|Year : 2015 | Volume
| Issue : 1 | Page : 1-13
Immunotherapy of viral warts: myth and reality
Mohamed El-Khalawany MD, ICDP - UEMS 1, Dalia Shaaban2, Soha Aboeldahab3
1 Department of Dermatology, Al-Azhar University, Cairo, Egypt
2 Department of Dermatology, Tanta University, Tanta, Egypt
3 Department of Dermatology, Sohag University, Sohag, Egypt
|Date of Submission||07-Jun-2015|
|Date of Acceptance||23-Jun-2015|
|Date of Web Publication||7-Aug-2015|
Department of Dermatology, Al-Azhar University, PO Box 32515, Al-Darasah, Cairo
Source of Support: None, Conflict of Interest: None
Immunotherapy has become one of the most important therapeutic tools for the treatment of warts. At present, immunotherapy for warts is usually limited to recalcitrant lesions that are not responding to conventional therapy. Although there are a lot of immunotherapeutic regimens, a minority seem to be really effective. Moreover, there is a lack of evidence-based data on their effectiveness. Common immunotherapeutic modalities used for the treatment of warts include contact sensitizers, imiquimod, intralesional interferon, and oral drugs such as levamisole, cimetidine, and zinc sulfate. Intralesional antigens such as MMR (measles, mumps, and rubella) vaccine, skin test antigens (mumps, Candida, and Trichophyton), BCG (bacillus Calmette-Guιrin) vaccine, and Candida antigen were reported as successful treatment modalities in various forms of warts. Moreover, intradermal injection of some vaccines such as purified protein derivatives was also reported as a successful regimen for the treatment of genital warts. Among the available options for treatment of warts, none is uniformly effective or viricidal. Moreover, in most cases their safety and efficacy has not been assessed in double-blind, controlled clinical trials, and thus the reproducibility of many of the listed treatments is difficult to evaluate and a possible placebo effect cannot be ruled out. In this report, the various forms of immunotherapy for warts are discussed and each regimen is evaluated in order to assess the efficacy of each form of treatment.
Keywords: Imiquimod, immunotherapy, interferon, vaccine, wart
|How to cite this article:|
El-Khalawany M, Shaaban D, Aboeldahab S. Immunotherapy of viral warts: myth and reality. Egypt J Dermatol Venerol 2015;35:1-13
|How to cite this URL:|
El-Khalawany M, Shaaban D, Aboeldahab S. Immunotherapy of viral warts: myth and reality. Egypt J Dermatol Venerol [serial online] 2015 [cited 2020 Jan 22];35:1-13. Available from: http://www.ejdv.eg.net/text.asp?2015/35/1/1/162451
| Introduction|| |
Warts are common viral infections on the skin and are prevalent worldwide. Warts are caused by the human papilloma virus (HPV), which has more than 100 strains; some of them are known to be premalignant. Although warts can appear at any age, they are more common in children and adolescents. The prognosis of warts cannot be predicted. In some patients they may spontaneously disappear, whereas others show persistence and progression with spreading to other body sites, leading to physical and emotional distress to the patients .
The treatment of warts depends on two main therapeutic options: the first is the conventional destructive and aggressive method, which includes treatment with chemical cautery, cryotherapy, electrocauterization, surgical excision, and laser ablation, and the second is immunotherapy, which is based on the activation of the immune system to deal with the virus and suppress its activity. Such therapy may be applied either topically or through intralesional injection or through systemic administration .
Selection of the most appropriate means of treatment is usually difficult because of the availability of various therapeutic lines and the immune status of the patients. There are many factors that should be considered before the treatment of the patients, such as age, sex, previous medical history, and the clinical characteristics of the warts. Patients with multiple warts or warts resistant to treatment are usually prone to have defective cell-mediated immune response .
In this report we will discuss and demonstrate the various immunotherapeutic agents used for the treatment of different types of warts in order to demonstrate the strengths and weaknesses of each treatment method. The immunotherapeutic agents were classified according to the mode of administration into three main categories - topical, intralesional, and systemic - as shown in [Table 1].
|Table 1: The most common immunotherapeutic methods for the treatment of warts|
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| Topical immunotherapy|| |
There are numerous topical immunomodulators that have been used for treating warts. Although topical mmunotherapy agents have their own clinical advantages over other destructive treatment modalities, an updated assessment of these agents is required.
Diphencyprone (DCP) was first synthesized in 1959. In view of its degradation by ultraviolet (UV) radiation, it is prepared as dilutions in acetone and marketed in brown UV-opaque bottles. It was first used for treatment of alopecia areata, and then used as a successful treatment method for resistant plantar warts .
DCP is a more potent contact sensitizer than dinitrochlorobenzene (DNCB) for the same concentration. However, DCP may be a safer option, as unlike DNCB it is not mutagenic in the Ames assay and not detectable in serum or urine after topical application. In the treatment of viral warts, sensitization to 2% DPC is carried out on a 1 cm 2 area of skin on the upper arm. After the warts are pared down, DPC is applied at a concentration of 0.1% (2% to the soles of the feet) .
There was wide variation in the intensity of the patients' reactions to different concentrations of DCP, which bore no relation to the eventual likelihood of clearing (e.g. some may clear after only one or two applications of 0.1% DCP with minimal or no symptoms, whereas others may fail to improve as the concentration of DCP is sequentially increased). Best results are achieved when diligent paring and keratolytic treatment are used in conjunction. It does, however, require repeated hospital visits and therefore is not a practical option for all individuals .
