SerpinB2 (aka plasminogen activator type 2) is well described as an extracellular inhibitor of urokinase-type plasminogen activator (uPA). However, the majority of SerpinB2 is retained intracellularly, and many uPA-independent activities have been reported for SerpinB2 suggesting an alternate function. This thesis explores the role of SerpinB2 in epithelial tumour cell lines, highlights the problems associated with various expression systems and argues that SerpinB2 has no role in growth or apoptosis of tumour cells. A potential role for immune modulation and angiogenesis is suggested in in vivo models. Previous research using SerpinB2 transfected, clonally selected tumour cell lines suggested that SerpinB2 regulates the retinoblastoma tumour suppressor protein (Rb) by binding and protecting Rb from degradation. Despite the use of two techniques under numerous conditions and positive controls, no significant interaction between SerpinB2 and Rb was found. SerpinB2 was reported to bind Rb through a PENF homology motif located within the SerpinB2 C-D interhelical loop region. The PENF homology motif was postulated to represent the motif responsible for binding to the C-pocket of Rb. Epstein Barr Virus nuclear antigen 6 (EBNA6) is a known Rb binding protein, which contains two predicted PENF homology motifs. However, mutation of the two PENF homology motifs within EBNA6 did not reduce Rb binding. Furthermore, the SerpinB2 PENF homology motif is actually not well conserved between SerpinB2 proteins from multiple species, whereas other regions of the SerpinB2 C-D loop show a high level of conservation. These data do not support a role for SerpinB2 and the PENF homology motif in Rb binding. SerpinB2 has been proposed to have a role in regulating growth and apoptosis. To further investigate this proposed phenotype of SerpinB2, SerpinB2 was expressed in a range of epithelial tumour lines using transient transfection. No change in growth, apoptosis or Rb levels were found. After ≈2-3 month antibiotic selection for the SerpinB2-expressing plasmid, SerpinB2 protein was lost without the loss of the transgene, indicating selective pressure against long-term SerpinB2 protein expression. To further investigate long-term SerpinB2 expression adenovirus and lentivirus vectors were used. Infection of tumour cell lines with adenovirus vectors expressing SerpinB2 resulted in reduced cell growth, characterised by increased p53 (but not Rb) levels and G2 arrest or apoptosis. When SerpinB2 expressing lentivirus vectors were used to transduce the same tumour cell lines, high levels of long-term expression of functional SerpinB2 was achieved. However, SerpinB2-expressing cell lines showed no differences in growth, proliferation, Rb levels, or apoptosis induced by a range of agents. Growth and apoptosis observed with adenovirus SerpinB2 had all the characteristics of adenovirus-associated toxicity, which has been reported previously for specific proteins. These experiments highlighted the problems associated with SerpinB2 expression systems and suggest that SerpinB2 expression per se is not toxic nor has a role in regulating Rb, growth and apoptosis. Screening of a number of tumour cell lines identified the HPV16 transformed cervical cancer line as expressing high levels of SerpinB2. SerpinB2 was located both extracellularly and intracellularly with a cytoplasmic and nuclear distribution. A high molecular weight SerpinB2 species was identified in CaSki cells and was shown to be the N-linked glycosylated species. Sequencing showed the protein to be Type A SerpinB2 and the protein was shown to form an inhibitory complex with uPA. An abundant low molecular weight SerpinB2 species was also identified in CaSki cell supernatants and appeared to be a proteolytic fragment of SerpinB2. Treatment of CaSki with PMA, TNFα and IFNγ increased SerpinB2 levels. Lentiviral based shRNA failed to significantly down regulate SerpinB2 expression and increasing SerpinB2 levels with lentiviral expression did not change growth, apoptosis, Rb levels or E7 transcription. Lentiviral expression of SerpinB2 in (normally SerpinB2 negative) HPV16 transformed SiHa cells, also failed to show changes in Rb levels or E7 transcription. CaSki thus express wild-type and functional SerpinB2, but no evidence could found that SerpinB2 effects HPV16 E7 transcription or Rb levels. The data presented identifies CaSki as valuable source of biologically functional SerpinB2. SerpinB2 expression in breast cancer cells has been associated with positive prognosis. Tubo, a SerpinB2-negative murine breast carcinoma cell line, was transduced with lentivirus expressing SerpinB2 and grown subcutaneously in BALB/c mice. SerpinB2 expressing tumours appeared red and were larger than control tumours. Furthermore, SerpinB2 expressing tumours had a ≈2 fold higher density of blood vessels when compared to Tubo and Tubo expressing EGFP. Mice carrying tumours expressing SerpinB2 also showed reduced anti-tumour IgG2 responses. These data suggest that a role for SerpinB2 in regulating angiogenesis and antitumour immunity. In conclusion, this thesis challenges the notion that SerpinB2 regulates Rb, cell cycle, and apoptosis and suggests a potential role for SerpinB2 in tumour angiogenesis and immunity.

