The cervix is the most sensitive area to the oncogenic action of human papillomavirus (HPV). Its transformation zone allows oncogenic agents (such as high-risk human papillomavirus [HR-HPV]) to have easier access to the base site of the reserve cells.¹

Several particularly vulnerable areas —including the glandular eversion area, also called ectropion or ectopic region— can be identified by their lack of a protective barrier.^1,2

Some studies show an essential correlation between the immunologic state and virus persistence wherein vaginal microbiome (VM), which is a determining factor in the modulation of the immunity system in the female genital area. Other factors also play their part, including ethnic origin, hormonal state, and other internal and external elements that influence the onset of HPV infections.^1,2

Nowadays, the hypothesis suggests that, for a host to develop medical conditions associated with HPV, an infection affecting base epithelial cells must be present, and such infection must specifically affect the stem cells found in the epithelium, either in the mucus or the skin.¹

It is known that the likelihood of persistence is related to the type of HPV. More than 150 subtypes have been identified, which can be classified into two groups based on their oncogenic potential: viruses with a low risk of progressing to cancerous lesions (LR-HPV) and viruses with a high risk of progressing to cervical cancer (HR-HPV). In the latter group, the main serotypes are 16, 18, and 31, which are most directly associated with the persistence of the virus. Persistent infection with an oncogenic or HR-HPV type is the main risk factor for detecting a cervical intraepithelial neoplasia (CIN) that can range from CIN 1 to CIN 3 and, ultimately, progress to cancer.^3,4

In the cervix, the transformation zone is preserved by a type of specialized cells known as reserve cells, and the HPV generally resides in an epithelial stem cell post-infection, which allows us to understand viral latency and reactivation; however, during latency, the virus may go unnoticed for the most common diagnosis technique.¹

The risk factors need to be assessed during the follow-up, where, unfortunately, a vast number of patients are lost, a situation that does not justify overtreatment but demands improving health strategies. In fact, up to 70% of women aged 30 years or younger affected by CIN 2 experience spontaneous regression, and colposcopists should increasingly be reluctant to subject young women to potentially harmful cervical treatments that are associated with recognized obstetric morbidity.⁵

In this article, a review is made of the adjuvants proposed for reepithelialization of the uterine cervix, improvement in the microbiota, and, as a secondary local immune effect, which may impact HPV infections.

Research has been conducted, and products have been developed to restore and promote cervical epithelium while stabilizing the microbiome and helping improve the local and immunity conditions to reduce the viral persistence locally since constant HPV infections increase the risk of developing pre-cancerous cervical lesions.

Although HPV itself cannot be treated, the high-grade lesions caused by an infection can and should be treated according to management guidelines.

We know that nearly all sexually active adults will develop an HPV infection at some point in their lives. Most of these infections are transient, with 90% clearing spontaneously or becoming inactive within 12 to 24 months after exposure to the virus. However, in some women, the infection persists, and there is a risk of developing precancerous lesions.³

HPV infection is not the only determining factor for clinical outcomes. Others—such as host immunity, the histological structure of the exocervix, and the state of the vaginal microbiota (VM)— can also be crucial for the progression or regression of the infection, even though their role is not yet well defined. Acting on some of these factors may offer an opportunity to enhance viral clearance and reduce risk.¹

For this reason, various therapies have been tested to promote cervical epithelization, regression of LSIL (low-grade squamous intraepithelial lesion), and/or prevent its progression and overtreatment.

We have reviewed imiquimod, interferon, vitamin D, probiotics, prebiotics, and symbiotics, as well as a variety of therapeutic combinations proposed as adjuvants to epithelial restoration, VM, and local immunomodulation.⁶

There are some epithelium restructuring agents to improve cervical conditions, including a vaginal gel based on Coriolus versicolor —a fungal agent— in combination with other ingredients, such as hyaluronic acid, α- and β-glucans, oligosaccharides, Centella asiatica, Azadirachta indica, and Aloe vera.⁷

Such combination has shown cell regeneration and improved VM moisturization and balance; thereby achieving several effects such as restructuring of the cervical epithelium, as well as improved VM and vaginal health. This combination has reduced local HPV infections. However, long-term follow-up trials are still needed, with larger study groups and comparative groups using estrogen-based gels and local microbicides.⁷