The potential side effect of DCP treatment is mainly blistering at the sensitization site. Distant or more widespread eczematous eruptions may also occur, either due to passive transfer of DCP or due to autoeczematization. Such reactions generally respond to a potent topical steroid. Other less frequent adverse effects include regional lymphadenopathy, contact urticaria, erythema multiforme-like reactions, and vitiligo .
Use in pregnancy is probably best avoided as it remains an unlicensed treatment modality. Patients should be followed up regularly and should not be allowed to self-treat. DCP should not be used to treat genital or facial warts, as the risk of passive transfer is high in the genital area, and bullous or eczematous side effects would be less acceptable on the face. DCP immunotherapy is particularly suitable for plantar, palmar, periungual, and digital warts .
The mechanism of action of DCP in the treatment of warts remains to be elucidated. Theories include alterations in cytokine levels, nonspecific inflammation causing wart regression, and binding of DPC to wart protein, inducing a specific immune reaction. It was suggested that topical DCP therapy may lead to the induction of long-term immunity to HPV, thus theoretically reducing the incidence of wart recurrence .
Diphenylcyclopropenone (DPCP) was first synthesized in 1959 as a potent contact allergen in humans and animals. Similar to DCP, UV radiation and heat cause degradation to DCP. Dilutions of DPCP are marketed in brown UV-opaque bottles to be stored at room temperature. DPCP is nonmutagenic at concentrations of 50 and 100 mg/ml and it does not appear to have significant systemic absorption after topical application .
DPCP is applied to warts (pared down where possible) at an initial concentration of 0.1% on fingers, periungual regions, palms, toes, and heel and 2% on the sole (the concentration of DPCP is adjusted to induce local erythema, pruritus, and/or vesiculation). The warts are then carefully covered with adhesive dressings for 48 h to avoid passive transfer of DPCP. When any reaction to DPCP has settled, patients are instructed to begin paring the warts and can use keratolytic agents .
Repeated treatments are carried out at intervals of 1-4 weeks. Treatment is stopped once there is no clinical sign of a residual wart. If there is failure to improve after six or more treatments despite eliciting an eczematous response, or if the maximum available concentration of DPCP (6%) has been used on several occasions without inducing an eczematous response, treatment is deemed to have failed and is discontinued .
The potential adverse effects are not inconsiderable. Unsightly pigmentation at the sensitization site occurs in many patients and therefore sensitization is best carried out on the inner upper arm. Local blistering at the treatment site and/or sensitization site is common during treatment. A number of patients may need to discontinue treatment or switch to another sensitizer because of repeated development of generalized eczema, or urticaria, occurring after each application .
The mechanism of action of DPCP for viral warts has been less thoroughly investigated. Theories include alterations in cytokine levels, nonspecific inflammation causing wart regression, and binding of DPCP to wart protein, inducing a specific immune reaction. The latter is supported by the fact that warts at untreated sites may sometimes improve or clear during treatment. DPCP induces a reversal of the CD4 : CD8 ratio such that CD8 cells predominate in a heavy epidermal and dermal inflammatory infiltrate. Immunotherapy with DPCP should not be used routinely as an initial therapy for warts, but is a possible alternative in selected patients with recalcitrant multiple warts .
DNCB was first found to be a potent contact allergen in 1912 and has a well-established role in the induction of experimental delayed-type hypersensitivity. In the last 30 years it has been widely used to induce hair growth in alopecia areata. It was also used in the treatment of variable skin diseases, including malignant and premalignant skin lesions .
DNCB is rapidly absorbed after topical application and its excretion is primarily renal. DNCB was reported to be mutagenic in the Ames assay and genotoxic by sister chromatid exchange in human skin fibroblasts. However, the related chemical dinitrofluorobenzene was shown not to be mutagenic, and the authors suggest that dinitrofluorobenzene may instead act as a tumor promoter. DNCB increases the incidence of complement-binding wart virus antibodies from 15% before treatment to 48% afterward, suggesting a role for humoral factors in the involution of viral warts .
For treatment of warts, the method of application of DNCB is similar to that used for DCP. In a placebo-controlled nonrandomized study by Muarrek et al. , the number of patients was insufficient to show any significant advantage with the active treatment. The duration of warts was not specified (warts of recent onset are more likely to resolve spontaneously) .
Squaric acid dibutylester
Squaric acid dibutylester (SADBE) is a universal topical sensitizer first synthesized and shown to be a potent contact allergen in 1979, and used for the treatment of AA. It is not mutagenic in the Ames assay but is considerably more expensive than the other contact sensitizers. Topical immunotherapy with SADBE represents an effective treatment in the management of multiple plantar and common warts. In the genital area the use of SADBE has some limitations, as it may be associated with a significant degree of irritation that produces considerable discomfort to the patient .
The immunomodulating action of SADBE in warts is still not certain. According to most authors, SADBE may work by inducing a type IV hypersensitivity response in an HPV-infected tissue, leading to wart destruction. Whether the contact sensitizer as a hapten attaches to wart protein inducing a specific immune reaction to wart antigen is unknown. Other theories postulate that the inflammation caused by SADBE determines the resolution of the warts through a nonspecific inflammatory reaction .
Method of application is similar for SADBE and DNCB. Lesions of warts that are hyperkeratotic are pared. SADBE sensitization is conducted using 2% SADBE on all nonfacial warts, followed by a minimum of 4 h occlusion, and site washing with soap and water when the bandages are removed. The initial session is followed every 2-3 weeks by 0.2% SADBE application to the residual lesions .
