Detection of human papillomavirus type 11 in a case of Buschke-Löwenstein tumor

ARTICLE

Auteur(s) : Yutaka Asato, Kiyohito Taira, Yu-ichi Yamamoto, Hiroshi Uezato

Division of Dermatology, Department of Medicine, University of the Ryukyu, 207 Uehara, Nishiara town, Okinawa, Japan (zip code: 903-0215)

accepté le 5 Decembre 2007

Buschke-Löwenstein tumor (BLT) was first described by Buschke and Löwenstein in 1925 [1] and was defined as “carcinoma-like condylomata acuminata of the penis” by Löwenstein in 1938 [2]. BLT is a slow-growing, locally destructive verrucous lesion without a malignant appearance in histology, and may occur elsewhere in the anogenital region.

In 1948, Ackerman reported that BLT could be regarded as a type of verrucous carcinoma (VC), as VC resembles BLT in clinical and histological appearance [3]. However, there are some reports that oppose this view [4, 5]. Therefore, confusion still exists over whether cauliflower-like tumors on the penis should be referred to as VC or BLT.

It is not fully understood, why the conyloma acuminatum (CA) develops into large tumors. There are many enigmatic factors in BLT and VC. For example, the HPV typically detected in BLT, which is a very malignant tumor, is not a high-risk type, such as genotype 16 or 18, but rather is a low-risk type, such as 6 or 11 [6, 7]. Moreover, the HPV-positivity rate in VC is lower than in common giant conyloma acuminatum (GCA) [8, 9]. To date, these questions remain unsolved, and several investigators have attempted to elucidate the etiology. Here, we report a case of BLT, in which HPV11 was observed, and discuss the possible mechanisms of BLT oncogenesis.

Case report

On clinical examination, a giant papillary tumor on the root of penis extending mostly throughout the frontal part was noted. The prepuce of the penis was also infiltrated by the tumor, which enveloped the whole of the glans (figure 1A). Magnetic resonance imaging (MRI) revealed no invasion into the dartos fascia or albuginea. Therefore, we decided to perform total excision of the tumor. Under general anesthesia, the tumor was excised with minimal margins at the deep layer of adipose tissue (figure 1B). For reconstruction, a mesh split-thickness skin graft closure was performed. The skin graft was taken from his thigh using an electric dermatome and was implanted on the excised lesion. Biological tissue adhesive (human blood factor XIII with fibrinogen) was injected under the skin graft in order to ensure tight adherence to the penis. The graft was successful and allowed to the patient to be discharged at 2 weeks after surgery.

Histological examination of the pubic and prepuce parts of the lesion was then performed. In the pubic part, the epidermis was elongated with a saw-toothed appearance and parts of the epidermis invaded into the deep layer of the dermis on low-power magnification. Although the nuclei were not atypical in high-power fields, koilocytes were observed in epidermis (figure 2A), while invasion of epidermis was not observed in the prepuce part, despite thick epidermal proliferation (figure 2B). On immunohistochemistry, anti-HPV polyclonal antibody staining (Dako Japan, Japan) was positive in koilocyte nuclei (figure 2C). Based on these results, we diagnosed the tumor as BLT.

We also performed genetic analysis of HPV using polymerase chain reaction (PCR) and direct sequencing methods. The L1 region of HPV was amplified as reported by Yoshikawa et al. [10] Tumor tissue DNA was extracted with a Tissue DNA Isolation Kit (Amersham Pharmanica Biotech, USA) in accordance with the manufacturer’s instructions. L1C1 (5’-CGTAAACGTTTTCCCTATTTTTTT-3’ 5609-5632 bp) and L1C2 (5’-CAATACAGAGTATTTAGGGTA-3’ 5841-5861 bp), were chosen as consensus primers for the Ll region. PCR was performed in a total volume of 50 μL, and each reaction mixture contained 400 ng of DNA template, each primer at 100 pM, dNTPs at 0.2 mM each, 1.25 units of EX taq polymerase, 10 mM Tris-HCL, pH 8.3, 50 mM KCl and 1.5 mM MgCl₂ (Takara, Japan). DNA was amplified using a Gene Amp® PCR SYSTEM 9700 (Applied Biosystems, USA) with initial denaturation at 94 °C for 1 min followed by 40 cycles of denaturation at 94 °C for 1 min, annealing at 50 °C for 1 min and extension at 72 °C for 1 min 30 s, with a final extension at 74 °C for 5 min. Each of PCR products was run on a 2.5% agarose gel, stained with ethidium bromide and visualized by UV illumination. PCR product was observed at 240 bp (figure 3A). This product was then purified using a QIAquick Gel Extracion Kit (Qiagen, USA), and was sequenced directly on an ABI PRISM^TM 310 automated sequencer (Applied Biosystems, USA) using the Big Dye terminator cycle sequencing ready reaction kit (Applied Biosystems, USA) (figure 3B). Alignment of DNA was edited with the software Genetyx Mac.ver 11.0 (Software Development Co. Ltd, Japan) and a homology search conducted using NCBI Nucleotide-nucleotide BLAST (http://www.ncbi.nlm.nih.gov/BLAST/) revealed the PCR product was homologous to HPV type 11.

