Topical imiquimod cream for the treatment of cervical intraepithelial neoplasia

Objectives

This is a protocol for a Cochrane Review (intervention). The objectives are as follows:

To assess the effectiveness and adverse effects of imiquimod in the treatment of women with cervical intraepithelial neoplasia.

Background

Description of the condition

Cervical cancer is one of the most common cancers, and a leading cause of mortality among women worldwide. In 2020, it accounted for an estimated 604,000 diagnoses and an estimated 341,000 deaths (Sung 2021; WHO 2021). Early detection and proper treatment of precancerous cervical lesions improve the survival of women (WHO 2022). Effective management of women with a precancerous lesion or a positive screening test is one of the three pillars of the global strategy to eliminate cervical cancer (WHO 2020).

The primary cause of cervical cancer and precancerous conditions is persistent and long‐lasting infection caused by one of the high‐risk types of human papillomavirus (HPV) – specifically the two most prevalent types, 16 and 18 – through sexual activity. Most HPV infections will resolve spontaneously, but a minority persist and lead to cervical pre‐cancer (WHO 2014). Following persistent HPV infection of the cervix, squamous intraepithelial dysplasia can develop, exhibiting abnormal cytological and histological differentiation, maturation, and stratification of squamous epithelium of the cervix (Muntean 2010).

Cervical precancerous lesions can be detected using screening methods, including Pap smear tests, HPV testing, co‐testing, liquid‐based cytology, and molecular tests (Mello 2022). A recent review reported that cervical cytology was the predominant screening method in 78% of 202 countries, while 48 of 139 countries advocated for primary HPV testing (Bruni 2022). The Bethesda system was established in 1991 for reporting cervical and vaginal cytologic diagnoses, and is commonly used to report abnormal cervical cytology (Luff 1991). The system categorizes squamous intraepithelial lesions (SIL) of the uterine cervix into atypical squamous cells of undetermined significance (ASCUS); atypical squamous cells, cannot exclude high‐grade SIL (ASC‐H); low‐grade SIL; low‐grade SIL, cannot exclude high‐gradeSIL (LSIL‐H); and high‐grade SIL (Alrajjal 2021)).

LSIL‐H cases have been found to overlap with low‐grade SIL and ASC‐H, and it has been suggested that women with LSIL‐H should be managed similarly to those with ASC‐H and high‐grade SIL (Alrajjal 2021). Before the implementation of the Bethesda system, many non‐standard terms were used to report cervical cytology, such as dysplasia, cervical intraepithelial neoplasia, and Papanicolaou classification, which were not standardized and lacked reproducibility (Nayar 2017). The relation between different cervical cytologic reporting system is described in Table 1.

1. The relationship between different cervical cytology reporting systems.

Papanicolaou class	Dysplasia system (WHO)	CIN system	The Bethesda system
Class I Class II	Normal Atypia	Normal	NILM Benign cellular changes ASC‐US
	Mild dysplasia or Mild dyskaryosis	CIN1	LSIL
Class III	Moderate dysplasia or Moderate dyskaryosis	CIN2	HSIL
Class IV	Severe dysplasia or Severe dyskaryosis or Carcinoma in‐situ	CIN3	HSIL
Class V	Invasive carcinoma	Invasive carcinoma	SCCA

ASC‐US: atypical squamous cell of undetermined significance; CIN: cervical intraepithelial neoplasia; HSIL: high‐grade intraepithelial lesion; LSIL: low grade intraepithelial lesion; NILM: negative for intraepithelial lesion or malignancy; SCCA: squamous cell carcinoma; WHO: World Health Organization.

During the histologic examination of cervical precancerous lesions, a continuum of histopathological changes in the transformation zone of the cervix are traditionally graded as cervical intraepithelial neoplasia (CIN) 1 (mild dysplasia), CIN 2 (moderate dysplasia), CIN 3 (severe dysplasia, or CIS (carcinoma in situ (IARC 2007; Kumar 2017)). Around 60% of CIN 1 lesions spontaneously regress to normal after one year, while CIN 2 and CIN 3 lesions have a higher risk of developing invasive cancer over several years (Mello 2022). Biomarkers play a crucial role in the diagnosis and management of cervical lesions. Overexpression of p16INK4A, a cyclin‐dependent kinase inhibitor, has been identified as a specific marker for dysplastic and neoplastic epithelial cells of the cervix, and immunohistochemical staining for p16INK4A can help differentiate between benign and malignant cervical lesions (Klaes 2001). Another biomarker, Ki‐67, is a marker of cellular proliferation, and is commonly used to assess the growth potential of cervical lesions (Sarma 2021). For reporting histopathology of squamous lesions of the lower anogenital tract, the CIN lesions are assimilated into two groups: (1) histologic low‐grade intraepithelial lesions including CIN 1/CIN 2 (p16‐negative); and (2) histologic high‐grade intraepithelial lesions covering CIN 2 p16‐positive and CIN 3 lesions. Increasing evidence suggests that treatment should be reserved for CIN 3, while CIN 2 lesions may be treated more conservatively (Kremer 2022). Without medical attention, it is estimated that 30% of high‐grade SIL will progress to cancer (McCredie 2008); however, the risk is dramatically reduced with treatment (Arbyn 2014). Previous studies found that recognition and treatment of CIN 2 and CIN 3 could reduce the incidence and mortality of invasive cervical cancer (Aareleid 1993; Andrae 2012). Annually, 1% to 2% of women worldwide are estimated to have CIN 2 plus lesions, with a 10% higher prevalence in women who are HIV‐positive (WHO 2013).