SerpinB2 (aka plasminogen activator type 2) is well described as an extracellular inhibitor of urokinase-type plasminogen activator (uPA). However, the majority of SerpinB2 is retained intracellularly, and many uPA-independent activities have been reported for SerpinB2 suggesting an alternate function. This thesis explores the role of SerpinB2 in epithelial tumour cell lines, highlights the problems associated with various expression systems and argues that SerpinB2 has no role in growth or apoptosis of tumour cells. A potential role for immune modulation and angiogenesis is suggested in in vivo models. Previous research using SerpinB2 transfected, clonally selected tumour cell lines suggested that SerpinB2 regulates the retinoblastoma tumour suppressor protein (Rb) by binding and protecting Rb from degradation. Despite the use of two techniques under numerous conditions and positive controls, no significant interaction between SerpinB2 and Rb was found. SerpinB2 was reported to bind Rb through a PENF homology motif located within the SerpinB2 C-D interhelical loop region. The PENF homology motif was postulated to represent the motif responsible for binding to the C-pocket of Rb. Epstein Barr Virus nuclear antigen 6 (EBNA6) is a known Rb binding protein, which contains two predicted PENF homology motifs. However, mutation of the two PENF homology motifs within EBNA6 did not reduce Rb binding. Furthermore, the SerpinB2 PENF homology motif is actually not well conserved between SerpinB2 proteins from multiple species, whereas other regions of the SerpinB2 C-D loop show a high level of conservation. These data do not support a role for SerpinB2 and the PENF homology motif in Rb binding. SerpinB2 has been proposed to have a role in regulating growth and apoptosis. To further investigate this proposed phenotype of SerpinB2, SerpinB2 was expressed in a range of epithelial tumour lines using transient transfection. No change in growth, apoptosis or Rb levels were found. After ≈2-3 month antibiotic selection for the SerpinB2-expressing plasmid, SerpinB2 protein was lost without the loss of the transgene, indicating selective pressure against long-term SerpinB2 protein expression. To further investigate long-term SerpinB2 expression adenovirus and lentivirus vectors were used. Infection of tumour cell lines with adenovirus vectors expressing SerpinB2 resulted in reduced cell growth, characterised by increased p53 (but not Rb) levels and G2 arrest or apoptosis. When SerpinB2 expressing lentivirus vectors were used to transduce the same tumour cell lines, high levels of long-term expression of functional SerpinB2 was achieved. However, SerpinB2-expressing cell lines showed no differences in growth, proliferation, Rb levels, or apoptosis induced by a range of agents. Growth and apoptosis observed with adenovirus SerpinB2 had all the characteristics of adenovirus-associated toxicity, which has been reported previously for specific proteins. These experiments highlighted the problems associated with SerpinB2 expression systems and suggest that SerpinB2 expression per se is not toxic nor has a role in regulating Rb, growth and apoptosis. Screening of a number of tumour cell lines identified the HPV16 transformed cervical cancer line as expressing high levels of SerpinB2. SerpinB2 was located both extracellularly and intracellularly with a cytoplasmic and nuclear distribution. A high molecular weight SerpinB2 species was identified in CaSki cells and was shown to be the N-linked glycosylated species. Sequencing showed the protein to be Type A SerpinB2 and the protein was shown to form an inhibitory complex with uPA. An abundant low molecular weight SerpinB2 species was also identified in CaSki cell supernatants and appeared to be a proteolytic fragment of SerpinB2. Treatment of CaSki with PMA, TNFα and IFNγ increased SerpinB2 levels. Lentiviral based shRNA failed to significantly down regulate SerpinB2 expression and increasing SerpinB2 levels with lentiviral expression did not change growth, apoptosis, Rb levels or E7 transcription. Lentiviral expression of SerpinB2 in (normally SerpinB2 negative) HPV16 transformed SiHa cells, also failed to show changes in Rb levels or E7 transcription. CaSki thus express wild-type and functional SerpinB2, but no evidence could found that SerpinB2 effects HPV16 E7 transcription or Rb levels. The data presented identifies CaSki as valuable source of biologically functional SerpinB2. SerpinB2 expression in breast cancer cells has been associated with positive prognosis. Tubo, a SerpinB2-negative murine breast carcinoma cell line, was transduced with lentivirus expressing SerpinB2 and grown subcutaneously in BALB/c mice. SerpinB2 expressing tumours appeared red and were larger than control tumours. Furthermore, SerpinB2 expressing tumours had a ≈2 fold higher density of blood vessels when compared to Tubo and Tubo expressing EGFP. Mice carrying tumours expressing SerpinB2 also showed reduced anti-tumour IgG2 responses. These data suggest that a role for SerpinB2 in regulating angiogenesis and antitumour immunity. In conclusion, this thesis challenges the notion that SerpinB2 regulates Rb, cell cycle, and apoptosis and suggests a potential role for SerpinB2 in tumour angiogenesis and immunity.

