- Email: [email protected]
Cutaneous Squamous Cell Carcinoma: A Smoking Gun but Still No Suspects
commentary
et al. (2006) Common loss-of-function variants of the epidermal barrier protein filaggrin are a major predisposing factor for atopic dermatitis. Nat Genet 38:441–6 Saaf AM, Tengvall-Linder M, Chang HY et al. (2008) Global expression profiling in atopic eczema reveals reciprocal expression of inflammatory and lipid genes. PLoS One 3:24 Sinclair C, O’Toole EA, Paige D et al. (2009) Filaggrin mutations are associated with ichthyosis vulgaris in the Bangladeshi population. Br J Dermatol 160:1113–5 Smith FJ, Irvine AD, Terron-Kwiatkowski A et al. (2006) Loss-of-function mutations in the gene encoding filaggrin cause ichthyosis vulgaris. Nat Genet 38:337–42s
Steinert PM, Candi E, Kartasova T et al. (1998) Small proline-rich proteins are cross-bridging proteins in the cornified cell envelopes of stratified squamous epithelia. J Struct Biol 122:76–85 Sugiura H, Ebise H, Tazawa T et al. (2005) Largescale DNA microarray analysis of atopic skin lesions shows overexpression of an epidermal differentiation gene cluster in the alternative pathway and lack of protective gene expression in the cornified envelope. Br J Dermatol 152:146–9 Vermeij WP, Backendorf C (2010) cornification proteins provide global between ROS detoxification and migration during wound healing. PLoS 5:e11957
Skin link cell One
See related article on pg 1745
Cutaneous Squamous Cell Carcinoma: A Smoking Gun but Still No Suspects John T. Schiller1 and Christopher B. Buck1 A viral etiology for cutaneous squamous cell carcinoma (cuSCC) has long been suspected, primarily on the basis of its dramatically increased incidence in immunocompromised individuals. In this issue, Arron and colleagues report a comprehensive hunt for viral gene transcription in cuSCC. Their findings show that it is very unlikely that any currently known virus is commonly responsible for the maintenance of this cancer. Journal of Investigative Dermatology (2011) 131, 1595–1596. doi:10.1038/jid.2011.151
Identification of a viral cause of a human cancer is desirable because it provides unique opportunities for intervention, particularly prevention. This is well illustrated by the recently introduced prophylactic vaccine to prevent infection by the sexually transmitted human papillomavirus (HPV) types most frequently associated with cervical and other anogenital and oral cancers. A common “smoking gun” trait of cancers with a well-documented viral etiology is
their increased incidence in immuno compromised individuals (Grulich et al., 2007). This is presumably attributable to impaired immunological control over what might otherwise be a harmless infection. Risk of nonmelanoma skin cancer, including cutaneous squamous cell carcinoma (cuSCC), is dramatically increased after immunosuppression, with a more than 50-fold increase in incidence among recipients of solid-organ transplants. It has therefore long been suspected that
Laboratory of Cellular Oncology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA
1
Correspondence: John T. Schiller, Laboratory of Cellular Oncology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA. E-mail: [email protected]
cuSCC may be caused by an infectious agent. Significant attention has been focused on cutaneous HPVs, particularly those from papillomavirus genusb (b-HPV), because they ubiquitously and chronically infect the skin and are clearly implicated in skin carcinogenesis in a rare genodermatosis known as epidermodysplasia verruciformis (Feltkamp et al., 2008). The conclusion that the association of mucosatropic HPVs with anogenital cancers is causal is supported by many lines of laboratory and epidemiological evidence. The consistent maintenance of the viral genome, as well as ongoing transcription of the viral oncogenes in essentially every cell of the tumor, is a central component of this causal inference. To date, the evidence suggesting a causal role for cutaneous HPVs in cuSCC in the general population has been mixed, at best (Nindl et al., 2007). b-HPV DNA is frequently detected in cuSCC, and some studies have detected higher serological responses to b-HPV virions in cases than in controls. However, viral DNA is also frequently detected in normal skin and, in studies that have evaluated viral genome copy number in cuSCC tumor specimens, this number was usually much less than one viral genome per tumor cell. Because these studies used HPV DNA and serological assays designed to evaluate only a few of the many dozens of known HPV species, they left open the possibility that an unstudied subset of HPV types not targeted by the assays was more strongly associated with cuSCC. In this issue, Arron and colleagues report their use of the powerful new technology of high-throughput mRNA sequencing to provide an unbiased and comprehensive assessment of the causal role of HPV gene expression in cuSCC. Using random primers, they amplified and sequenced a representative fraction of the total transcriptosome from 31 cuSCC and 8 patient-paired normal skin biopsies, with an impressive mean count of 3.5 million reads per sample. The sequence reads, averaging 54 nucleo tides in length, were compared with a database of all known viruses. Only www.jidonline.org 1595
commentary
one SCC showed evidence of HPV transcripts (HPV92, a b-HPV) and only at very low levels (0.0001% of total reads). In important controls for the technical aspects of the study, HPV16 transcripts were detected relatively abundantly in a cervical SCC and a periungual SCC positive for HPV16 DNA (0.02% and 0.06% of total reads, respectively). Transcripts of other eukaryotic virus were not detected with any substantial frequency. Therefore, the results strongly support the important conclusion that known virus species are not consistent contributors to the maintenance of cuSCCs.