Description of the intervention

The standard management for CIN lesions involves surgical interventions, including cervical excision, ablation, or cryotherapy procedures, such as large loop excision of the transformation zone (LLETZ), cold knife conization, and laser conization, depending on the staging of the lesion and available resources (Prendiville 2017; Guido 2014). Despite success rates of approximately 90%, adverse effects, such as subfertility (Bevis 2011; Jin 2014), spontaneous preterm birth, premature rupture of the membranes, chorioamnionitis, low birth weight, admission to neonatal intensive care, and perinatal mortality are increased after treatment with excisional methods (Kyrgiou 2016).

Non‐surgical, less invasive treatment options may address the limitations of current treatment models; as adjuctive treatments, they may also improve the efficacy of the recommended treatment (Desravines 2020). Research has been conducted on topical therapies, which allow self‐application by women, as a medical alternative for the treatment of CIN lesions and HPV infection (Maiman 1999; Rahangdale 2014).

Imiquimod (IMQ) is an immune response modifier that was approved in 1997 for topical application for external genital and perianal warts (Sauder 2003). IMQ stimulates innate and acquired immune responses, which consequently lead to inflammatory cell infiltration within the field of drug application, followed by apoptosis of diseased tissue. It does not have direct antiviral activity (Miller 1999). However, due to its ability to stimulate the host’s immune system, IMQ has been recognized for its off‐label use potential in over 60 unlisted conditions (Ganjian 2009; Skinner 2002). Several intervention studies analyzed the effectiveness of IMQ in HPV‐related diseases, including precancerous lesions of the genital tract (vulval, vaginal, and cervical intraepithelial neoplasia (Desravines 2020; De Witte 2015)). Systematic reviews have also shown that IMQ can be effective for high‐grade vaginal and vulval intraepithelial neoplasia (Inayama 2021; Lawrie 2016; Voss 2022).

For precancerous conditions of the cervix, clinical trials have examined the effectiveness of local application of IMQ cream in women with CIN 2+ as either a stand‐alone treatment option, or as an adjunct to excision or ablation; these trials found HPV clearance, histologic regression, recovery from high‐grade SIL, and reduction of recurrent dysplasia rate (Cokan 2021; Fonseca 2021; Grimm 2012; Hendriks 2022; Pachman 2012; Polterauer 2022).

The recommended regimen of IMQ for listed conditions includes topical application of 1% to 5% concentration for anogenital warts, and topical application of 5% concentration for cutaneous molluscum contagiosum (Banerjee 2017; van der Wouden 2017). However, consensus has not yet been reached on the optimum dosage and route for IMQ therapy in CIN lesions. The common practice in clinical trials is to use IMQ in concentrations of 1% to 5%, applied locally to the cervix, three to seven times per week, for 8 to 16 weeks (Dolin 2015). In 2010, another regimen, which used self‐applied vaginal suppositories containing 6.25 mg of IMQ for 16 weeks, was evaluated (Grimm 2012).

How the intervention might work

IMQ is a low‐molecular‐weight imidazoquinoline heterocyclic amine, which triggers a local immune response that is effective against both cancer cells and viruses (Miller 1999). Although it has no direct antiviral effects, it shows antiviral and antitumor effects in vivo by inducing cytokines and enhancing cell‐mediated cytolytic antiviral activity. IMQ stimulates the innate immune response by inducting cytokines, and the cellular arm of acquired immunity by inducting interferon‐alpha, interferon‐gamma, and interleukin‐12 (Sauder 2000). Schön 2003 found that IMQ induced apoptosis in all squamous cell carcinoma cell lines, and it is a safe and effective treatment for usual type vulvar intraepithelial neoplasia (VIN), which is pathophysiologically comparable to CIN (van Seters 2008). A 2015 Cochrane review that evaluated five randomized controlled trials involving 297 women, concluded that topical IMQ appears to be a safe and effective treatment for high‐grade VIN, even though local side effects may necessitate dose reductions. Long‐term follow‐up data are needed to assess effects on progression to vulval cancer (Lawrie 2016; Pepas 2015).

Previous studies showed improvements in women with CIN lesions who were treated with IMQ for histological regression, recovery from high‐grade SIL, change in HPV status, and reduction of recurrent dysplasia rate (Cokan 2021; Fonseca 2021; Grimm 2012; Hendriks 2022; Pachman 2012; Polterauer 2022).

