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Photodynamic Therapy With Topical Metatetrahydroxychlorin (Fosgel) Is Ineffective for the Treatment of Anal Intraepithelial Neoplasia, Grade III

2009; Lippincott Williams & Wilkins; Volume: 52; Issue: 1 Linguagem: Inglês

10.1097/qai.0b013e3181b05f93

1944-7884

Eric M. van der Snoek, Arien Amelink, Marc van der Valk, Jan C. den Hollander, Jan G. den Hollander, Frank P. Kroon, R. Vriesendorp, Harald Neumann, Dominic J. Robinson,

Nonmelanoma Skin Cancer Studies

To the Editor: The incidence of anal cancer and preneoplastic anal lesions as anal intraepithelial neoplasia (AIN) in western European countries and the United States has increased in recent decades.1 Recent studies suggest that the incidence of anal cancer is even as high as 70-100 cases per 100,000 per year among HIV-positive men who have sex with men. Prevalences between 21% and 59% of AIN diagnosed in HIV-positive men who have sex with men have been reported.2 Persistent high-risk human papillomavirus (HPV) infection is an important risk factor for AIN and anal cancer because of its possibility to trigger carcinogenic development. Recent studies showed that highly active antiretroviral therapy does not improve the clinical course of an HPV infection nor decreases HPV persistence or the incidence of AIN.3 Long lasting suppressed cellular immunity in HIV-HPV-coinfected individuals may increase the prevalence of anogenital malignancies. There is currently no accepted standard management of AIN.4 For severe dysplasia or AIN III, radiotherapy has been used. However, long-term side effects as anorectal dysfunction after radiotherapy (local scarification, urgency of defecation, and faecal incontinence) were observed in up to 40% of all patients.5 The use of photodynamic therapy (PDT) is now widely accepted in the dermato-oncological treatment of (pre)malignant skin lesions. Topically applied porphyrin precursors lead to the accumulation of protoporphyrin IX (PpIX) in atypical cells.6 After a suitable interval, illumination with light of the appropriate wavelength leads to the generation of reactive oxygen species that causes cell necrosis, apoptosis, and vascular damage. PDT gives no unacceptable scars, fibrosis of the skin, or hyposensibilisation.7 Considering the well-known side effects of PDT with porphyrin precursors, itching and often very significant local pain during and after illumination, we decided to explore the use of other photosensitisers. Fosgel is a liposomal formulation of meta-tetrahydroxyphenylchlorin (mTHPC; Foscan) that can be applied locally. Foscan is known to be one of the most potent photosensitizers, as relatively small drug doses and light fluences are required to achieve therapeutic response. Fosgel-PDT has been shown to be less painful than PDT using porphyrin precursors.8 Outpatients were requited from 4 hospitals between February 2007 and March 2009 and invited to anal screening and underwent high-resolution anoscopy with application of acetowhite. Diagnostic biopsies were taken from perianal or intra-anal clinically suspicious lesions to confirm clinical AIN. All patients with histological proven AIN III were included in this study. Participants gave written informed consent, and the local hospital ethics committee approved the study. Histopathological classification was according to the revised Bethesda classification. AIN III was regarded as severe dysplasia. PDT was performed 8 hours after the topical application of a thermosetting formulation of meta-tetrahydroxyphenylchlorin, (Fosgel) containing 0.75 mg/mL mTHPC (Biolitec AG, Jena, Germany). Fosgel was applied to a thickness of approximately 3 mm with a 10 mm margin. Before illumination, excess gel was thoroughly wiped from the lesion and margin. Lesions were illuminated using 652 nm light from a diode laser (Biolitec AG). The lesion and margin were illuminated at a fluence rate of 50 mWcm−2 to a fluence of 20 Jcm−2. For intra-anal lesions, the illumination was performed using a cylindrical inflatable applicator. An in situ measured fluence of 20 J cm−2 was delivered to the surface of the anal cavity at 50 mWcm−2 using a 5 cm inflatable balloon and a 5 cm diffuser. All patients were treated twice with a 7-day interval between treatments. The second treatment session involved the reapplication of Fosgel. In 3 lesions, we performed a noninvasive optical determination of the intensity of mTHPC fluorescence during therapy. Our optical technique, fluorescence differential path-length spectroscopy, described elsewhere, has been validated in optical phantoms and applied in vivo in various tissues.9 The measurement involves placing a probe of approximately 3 mm diameter in contact with the tissue. Reflectance and fluorescence spectra are recorded via 2 optical fibres under white light and 405 nm laser excitation. Light is collected from the fibres and fed into 2 spectrometers that, after an appropriate calibration, yield quantitative fluorescence spectra. The interrogation depth of fluorescence differential path-length spectroscopy used here is approximately 400 μm. The acquisition of reflectance spectra allows the hemodynamic parameters (blood saturation/blood volume) of tissue to be determined. A 3-mm punch biopsy was recovered from the site of the optical measurement and frozen for confocal fluorescence microscopy. Care was taken to make sure that these tissue volumes were coincident and biopsy sites were swabbed until bleeding had stopped before PDT. Biopsies were sectioned at 3 depths in sections of 50 μm thick. A 2-μm optical section was imaged at the centre of the lesion section under 405 nm excitation. Spectral imaging was performed on each optical slice. Background and reference data were recorded for background and flat field corrections. Basis spectra for tissue autofluorescence and mTHPC