|
It is unlikely that a known virus is responsible for maintenance of cuSCC.
The study leaves open the possibility that a virus may be involved in the initiation, but not the maintenance, of cuSCCs. Because b-HPVs are known to encode candidate oncoproteins that interfere with proapoptotic signaling in epithelial cells, they might prevent UV-induced apoptosis, thereby allowing keratinocytes with UV-induced mutations to survive and progress to carcinomas (Jackson et al., 2000). Although this type of hit-and-run mechanism is often dismissed, it has been clearly documented in experimental systems. In the most relevant example, bovine papillomavirus type 4 (BPV4) induces alimentary tract papillomas that can progress to cancer in experimentally challenged cattle. Although the initial papillomatous precursor lesions contain BPV4 DNA, the malignant carcinomas are devoid of the viral DNA (Campo et al., 1994). If this mechanism were to function in cuSCC, it would imply that there is strong biological or immunological selection for loss of viral gene expression during carcinogenic progression. It is unclear why this would be the case for cutaneous, but not genital, neoplasia.
Unfortunately, compelling evidence for hit-and-run etiology is difficult to obtain in clinical studies. Establishing such a connection would probably require the unequivocal identification of the critical cuSCC precursor lesion and the demonstration that the precursor usually contained nucleic acid sequences of a particular virus or group of viruses. Serological studies might also be useful for the establishment of hit-and-run etiologies because antibody responses are often very durable and can serve as a fossil record of infection, even if the malignant lesion does not ultimately contain nucleic acid sequences of the etiologically responsible pathogen. Unfortunately, serological investigations have the drawback that they require individual study of known viral serotypes. Furthermore, serological results could reflect indirect effects. For example, the increased serological responsiveness of cuSCC patients against b-HPVs could theoretically reflect a generalized loss of immunological control over multiple types of viral infections, including HPVs, and an unknown pathogen that is the true culprit underlying cuSCC. The work by Arron and colleagues (2011) also leaves open the possibility that cuSCC is caused by an as-yet unknown virus with too little similarity to known viruses to be detected in BLAST homology searches. Interesting new human skin viruses continue to be identified, as exemplified by four recently discovered cutaneous polyomaviruses—Merkel cell polyoma virus (Feng et al., 2008), HPyV6 and HPyV7 (Schowalter et al., 2010), and trichodysplasia spinulosa-associated polyomavirus (van der Meijden et al., 2010)—and a number of recently identified cutaneous HPVs (particularly of the highly diverse papillomavirus genus-g; Kohler et al., 2011). It is likely that the explosive increase in the use of high-throughput sequencing technologies will uncover additional new cutaneous viruses, perhaps some from currently unknown virus families. An attractive aspect of Arron and colleagues’ work is that it provides a large, valuable, and publically accessible data set (deposited in the National
1596 Journal of Investigative Dermatology (2011), Volume 131
Center for Biotechnology Information Sequence Read Archive) that includes more than 1 million unidentified transcript reads with no clear homology to known human sequences. These unknown reads could be a useful hunting ground for future studies aimed at establishing possible associations between newly discovered viral isolates and the maintenance of cuSCC. In other words, the fact that the usual viral suspects previously hypothesized to underlie cuSCC have not been caught at the crime scene should not be taken to mean that the case is closed. CONFLICT OF INTEREST
The authors state no conflict of interest.