A recent Phase 2 randomized controlled trial found that in the group receiving IMQ, 63% of women (32 of 51) showed histologic regression six months after treatment, and 37% of women (19 of 51) achieved complete histologic remission (Polterauer 2022).

An earlier Phase 2 randomized controlled trial, conducted in 2017, compared the effectiveness of IMQ treatment followed by a loop electrosurgical excision procedure to a control group that received surgery alone. One woman (N = 40) in the control group and one woman in the experimental group (N = 46) experienced progression of the lesion to invasive squamous cell carcinoma. In the intention‐to‐treat (ITT) analysis, histologic regression occurred in 53.3% of the specimens in the experimental group compared to 22.5% in the control group. Persistent high‐grade SIL was observed in 44.5% of the specimens in the experimental group compared to 75% in the control group (Fonseca 2021).

HPV clearance rates vary across studies. In Grimm 2012, HPV clearance was higher in the IMQ group (60%) compared with the placebo group (14%). However, in a more recent randomized controlled trial, HPV clearance was lower in the IMQ group (43.1%) compared to the group that received large loop excision of the transformation zone (LLETZ; 64.3%), with ITT analysis (Polterauer 2022).

Common adverse events associated with IMQ therapy included mild to moderate systemic and local effects: headache, fever, fatigue, myalgia, vaginal discharge, vaginal bleeding, vulvar pruritus, pain from vulvar and vaginal inflammation (Grimm 2012; Lin 2012; Pachman 2012). However, one case series reported more severe side effects, including hyponatremia, leukopenia, and corneal erosion (Wouters 2019). Two case reports documented effluvium after using imiquimod therapy (Borst 2019; Cokan 2022).

A recent randomized controlled trial assigned 104 women between the ages of 18 and 40, who had histological high‐grade SIL (CIN 2 p16+ and CIN 3) to receive either IMQ or LLETZ treatment (52 women in each group). Side effects were experienced by 88.5% of women (46 of 52); moderate side effects were reported by 38.5% (20 of 52), and severe side effects by 13.5% (7 of 52). Nine women discontinued treatment completely, most of them within the first 10 weeks, primarily due to side effects. The occurrence of side effects decreased slightly over the follow‐up period, with a reduction in overall occurrence and moderate side effects. Common Terminology Criteria for Adverse Events (CTCAE) grade 3 side effects, i.e. severe or medically significant but not immediately life‐threatening, hospitalization, or prolongation of hospitalization indicated, disabling, limiting self‐care activities of daily life, were resolved by the 20‐week follow‐up (Cokan 2021).

There is limited evidence supporting different levels of effectiveness of IMQ in specific populations with CIN. Most studies on IMQ for CIN include women from different populations. A multicenter, non‐randomized controlled trial with 123 participants (N = 61 in the IMQ group and N = 62 in the LLETZ group) conducted a subgroup analysis of responders versus non‐responders in the IMQ group. None of the potential predictors (nulliparity, single sexual partner, previous abnormal cytology, CIN 2 at diagnosis, smoking, number of imiquimod applications, HPV 16/18 infection at baseline) predicted the outcome of IMQ treatment (Hendriks 2022).

Why it is important to do this review

While there are concerns that the standard treatment of surgical excision may potentially impact pregnancy outcomes (e.g. preterm delivery, low birth weight, and premature rupture of membranes), IMQ shows potential as a treatment option for low‐ and middle‐income countries with a clinician shortage and limited access to specialized healthcare, despite limitations related to its availability and storage. Clinical trials that evaluated the effects of IMQ in women with cervical cancer‐related conditions found varying levels of effectiveness (Cokan 2021; Fonseca 2021; Grimm 2012; Hendriks 2022; Pachman 2012; Polterauer 2022). A review published in 2015, on IMQ treatment for vulvar, vaginal, and cervical intraepithelial neoplasia, did not reach a conclusion about effectiveness, due to limited evidence at the time (De Witte 2015). A more recent narrative review, which covered various topical treatments used for CIN lesions, found that IMQ was the most promising agent of the immunomodulators they reviewed, but lacked conclusive evidence (Desravines 2020). Randomized controlled trials that evaluated the effectiveness of IMQ in women with high‐grade SIL have recently been published (Cokan 2021; Fonseca 2021; Polterauer 2022). Recently published reviews did not assess the risk of bias, or GRADE the evidence (Inayama 2023; Mutombo 2019; van de Sande 2023). In our study, we plan to complete both of these assessments to enhance the reliability and transparency of the evidence evaluation.

Objectives

To assess the effectiveness and adverse effects of imiquimod in the treatment of women with cervical intraepithelial neoplasia.

Methods

Criteria for considering studies for this review

Types of studies

We will include randomized controlled trials (RCTs) and cluster‐randomized trials. We will include the data from both full‐text publications and abstracts that offer sufficient information, such as population and sample details, clear methodology, outcome variables, results, and measurement methods. If the abstract does not provide sufficient information, we will contact the study authors to request the full information. Studies published in the abstract will only be eligible for inclusion if the study authors confirm in writing that the data to be included in the review have come from the final analysis and will not change. In studies with only a subset of eligible participants, we will include the data of the population that meet the review criteria.