fluorescence were fit to the data to determine the distribution of mTHPC fluorescence. Clinical evaluation took place 2 weeks after the second treatment session. In case of any doubt with regard to clinical improvement, a biopsy was taken for additional histological evaluation. Nine patients were included in this study. Three HIV-positive men had perianal bowenoid AIN III and the other 4 HIV-positive males were diagnosed with intra-anal AIN III. Of these 7 males, 3 used highly active antiretroviral therapy and 2 had undetectable HIV RNA. In addition, 2 HIV-negative women were included. One patient was diagnosed with perianal Bowen's disease and the other patient had erythroplakic perianal AIN III. Four patients had been treated previously by means of incomplete excision, topical 5-FU or imiquimod creme. In all cases, therapy was well tolerated. During illumination, all patients reported sensation for all lesions described as a slight itch/discomfort or slight pain. This sensation was most severe within 30 seconds of the start of the illumination and diminished or ended immediately after the illumination. Quantitative fluorescence and reflectance spectroscopy showed high mTHPC signals before therapy. At the end of the illumination period, lesions were shiny and there was a clear reddening of lesions. The Visual Analogue Scale for pain reported during the first treatment had a median of 0.2 (range 0-6.1). The second illumination was not significantly more painful; median of 0.5 (range 0-3.8). Two weeks after the second treatment, there was no clinical (9 of 9) or histological (3 of 3) improvement. Figure 1 shows typical reflectance and fluorescence spectra, before and after therapy. Spectra show intense mTHPC fluorescence before illumination (λmax = 654 nm). There were small intralesion variations in fluorescence (SD: 18%) but large differences between lesions (SD: 76%). During illumination, there is rapid photobleaching of mTHPC such that after the delivery of 20 Jcm−2, 19% of the initial mTHPC intensity remains. The high concentration of mTHPC in the lesions is also illustrated by the characteristic combination mTHPC absorption and fluorescence in reflectance spectra. Interestingly, spectroscopy data showed tissue vasculature responds to therapy. Before PDT, there is less blood within the interrogation volume. Immediately after PDT, the blood volume increased by a factor that was an average of 9 times that before PDT. The percentage of saturated blood in the lesions also increased. Figure 1 also shows a confocal fluorescence image of a biopsy sliced parallel to its depth axis. mTHPC fluorescence and tissue autofluorescence are displayed as red and green, respectively. The fluorescence from mTHPC is predominately located in the very superficial layers of lesions and is confined to the disordered stratum corneum and the very superficial layers of AIN III. An analysis of the mTHPC distribution in 3 sections from each biopsy showed mTHPC present only in the uppermost layers of tissue. At depths beyond 60 μm, the fluorescence intensity is not significantly different from tissue autofluorescence.FIGURE 1: Reflectance and fluorescence spectra before and after illumination and a confocal fluorescence section AINIII before PDT. Red shows mTHPC fluorescence.Our study shows that PDT with the use of topical mTHPC in a liposome formulation (Fosgel) was ineffective for the treatment of AIN III. Although we found no evidence for a clinical or a histologically positive response after therapy, we were able to confirm that mTHPC was present in high concentrations before illumination. We were also able to show that mTHPC is rapidly photobleached. It is clear that, locally, large amounts of singlet oxygen are being generated. It is interesting that the absorption maximum of mTHPC, used in our spectral analysis, is almost identical to that of mTHPC dissolved in ethanol (Q band: λmax = 650.5 nm) and shorter than that normally acquired in vivo (652.9 nm). This is an indication that mTHPC remains associated with the solvent used. Our results concerning the penetration of mTHPC after topical Fosgel application are not in agreement with preclinical data reported in chemically induced/ultraviolet-promoted squamous cell carcinomas in mouse models.8 It is possible that there are significant differences in the structure of skin of these models and that of AIN III that might explain the absence of mTHPC to a greater depth in AIN III. It is interesting to note that despite the superficial localization of mTHPC, the generation of singlet oxygen in this volume of tissue induced sensation during therapy. This and the fact that reflectance data show an increase in blood content after illumination show that there is a clear response after PDT. However, we conclude that the limited penetration of mTHPC in AIN III restricts the clinical utility of topical Fosgel-PDT for the treatment of AIN III. Ongoing studies exploring intravenous photosensitisers or topical ALA are warranted. Systemic PDT using metatetrahydroxychlorin (Foscan) has proven to be very effective in the treatment of extensive vulval intraepithelial neoplasia.10 Eric M. van der Snoek, MD, PhD* Arien Amelink, PhD† Marchina E. van der Ende, MD, PhD‡ Jan C. den Hollander, MD§ Jan G. den Hollander, MD, PhD‖ Frank P. Kroon, MD, PhD¶ Robert Vriesendorp, MD, PhD# H. A. Martino Neumann, MD, PhD* Dominic J. Robinson, PhD*† *Department of Dermatology †Department of Radiation Oncology, Center for Optical Diagnostics and Therapy ‡Department of Internal Medicine §Department of Pathology Erasmus Medical Center Rotterdam, The Netherlands ‖Department of Internal Medicine, Maasstad Hospital, Rotterdam, The Netherlands ¶Department of Infectious Diseases, Leiden University Medical Center, The Netherlands #Department of Internal Medicine, Medical Center Haaglanden The Hague, The Netherlands