Clinical resolution and length of treatment in patients treated by applications to remote areas was comparable to when SADBE was applied directly to lesions. This substantiates the efficacy and safety of topical SADBE in the treatment of multiple recurrent/recalcitrant anogenital warts, and also indicates a possible systemic effect of contact immunotherapy, suggesting that the mechanism of action of SADBE could be more than a nonspecific inflammatory reaction or a local cell-mediated process triggered by a nonwart antigenic immune stimulus .
There are no significant adverse reactions that could be attributed to topical immunotherapy with SADBE. Therefore, topical immunotherapy using SADBE could be considered an efficient and relatively painless treatment method for refractory common warts, and an effective alternative to traditional treatment modalities that are destructive to tissues, including cryotherapy. In combination treatment with TCA 50%, SADBE significantly improves the consistency and speed of clearance of warts .
5-Fluorouracil (5-FU) is a fluorinated pyrimidine antimetabolite that functions as an antineoplastic agent by blocking DNA synthesis. Once administered, the drug is concentrated especially on neoplastic tissue. This antimetabolite drug is most commonly used to treat a variety of skin neoplasms and precancerous lesions, such as actinic keratosis. It has been used by clinicians for treating urethral condylomata since the early 1990s .
5-FU has been used as a cream or injected solution in concentrations between 1 and 5% for treatment of genital warts and has been tried with variable results as an adjuvant to laser therapy in severe papillomavirus-associated vulval disease. The antineoplastic and antimetabolite properties of 5-FU that inhibit DNA and RNA synthesis are believed to be the mechanism that stops wart proliferation . Evidence from the studies we reviewed showed that 5-FU had better results for cure than placebo or no treatment, than treatment with meta-cresol sulfonic acid, and treatment with podophyllin 2, 4, or 25%. No statistical difference was found when 5-FU was compared with CO 2 laser treatment, and results were poor when 5-FU was compared with 5-FU+IFN-2α (high dose) or 5-FU+CO 2 laser IFN-2α (high dose) .
It was found that 5-FU plus salicylic acid treatment gave a significantly better cure rate than treatment with 5-FU alone . However, in another report, two different concentrations of 5-FU were compared with standard topical SA/LA for mosaic plantar warts (warts on the feet). The cure rates for all three treatments were close to 50% and were not significantly different. A more recent trial of 40 participants compared 5-FU with tape occlusion and found that there was a high cure rate at 6 months in the 5-FU group at 85%, compared with 10% in the tape occlusion group .
Compared with placebo after cryotherapy, 5-FU showed a cure rate of 30 versus 43% in the placebo group. This was not statistically significant and implies no additional benefit of 5-FU . However, Artese compared 5-FU with cautery and found that cure rates were better in the 5-FU group (85 vs. 66%) .
Side effects of 5-FU include onycholysis (nail detachment) (especially when it was used for warts near the nails), blistering, and moderate/severe pain. Unexpectedly, the previous reports on cryotherapy combined with either podophyllin cream or 5-FU showed no increment in the cure rate, compared with cryotherapy alone. No specific explanations about the disappointing outcomes of topical agents were provided in both previous studies, and the authors merely suggested that the insufficient sample size might be a reason. 5-FU may be a successful adjuvant to cryotherapy in reducing the number of agonizing cryotherapy sessions .
Few studies have examined the effect of topical bacillus Calmette-Guérin (BCG) on condylomata acuminata and genital warts. External application of BCG led to the immediate clearance of genital warts within 6 weeks in six of 10 patients at a follow-up of 9.2 months. Topical BCG may also lead to greater cellular reactivity and the consecutive elimination of HPV in humans, but the mode of action must clearly be elucidated further. It was concluded that immunotherapy with BCG is an accepted treatment and appears to reduce recurrence in patients with condylomata acuminata . Local immunotherapy of genital warts with viable BCG was found to decrease the annual recurrence in patients with refractory recurrent condylomata acuminata. Topical BCG was also used as a paste in combination with salicylic acid in subtherapeutic dose, and this resulted in significantly better outcome .
Viable BCG as immunotherapy in warts was also used as an intradermal injection in 81 patients, with complete response in 39.7% . Viable BCG was tried by Salem et al. to treat common and plane warts in children, particularly with small and recent-onset verruca. The mechanism of action of this type of immunotherapy depends on the activation of CD4 lymphocytes and an increase in cytokines such as interleukin (IL)-1, IL-2, and tumor necrosis factor α (TNF-α). IL-1 and TNF-α have been shown to have antiviral effects on HPV through the downregulation of its gene transcription .
Imiquimod is a member of a novel class of synthetic immune response modifiers that has antiviral and antitumor activity. It was FDA approved for treatment of external genital and perianal warts in 1997. It is more suitable for wart area 0.5-1.0 cm in diameter and for moist nonkeratinized warts. It has more recently been approved for the treatment of nonhypertrophic actinic keratoses and superficial basal cell carcinomas. Others have reported its effectiveness in the treatment of recalcitrant plantar, periungual, and subungual warts .
Imiquimod is a toll-like receptor (TLR) agonist. It binds to TLR7, which activates dendritic cells, macrophages, and monocytes that stimulate a complicated cascade of reactions in the cytoplasm, leading to activation of nuclear factor κB (NF-κB), and eventually the production of cytokines and chemokines outside of the cell. The most important of these cytokines and chemokines include interferon-α (IFN-α), IFN-g, ILs 1, 6, 8, 10, and 12, and TNF-α .