If the necessary data are not reported in the included studies, we will contact the study authors for more information. We plan to include non‐English articles, and will translate them using Google Translation.

We will exclude quasi‐randomized trials from our review. Studies in which the allocation of participants to treatment groups is not truly random, such as those based on alternate days of the week or the hospital record number, will be defined as quasi‐randomized trials. We will exclude cross‐over trials, and observational studies. We will exclude letters to the editor, conference records, and literature reviews from our review.

Types of participants

We will include studies with women with any grade of cervical intraepithelial neoplasia (CIN) lesions, and of any age. We will consider studies that used histopathological, cytological, or other recognized diagnostic criteria for CIN, provided the studies provide clear descriptions of the specific diagnostic method used.

Types of interventions

The intervention will be the prescription of IMQ in any formulation (vaginal suppository or topical cream application to the cervix), any dose, any schedule, any applicator (self‐administered or provider administered), and in any setting (both inpatient and outpatient). We will compare the use of IMQ with placebo, no intervention, or standard excisional treatment used in the studies. We will compare different regimens of IMQ cream.

Types of outcome measures

We will include all eligible studies, regardless of the outcomes reported. We will include any duration of follow‐up defined by the authors. However, we will prioritize the six‐month post‐completed treatment follow‐up visit as the primary time point for all outcomes. We will consider other relevant time points if reported, but we may not report them in the summary of findings tables. We will discuss notable findings or differences between those time points in the review.

Primary outcomes

• Regression of CIN

• Discontinuation of treatment due to side effects

Regression of CIN will be defined as complete remission or remission to CIN 1. We will consider regression of CIN when the included studies report a reduction in the severity or extent of CIN lesions, based on histopathological, cytological, or other established diagnostic criteria. The criteria for regression must be clearly defined by the authors, including duration.

We will consider discontinuation of treatment due to side effects when the included studies report instances when the women stopped or interrupted the treatment protocol specifically due to adverse reactions or unwanted effects.

Secondary outcomes

• Progression of CIN to higher grades of CIN, or cancer

• Grade 2+ pain from vaginal and vulvar inflammation

• Persistent dysplasia

• Persistent high‐risk human papillomavirus (HPV)

• Any grade 2+ side effect

We will consider progression of CIN when the included studies report an increase in the severity or extent of CIN lesions, transitioning to higher grades of CIN, or the development of cervical cancer. Progression should be based on histopathological examination or other established diagnostic criteria, and the criteria for progression, including duration, should be clearly defined by the trial authors.

We will consider grade 2+ pain from vaginal and vulvar inflammation when the included studies report moderate to severe pain levels associated with inflammation of the vaginal and vulvar tissues. Pain assessment should be based on standardized pain scales, such as the numeric rating scale (NRS) or visual analog scale (VAS).

We will consider persistent dysplasia when the included studies report the continued presence of abnormal cellular changes in the cervical epithelium over time. Persistence should be based on repeated histopathological evaluations or assessments, with the criteria for persistence clearly defined by the trial authors.

We will consider persistent high‐risk HPV when the included studies report the continued presence of high‐risk types of HPV in the cervix over an extended period. Persistence should be determined through molecular techniques, such as polymerase chain reaction (PCR) or other valid HPV genotyping methods, with the specific HPV type clearly specified, and the duration of persistence defined by the trial authors.

We will consider grade 2+ side effects when the included studies report moderate to severe adverse events that required medical intervention or management, or unwanted effects experienced by participants. Side effects should be assessed and graded based on standardized scales, such as the Common Terminology Criteria for Adverse Events (CTCAE).

Search methods for identification of studies

Electronic searches

We will systematically search the following electronic bibliographic databases:

• Cochrane Central Register of Controlled Trials (CENTRAL; current issue) in the Cochrane Library;

• Medline Ovid (1946 to search date);

• PubMed (1971 to search date);

• Scopus Elsevier (search date);

• CINAHL EBSCO (Cumulative Index to Nursing and Allied Health Literature; 1982 to search date);

• LILACS VBS (Latin American and Caribbean Health Science Information database; 1982 to search date).

The search strategies for these databases are presented in Appendix 1. We will not limit the search results for publication year or language.

We will also search for trial registrations and ongoing trial reports in ClinicalTrials.gov (clinicaltrials.gov/) and the World Health Organization (WHO) International Clinical Trials Registry Platform (ICTRP; apps.who.int/trialsearch) using the search strategies described in Appendix 2.

Searching other resources

We will screen the reference lists of included studies and relevant systematic reviews identified during the search to identify additional studies. We will also search for gray literature, such as dissertations, and will contact authors of included studies for more information if needed.

Data collection and analysis

We will collect and analyze the data according to guidance in the Cochrane Handbook for Systematic Reviews of interventions (Higgins 2023).