Thesis (Ph. D.)--University of Washington, 2007. / Includes bibliographical references (leaves 79-99).

Indiana University-Purdue University Indianapolis (IUPUI) / DNA is subject to various types of damage that can impair cellular function or cause cell death. DNA damage blocks normal cellular processes such as replication and transcription and can have catastrophic consequences for the cell and for the organism. It has long been thought that the G1/S cell cycle checkpoint allows time for DNA repair by delaying S-phase entry. The p53 tumor suppressor pathway regulates the G1/S checkpoint by regulating the cyclin-dependent kinase inhibitor p21Waf1/Cip1, but p53 also regulates the nucleotide excision DNA repair protein XPC. Here, using p53-null cell lines we show that additional mechanisms stabilize XPC protein and promote NER in concert with the G1/S checkpoint. At least one mechanism to stabilize and destabilize XPC involves ubiquitin-mediated degradation of XPC, as the ubiquitin ligase inhibitor MG-132 blocked XPC degradation. The retinoblastoma protein, RB, in its unphosphorylated form actually stabilized XPC and promoted NER as measured by host-cell reactivation experiments. The data suggest that XPC protein and XPC-mediated NER is tightly linked to the G1/S checkpoint even in cells lacking functional p53.

The proper maintenance of the pathways governing cell growth is critical to ensure cell survival and DNA fidelity. Much of our understanding of how the cell cycle is regulated comes from studies examining the relationship between DNA viruses and the mechanisms of cell proliferation control. There are numerous examples demonstrating that viruses can alter the host cell environment to their advantage. In particular, the small DNA tumor viruses, which include adenovirus, simian-virus 40 (SV-40), and human papillomavirus (HPV), can modulate the host cell cycle to facilitate viral DNA replication. Due to the fact that these viruses infect quiescent, non-cycling cells and lack the necessary enzymes and resources to replicate their DNA (e.g. DNA polymerase), the small DNA tumor viruses must activate the host cell replication machinery in order to expedite viral DNA replication. The capacity of these viruses to perturb normal cell proliferation control is dependent upon their oncogene products, which target p53 and members of the Retinoblastoma (RB) family of proteins and inactivate their respective functions. By targeting these key cell cycle regulatory proteins, the small DNA tumor viruses induce the infected host cells to enter S-phase and activate the components involved with host cell DNA synthesis thereby generating an environment that is conducive to viral DNA replication. In contrast, the larger, nuclear-replicating DNA viruses such as those from the family Herpesviridae, do not share the same stringent requirement as the small DNA viruses to induce the infected host cell to enter S-phase. The herpesviruses encode many of the components to stimulate nucleotide biosynthesis and the necessary factors to facilitate virus DNA replication including a viral DNA polymerase and other accessory factors. Additionally, many herpesviruses encode gene products that arrest the host cell cycle, in most instances, prior to the G1/S transition point. Inducing cells to growth arrest appears to be a prerequisite for the replication of most herpesviruses. However, in addition to encoding factors that inhibit the cell cycle, many herpesviruses encode proteins that can promote cell cycle progression in a manner similar to the small DNA tumor virus oncoproteins. By targeting members of the RB family and p53 protein, the herpesvirus proteins induce S-phase and activate S-phase associated factors that playa role in DNA replication. In this manner, the herpesviruses may promote an environment that is favorable for DNA replication. Consistent with the other herpesviruses, human cytomegalovirus (HCMV)induces human fibroblasts to growth arrest. However, in other cell types, virus infection causes cells to enter S-phase. In addition, HCMV replication requires several cellular factors that are present only during S-phase. Furthermore, HCMV induces the activation of S-phase-associated