For treatment of anogenital warts, it is applied three times a week at bedtime, followed by 4 days of rest or every other day (three times a week). It should be washed with soap and water 6-10 h after application. Maximum duration of treatment is 16 weeks. It is more effective in women than in men. Local reactions like erythema often occur (in 33-80%). Flu-like symptoms, fatigue, diarrhea, fever, skin blistering, erosion, excoriation, flaking, edema, paresthesia, pruritus, burning, tenderness, stinging, crusting, and superficial ulcer may occur .
Combined procedure using imiquimod cream after ablation of anogenital warts (by electrocautery, liquid nitrogen, or laser therapy) might be favorable compared with any of the monotherapies. Moreover, treatment with imiquimod followed by excision of the remaining warts resulted in a lower recurrence rate compared with surgery alone. Imiquimod was successfully used in the treatment of nongenital warts (common warts, periungual, plantar warts, and recalcitrant verruca plana) with low recurrence rates. It can be used once daily for 4 weeks. The results are better when imiquimod cream is used, under occlusion or after treatments that weaken the stratum corneum and facilitate its penetration .
Activated vitamin D
Maxacalcitol, an active vitamin D3 analog, has been used for the treatment of psoriasis vulgaris and palmoplantar keratosis. It is well known that vitamin D3 analogs have some biological actions in epidermal cells, such as the regulation of cell proliferation and differentiation and the modulation of cytokine production. Recent evidence demonstrates an effect on cell death, tumor invasion, and angiogenesis, rendering it a candidate agent for cancer regulation . Egawa and Ono  reported the effectiveness of a topical vitamin D3 derivative for condylomata acuminata on the corona and glans in a 74-year-old man with simple application twice a day for 4 months.
Treatment with topical vitamin D3 derivatives has been reported to be effective for HPV-infected verruca. The effect of vitamin D3 derivatives on verruca was speculated to be derived from its potential to regulate epidermal cell proliferation and differentiation and to modulate cytokine production. An important observation was reported, which suggested that TLR activation of human macrophages upregulated the expression of vitamin D receptor and vitamin D-1-hydroxylase genes, leading to induction of the antimicrobial peptide. This suggests an association of TLRs and vitamin D-mediated innate immunity. Thus, imiquimod and vitamin D3 derivatives, to some extent, may share the same antiviral efficacy .
A half-day occlusive dressing therapy (ODT) of a vitamin D3 derivative was used successfully to treat recalcitrant warts. A new focus of interest is the tissue levels of vitamin D3 derivatives, particularly in relation to local cellular growth regulation. Inaba et al.  suggested the possibility that the combination of maxacalcitol ointment and salicylic acid sticking plaster can be an effective alternative to traditional destructive modalities, including cryotherapy, as a treatment for warts. The effect is increased by ODT, whereas a simple application has a minimal effect. Therefore, salicylic acid sticking plasters were used both to soften the hyperkeratotic warts and to increase the tissue levels of maxacalcitol in the warts .
Rind et al.  reported an infant case with an anogenital wart on the anus, which was successfully treated with calcipotriene ointment, a vitamin D3 derivative. It took 4 months for the treatment of the anogenital wart. Topical vitamin D3 derivatives were successfully used to treat recalcitrant warts in three immunocompromised patients with systemic lupus erythematosus, Crohn's disease, and human T-lymphotropic virus type 1. It was also successfully used to treat resistant warts in a renal transplant recipient . No adverse effects or abnormal serum test results, including elevated serum level of Ca 2+ , were observed in most of studies. Therefore, it was suggested that the ODT of vitamin D3 derivatives offers a safe, effective alternative form of treatment for recalcitrant warts .
Topical sinecatechins ointment 15% is the first FDA-approved botanical drug for treating anogenital warts. Sinecatechins are a standardized extract of green tea leaves from Camellia sinensis containing polyphenols, particularly catechins (>85%). It contains eight different catechins and other green tea components. The main catechin in sinecatechins ointment is epigallocatechin gallate, which has the highest biological activity. The mechanism of action of sinecatechins ointment 15% in the resolution of external genital warts (EGWs) is not completely known .
The direct action of catechins is scavenging the reactive oxygen-free radicals. Catechins may exert indirect antioxidant activity through inhibitory effects on transcription factors (e.g. NF-κB, activator protein-1) and also inhibits the activity of enzymes that increase oxidative stress (e.g. lipoxygenases, cyclooxygenases, and inducible nitric oxide). Epigallocatechin gallate also induces expression of endogenous antioxidant systems. Catechins also check the proliferation of cells by inhibiting kinases and promoting apoptosis. In addition, catechins induce an immune-stimulating effect by increasing the activation of macrophages, lymphocytes, Langerhans cells, and induction of cytokines (IL-1b, TNF-α, IFN-g), thereby eliciting cell-mediated immunity against HPV. However, it is unknown to what extent this activity affects the clearance of EGWs in humans .
Sinecatechins ointment should be applied three times daily to all EGWs. Treatment should continue until all EGWs are cleared, but for no longer than 16 weeks (4 months), and involves placing approximately 0.5 cm of ointment on the fingertip and covering the entire wart(s), leaving a thin layer. The treatment areas do not need to be washed before the next application of ointment. Local skin reactions are the most commonly reported adverse reactions. These can include redness, burning, pain, itching, and swelling on and around the treatment site(s). Local reactions typically peak early after beginning of therapy and decrease over the course of treatment .
