® arm of the original study [21] and 179 out of 201 of the Mirasol ® arm were included in this analysis. Other 5 studies were judged at unclear risk of attrition bias, and the remaining 13 (68%) studies were judged at low risk of bias. Risk of bias for selective reporting or other potential source of bias was present in 3 studies (high risk in 1 case, unclear risk in 2), 2 of which available only as abstracts [32,34].

Effects of Interventions
See Supplementary Table S1, data and analyses.

Outcomes
The outcomes reported (Table 2) were bleeding events, adverse events, number of patients with acute transfusion reactions, platelet count increment (CI) and corrected count increment (CCI), number of patients with platelet refractoriness and alloimmunization, number of platelet transfusions/patient, and number of RBC transfusions/patient.
In the overall analysis and in subgroup analyses, participants who received PR-PLT transfusions had a lower 24 h CCI. In the overall analysis and in the subgroup of Intercept ® and Mirasol ® trials, the no. of patients with PLT refractoriness was significantly higher in PR-PLT group compared to St-PLT. No statistically significant between-groups difference was observed in a single trial with Theraflex ® .  2 Downgraded twice for ROB (differences in the definition and assessment of overall adverse events) and inconsistency (substantial heterogeneity). 3 Downgraded for imprecision because most of the trials were underpowered to detect the occurrence of rare outcomes. * The basis for the assumed risk is the mean control group risk across studies. The corresponding risk (and its 95% confidence interval) is based on the assumed risk in the comparison group and the relative effect of the intervention (and its 95% CI).

Risk of Bias in Included Studies
Fourteen out of 19 reports were at high or unclear risk of bias for 1 or more domai 6 were at high risk of bias in 1 domain, and 10 were at unclear risk of bias in 1 or m domains (Figure 2). Five reports [18,20,21,28] were at low risk of bias in all the domains. We assesse studies as being at unclear risk of selection bias because they were unclear about the ra dom sequence generation and the allocation concealment, while 14 studies were at l risk of selection biases. There were 4 open-label trials, and they were graded as high r of performance bias (blinding of participants and personnel). Six studies were graded unclear risk of detection bias due to the fact that they did not provide information to p mit judgement about "high" or "low" risk of bias related to the blinding of participa and personnel. Nine studies were reported as double blind. Fourteen studies (73.6%) w graded at low risk of detection bias due to the fact that the assessor was blinded to tre ment allocation, whereas 5 studies were graded at unclear risk of detection bias becau they did not provide information to permit judgement about "high" or "low" risk of b related to the blinding of outcome assessors. One trial [33] was judged at high risk of trition bias because only 179 out of the 237 patients enrolled in the Intercept ® arm of original study [21] and 179 out of 201 of the Mirasol ® arm were included in this analy Other 5 studies were judged at unclear risk of attrition bias, and the remaining 13 (68

Bleeding Events
Bleeding events at more than 7 days were reported as any bleeding events (WHO grades 1 to 4), clinically significant bleeding (WHO grade ≥ 2), and severe bleeding (WHO grade ≥ 3) (Figure 3).

Adverse Events
Adverse events complicating platelet transfusion were reported as any adverse events and/or serious adverse events, as defined in individual studies, occurring during the study and follow-up periods (Figure 4).

Adverse Events
Adverse events complicating platelet transfusion were reported as any adverse events and/or serious adverse events, as defined in individual studies, occurring during the study and follow-up periods (Figure 4).  Several studies reported also acute mild transfusion reactions (e.g., rigors, fever, skin rash, and urticaria). Any adverse event was reported in 11 trials (12 records, 3445 patients). No between-groups difference in the occurrence of any adverse event was found in the 6 Intercept ® trials (RR 1.01; 95% CIs, 0.89/1.15; p = 0.84). By contrast, a higher incidence of adverse events in the PR-PLTs compared with the St-PLTs was found in the overall analysis (RR 1.09; 95% CIs, 1.01/1.19; p = 0.03) and in a subgroup analysis of Mirasol ® trials ((RR 1.16; 95% CIs, 1.04/1.30; p = 0.006). The quality of the evidence was graded as low due to heterogeneity and risk of bias. Serious adverse events were reported in 13 studies (3247 patients), 7 with Intercept ® (2078 patients), 5 with Mirasol ® (1027 patients), and 1 with Theraflex ® (142 patients). No between-groups differences in the occurrence of a serious adverse event was found in the overall analysis (RR 1.01; 95% CIs, 0.82/1.24) and in a subgroup analysis of Intercept ® trials (RR 1.08; 95% CIs, 0.86/1.37), a Mirasol ® trial (RR 0.78; 95% CIs, 0.49/1.25), and a Theraflex ® trial (RR not estimable because no events were reported in either group): moderate quality of evidence due to imprecision because most of the trials were underpowered to detect the occurrence of rare outcomes. Acute transfusion reactions, as defined among the adverse events or the serious adverse events (e.g., rigors, fever, skin rash, and urticaria) were reported in 7 trials, and the occurrence did not differ between groups (RR 0.95; 95% CIs, 0.62/1.47).