Selection of studies

We will use Covidence, a web‐based software that helps review authors to independently screen records, store the search results, and identify and remove any duplicate records (Covidence). Two review authors (HYO and LTS) will independently screen the titles and abstracts of the remaining records, and select potentially relevant studies for full‐text review. We will proceed with these two screening processes using Covidence. Any disagreements will be resolved by discussion. If necessary, a third review author will be invited for consultation.

We will contact the trial authors to request any missing data. We will present the number of records excluded at each stage of the screening process in a PRISMA flow diagram. For the full‐text screening process, we will provide reasons for any articles excluded.

Data extraction and management

We will develop a data extraction form using Microsoft Excel, which will be pretested and checked for accuracy. When information is unclear, we will contact the study authors to request further details. We will extract the following data from the included studies.

• Author, year of publication and language

• Country

• Setting

• Inclusion and exclusion criteria

• Study design, methodology

• Study population (total number of participants in each group, their characteristics, age, comorbidity, and other baseline characteristics)

• Number of intervention groups

• Intervention details (type of intervention, description of intervention, frequency and duration of intervention)

• Control details (other reported information besides no active intervention)

• Duration of follow‐up

• Outcomes: outcome definition, unit of measurement, scale of measurement, number of participants allocated to each intervention arm, and number of participants at the end of the study

Two review authors (HYO, LTS) will independently extract the data. Differences between review authors will be resolved by discussion, or by appeal to a third review author if necessary (KLS or KNT).

Assessment of risk of bias in included studies

Two reviewers (HYO and LTS) will independently assess the risk of bias for each study using the Cochrane RoB 2 tool, available at riskofbias.org, and guidance in chapter 8 of the Cochrane Handbook (Higgins 2023; Sterne 2019). Any disagreement will be resolved by discussion or by involving a third reviewer.

We will examine the effect of assignment to preserve the benefits of randomization. We will examine the effect of assignment to the interventions at baseline, regardless of whether the interventions are received as intended (intention‐to‐treat effect). We will assess the risk of bias for all primary outcomes. We will use the following five domains to assess the risk of bias in the individual randomized trials (Higgins 2023; Sterne 2019).

• Bias arising from the randomization process

• Bias due to deviations from the intended interventions

• Bias due to missing outcome data

• Bias in measurement of the outcome

• Bias in the selection of reported results

For cluster‐randomized trials, we will add the assessment of bias arising from the identification or recruitment of participants into clusters (Eldridge 2021).

Possible answers to the signaling questions are yes, probably yes, probably no, no, and no information. After this step, we will judge each domain according to the algorithm result as low risk of bias, some concerns, or high risk of bias for each outcome. The overall risk of bias judgment will be the least favorable assessment across the domains. The three levels of judgment for an overall rating are:

• Low risk of bias: the trial is judged to be at low risk of bias for all domains for this result.

• Some concerns: the trial is judged to raise some concerns in at least one domain for this result, but is not at high risk of bias for any of the remaining domains.

• High risk of bias: the trial is either judged to be at high risk of bias in at least one domain for this result, or the study is judged to have some concerns about multiple domains in a way that substantially lowers confidence in the result.

We will summarize our findings in a risk of bias graph and risk of bias summary table.

Measures of treatment effect

Dichotomous data

We will present the treatment effects of outcomes as risk ratios (RRs) with 95% confidence intervals (CI).

Continuous data

We will present the treatment effects of outcomes as mean difference (MD) with 95% CI if the outcomes were measured in the same way between trials. We will calculate a standardized mean difference (SMD) with 95% CI if trials measured the same outcome but used different measurements. We will interpret SMD results as follows (Schünemann 2023):

• SMD less than 0.40 = small effect;

• SMD greater than 0.40 and less than 0.69 = medium effect;

• SMD 0.70 or greater = large effect.

Time‐to‐event data

We will calculate hazard ratios (HRs) with 95% CI for time‐to‐event data for the discontinuation rate, if the included studies reported these data.

Unit of analysis issues

Cluster‐randomized trials

For cluster‐randomized trials, the unit of analysis will be clusters instead of individual women. We will adjust the clustering effects by taking the relative variability within and between clusters, the intracluster (or intraclass) correlation coefficient (ICC), into account in the analysis. When the ICC is not available in the published reports, we will contact the trial authors to request it, or estimate the ICC using guidance from chapter 23 of the Cochrane Handbook (Higgins 2023).

Trials with multiple intervention groups

We will include trials with more than two intervention arms. If we include multiple arms in the same analysis, we will divide the number of events and total participants from the same control or comparator group for each comparison to avoid double‐counting.

Dealing with missing data

We will not impute missing outcome data for the primary outcomes. If data are missing, or only imputed data are reported, we will contact trial authors to request the required data. When possible, we will use the RevMan calculator to obtain the required information, e.g. missing standard deviations of continuous outcome, by using relevant statistics, such as CIs, standard errors, t‐ or z‐statistics.

Assessment of heterogeneity

We will assess the included studies for clinical diversity by considering differences in the participants, setting, interventions, and outcomes assessed. We will consider age, stage of CIN, and immunosuppressive condition.