events as well as the increased expression of numerous S-phase genes following infection. HCMV encodes two immediate early (IE) gene products, IE1-72 and IE2-86, which can interact with members of the RB family of proteins. Additionally, the IE2-86 protein can bind to and inhibit p53 protein function. Given the functional resemblance between the HCMV IE proteins and the oncoproteins of the small DNA tumor viruses, we hypothesized that expression of the HCMV IE proteins could modulate cell cycle control. Specifically, we determined that expression of either IE1-72 or IE2-86 can induce quiescent cells to enter S-phase and delay cell cycle exit following serum withdrawal. Moreover, IE2-86 mediates this effect in the presence or absence of p53, whereas IE1-72 fails to do so in p53-expressing cells. Furthermore, both IE1-72 and IE2-86 induce p53 protein accumulation that is nuclear localized. Because IE1-72 fails to promote S-phase entry in cells expressing p53 and induces p53 protein levels, the mechanism by which IE1-72 alters p53 levels was examined. IE1-72 elevates p53 protein levels by inducing both p19ARF protein and an ATM-dependent phosphorylation of p53 at Ser15. IE1-72 also promotes p53 nuclear accumulation by abrogating p53 nuclear shuttling. As consequence of this IE1-72-mediated increase in p53 levels, p21 protein is induced leading to a p21-dependent growth arrest in cells expressing IE1-72. These findings demonstrate that the HCMV IE proteins can alter cell proliferation control and provide further support to the notion that HCMV, through the expression of its IE proteins, induces S-phase and factors associated with S-phase while blocking cell DNA synthesis, to possibly generate an environment that is suitable for viral DNA replication.

INTRODUÇÃO: O melanoma acral lentiginoso (MAL) tem freqüência expressiva entre os casos de melanoma observados no nosso meio e difere dos outros tipos clinicopatológicos de melanoma cutâneos por não ter a exposição solar como fator predisponente. Poucos trabalhos da literatura enfocam as alterações dos genes supressores de tumores e a expressão de suas proteínas nas lesões de MAL. Por esses motivos propusemo-nos a realizar um estudo retrospectivo visando uma melhor compreensão das proteínas envolvidas na via p16 INK4a /ciclina D1/CDK4/pRb do ciclo celular, no MAL em casuística brasileira. MÉTODOS: Através de técnica de imunoistoquímica foi demonstrada a expressão das proteínas p16, ciclina D1, CDK4 e pRb em 32 lesões de MAL. Comparou-se a freqüência de expressão desses marcadores nos tumores delgados (espessura de até 1,0mm) e espessos (mais de 1,0 mm de espessura), assim como de acordo com a presença ou ausência de ulceração. RESULTADOS: Houve expressão de p16 em 78% das lesões, de ciclina D1 em 61,5%, CDK4 em 84% e pRb em 85% dos MAL analisados. O padrão de imunomarcação das células neoplásicas foi nuclear para todos os anticorpos. Expressão nuclear associada à citoplasmática foi observada em 76% dos tumores positivos para p16 e 81% dos casos positivos para CDK4. Não houve diferenças significativas entre as freqüências de expressão de cada um dos marcadores quando comparados de acordo com sua espessura (delgados e espessos) e presença de ulceração. CONCLUSÕES: A expressão de ciclina D1 e CDK4, nos melanomas acrais lentiginosos, pode ser considerada aberrante e reflexo de provável alteração na via supressora de tumores p16/ciclinaD1/CDK4/pRb. A expressão tecidual de p16 e pRb, demonstrada pela técnica imunoistoquímica, pode não refletir as prováveis alterações da via supressora de tumores estudada. Esses marcadores, isoladamente, não devem ser considerados como fatores prognósticos no melanoma acral lentiginoso / INTRODUCTION: Acral lentiginous melanoma (ALM) has an expressive frequency between melanoma cases in our country, and it differs from the others clinical pathological types of cutaneous melanoma, because it does not have sun exposure as a predisposing factor. There are few publications in the literature addressing the expression of tumor suppressor pathway proteins in ALM. For these reasons we proposed to carry out a retrospective study aiming at a better understanding of p16 INK4a /cyclin D1/CDK4/pRb pathway