Platelet Refractoriness and Platelet Alloimmunization
There was heterogeneity in the definition of refractoriness (e.g., 2 successive 1 or 24 h CCIs below 7.5 × 10 3 or 4.5/5 × 10 3 ), and in some of the selected trials, subjects with a previous history of clinical refractoriness to platelet transfusions were not eligible for inclusion in the analysis. Ten reports (2380 participants) reported the number of patients experiencing platelet refractoriness. Participants who received pathogen-reduced platelet transfusions had an increased risk of developing platelet refractoriness in the overall analysis (RR 2.59: 95% CIs, 1.98/3.39; p < 0.00001) and in a subgroup analysis of 6 Intercept ® trials (RR 2.85; 95% CIs, 1.96/4.15; p < 0.00001) and 3 Mirasol ® trials (RR 2.46; 95% CIs, 1.61/3.76; p < 0.0001); in the Theraflex ® trial, there was a trend towards a higher refractoriness in PR-PLT recipients compared with St-PLTs, but the difference was not statistically significant (RR 1.81; 95% CI, 0.71/4.64; p = 0.22) (Figure 7). The quality of the evidence was graded as high.   Platelet refractoriness and refractoriness specifically due to alloimmunization were reported in 11 reports (2628 participants) (Figure 8). Similarly to the refractoriness analysis, patients who received PR-PLT transfusions had an increased risk of developing platelet refractoriness and alloimmunization in the overall analysis (RR 1.77; 95% CIs, 1.47/2.13; p < 0.00001) and in a subgroup analysis of 7 Intercept ® trials (RR 1.61; 95% CIs, 1.28/2.02; p < 0.0001) and of 3 Mirasol ® trials (RR 2.14; 95% CIs, 1.50/3.07; p < 0.0001); in the Theraflex ® trial, there was a trend towards a higher refractoriness in PR-PLTrecipients compared with St-PLTs, but the difference was not statistically significant (RR 1.77; 95% CI, 0.74/4.24; p = 0.20). The quality of the evidence was graded as high.

Discussion
The main aims of this systematic review and meta-analysis are to comprehensively evaluate the efficacy and safety of platelets treated with currently available PRTs, especially by comparing the treated products with the standards in terms of reduction of bleeding and transfusion-related adverse reactions.
We included in this review 19 RCTs evaluating 4332 patients receiving platelet transfusions treated with three different PRTs. The majority of the participants were patients with hematological malignancies. On average, compared with St-PLTs, PR-PLT transfusion does not increase the occurrence of bleeding events, although a slight increase in the occurrence of severe bleeding events was observed in the overall comparison. The quality of evidence for these outcomes was from moderate to high. No between-groups difference in the occurrence of serious adverse events was observed, in the overall analysis, and in subgroup analyses of Intercept ® , Mirasol ® , and Theraflex ® trials (moderate quality of evidence). In the overall analysis and in the subgroup of Mirasol ® trials, but not in the subgroup analysis of Intercept ® trials, overall adverse events were more commonly observed in PR-PLT recipients compared with St-PLT recipients; the quality of the evidence was graded as low due to risk of bias (differences in the definition and assessment of overall adverse events) and inconsistency (substantial heterogeneity between trials). On average, PR-PLT recipients had a lower 1 and 24 h CI and a lower 1 and 24 h CCI (moderate quality of evidence due to between-trials heterogeneity). There was also high quality of evidence that the number of patients with platelet refractoriness and the number of patients with