Coriolus versicolor is a fungus whose biomass acts as a non-specific immunomodulator. The immunostimulatory effect of Coriolus is due to the beta-glucans and proteoglycans it contains (polysaccharide [PSK] and polysaccharopeptide [PSP]), which stimulate the effect of natural killer (NK) cells and increase the number of T lymphocytes.⁸

The immunological activity of these molecules is achievable when the peptides have a molecular size above 10kD (mainly 100 kD).^8,9 In particular, PSP can activate an effective tumoricidal activity by expressing TNF-a inhibits tumor cell proliferation.⁸ The immunostimulatory effect is enhanced by the content of specific secondary molecules such as IL-α and IL-β.⁸

In recent years, it has been observed that Coriolus versicolor favors the repair of low-grade cervical lesions related to HPV, which is expected to reduce the persistence of HPV infection, as shown in the PALOMA study.⁷

Likewise, a trial published in 2023 reported the outcomes of using Coriolus versicolor gel to repair HPV-dependant LSIL in HPV-positive women ≥25 years old with abnormal cytology - ASC-US (atypical squamous cells of undetermined significance) and LSIL with matching colposcopies.⁵

The gel was applied using a vaginal cannula for 21 days continuously in the first month. Then, one cannula was used every other day for five months, so this treatment was applied for a total of six months, with some instances where a second treatment cycle was prescribed for six additional months.⁵

Follow-up was performed through cytology and colposcopy, clearance of HPV, tolerability, and level of satisfaction, which was graded at 7.9 and 7.5 out of 10 by months 6 and 12, respectively. This technique was shown to have higher regression rates for LSIL among women treated with the Coriolus versicolor vaginal gel when compared to women who were only followed up for spontaneous regression.⁵

It should be noted that risk factors and the state of the immune system play an essential role in the persistence or clearance of the infection. The combination of Coriolus versicolor extract and neem stimulates the immune system to respond locally.^7,10

Another adjuvant described in the restoration of epithelium and VM is carboxymethyl-beta-glucan in patients infected by HPV, LSIL or CIN 1. The mechanism of action of carboxymethyl-beta-glucan contributes to the maintenance and restoration of the vaginal microbiota through a specific prebiotic effect: providing an energy source for Lactobacillus sp., thereby promoting its growth and dominance in the vaginal microenvironment.

A trial with this adjuvant¹¹ was performed in 181 women who were carriers of HPV or CIN 1. It was a case-control study that evaluated Pap smear, HPV screening, vaginal health under Amsel criteria, both colposcopy and biopsy, with the trial population divided into two groups during the 6-month follow-up.

Group A received the carboxymethyl-beta-glucan gel (one daily application for 20 days per month for three months), and Group B was the control group. Group A achieved:

• significant improvement in the ectopy pattern, with a larger number of cases presenting metaplasia;
• significant improvement in pH levels;
• negative Swift test and stoppage of leukorrhea, with 37.1% of the cases obtaining a negative Pap smear and 39.9%, respectively;
• negative HPV versus 15.2% and 16.5%, in the treatment and control branches, respectively;
• negative colposcopies, with a reduced number of cervical lesions.

All of this data concludes that a positive impact was achieved to help reduce the risk of persistence and progression of the low-grade intraepithelial lesions.¹¹

Another combination of extracts used to promote remission of LSIL is the mixture of Echinacea angustifolia, Echinacea purpurea, and hyaluronic acid.⁹ The roots and extracts of E. angustifolia and E. purpurea have shown anti-inflammatory activity and immunomodulation effects by inhibiting the expression of hallmark pro-inflammatory cytokines such as interleukin 6 and 8, favoring viral clearance such as HPV infection.¹¹

Echinacea is a botanical genus belonging to the family Asteraceae, consisting of nine species. The three most sought-after species are Echinacea angustifolia, Echinacea pallida, and Echinacea purpurea.¹² The main substances found in Echinacea are phenolic compounds such as caffeic acid, alkamides (echinacein), and high-molecular-weight polysaccharides (such as arabinogalactan and fucogalactoxyloglucans). It can help support immune function.¹²