To assess statistical heterogeneity, we will visually inspect the CIs of the treatment effects in the forest plots, and use a Chi² test (P ≤ 0.10). We will use the I² statistic to quantify statistical heterogeneity. We will interpret the I² statistic according to the Cochrane Handbook as follows (Deeks 2023).

• 0% to 40% might not be important

• 30% to 60% may represent moderate heterogeneity

• 50% to 90% may represent moderate heterogeneity

• 75% to 100% may represent considerable heterogeneity

If we identify statistical heterogeneity, we will explore possible sources through subgroup analyzes.

Assessment of reporting biases

We will assess reporting biases for primary outcomes using funnel plots if the meta‐analysis includes more than 10 studies. We will assess funnel plot asymmetry visually.

Data synthesis

We will conduct meta‐analyzes using a random‐effects model for all primary and secondary outcomes for which at least two trials are available. If there is clinical or considerable statistical heterogeneity (I² > 75%), we will synthesize the findings narratively, and present the results using forest plots without calculating an overall estimate. We will use Review Manager (RevMan) to carry out the statistical analyses (RevMan 2024). We will include all eligible studies, regardless of their risk of bias, and undertake a sensitivity analysis to assess the impact of studies at high risk of bias.

Subgroup analysis and investigation of heterogeneity

When we identify substantial heterogeneity among primary outcomes, we will undertake subgroup analyzes to explore sources of heterogeneity. We will evaluate relative effects and assess model fit for the following factors for the primary outcomes.

• Different stages of CIN (CIN 1/CIN 2/CIN 3)

• Immunosuppressive condition (Yes/No)

• Formulation of IMQ (vaginal suppository or topical cream application to the cervix)

• Doses of intervention

• Setting (inpatient or outpatient settings)

We will compare subgroups using the formal test for subgroup differences in RevMan.

We will review the relevant studies to assess the feasibility and availability of data to explore HPV clearance based on genotype. Based on our findings, we will determine whether to include it as a subgroup analysis in our study.

Sensitivity analysis

We will undertake sensitivity analyzes for all primary outcomes to assess the impact of the following factors on the results.

• Repeating the analysis excluding trials considered to be at overall high risk of bias

• Repeating the analysis excluding trials presented only as an abstract

• Repeating the analysis excluding cluster‐RCTs

Summary of findings and assessment of the certainty of the evidence

We will use the GRADE approach to evaluate the certainty of the evidence for all outcomes, as outlined in the Handbook for Grading the Quality of Evidence and the Strength of Recommendations using the GRADE Approach (Schünemann 2013). The GRADE approach uses five considerations (study limitations, consistency of effect, imprecision, indirectness, and publication bias) to assess the certainty of the body of evidence for specific outcomes. We will use GRADEpro GDT software to present the key findings by creating a summary of findings table for each comparison of interventions (GRADEpro GDT). Two review authors (HYO and LTS) will independently assess the certainty of the evidence, resolving disagreements by discussion with all review authors. We will present outcomes at the six‐month post‐complete treatment time point.

We will create a summary of findings table comparing the treatment effect of IMQ with placebo or no treatment, or with standard excisional treatment for the following seven outcomes.

• Regression of CIN

• Discontinuation of treatment due to side effects

• Progression of CIN to higher grades of CIN, or to cancer

• Grade 2+ pain from vaginal and vulvar inflammation

• Persistent dysplasia

• Persistent high‐risk HPV

• Any grade 2+ side effects

Appendices

Appendix 1. Search methods for bibliographic databases

CENTRAL

1. MeSH descriptor: [Squamous Intraepithelial Lesions] explode all trees
2.cervi* NEAR/3 ("Squamous Intraepithelial Lesions" OR "Squamous Intraepithelial Lesions”)
3.MeSH descriptor: [Uterine Cervical Dysplasia] explode all trees
4. MeSH descriptor: [Uterine Cervical Neoplasms] explode all trees
5. cervi* NEAR/3 (cancer* OR tumor* OR tumour* OR neoplas* OR malignan* OR carcinom* OR intraepithel* OR epithel* OR intra‐epithel* OR dysplasia OR pre‐cancer* OR precancer* OR benign OR "human papilloma virus" OR "human papilloma viruses" OR "human papillomavirus" OR "HPV" OR "papillomaviridae")
6. (cervi* OR pap OR vagina*) NEAR/3 (screen* OR test* OR diagno* OR identif* OR detect* OR smear* OR "cytology" OR "cytobrush" OR "visual inspection")
7. CIN*
8. #1 OR #2 OR #3 OR #4 OR #5 OR #6 OR #7 
9. MeSH descriptor: [Imiquimod] explode all trees
10. "Zyclara" OR "S 26308" OR "S‐26308" OR "Aldara" OR "1‐Isobutyl‐1H‐imidazo(4,5‐c)quinolin‐4‐amine" OR "R‐837" OR "R837" OR "R 837" OR "Imiquimod" OR ("immune" AND "cream")
11. #9 OR #10
12. #8 AND #11 
13. #12 (in Trials)