involvement in ALM lesions. METHODS: Expression of p16, cyclin D1, CDK4 and pRB in 32 ALM lesions was displayed by immunohistochemistry. We compared the frequency of the expression of these markers in thin (thickness up to 1.0 mm) and thick lesions (thickness above 1.0 mm), as well as in accordance with the presence or absence of ulceration. RESULTS: 76% of the tumors were positive to p16 and 81% of the cases were positive to CDK4. There was no significant difference between the frequency of the expression of each marker when comparing in accordance with the thickness (thin and thick lesions) and the presence of ulceration. CONCLUSIONS: The expression of cyclin D1 and CDK4 in acral lentiginous melanoma may be considered aberrant and as reflex of a probable alteration in the p16/cyclin D1/CDK4/pRb tumor suppressor pathway. p16 and pRb tumor expression displayed by immunohistochemistry may not reflect probable alterations of the tumor suppressor pathway studied. The expression of these proteins, per se, may not be considered as prognostic factor in acral lentiginous melanoma

INTRODUÇÃO: O melanoma acral lentiginoso (MAL) tem freqüência expressiva entre os casos de melanoma observados no nosso meio e difere dos outros tipos clinicopatológicos de melanoma cutâneos por não ter a exposição solar como fator predisponente. Poucos trabalhos da literatura enfocam as alterações dos genes supressores de tumores e a expressão de suas proteínas nas lesões de MAL. Por esses motivos propusemo-nos a realizar um estudo retrospectivo visando uma melhor compreensão das proteínas envolvidas na via p16 INK4a /ciclina D1/CDK4/pRb do ciclo celular, no MAL em casuística brasileira. MÉTODOS: Através de técnica de imunoistoquímica foi demonstrada a expressão das proteínas p16, ciclina D1, CDK4 e pRb em 32 lesões de MAL. Comparou-se a freqüência de expressão desses marcadores nos tumores delgados (espessura de até 1,0mm) e espessos (mais de 1,0 mm de espessura), assim como de acordo com a presença ou ausência de ulceração. RESULTADOS: Houve expressão de p16 em 78% das lesões, de ciclina D1 em 61,5%, CDK4 em 84% e pRb em 85% dos MAL analisados. O padrão de imunomarcação das células neoplásicas foi nuclear para todos os anticorpos. Expressão nuclear associada à citoplasmática foi observada em 76% dos tumores positivos para p16 e 81% dos casos positivos para CDK4. Não houve diferenças significativas entre as freqüências de expressão de cada um dos marcadores quando comparados de acordo com sua espessura (delgados e espessos) e presença de ulceração. CONCLUSÕES: A expressão de ciclina D1 e CDK4, nos melanomas acrais lentiginosos, pode ser considerada aberrante e reflexo de provável alteração na via supressora de tumores p16/ciclinaD1/CDK4/pRb. A expressão tecidual de p16 e pRb, demonstrada pela técnica imunoistoquímica, pode não refletir as prováveis alterações da via supressora de tumores estudada. Esses marcadores, isoladamente, não devem ser considerados como fatores prognósticos no melanoma acral lentiginoso / INTRODUCTION: Acral lentiginous melanoma (ALM) has an expressive frequency between melanoma cases in our country, and it differs from the others clinical pathological types of cutaneous melanoma, because it does not have sun exposure as a predisposing factor. There are few publications in the literature addressing the expression of tumor suppressor pathway proteins in ALM. For these reasons we proposed to carry out a retrospective study aiming at a better understanding of p16 INK4a /cyclin D1/CDK4/pRb pathway involvement in ALM lesions. METHODS: Expression of p16, cyclin D1, CDK4 and pRB in 32 ALM lesions was displayed by immunohistochemistry. We compared the frequency of the expression of these markers in thin (thickness up to 1.0 mm) and thick lesions (thickness above 1.0 mm), as well as in accordance with the presence or absence of ulceration. RESULTS: 76% of the tumors were positive to p16 and 81% of the cases were positive to CDK4. There was no significant difference between the frequency of the expression of each marker when comparing in accordance with the thickness (thin and thick lesions) and the presence of ulceration. CONCLUSIONS: The expression of cyclin D1 and CDK4 in acral lentiginous melanoma may be considered aberrant and as reflex of a probable alteration in the p16/cyclin D1/CDK4/pRb tumor suppressor pathway. p16 and pRb tumor expression displayed by immunohistochemistry may not reflect probable alterations of the tumor suppressor pathway studied. The expression of these proteins, per se, may not be considered as prognostic factor in acral lentiginous melanoma