REFERENCES Arron ST, Ruby JG, Dybbro E et al. (2011) Transcriptome sequencing demonstrates that human papillomavirus is not active in cutaneous squamous cell carcinoma. J Invest Dermatol 131:1745–53 Campo MS, O’Neil BW, Barron RJ et al. (1994) Experimental reproduction of the papilloma– carcinoma complex of the alimentary canal in cattle. Carcinogenesis 15:1597–601 Feltkamp MC, de Koning MN, Bavinck JN et al. (2008) Betapapillomaviruses: innocent bystanders or causes of skin cancer. J Clin Virol 43:353–60 Feng H, Shuda M, Chang Y et al. (2008) Clonal integration of a polyomavirus in human Merkel cell carcinoma. Science 319: 1096– 100 Grulich AE, van Leeuwen MT, Falster MO et al. (2007) Incidence of cancers in people with HIV/AIDS compared with immunosuppressed transplant recipients: a meta-analysis. Lancet 370:59–67 Jackson S, Harwood C, Thomas M et al. (2000) Role of Bak in UV-induced apoptosis in skin cancer and abrogation by HPV E6 proteins. Genes Dev 14:3065–73 Kohler A, Gottschling M, Manning K et al. (2011) Genomic characterization of ten novel cutaneous human papillomaviruses from keratotic lesions of immunosuppressed patients. J Gen Virol; e-pub ahead of print 6 April 2011 Nindl I, Gottschling M, Stockfleth E (2007) Human papillomaviruses and non-melanoma skin cancer: basic virology and clinical manifestations. Dis Markers 23:247–59 Schowalter RM, Pastrana DV, Pumphrey KA et al. (2010) Merkel cell polyomavirus and two previously unknown polyomaviruses are chronically shed from human skin. Cell Host Microbe 7:509–15 van der Meijden E, Janssens RW, Lauber C et al. (2010) Discovery of a new human polyomavirus associated with trichodysplasia spinulosa in an immunocompromized patient. PLoS Pathog 6:e1001024
et al. (2006) Common loss-of-function variants of the epidermal barrier protein filaggrin are a major predisposing factor for atopic dermatitis. Nat Genet 38:441–6 Saaf AM, Tengvall-Linder M, Chang HY et al. (2008) Global expression profiling in atopic eczema reveals reciprocal expression of inflammatory and lipid genes. PLoS One 3:24 Sinclair C, O’Toole EA, Paige D et al. (2009) Filaggrin mutations are associated with ichthyosis vulgaris in the Bangladeshi population. Br J Dermatol 160:1113–5 Smith FJ, Irvine AD, Terron-Kwiatkowski A et al. (2006) Loss-of-function mutations in the gene encoding filaggrin cause ichthyosis vulgaris. Nat Genet 38:337–42s
Steinert PM, Candi E, Kartasova T et al. (1998) Small proline-rich proteins are cross-bridging proteins in the cornified cell envelopes of stratified squamous epithelia. J Struct Biol 122:76–85 Sugiura H, Ebise H, Tazawa T et al. (2005) Largescale DNA microarray analysis of atopic skin lesions shows overexpression of an epidermal differentiation gene cluster in the alternative pathway and lack of protective gene expression in the cornified envelope. Br J Dermatol 152:146–9 Vermeij WP, Backendorf C (2010) cornification proteins provide global between ROS detoxification and migration during wound healing. PLoS 5:e11957
Skin link cell One
See related article on pg 1745
Cutaneous Squamous Cell Carcinoma: A Smoking Gun but Still No Suspects John T. Schiller1 and Christopher B. Buck1 A viral etiology for cutaneous squamous cell carcinoma (cuSCC) has long been suspected, primarily on the basis of its dramatically increased incidence in immunocompromised individuals. In this issue, Arron and colleagues report a comprehensive hunt for viral gene transcription in cuSCC. Their findings show that it is very unlikely that any currently known virus is commonly responsible for the maintenance of this cancer. Journal of Investigative Dermatology (2011) 131, 1595–1596. doi:10.1038/jid.2011.151
Identification of a viral cause of a human cancer is desirable because it provides unique opportunities for intervention, particularly prevention. This is well illustrated by the recently introduced prophylactic vaccine to prevent infection by the sexually transmitted human papillomavirus (HPV) types most frequently associated with cervical and other anogenital and oral cancers. A common “smoking gun” trait of cancers with a well-documented viral etiology is