Discussion

However, there is still some confusion between BLT and VC. Some reports regard these lesions as distinct entities [4, 5]. In fact, the HPV-positivity rate of VC is lower than that of general GCA [8, 9], with some reports stating that only 12% of cases (3/26) are positive [8, 9, 11, 12]. Another report found that VC of the vulva is a rare HPV-negative neoplasm [13]. As BLT was originally reported as a type of GCA, it should be an HPV-associated tumor. These results are thus inconsistent with BLT. Therefore, Clare et al. believed that BLT was a low-risk HPV (type 6 or 11)-positive tumor, and considered that the presence of HPV may be useful in differentiating BLT from VC [14]. As HPV type 11 was also detected in the present patient, we diagnosed BLT rather than VC.

It is unclear why CA grows into giant tumors such as BLT, but there are three hypotheses. The first hypothesis relates to oncogenic viruses. Some reports have suggested that the presence of high-risk HPV type 16 or 18 within a condyloma already containing HPV type 6 or 11 may be important for the development of BLT; however, PCR did not detect high-risk HPV in our case [15]. Other reports postulated that mutations within the E6 and E7 coding regions of low-risk HPV (oncogenes in high-risk HPV) may alter the oncoproteins and be related to oncogenesis [16]. Heck et al. revealed that the substitution of a single amino acid at position 22 in HPV type 6 (replacing glycine with aspartic acid) was responsible for enhanced pRB binding affinity and increased potential to cooperate with ras in the transformation of primary rodent cells [17]. Sang et al. also reported that subtle amino acid changes in HPV type 6 E7 proteins may result in HPV type 16 oncoprotein-like transforming activities in vitro [16].These reports demonstrate the possibility that viral mutations are related to carcinogenesis. On the other hand, Grassmann et al. attempted to perform sequencing of the E6 and E7 regions from a number of clinical samples in order to detect specific mutations for oncogenesis, but identified few unique codes in individual samples, although several amino acid substations were noted, particularly in the E6 protein.

The second hypothesis relates to host immunity. Cuesta et al. reported that VC in human immunodeficiency virus (HIV)-infected patients is associated with HPV [18]. This supports the notion that concurrent immune deficiency resulting from HIV infection may play a role in promoting the effects of HPV in the pathogenesis of genito-anal cancer. Our patient was negative for HIV and human T-lymphotropic virus 1 (HTLV-1). HTLV-1 is a similar retrovirus to HIV and is endemic in South America, Central Africa and southwestern parts of Japan, including Okinawa. These two human retroviruses (HIV and HTLV-1) differ greatly. HIV vigorously replicates in vivo, while HTLV-1 has long resided in the human body. It is unclear whether HTLV-1 may have influenced tumorigenesis in our patient. However, other virus-associated tumors, such as Kaposi’s sarcoma (KS) caused by human herpes virus 8 (HHV-8), which is ordinarily observed in AIDS patients, are frequently reported in Okinawa, even in patients who are not HIV carriers [19]. At least one report has suggested that that HTLV-1 infection may be related to HHV-8 infection and raises the risk of KS [20]. Such reports suggest the need for further research into HPV.

The third hypothesis relates to penile hygiene. Some reports have suggested that uncircumcised males are inclined to higher rates of BLT when compared with circumcised males, and that a higher incidence of circumcision is a factor in the prevention of BLT [20]. The present patient exhibited phimosis, and thus penile hygiene may have played a role in oncogenesis.

Unfortunately, there was insufficient data to conclusively identify the pathogenesis of BLT. However, we were able to discuss the roles of various possible factors in oncogenesis. We need to follow future reports more closely.

Acknowledgements

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