Sinecatechins ointment is approved for use in women and men aged 18 years and older. The safety and efficacy of use in pediatric patients has not been established, and it is not known whether sinecatechins ointment 15% is excreted into breast milk . In addition, sinecatechins ointment 15% is not recommended for individuals who are infected with HIV, are immunocompromised, or have genital herpes, as safety and efficacy in these populations has not been established. Not only do sinecatechins have a wide spectrum of action but they are also associated with a low recurrence rate (6.5%). Hence, it could be a better option for treatment of anogenital warts in future. Sinecatechins yields a lower cost of treatment compared with imiquimod .
| Intralesional therapy|| |
Intralesional immunotherapy utilizes the ability of the immune system to mount a delayed-type hypersensitivity response to various antigens and also the wart tissue. This therapy has been found to be associated with the production of Th1 cytokines that activate cytotoxic and natural killer cells to eradicate HPV infection. This clears not only the local warts but also distant warts, unlike traditional wart therapies .
Many authors have used different immunotherapeutic agents for intralesional injection. These include Candida antigen, mumps antigen, trichophytin skin test antigen, BCG vaccine, measles, mumps, and rubella (MMR) vaccine, Mycobacterium w (Mw) vaccine, and IFN-α and IFN-g injection. The use of these agents is based on the fact that there is a high prevalence of immunity to these antigens in the general population. It has been found that older individuals (>40 years) are less likely to respond to this modality as compared with younger individuals on account of the less robust immunological responses with increasing age .
There are two different approaches used by various authors. In the first approach, the individual is injected with an intradermal antigen on the volar aspect of the forearm and the delayed hypersensitivity response is assessed as erythema and induration 48-72 h later. Responders (with erythema and induration of 5 mm in diameter) can use the therapy. Then the wart, usually the largest one, is injected with the Candida or mumps antigen in a volume determined by the size of the test reaction .
Some authors have directly injected the antigen into the largest wart without performing a preliminary intradermal testing. Warts are injected intralesionally using an insulin syringe. The syringe is held parallel with the skin surface with the bevel facing upward. This therapy is repeated every 3 weeks until complete clearance of the warts or for a maximum of three treatments without response .
Bleomycin, an antibiotic derived from Streptomyces verticillus, has an antitumor, antibacterial, and antiviral activity that may be related to its ability to bind with DNA, causing bleomycin strand scission and elimination of pyrimidine and purine bases. The bleomycin hydrolase enzyme, which is known to inactivate bleomycin, is normally found in all body tissues but is present in very small amounts in skin. Thus, after injecting bleomycin intralesionally, a significant amount of the active drug is available at the site .
Intralesional bleomycin has been used for the treatment of warts since the 1970s. Numerous reports have been published on the use of intralesional bleomycin for the treatment of recalcitrant warts with cure rates ranging from 14 to 99%. Intralesional bleomycin was found to be very effective in treating warts particularly in periungual and palmoplantar areas .
Bleomycin for injection is available in vials containing 15 mg powder. It is diluted first with 5 ml distilled water to prepare the stock solution, which can be stored for 60 days at 4-8°C. Two parts of 2% lignocaine and one part of the bleomycin stock solution are taken in a tuberculin syringe, so that the final concentration becomes 1 mg/ml. Each wart and the adjacent skin is cleansed with isopropyl alcohol. Superficial paring is done to remove the callus surrounding the wart; bleeding points are not reached and the fresh solution is injected strictly intralesionally until blanching of the lesion occurs .
The amount injected depends on the size of warts: warts up to 5 mm, those measuring 10 mm, and warts measuring more than 10 mm received 0.2, 0.5, and 1.0 ml, respectively. The total volume injected at one treatment sitting is limited to 2 ml, and the injection into a single wart is limited to 1 ml. After 2 weeks of bleomycin injection, a black, ecchymosed eschar developed, which is pared, and residual warts if present are injected a second time .
Bleomycin application was mostly done by drug injection into the lesion through syringes, but other methods such as the use of a dermojet, prick method, and dermatographic method can also be applied. The use of a pulsed dye laser before bleomycin injection has been reported by some studies. Patients are usually followed up weekly during the first month, and then fortnightly up to 3 months. Routine hemogram, liver function tests, renal function tests, and radiograph of the chest should be done before and after 3 months of treatment .
The use of intralesional bleomycin has not been approved by the US FDA. For a very low dose (1 mg/ml), no systemic side effects have been observed. However, local signs such as necrosis, pain, scaring, pigment change, Raynaud's phenomenon, and nail dystrophy may occur in some cases. Bleomycin should be avoided in children, pregnant women, and patients with peripheral vascular disease .
Interferon (α and g)
IFN-α is a low-molecular-weight glycoprotein produced by cell types that inhibit viral replication and tumor growth. IFN-α-2b is an intralesional IFN approved by the FDA for the treatment of genital warts, requiring twice-weekly injection for 3 weeks for optimal results . IFN-g, produced by activated T cells, is more potent in its antiproliferative activity than IFN-α and IFN-b. IFN-g appears to be the more favorable IFN to evoke cellular immune responses to resistant warts .
In addition to its ability to enhance the cytotoxic activity of NK cells and macrophages, IFN-g has been shown to induce synthesis of TNF receptors in some cells, thereby increasing their sensitivity to cytostatic or cytolytic activity of TNF. It is therefore possible that exogenously administered IFN-g may increase TNF receptors on virus-infected keratinocytes, thereby rendering them more sensitive to TNF released from activated T cells and macrophages .
Intralesional injection of IFN eradicates 40-60% of warts. Side effects include an elevation of temperature, chills, fatigue, diarrhea, headache, nausea, and vomiting (flu-like symptoms). The severity of these symptoms appears to be dose related .