No Brasil, como no mundo, o carcinoma epidermóide bucal está entre os dez tipos mais comum de câncer e acomete mais de 13 mil pessoas por ano. Apesar de ser um sério problema devido a sua morbidade e mortalidade, alguns casos desta doença têm um comportamento biológico menos agressivo. A proteína ciclina D1, depois que forma complexos com as proteínas CDK4 e CDK6, tem como principal função fosforilar a proteína Retinoblastoma. Após sua fosforilação, a proteína libera um fator de transcrição, o E2F, que leva a célula à progressão da fase G1 para fase S do ciclo celular. A proteína c-jun, que faz parte do fator de transcrição AP-1, tem participação ativa no ciclo celular, principalmente durante a transcrição da fase G0 a G1. O gene retinoblastoma é um supressor de tumor. Este gene codifica uma fosfoproteína nuclear, que recebe o mesmo nome. Essa proteína regula o ciclo celular através de múltiplas funções. Também regula outros processos que afetam a proliferação celular, a diferenciação terminal e a apoptose. O HPV é um vírus de DNA que é encontrado em vários tipos de câncer e é o principal agente etiológico do carcinoma de colo uterino. Este trabalho comparou a expressão das proteínas ciclina D1, c-jun e do retinoblastoma em carcinomas epidermóides de baixo e alto grau de malignidade e tentou analisar se o HPV é um fator etiológico desta neoplasia. Apesar das lesões de baixo grau de malignidade expressarem as proteínas num maior número de células que as lesões de alto grau, só houve diferença estatística, entre os dois grupos estudados, para a proteína do retinoblastoma. Não foi encontrado o DNA do HPV em nenhum dos casos estudados. De acordo com este trabalho e com a literatura, a proteína do retinoblastoma é expressa em um número menor de células em carcinomas epidermóides bucais mais agressivos e o HPV não é um agente etiológico de todos os casos desta doença / In Brazil, as in the world, the oral squamous cell carcinoma is among the ten more common types of cancer e affects more than 13 thousand of people by year. Even though it is a serious problem due to its morbidity and mortality, some cases of this disease have a less aggressive biological behavior. The cyclin D1 protein after it forms complexes with the CDK4 and CDK6 proteins has as main function phosphorylate the Retinoblastoma protein. After its prosphorylation, the protein releases a transcription factor, the E2F, that leads the cell to the progression from the phase G1 to the phase S of the cell cycle. The c-jun protein, that is part of the transcription factor AP-1, has active participation in the cell cycle, mainly during the transcription from the phase G0 to G1. The retinoblastoma gene is a tumour suppressor. This gene codifies a nuclear phosphoprotein that receives the same name. This protein regulates the cell cycle through multiple functions. It also regulates other processes that affect the cell proliferation, the terminal differentiation and apoptosis. The HPV is a DNA virus that is found in many types of cancer and is the main etiological agent of the cervical cancer. This study compared the protein expression of the cilin D1, c-jun and retinoblastoma in low and high grade squamous cell carcinoma and tried to analyze if the HPV is a etiological factor for this neoplasm. In spite of the low grade of malignancy lesions express the protein in a greater number of cells than in the high grade lesion, there only was statistical difference, among the two studied groups, for the retinoblastoma protein. It was not found DNA of the HPV in any of the studied cases. According with this study and with the literature the retinoblastoma protein is expressed in a lower number of cells in the more aggressive oral squamous cell carcinomas and the HPV is not the etiological agent in all of the cases of this disease.