their increased incidence in immuno compromised individuals (Grulich et al., 2007). This is presumably attributable to impaired immunological control over what might otherwise be a harmless infection. Risk of nonmelanoma skin cancer, including cutaneous squamous cell carcinoma (cuSCC), is dramatically increased after immunosuppression, with a more than 50-fold increase in incidence among recipients of solid-organ transplants. It has therefore long been suspected that
Laboratory of Cellular Oncology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA
1
Correspondence: John T. Schiller, Laboratory of Cellular Oncology, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA. E-mail: [email protected]
cuSCC may be caused by an infectious agent. Significant attention has been focused on cutaneous HPVs, particularly those from papillomavirus genusb (b-HPV), because they ubiquitously and chronically infect the skin and are clearly implicated in skin carcinogenesis in a rare genodermatosis known as epidermodysplasia verruciformis (Feltkamp et al., 2008). The conclusion that the association of mucosatropic HPVs with anogenital cancers is causal is supported by many lines of laboratory and epidemiological evidence. The consistent maintenance of the viral genome, as well as ongoing transcription of the viral oncogenes in essentially every cell of the tumor, is a central component of this causal inference. To date, the evidence suggesting a causal role for cutaneous HPVs in cuSCC in the general population has been mixed, at best (Nindl et al., 2007). b-HPV DNA is frequently detected in cuSCC, and some studies have detected higher serological responses to b-HPV virions in cases than in controls. However, viral DNA is also frequently detected in normal skin and, in studies that have evaluated viral genome copy number in cuSCC tumor specimens, this number was usually much less than one viral genome per tumor cell. Because these studies used HPV DNA and serological assays designed to evaluate only a few of the many dozens of known HPV species, they left open the possibility that an unstudied subset of HPV types not targeted by the assays was more strongly associated with cuSCC. In this issue, Arron and colleagues report their use of the powerful new technology of high-throughput mRNA sequencing to provide an unbiased and comprehensive assessment of the causal role of HPV gene expression in cuSCC. Using random primers, they amplified and sequenced a representative fraction of the total transcriptosome from 31 cuSCC and 8 patient-paired normal skin biopsies, with an impressive mean count of 3.5 million reads per sample. The sequence reads, averaging 54 nucleo tides in length, were compared with a database of all known viruses. Only www.jidonline.org 1595
commentary
one SCC showed evidence of HPV transcripts (HPV92, a b-HPV) and only at very low levels (0.0001% of total reads). In important controls for the technical aspects of the study, HPV16 transcripts were detected relatively abundantly in a cervical SCC and a periungual SCC positive for HPV16 DNA (0.02% and 0.06% of total reads, respectively). Transcripts of other eukaryotic virus were not detected with any substantial frequency. Therefore, the results strongly support the important conclusion that known virus species are not consistent contributors to the maintenance of cuSCCs.
|
It is unlikely that a known virus is responsible for maintenance of cuSCC.
The study leaves open the possibility that a virus may be involved in the initiation, but not the maintenance, of cuSCCs. Because b-HPVs are known to encode candidate oncoproteins that interfere with proapoptotic signaling in epithelial cells, they might prevent UV-induced apoptosis, thereby allowing keratinocytes with UV-induced mutations to survive and progress to carcinomas (Jackson et al., 2000). Although this type of hit-and-run mechanism is often dismissed, it has been clearly documented in experimental systems. In the most relevant example, bovine papillomavirus type 4 (BPV4) induces alimentary tract papillomas that can progress to cancer in experimentally challenged cattle. Although the initial papillomatous precursor lesions contain BPV4 DNA, the malignant carcinomas are devoid of the viral DNA (Campo et al., 1994). If this mechanism were to function in cuSCC, it would imply that there is strong biological or immunological selection for loss of viral gene expression during carcinogenic progression. It is unclear why this would be the case for cutaneous, but not genital, neoplasia.