MEDLINE Ovid

1. exp Squamous Intraepithelial Lesions/
2. exp Uterine Cervical Dysplasia/
3. exp Uterine Cervical Neoplasms/
4. (cervi* ADJ3 ("Squamous Intraepithelial Lesion" or "Squamous Intraepithelial Lesions”)).af.
5. (cervi* ADJ3 (cancer* OR tumor* OR tumour* OR neoplas* OR malignan* OR carcinom* OR intraepithel* OR epithel* OR intra epithel* OR dysplasia OR pre cancer* OR precancer* OR benign OR "human papilloma virus" OR "human papilloma viruses" OR "human papillomavirus" OR "HPV" OR "papillomaviridae")).af.
6. ((cervi* OR pap OR vagina*) ADJ3 (screen* OR test* OR diagno* OR identif* OR detect* OR smear* OR "cytology" OR "cytobrush" OR "visual inspection")).af.
7. CIN*.af.
8. 1 or 2 or 3 or 4 or 5 or 6 or 7
9. exp Imiquimod/
10. ("Zyclara" or "S 26308" or "S‐26308" or "Aldara" or "1‐Isobutyl‐1H‐imidazo(4,5‐c)quinolin‐4‐amine" or "R‐837" or "R837" or "R 837" or "Imiquimod" or ("immune" ADJ3 "cream")).af.
11. 9 or 10
12. randomized controlled trial.pt. OR controlled clinical trial.pt. OR randomized.ab. placebo.ab. OR drug therapy.fs. OR randomly.ab. OR trial.ti. OR groups.ab.
13. 8 and 11 and 12

Note: Line #12 is the Cochrane Highly Sensitive Search Strategy for identifying randomized trials in MEDLINE: sensitivity‐maximizing version (2008 edit) (Lefebvre 2022); Available from: https://training.cochrane.org/handbook/current/chapter‐04

PubMed

Search Query
1. Squamous Intraepithelial Lesions[MeSH Terms]
2. cervi* AND (“Squamous IntraepithelialLesions” OR "Squamous Intraepithelial Lesions”)
3. Uterine Cervical Neoplasms[MeSH Terms]
4. Uterine Cervical Dysplasia[MeSH Terms]
5. cervi* AND (cancer* OR tumor* OR tumour* OR neoplas* OR malignan* OR carcinom* OR intraepithel* OR epithel* OR intra‐epithel* OR dysplasia OR pre‐cancer* OR precancer* OR benign OR "human papilloma virus" OR "human papilloma viruses" OR "human papillomavirus" OR "HPV" OR "papillomaviridae")
6. (cervi* OR pap OR vagina*) AND (screen* OR test* OR diagno* OR identif* OR detect* OR smear* OR "cytology" OR "cytobrush" OR "visual inspection")
7. CIN*
8. #1 OR #2 OR #3 OR #4 OR #5 OR #6 OR #7
9. Imiquimod[MeSH Terms]
10. "Zyclara" OR "S 26308" OR "S‐26308" OR "Aldara" OR "1‐Isobutyl‐1H‐imidazo(4,5‐c)quinolin‐4‐amine" OR "R‐837" OR "R837" OR "R 837" OR "Imiquimod" OR ("immune" AND "cream")
11. #9 OR #10
12. randomized controlled trial [pt]
13. controlled clinical trial [pt]
14. randomized [tiab]
15. placebo [tiab]
16. drug therapy [sh]
17. randomly [tiab]
18. trial [tiab]
19. groups [tiab]
20. #12 OR #13 OR #14 OR #15 OR #16 OR #17 OR #18 OR #19
21. #8 AND #11 AND #20

Note: Lines #12 to #19 are the Cochrane Highly Sensitive Search Strategy for identifying randomized trials in MEDLINE: sensitivity‐maximizing version (2008 edit) (Lefebvre 2022); Available from: https://training.cochrane.org/handbook/current/chapter‐04

Scopus

1. TITLE‐ABS‐KEY ( cervi* W/3 ( cancer* OR tumor* OR tumour* OR neoplas* OR malignan* OR carcinom* OR intraepithel* OR epithel* OR intra‐epithel* OR dysplasia OR pre‐cancer* OR precancer* OR benign OR "human papilloma virus" OR "human papilloma viruses" OR "human papillomavirus" OR "hpv" OR "papillomaviridae" ) )
2. TITLE‐ABS‐KEY ( "zyclara" OR "s 26308" OR "s‐26308" OR "aldara" OR "1‐isobutyl‐1h‐imidazo(4,5‐c)quinolin‐4‐amine" OR "r‐837" OR "r837" OR "r 837" OR "imiquimod" OR ( "immune" W/3 "cream" ) )
3. TITLE‐ABS‐KEY ( ( ( "randomized" OR "randomised" ) W/3 ( "controlled trial" OR "controlled trials" ) ) OR ( ( "randomized" OR "randomised" ) W/3 ( "control trial" OR "control trials" ) ) OR "rct" OR "rcts" OR "controlled clinical trial" OR "controlled clinical trials" OR "placebo" OR "randomly" OR allocate* )
4. #1 AND #2 AND #3