No Brasil, como no mundo, o carcinoma epidermóide bucal está entre os dez tipos mais comum de câncer e acomete mais de 13 mil pessoas por ano. Apesar de ser um sério problema devido a sua morbidade e mortalidade, alguns casos desta doença têm um comportamento biológico menos agressivo. A proteína ciclina D1, depois que forma complexos com as proteínas CDK4 e CDK6, tem como principal função fosforilar a proteína Retinoblastoma. Após sua fosforilação, a proteína libera um fator de transcrição, o E2F, que leva a célula à progressão da fase G1 para fase S do ciclo celular. A proteína c-jun, que faz parte do fator de transcrição AP-1, tem participação ativa no ciclo celular, principalmente durante a transcrição da fase G0 a G1. O gene retinoblastoma é um supressor de tumor. Este gene codifica uma fosfoproteína nuclear, que recebe o mesmo nome. Essa proteína regula o ciclo celular através de múltiplas funções. Também regula outros processos que afetam a proliferação celular, a diferenciação terminal e a apoptose. O HPV é um vírus de DNA que é encontrado em vários tipos de câncer e é o principal agente etiológico do carcinoma de colo uterino. Este trabalho comparou a expressão das proteínas ciclina D1, c-jun e do retinoblastoma em carcinomas epidermóides de baixo e alto grau de malignidade e tentou analisar se o HPV é um fator etiológico desta neoplasia. Apesar das lesões de baixo grau de malignidade expressarem as proteínas num maior número de células que as lesões de alto grau, só houve diferença estatística, entre os dois grupos estudados, para a proteína do retinoblastoma. Não foi encontrado o DNA do HPV em nenhum dos casos estudados. De acordo com este trabalho e com a literatura, a proteína do retinoblastoma é expressa em um número menor de células em carcinomas epidermóides bucais mais agressivos e o HPV não é um agente etiológico de todos os casos desta doença / In Brazil, as in the world, the oral squamous cell carcinoma is among the ten more common types of cancer e affects more than 13 thousand of people by year. Even though it is a serious problem due to its morbidity and mortality, some cases of this disease have a less aggressive biological behavior. The cyclin D1 protein after it forms complexes with the CDK4 and CDK6 proteins has as main function phosphorylate the Retinoblastoma protein. After its prosphorylation, the protein releases a transcription factor, the E2F, that leads the cell to the progression from the phase G1 to the phase S of the cell cycle. The c-jun protein, that is part of the transcription factor AP-1, has active participation in the cell cycle, mainly during the transcription from the phase G0 to G1. The retinoblastoma gene is a tumour suppressor. This gene codifies a nuclear phosphoprotein that receives the same name. This protein regulates the cell cycle through multiple functions. It also regulates other processes that affect the cell proliferation, the terminal differentiation and apoptosis. The HPV is a DNA virus that is found in many types of cancer and is the main etiological agent of the cervical cancer. This study compared the protein expression of the cilin D1, c-jun and retinoblastoma in low and high grade squamous cell carcinoma and tried to analyze if the HPV is a etiological factor for this neoplasm. In spite of the low grade of malignancy lesions express the protein in a greater number of cells than in the high grade lesion, there only was statistical difference, among the two studied groups, for the retinoblastoma protein. It was not found DNA of the HPV in any of the studied cases. According with this study and with the literature the retinoblastoma protein is expressed in a lower number of cells in the more aggressive oral squamous cell carcinomas and the HPV is not the etiological agent in all of the cases of this disease.

Previous studies suggest that the IGF-I stimulates the elastin gene transcription through the unique responsive sequence on the elastin promoter, which is a putative retinoblastoma control element (RCE). This site interacts with (Sp)-family transcription factors whose delivery is mediated by the retinoblastoma protein (Rb). Since Rb (phosphorylated on serine 780) has been implicated in the initiation of the cell cycle, we speculated that a different phosphorylation of Rb might determine Rb involvement in elastogenesis. Obtained results demonstrated that, IGF-I-induced elastogenic signaling pathway in human dermal fibroblasts includes activation of cyclinE/cdk2 causing a site specific phosphorylation of Rb on threonine 821. This permits the sequestration of Sp1 by Rb before it could bind the elastin promoter, thereby allowing the elastin gene transcription. We also found that blocking of H-Ras in Costello syndrome fibroblasts (characterized by heightened proliferation and impaired elastogenesis), selectively down-regulated Rb phosphorylation on serine 780 and normalized impaired elastogenesis.