Candida albicans antigen
Candida antigen has been documented as an effective mode of treatment for warts since 1979. Intralesional candida injections were compared with cryotherapy in the treatment of warts. Complete resolution of warts treated with candida injections was observed in 74% patients. Intralesional candida immunotherapy has also been tried in children with recalcitrant warts with a response rate of 47% for the treated wart and 34% for all body warts .
The effectiveness of candida immunotherapy has been described even for genital warts. Adverse effects that have been reported with candida immunotherapy include febrile reactions, myalgia, pain, erythema, and edema at the injection site, and painful purple digit syndrome. However, this form of therapy is inexpensive .
Measles, mumps, and rubella vaccine
Intralesional immunotherapy using MMR vaccine has the potential advantages of clearance of both treated and untreated distant warts without scarring, a presumed low rate of recurrence, and a high safety profile. Although the mechanism of effectiveness of intralesional injection of MMR vaccine and antigens has not yet been known, it seems that nonspecific inflammatory response to the antigens is the major mechanism of immunotherapy .
MMR vaccine was evaluated as a treatment option for cutaneous warts. Nofal and Nofal  reported complete clearance of warts in 81.4% of patients in the MMR group compared with 27.5% of patients in the placebo group. Injection-site pain and flu-like symptoms were the only side effects noted . In their study, Gamil et al.  noticed a statistically significant inverse relationship between the duration of warts and the therapeutic response to MMR vaccine. This finding may be explained by the higher viral load expected to increase with the longer duration of the warts, and by the increase of some soluble factors that block the local expression of cellular immunity against HPV in warts of long duration .
Lipid garlic extract
Components of garlic have been shown to have antiviral activity and to inhibit cellular proliferation of virally infected cells. Raw garlic cloves can be rubbed onto the wart nightly, followed by occlusion. The application of chloroform extracts of garlic was reported to result in the complete resolution of cutaneous warts with no recurrence after 3-4 months ,,.
Killed Mw vaccine was first developed in India from a nonpathogenic, rapidly growing, atypical Mycobacterium belonging to Runyon class IV. It was approved as an immunotherapeutic adjunct to multidrug therapy of multibacillary leprosy in India. It is strongly antigenic and generates robust cytokine (IL-2, IFN-g) and T-cell responses. Its nomenclature was later changed to Mycobacterium indicus pranii .
Mw vaccine is available in a multidose vial of 0.5 ml containing 500 million heat-killed bacilli in a buffer solution. A sensitizing dose of the vaccine is administered intradermally at a dose of 0.1 ml over a deltoid area of each shoulder using an insulin syringe. After 2 weeks, the injected sites are examined for an immune response manifesting as persistent erythema or a nodule. In sensitized patients, 0.1 ml or less of the vaccine is injected intraepidermally or into the superficial dermis in two to four warts. Larger warts are preferred for injection. The injections are repeated at 2-weekly intervals until complete resolution or a total of 10 injections are administered, whichever is earlier .
Mw vaccine appears to be an efficacious and safe treatment modality for extensive, long-standing/resistant cutaneous warts. Therapy was relatively well tolerated. Intralesional Mw vaccine on patients with external anogenital warts (88.9%) achieved complete clinical clearance after a mean treatment duration of 5.9 weeks. Adverse effects include fever, pain, sterile pustule at the injection site, and paraesthesias in limbs distal to the injected warts .
Bacillus Calmette-Guérin vaccine
BCG was introduced as a prophylactic agent against tuberculosis as well as in the treatment of alopecia areata and recurrent oral aphthosis. The mechanism of action could be explained on the basis of stimulating macrophages, T and B lymphocytes, and NK cell function that might help in resolution of viral warts .
Intradermal BCG vaccine has also been evaluated for warts and has been found to produce complete clinical remission in 39.7% of patients. Salem et al.  evaluated topical BCG vaccine for warts in children and noted a complete response in 65% patients with common warts and 45% with plane warts.
Sharquie et al.  observed that there were no local or systemic adverse effects with the use of BCG vaccine in the treatment of viral warts; however, they noticed some signs accompanying the regression of viral warts, such as itching in 100% of patients, increased size of the lesions in 80%, and some lesions becoming tender in 30%. Treatments with tuberculin and BCG vaccine are likely to be cheap. This could be a very useful modality of treatment of warts in poor and developing countries .
| Systemic therapy|| |
Activation of the immune system by systemic treatment could be achieved through different administrations. In this part we show the treatments that activate the immune system away from the site of the lesions. Oral therapies include cimetidine, zinc sulfate, and levamisole. Other routes of administration include acupuncture, intradermal injection, and auto-inoculation in remote sites .
Cimetidine is a well-known H2 receptor antagonist that was approved in 1977 for treating duodenal ulcer disease and hypersecretory states of gastric acid. In dermatological diseases, cimetidine was used as an adjuvant therapy in some allergic conditions that are characterized by increased histamine release, such as urticaria, mastocytosis, and different eosinophilic dermatoses. Moreover, cimetidine was proven to have an immunomodulator effect by blocking H2 receptors from suppression T cells, as well as to increase the proliferation of lymphocytes, inhibit the function of suppressing T cells, and enhance the reactivity of skin tests .
Because of its immunomodulating effect, cimetidine was successfully used to stimulate the immune system of patients with T-cell immunodeficiency, mostly skin cancer reported as malignant melanoma, and cutaneous viral infections such as molluscum contagiosum, epidermodysplasia verruciformis, herpes zoster, herpes simplex, and warts . The most common indications for cimetidine in the treatment of warts include childhood warts, and recalcitrant warts in adults. Other indications that were also reported included periungual warts, genital warts, and multiple warts .