Unfortunately, compelling evidence for hit-and-run etiology is difficult to obtain in clinical studies. Establishing such a connection would probably require the unequivocal identification of the critical cuSCC precursor lesion and the demonstration that the precursor usually contained nucleic acid sequences of a particular virus or group of viruses. Serological studies might also be useful for the establishment of hit-and-run etiologies because antibody responses are often very durable and can serve as a fossil record of infection, even if the malignant lesion does not ultimately contain nucleic acid sequences of the etiologically responsible pathogen. Unfortunately, serological investigations have the drawback that they require individual study of known viral serotypes. Furthermore, serological results could reflect indirect effects. For example, the increased serological responsiveness of cuSCC patients against b-HPVs could theoretically reflect a generalized loss of immunological control over multiple types of viral infections, including HPVs, and an unknown pathogen that is the true culprit underlying cuSCC. The work by Arron and colleagues (2011) also leaves open the possibility that cuSCC is caused by an as-yet unknown virus with too little similarity to known viruses to be detected in BLAST homology searches. Interesting new human skin viruses continue to be identified, as exemplified by four recently discovered cutaneous polyomaviruses—Merkel cell polyoma virus (Feng et al., 2008), HPyV6 and HPyV7 (Schowalter et al., 2010), and trichodysplasia spinulosa-associated polyomavirus (van der Meijden et al., 2010)—and a number of recently identified cutaneous HPVs (particularly of the highly diverse papillomavirus genus-g; Kohler et al., 2011). It is likely that the explosive increase in the use of high-throughput sequencing technologies will uncover additional new cutaneous viruses, perhaps some from currently unknown virus families. An attractive aspect of Arron and colleagues’ work is that it provides a large, valuable, and publically accessible data set (deposited in the National
1596 Journal of Investigative Dermatology (2011), Volume 131
Center for Biotechnology Information Sequence Read Archive) that includes more than 1 million unidentified transcript reads with no clear homology to known human sequences. These unknown reads could be a useful hunting ground for future studies aimed at establishing possible associations between newly discovered viral isolates and the maintenance of cuSCC. In other words, the fact that the usual viral suspects previously hypothesized to underlie cuSCC have not been caught at the crime scene should not be taken to mean that the case is closed. CONFLICT OF INTEREST
The authors state no conflict of interest.
REFERENCES Arron ST, Ruby JG, Dybbro E et al. (2011) Transcriptome sequencing demonstrates that human papillomavirus is not active in cutaneous squamous cell carcinoma. J Invest Dermatol 131:1745–53 Campo MS, O’Neil BW, Barron RJ et al. (1994) Experimental reproduction of the papilloma– carcinoma complex of the alimentary canal in cattle. Carcinogenesis 15:1597–601 Feltkamp MC, de Koning MN, Bavinck JN et al. (2008) Betapapillomaviruses: innocent bystanders or causes of skin cancer. J Clin Virol 43:353–60 Feng H, Shuda M, Chang Y et al. (2008) Clonal integration of a polyomavirus in human Merkel cell carcinoma. Science 319: 1096– 100 Grulich AE, van Leeuwen MT, Falster MO et al. (2007) Incidence of cancers in people with HIV/AIDS compared with immunosuppressed transplant recipients: a meta-analysis. Lancet 370:59–67 Jackson S, Harwood C, Thomas M et al. (2000) Role of Bak in UV-induced apoptosis in skin cancer and abrogation by HPV E6 proteins. Genes Dev 14:3065–73 Kohler A, Gottschling M, Manning K et al. (2011) Genomic characterization of ten novel cutaneous human papillomaviruses from keratotic lesions of immunosuppressed patients. J Gen Virol; e-pub ahead of print 6 April 2011 Nindl I, Gottschling M, Stockfleth E (2007) Human papillomaviruses and non-melanoma skin cancer: basic virology and clinical manifestations. Dis Markers 23:247–59 Schowalter RM, Pastrana DV, Pumphrey KA et al. (2010) Merkel cell polyomavirus and two previously unknown polyomaviruses are chronically shed from human skin. Cell Host Microbe 7:509–15 van der Meijden E, Janssens RW, Lauber C et al. (2010) Discovery of a new human polyomavirus associated with trichodysplasia spinulosa in an immunocompromized patient. PLoS Pathog 6:e1001024