Discussion
The main aims of this systematic review and meta-analysis are to comprehensively evaluate the efficacy and safety of platelets treated with currently available PRTs, especially by comparing the treated products with the standards in terms of reduction of bleeding and transfusion-related adverse reactions.
We included in this review 19 RCTs evaluating 4332 patients receiving platelet transfusions treated with three different PRTs. The majority of the participants were patients with hematological malignancies. On average, compared with St-PLTs, PR-PLT transfusion does not increase the occurrence of bleeding events, although a slight increase in the occurrence of severe bleeding events was observed in the overall comparison. The quality of evidence for these outcomes was from moderate to high. No between-groups difference in the occurrence of serious adverse events was observed, in the overall analysis, and in subgroup analyses of Intercept ® , Mirasol ® , and Theraflex ® trials (moderate quality of evidence). In the overall analysis and in the subgroup of Mirasol ® trials, but not in the subgroup analysis of Intercept ® trials, overall adverse events were more commonly observed in PR-PLT recipients compared with St-PLT recipients; the quality of the evidence was graded as low due to risk of bias (differences in the definition and assessment of overall adverse events) and inconsistency (substantial heterogeneity between trials). On average, PR-PLT recipients had a lower 1 and 24 h CI and a lower 1 and 24 h CCI (moderate quality of evidence due to between-trials heterogeneity). There was also high quality of evidence that the number of patients with platelet refractoriness and the number of patients with platelet refractoriness and alloimmunization were significantly higher in PR-PLT recipients compared with St-PLT recipients. In the same way, PR-PLT recipients had a higher number of platelet transfusions and RBC transfusions compared with St-PLT recipients, with a shorter transfusion time interval.
Similar to the Cochrane review [35], our review reports a significant reduction of the 1 and 24 h CI and CCI in PR-PLT recipients compared with St-PLTs recipients, as well as an increased platelet transfusion and RBC demand. However, unlike the Cochrane review, we did not observe an increase in overall bleeding events (RR 1.03; 95% CI, 0.85 to 1.24), but a slight increase in severe bleeding events in the overall analysis (RR 1.16; 95% CIs, 1.02/1.32); also, we graded the evidence about serious bleeding events as high level of certainty, compared with the moderately low quality found in the Estcourt et al. review [35]. Moreover, unlike the Cochrane review, we did not include all-cause mortality and attributable mortality (e.g., due to infection or bleeding) among the outcomes, because there was no evidence in the trials considered that overall mortality and attributable mortality were related to platelet transfusions, but rather to the underlying clinical conditions (most of the participants were patients with thrombocytopenia from oncological diseases).
In conclusion, the results of our review show that the treatment of blood components with PRTs is not implicated in serious adverse events in the recipient. In particular, the treatment of platelets does not have a major effect on the increase in bleeding events. However, treatment with PRTs may require a greater number of transfusions in shorter time intervals and may be implicated in an increase in platelet refractoriness and alloimmunization. To better understand and define the adverse events and any limitations related to the treatment with PRTs, it is important to conduct further investigations in this regard also through a comparative analysis of the different PRTs.

Materials and Methods
This systematic review was conducted according to recommended PRISMA checklist guidelines [37]. The protocol has been registered in PROSPERO (registration number CRD42022320422), the international prospective register of systematic reviews. The review is aimed at evaluating the safety and effectiveness of PR-PLTs in people undergoing platelet transfusions. We included RCTs comparing the transfusion of PR-PLTs with St-PLTs. Three different types of pathogen reduction technologies were considered, including the Intercept ® , Mirasol ® , and Theraflex ® systems.

Search Strategy
A computer-assisted literature search of the Medline (through PubMed), Embase, Scopus, Ovid, and Cochrane Library was performed (latest search in February 2022) to identify RCTs evaluating pathogen reduction technologies. A combination of the following text words was used: platelet AND pathogen reduction, pathogen reduction platelet, Mirasol ® platelet, Intercept ® platelet, Theraflex ® platelet. In addition, we checked the reference lists of the most relevant items (original studies and reviews) in order to identify potentially eligible studies not captured by the initial literature search. For the search, no restriction on language was applied.