CINAHL

S1. TX (cervi* N3 ("Squamous Intraepithelial Lesions" or "Squamous Intraepithelial Lesions”))
S2. TX (cervi* N3 (cancer* OR tumor* OR tumour* OR neoplas* OR malignan* OR carcinom* OR intraepithel* OR epithel* OR intra‐epithel* OR dysplasia OR pre‐cancer* OR precancer* OR pre‐cancer* OR benign OR "human papilloma virus" OR "human papilloma viruses" OR "human papillomavirus" OR "HPV" OR "papillomaviridae"))
S3. TX ((cervi* OR pap OR vagina*) N3 (screen* OR test* OR diagno* OR identif* OR detect* OR smear* OR "cytology" OR "cytobrush" OR "visual inspection"))
S4. TX CIN*
S5. S1 OR S2 OR S3 OR S4
S6. TX "Zyclara" OR "S 26308" OR "S‐26308" OR "Aldara" OR "1‐Isobutyl‐1H‐imidazo(4,5‐c)quinolin‐4‐amine" OR "R‐837" OR "R837" OR "R 837" OR "Imiquimod" OR ("immune" N3 "cream")
S7. TX (("randomized" OR "randomised") N3 ("controlled trial" OR "controlled trials")) OR (("randomized" OR "randomised") N3 ("control trial" OR "control trials")) OR "RCT" OR "RCTs"
S8. TX "controlled clinical trial" OR "controlled clinical trials"
S9. TX "placebo" OR "randomly" OR allocate*
S10. S7 OR S8 OR S9
S11. S5 AND S6 AND S10

LILACS

(( cervi* AND ( cancer* OR tumor* OR tumour* OR neoplas* OR malignan* OR carcinom* OR intraepithel* OR epithel* OR intra‐epithel* OR dysplasia OR pre‐cancer* OR precancer* OR benign OR "human papilloma virus" OR "human papilloma viruses" OR "human papillomavirus" OR "hpv" OR "papillomaviridae" ) )) AND (( "zyclara" OR "s 26308" OR "s‐26308" OR "aldara" OR "1‐isobutyl‐1h‐imidazo(4,5‐c)quinolin‐4‐amine" OR "r‐837" OR "r837" OR "r 837" OR "imiquimod" OR ( "immune" AND "cream" ) )) AND (( ( ( "randomized" OR "randomised" ) AND ( "controlled trial" OR "controlled trials" ) ) OR ( ( "randomized" OR "randomised" ) AND ( "control trial" OR "control trials" ) ) OR "rct" OR "rcts" OR "controlled clinical trial" OR "controlled clinical trials" OR "placebo" OR "randomly" OR allocate* ))

Appendix 2. Search methods for ClinicalTrials.gov and WHO ICTRP

ClinicalTrials.gov

Imiquimod | Interventional Studies | Intraepithelial Lesion, Squamous

ICTRP

Imiquimod AND Squamous AND Intraepithelial AND Lesion*

Contributions of authors

Htun Yadanar Oo (HYO), Le Thandar Soe (LTS), Swe Mar Myint Lwin (SMML), Aye Lei Thu (ALT), Khin San Myint (KSM), Khaing Nwe Tin (KNT) and Kyaw Lwin Show (KLS) conceived the idea for this protocol.

HYO, LTS, SMML, ALT, KSM, KNT, KLS, Siwanon Rattanakanokchai (SR), Jen Sothornwit (JS), Apiwat Aue‐aungkul (AA), Porjai Pattanittum (PP), Chetta Ngamjarus (CN), Nampet Jampathong (NJ), and Pisake Lumbiganon (PL) designed the protocol.

HYO, LTS, SMML, ALT, KSM, KNT, KLS, SR, JS, AA, PP, CN, NJ, and PL provided methodological perspective.

HYO, LTS, KNT, SR, PP, and CN provided statistical advice and input.

HYO, LTS, SMML, ALT, and KSM drafted the protocol.

All authors reviewed the protocol, provided critical feedback, and approved the final version of this protocol.

Sources of support

Internal sources

• Department of Obstetrics and Gynaecology, Faculty of Medicine, Khon Kaen Universiy, Thailand

  The review authors are employed by the institution.

• Department of Epidemiology and Biostatistics, Faculty of Public Health, Khon Kaen University, Thailand

  The review authors are employed by the institution.

• Cochrane Thailand, Thailand

  Technical support

• Maternal and Reproductive Health Division, Department of Public Health, Ministry of Health, Myanmar

  The review authors are employed by the institutions indicated by their respective affiliations, except where otherwise stated.

• University of Public Health, Yangon, Myanmar

  The review authors are employed by the institutions indicated by their respective affiliations, except where otherwise stated.

• Department of Medical Research, Myanmar, Myanmar

  The review authors are employed by the institutions indicated by their respective affiliations, except where otherwise stated.

External sources

• none, Other

  none received

Declarations of interest

All authors declare that they have no conflict of interest.

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