Hyaluronic acid is one of the body's main defense mechanisms. It is found between the cells of tissues, acting as a protector of cellular integrity. When hyaluronidase levels are high, this enzyme breaks down the structure of hyaluronic acid, which quickly loses viscosity and stops acting as a protective layer for the cells, potentially compromising immune function. Additionally, it has been shown that using Echinacea prevents the enzymes’s dissolution of hyaluronic acid.^11,13

A blinded, parallel-group, randomized trial¹¹ with a control group was conducted in a tertiary-care colposcopy unit. Three research branches (A, B, and C) were created:

• In group A, hyaluronic acid-based soft gel vaginal capsules (Tiagin soft gel, IDI Farmaceutici, Italy) were used once daily for ten days throughout three months, with an adjuvant therapy consisting of a supplement of E. angustifolia (100 mg), E. purpurea (100 mg), polyphenols (4 mg), vitamin C (40 mg), zinc (5 mg), copper (0.5 mg) (Normoimmuno, ADL Farmaceutici SRL, Italy) administered twice a day for three months.
• In group B, hyaluronic acid-based gel vaginal capsules (Tiagin soft gel, IDI Farmaceutici, Italy) were administered once per day for ten days throughout three months.
• Group C received E. angustifolia (100 mg), E. purpurea (100 mg), polyfpolyphenols (4 mg), vitamin C (40 mg), zinc (5 mg), copper (0.5 mg) (Normoimmuno, ADL Farmaceutici SRL, Italy) twice per day for three months.

The primary outcomes found were regression of CIN 1 in each treatment cohort, at 3, 6, and 12 months after diagnosis, with a more significant improvement in group A at 6 and 12 months. Secondary outcomes included changes in epithelialization, cytology, or Pap smear as well as in the colposcopy, hence, in overall vaginal health indexes (VHIs).¹¹

This study concludes that oral supplementation with Echinacea extracts, zinc, vitamin C, and polyphenols, together with vaginal use of hyaluronic acid-based soft gel, may increase the regression of LSIL/CIN 1 by significantly increasing the clearance of HPV lesions. However, the population used in this study is not significant numerically; thus, further trials are required to reach more consistent outcomes.¹¹

Other studies have shown outcomes mediated by the effect of HR-HPV clearance, as well as from favoring the healing of visible cytological findings (ASC-US, LSIL, ASC-H [atypical high-grade squamous intraepithelial lesion], HSIL [high-grade squamous intraepithelial lesion]), by administering treatment with a vaginal gel containing silicate dioxide, citric acid, and selenite, after six months of use.^6,14

The mechanism of action of silicon dioxide, citric acid, and selenite involves the adhesion of pathogens, inhibition/neutralization of pathogens (antioxidant effect), and promotion of epithelial regeneration.¹⁴

The highly dispersed and micronized particles of silicon dioxide adsorb pathogens (bacteria, viruses, fungi, cellular debris, irritating particles) from the surface of the cervix. Silicon dioxide inhibits these pathogens and, therefore, prevents their proliferation and spread over the surface of the cervix and in the vagina.^14,15

Once the pathogens are inhibited, they are neutralized by preventing the release of their free radicals, taking advantage of a potentially pro-oxidant action of the adsorbed pathogens through the antioxidant properties of the patented combination of sodium selenite and citric acid. It also stops the inflammatory process of the cervical epithelium and vaginal mucosa from potentially irritating pathogens. It promotes spontaneous remission by adjusting the environment of the cervical and vaginal secretions during the psychologically stressful period of "watchful waiting".¹⁴

Those trials found an improvement in the treatment group regarding cytological findings with Pap smear in 80.9% of the subjects (ASC-US, LSIL, ASC-H, HSIL), as well as HPV clearance in 53% of cases after three months of gel administration.¹⁵

In patients who were positive for p16/Ki67, the reduction was from 75% to 5.3%. Compared with the non-treatment group, the improvement percentage was minimal, from a baseline of 91.5% to 75.2% at six months.^6,15 No serious adverse effects were reported across any adjuvant therapy studies.¹⁴

Regarding cervical cancer, oxidative stress or antioxidant deficiency is frequently reported to be associated with increased production of reactive oxygen species, which ultimately cause DNA damage in cervical cells. Under these conditions, cervical cells become vulnerable to HPV infection and the potential development of cancer.¹⁶

Therefore, in recent decades, the intake of antioxidants from natural foods has received much attention due to their preventive role in this type of cancer. For example:

• Frequent consumption of vitamin A has been shown to reduce the risk of HPV infection by 10%.¹⁷
• Vitamin D, due to its anti-inflammatory activity and improvement of insulin resistance, has been shown to be useful in improving clinical and metabolic symptoms in patients with HPV infection.¹⁵
• Vitamin C can reduce HPV infection and cervical cancer through the induction of apoptosis. Similarly, some polyphenols have been shown to inhibit HPV proliferation by inducing apoptosis, causing cell cycle arrest, and inhibiting DNA synthesis.¹⁸

It is necessary to thoroughly analyzed the trials using adjuvants, increase the number of subjects in the study groups, perform a longer-term follow-up on each case, and continue to carry out randomized comparative studies. The design of the relevant protocols should be monitored too.

All the previous considerations pose a real problem for cervical screening programs, especially if the HPV test is used first. Many healthy and asymptomatic women will have a positive result and suffer anxiety because of it, even though they are at relatively low risk of developing cervical cancer or needing treatment for CIN.

In some cases, particularly if subsequent tests are normal or low-grade, conservative management is advised, even after referral for colposcopy. Since this approach can take several months, it may be challenging to manage for both the patient and physicians. Therefore, in these circumstances, a safe and non-surgical treatment is needed to facilitate virus elimination.

When faced with a positive diagnosis of HPV and abnormal cytology, many women go through negative emotional responses, including long-term psychological distress. One study showed that a positive diagnosis of HPV was associated with annoyance, shame, shock, and fear of cancer.¹⁹

Adverse responses were categorized into eight cognitive constructs:

1. low self-control
2. confusion
3. worries related to cancer
4. worries related to interpersonal relations
5. concerns of a sexual nature
6. uncertainty
7. stigma
8. lack of confidence

The responses can also be classified according to six behavioral constructs¹⁹:

1. relationship problems
2. social impact
3. unwillingness to disclose the diagnosis
4. idiosyncratic prevention
5. indirect clinical interaction
6. changes in sexual practices

Clinicians, epidemiologists, and health-related decision-makers should consider the impact of mental health in this area to mitigate adverse psychological consequences since this is crucial to maximize the repetition of screening and follow-up trials or to provide the necessary treatment.

The emotional impact suffered by a woman when she receives the news of an HPV infection may adversely affect her self-esteem, confidence, and even her interest in romantic relationships. Frequently, the increased psychological stress may lead to a higher potential of acquiring viral infections, especially infections mediated by IL-6.²⁰

The influence that emotional stress has in reducing cell apoptosis may result in immunosuppression. For instance, in cases of acute stress, the immune system becomes blocked more often. At the same time, lymphocyte count drops and the ratio of Th1/Th2 cells becomes altered, thereby suppressing cytokines Th1 and causing the reaction to evolve towards Th2 cells, increasing the risk of developing high viral replication.²⁰

Psychological stress is a crucial factor involved in the disease caused by HPV and has been seen to increase more when inconsistencies are found during clinical tests or when social support is absent, with personal coping mechanisms and the sensation of social stigma also playing a role in the infection process.^19,21

Symmons' trial²² found that, when facing a positive result in a VPH test, regardless of the patient's education level (categorized into high and low), women had unanswered questions about the outcomes and the protocol followed during the screening.

Moreover, the service received by the patients did not fulfill screening expectations, especially in terms of the content of the letters and the delivery of the outcomes, which patients deemed unclear, and this lack of clarity was accentuated among women in the low-education category.²⁰

Based on these results, the need to continue working in specific areas becomes evident, beginning by training healthcare professionals to gain better communication skills when talking to patients²³ and setting educational guidelines led by decision-makers, particularly concerning the goals stipulated by the World Health Organization (WHO) to eliminate cervical cancer.²⁴

The healthcare team in charge of a patient should be made aware of the significant emotional impact that the information provided and the treatment of women carrying HPV infections will have in their lives and their close social network, aiming at the creation of a professional-patient partnership based on mutual respect and objective information.

Patients should not feel afraid of interacting with healthcare professionals; instead, they should see them as a source of education, values and ethical principles, as a therapist who will guide and accompany them. As such, appropriate training should be provided to help these professionals determine the most adequate interventions throughout the different stages of the disease.