Data Collection and Analysis
For each RCT included in the systematic review, the following data were extracted by two reviewers (MC and IP) independently: first author, year of publication, details of intervention in study and control group, sample size, pathogen reduction technology used, control group, outcome measurements, and main results. Measures of treatment effect were mean differences (MD) together with 95% confidence intervals (CI) for continuous outcome measures and risk ratio (RR) with 95% CI for dichotomous outcomes. For continuous outcomes, the score had to be reported as mean and standard deviation (SD); when studies reported other dispersion measures, such as median and range, or standard error (SE) of the mean or 95% CI of the mean, we calculated the mean and SD from these measures in order to perform the relevant meta-analytical pooling [38,39]. Disagreement was resolved by consensus and by the opinion of a third reviewer (FM), if necessary.
The study weight was calculated using the Mantel-Haenszel method. We assessed statistical heterogeneity using t2, Cochran's Q, and I 2 statistics. The I 2 statistic describes the percentage of total variation across trials that is due to heterogeneity rather than sampling error. In the case of not important heterogeneity (I 2 < 40), studies were pooled using a fixed-effects model. Where values of I 2 were >40, a random-effects analysis was undertaken. All calculations were performed using Excel and RevMan 5.4.

Outcomes
The outcomes included in the analysis were: bleeding events (any bleeding event, significant bleeding, and serious bleeding) mainly using two bleeding scales: WHO Bleeding Scale Grades 0 to 4 and Common Terminology Criteria for Adverse Events (CTCAE) Grades 1 to 5 or equivalent; adverse events graded for clinical severity (any adverse event, serious adverse event, acute transfusion reactions); platelet CI and CCI at 1 and 24 h; number of patients with platelet refractoriness and number of patients with platelet refractoriness and alloimmunization; number of platelet transfusions/participant and number of red blood transfusions/participant; and platelet transfusion interval (day of the next platelet transfusion).

Subgroup Analyses
We undertook subgroup analyses according to type of PRT methodology used (Intercept ® , Mirasol ® , and Theraflex ® ).

Assessment of Risk of Bias in Included Studies
Two review authors (MC, IP) independently assessed the risk of bias of each included study following the domain-based evaluation described in the Cochrane Handbook for Systematic Reviews of Interventions [39]. They discussed any discrepancies and achieved consensus on the final assessment. The Cochrane 'risk of bias' tool addresses six specific domains: sequence generation, allocation concealment, blinding, incomplete data, selective outcome reporting, and other issues relating to bias. We have presented our assessment of risk of bias using two 'risk of bias' summary figures: (1) a summary of bias for each item across all studies and (2) a cross tabulation of each trial by all of the 'risk of bias' items ( Figure 2).
We used the principles of the GRADE system to assess the quality of the body of evidence associated with specific outcomes, and constructed a 'summary of findings' table (Table 2) using RevMan 5 [40].
These tables present key information concerning the certainty of the evidence, the magnitude of the effects of the interventions examined, and the sum of available data for the main outcomes [39]. The 'summary of findings' tables also include an overall grading of the evidence related to each of the main outcomes using the GRADE approach, which defines the certainty of a body of evidence as the extent to which one can be confident that an estimate of effect or association is close to the true quantity of specific interest. The certainty of a body of evidence involves consideration of within-trial risk of bias (methodological quality), directness of evidence, heterogeneity, precision of effect estimates, and risk of publication bias [41].
When evaluating the 'risk of bias' domain, we downgraded the GRADE assessment when we classified a study as being at high risk of bias for one or more of the following domains: selection, attrition, performance, detection, reporting, and other bias, or when the 'risk of bias' assessment for selection bias was unclear (this was classified as unclear for either the generation of the randomization sequence or the allocation concealment domain).
We have presented the following outcomes in the 'summary of findings' table: bleeding events, adverse events, and platelet CI and CCI at 1 and 24 h; number of patients with platelet refractoriness and number of patients with platelet refractoriness and alloimmunization; and number of platelet transfusions/participant and number of red blood transfusions/participant.

Conclusions and Perspectives
Blood components treated with PRTs are not implicated in serious adverse events, and PR-PLTs do not have a major effect on the increase in bleeding events. However, treatment with PRTs may require a greater number of transfusions in shorter time intervals and may be implicated in an increase in platelet refractoriness and alloimmunization. However, to better understand and define the clinical effectiveness and safety of PRTs, further investigations directly comparing different pathogen reduction techniques and their use in non-hemato-oncological patients are required.