Although the cure rate of warts with cimetidine therapy is controversial because of variable outcomes in different studies, it seems that the use of cimetidine is generally of limited efficacy and its success is attributed to factors other than its immunomodulating effect. Rogres et al.  reported a cure rate of 26% compared with 24% in the placebo group, whereas Yilmaz et al.  obtained a cure rate of 32% with cimetidine compared with 30.7% with placebo. These results show that cimetidine is no more effective than placebo in the treatment of patients with common warts. However, successful results were obtained with Chern and Cheng , who reported a complete clearance of recalcitrant periungual warts in a 12-year-old girl after a high dose of cimetidine.
In comparison with other systemic therapies, cimetidine was also found to be less effective than zinc sulfate in the treatment of multiple and recalcitrant warts either in children or in adults. Moreover, Cohler and Schaffer  also reported a successful treatment of massive anogenital warts in a 2-year-old boy after treatment with cimetidine combined with imiquimod. These reports suggested that the therapeutic response to cimetidine may be related to the high dose or combination with other immunomodulators .
Levamisole is a well-known antihelminthic drug that was originally used to treat worm infestations in both humans and animals. Although levamisole was pulled from the market in USA and Canada, it is still used in many other countries because of immunomodulator actions. In dermatologic diseases, levamisole has been successfully used in the treatment of parasitic, viral, and bacterial infections such as leprosy. It has also been used in combination with other drugs for treating a number of dermatologic disorders - e.g. in combination with prednisolone for treating lichen planus, erythema multiforme, and aphthous ulcers of the mouth .
Adverse affects of levamisole are usually mild and infrequent. The common side effects include rash, nausea, abdominal cramps, taste alteration, alopecia, arthralgia, and a flu-like syndrome. Although agranulocytosis is considered the most serious side effect of levamisole, it rarely occurs and it is reversible when treatment is discontinued .
The use of levamisole in the treatment of warts was controversial and showed variable results. Schou and Helin  did not recommend the use of levamisole in the treatment of warts after failure to achieve a significant improvement in 49 patients with common warts and in 50 patients with venereal warts. Treating with a dose of 150 mg daily for 3 days every second week failed to show enhanced wart regression compared with the control group . Similar conclusion was also proposed by Saúl et al.  after failure to achieve a significant improvement, compared with the placebo group, even with the use of a different dose of levamisole (2.5 mg/kg, 2 days a week, for 8 weeks).
In a more recent study by Amer et al.  levamisole was administrated at a dose of 5 mg/kg body weight on 3 consecutive days every 2 weeks for a period of 5 months. This regimen showed a significant cure rate in plane and common warts but not in plantar warts . More efficacy was obtained when levamisole was combined with other immunomodulating drugs such as cimetidine. Parsad et al. , reported the effectiveness of a combination between levamisole and cimetidine in both adults and children.
The postulated mechanism of the immunomodulator action of levamisole was proposed by Moncada and Rodrigues  who postulated that patients with multiple warts have defective cell-mediated mechanisms as far as the E-rosette count is lower than that in controls. Levamisole increases E-rosette formation in vitro, which is useful in the treatment of these patients .
Thus, the role of levamisole in the treatment of warts is still limited and not strongly proved. The administrative dose, the therapeutic regimen, the type of wart, and the immune status of the patients are the most related factors that could influence the response to levamisole.
Zinc is considered one of the most essential elements and micronutrients for humans. It is used as a common therapeutic product in both topical and oral forms. In dermatology, zinc was used in the treatment of a wide variety of skin diseases, including infections (such as leishmaniasis, leprosy, and dermatophytosis), inflammatory conditions (such as acne vulgaris, psoriasis, and eczema), and hair and nail disorders (such as alopecia, seborrheic dermatitis, and erosive pustular dermatosis of the scalp) .
The use of zinc in the treatment of warts was proved in many studies either in its topical form or as a systemic therapy. Sharquie et al.  studied the efficacy of topical zinc sulfate in the treatment of viral warts. In their pilot clinical trial, 10 patients with plane warts were treated with 10% zinc sulfate solution three times daily for a period of 4 weeks. They observed complete clearance in 80% of patients. The authors also reported the results of a double-blind clinical trial in the same study comprising 50 patients with common warts and 40 patients with plane warts treated with topical 10 and 5% zinc sulfate solution applied three times daily for 4 weeks while distilled water was used as a control. Complete clearance of plane warts was noticed in 85.7 and 42.8% of cases from topical 10 and 5% zinc sulfate solution, respectively, whereas complete clearance was also seen in 11 and 5% of patients with common warts, respectively, which was statistically insignificant .
In another study, Khattar et al.  in a randomized double-blind controlled trial of 44 patients observed that topical 20% zinc oxide was more effective than keratolytic ointments containing salicylic acid (15%) and lactic acid (15%) for the treatment of warts. A complete cure was observed after 3 months in 50% of patients with common warts in the zinc oxide group in comparison with the keratolytic group (42%) .
With regard to oral therapy, Al-Gurairi et al.  in a study included 80 patients with resistant viral warts (common, plantar, and plane); 40 patients were treated with oral zinc sulfate at a dose of 10 mg kg/daily up to 600 mg/day and the other 40 patients were given a placebo oral treatment in the form of glucose. They reported that complete clearance of warts was observed in 20 patients (86.9%) after 2 months of treatment. Three patients (13.3%) failed to respond to the treatment after 2 months of therapy .
In other studies, Mun et al.  reported a clearance rate of 50% with the same dose of oral zinc sulfate after 2 months. Moreover, oral zinc (10 mg/kg/day) has also been reported to clear recalcitrant warts in a patient with epidermodysplasia verruciformis in 12 weeks . With the same dose of 10 mg/kg/day of zinc sulfate (maximum 600 mg/day) for 3 months, Stefani et al.  reported complete clearance in five patients (one patient did not show modifications in lesions and one patient did not complete treatment because of nausea and vomiting). They suggested that a dose of zinc sulfate at 10 mg/kg/day seems to be more effective than cimetidine for the treatment of children and adults with multiple and difficult-to-handle warts. Measurement of serum zinc in patients with persistent warts was recommended by some investigators, as zinc deficiency is associated with persistent, progressive, or recurrent viral warts. Although the previous studies suggested that topical or oral zinc can be a useful therapeutic modality for warts especially in children, it is still believed that zinc may be effective in only selected populations with zinc deficiency .
Acupuncture is probably the most popular alternative therapy practiced in the USA, Europe, and many Asian countries. In addition to the analgesic effect of acupuncture, an increasing number of studies have demonstrated that acupuncture treatment can control autonomic nerve system functions such as blood pressure regulation, Sphincter Oddi More Details relaxation, and immune modulation. Although a limited number of control studies have assessed the efficacy of acupuncture, increasing clinical evidence supports that acupuncture treatment is effective for various immunological diseases including allergic disorders, infections, autoimmune diseases, and immunodeficiency syndromes .
The use of acupuncture in the treatment of warts was proposed by Ning et al.  in a single-blind study that included 60 patients with flat warts. Patients were classified into two groups; those in the first group were treated with weekly auricular acupuncture, whereas the other group was treated with 0.1% tretinoin ointment. The acupuncture group showed complete clearance of lesions in 53.33% of patients compared with 3.33% in the tretinoin group, with a statistically significant difference between the two groups (P<0.01). Although these results suggested that auricular acupuncture may be a viable alternative for the treatment of flat warts, the authors recommended larger randomized studies to fully evaluate auricular acupuncture in relation to conventional treatments .
In another study, Yuan et al.  assessed the clinical therapeutic effect of acupoint injection of BCG polysaccharide nuclear acid (BCG-PSN) in the treatment of condyloma acuminatum. The study included 200 patients who were divided into 4 groups; the treatment group (group A) was treated with acupoint injection of BCG-PSN; control group I (group B) was treated with intramuscular injection of BCG-PSN; control group II (group C) was treated with intramuscular injection of IFN; and the blank control group (group D) was administered no treatment. They found that the cure rate of 94.3% in group A was significantly higher than the 78.0% cure rate in group B, 80.4% in group C, and 78.2% in group D, with significant differences (P<0.05). The percentage of CD4+ increased, that of CD8+ decreased, the CD4+/CD8+ ratio increased, and NK cell activity increased. They suggested that the cellular immunoregulatory action is one of the mechanisms of this therapy that help in treatment and prevent relapse of condyloma acuminatum .
Although there is almost universal agreement that the quality of clinical trials of acupuncture is poor, the therapeutic effect of such modality seems to be promising in the treatment of warts. However, the low number of studies and the lack of data from other world areas and different populations limit the proper evaluation of this method.
Purified protein derivative
Purified protein derivative (PPD) is an extract of Mycobacterium tuberculosis and is used for testing exposure to tuberculin protein, either from a previous vaccination or from the environment. It contains live, attenuated Mycobacterium bovis . Eassa et al.  reported the role of intradermal injection of PPD in the treatment of anogenital warts in pregnant women. The study included 40 pregnant women treated with weekly intradermal injections of PPD given in the forearms. The overall improvement was 85% and it was related to the extent of tuberculin reactivity. Nineteen (47.5%) patients demonstrated complete clearance, 15 (37.5%) had partial response, and three (7.5%) had minimal response. Only three (7.5%) patients did not respond to treatment. Reported side effects in this study were minimal and insignificant .
Abd-Elazeim et al.  used PPD as an intralesional method in a recent controlled study that included 40 patients with recalcitrant multiple common warts. Complete response was achieved in 75% after 5.8 ± 0.7 sessions. They also found a statistically significant increase in IL-12. However, more studies are required for full assessment of this method.
As no single therapy has been found to be efficacious and cosmetically acceptable in the majority of patients with warts, Srivastava and Bajaj  suggested the use of autowart injection as a treatment option for extensive and recalcitrant warts. The technique was based on removing 3-4 mm of wart tissue by radiocautery and placing on sterile gauge. The tissue was then crushed in pastel and mortar with distilled water. A fine suspension was prepared and taken into a sterile disposable syringe and injected into the gluteal region. They found a response rate up to 89% (66.03% with complete resolution and 22.64% with partial improvement), whereas 11.32% had no improvement .
With another modified technique, Nischal et al.  used the pared tissue of the warts for autoimplantation in subcutaenous tissue and they also reported a response greater than 75%. These results suggested that autowart injection is an effective and noninvasive therapeutic option in extensive and recalcitrant warts. However, the rare available studies in the literature increase the difficulty to evaluate this therapeutic option.
| Conclusion|| |
Immunotherapy has become one of the most essential therapeutic modalities in warts. The need for such treatment is restricted not only to recalcitrant or multiple lesions but also to most of the treated cases. Although not all immunotherapeutic agents are fully studied, the variable methods of application, easy applicability, suitability for children, and its role in decreasing relapse have encouraged their use either alone or in combination with other conventional methods. From our review, we believe that immunotherapy as a single therapy is not very effective for the treatment of warts and that combination with conventional methods is most appropriate.
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Conflicts of interest
There are no conflicts of interest.
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