Lesiones metastásicas a glándula tiroides. Una serie de casos / Metastatic lesions to the thyroid gland. Case series

Las lesiones metastásicas representan hasta un 3 % de los tumores malignos de la glándula tiroides. La mayoría de los casos se originan de tumores de células renales y de pulmón. El abordaje diagnóstico implica una alta sospecha clínica en pacientes con primarios conocidos, sin embargo, puede ser la manifestación inicial de una enfermedad maligna extensa no diagnosticada hasta en un 20 % a 40 % de los pacientes. La biopsia por aguja fina ha demostrado buen rendimiento para el diagnóstico de los nódulos metastásicos. El pronóstico y la opción del tratamiento quirúrgico dependen del control local del primario y del estado de la enfermedad sistémica asociada, por lo tanto, debe ser individualizado. Por lo general, hasta un 80 % de los pacientes con compromiso de la tiroides tienen enfermedad metastásica multiorgánica, y la intención del tratamiento quirúrgico es con fines paliativos para prevenir las complicaciones derivadas de la extensión local de la enfermedad a las estructuras del tracto aerodigestivo superior en el cuello. Se presenta a continuación, una serie de seis casos de pacientes con lesiones metastásicas a glándula tiroides con primarios en riñón, mama y de melanomas

Metastatic lesions represent up to 3% of malignant tumors of the thyroid gland. Most cases originate from lung and renal cell tumors. The diagnostic approach implies a high clinical suspicion in patients with known primaries, however, it can be the initial manifestation of an extensive undiagnosed malignant disease in up to 20% to 40% of patients. Fine-needle biopsy has shown good performance for the diagnosis of metastatic nodules. The prognosis and the option of surgical treatment depend on the local control of the primary condition and the state of the associated systemic disease, therefore it must be individualized. In general, up to 80% of patients with thyroid involvement have multi-organ metastatic disease and surgical treatment is intended to be palliative to prevent complications resulting from local extension of the disease to structures of the upper aerodigestive tract in the neck. A case series of six patients with metastatic lesions to the thyroid gland with primaries in the kidney, breast and melanomas is presented below

Morphine promotes migration and lung metastasis of mouse melanoma cells

Abstract Background Morphine is an analgesic agent used for cancer pain management. There have been recent concerns that the immunosuppressant properties of morphine can also promote cancer metastasis. Morphine is an agonist for toll like receptor 4 (TLR4) that has a dual role in cancer development. The promotor or inhibitor role of morphine in cancer progression remains controversial. We investigated the effects of morphine on migration and metastasis of melanoma cells through TLR4 activation. Methods Mouse melanoma cells (B16F10) were treated with only morphine (0, 0.1, 1, and 10 μM) or in combination with a TLR4 inhibitor (morphine10 μM +CLI-095 1μM) for either 12 or 24 hours. Migration of cells was analyzed by transwell migration assays. Twenty C57BL/6 male mice were inoculated with B16F10 cells via the left ventricle of the heart and then randomly divided into two groups (n = 10 each) that received either morphine (10 mg.kg−1, sub-q) or PBS injection for 21 days (control group). Animals were euthanized and their lungs removed for evaluation of metastatic nodules. Results Morphine (0.1, 1, and 10 μM) increased cell migration after 12 hours (p < 0.001) and after 24 hours of treatment with morphine (10 μM) (p < 0.001). Treatment with CLI-095 suppressed migration compared to cells treated with morphine alone (p < 0.001). Metastatic nodules in the morphine-treated group (64 nodules) were significantly higher than in the control group (40 nodules) (p < 0.05). Conclusion Morphine increases the migration and metastasis of mouse melanoma cells by activating TLR4.

Ultrassonografia de alta frequência e Doppler no diagnóstico, caracterização e seguimento dos tumores cutâneos / High-frequency ultrasonography and Doppler in the diagnosis, characterization and follow-up of skin tumors

A ultrassonografia (US) é uma ferramenta diagnóstica amplamente utilizada na medicina há mais de 50 anos. Na dermatologia, o uso desta técnica vem crescendo, principalmente após o desenvolvimento de dispositivos de maior frequência (acima de 20 MHz), que possibilitaram a avaliação mais detalhada da pele e seus anexos. Apesar da ampla gama de informações que podem ser obtidas através deste exame e da necessidade de métodos não invasivos que auxiliem no diagnóstico, planejamento terapêutico e seguimento dos tumores cutâneos, a falta de uniformidade nos critérios avaliados e parâmetros ultrassonográficos ainda dificulta o uso rotineiro desta técnica de imagem na oncologia cutânea. Objetivos: Descrever as características ultrassonográficas dos tumores cutâneos no modo B e Doppler; comparar com técnicas já estabelecidas, dermatoscopia e histologia; e avaliar características morfológicas no modo B e da vascularização ao Doppler que possam diferenciar nevos e melanomas. Material e métodos: Análise prospectiva de lesões suspeitas de câncer de pele de pacientes provenientes do A.C.Camargo Cancer Center. As lesões foram submetidas à, dermatoscopia, ultrassonografia de alta frequência (USAF) e Doppler para posterior correlação com parâmetros anatomopatológicos. Resultados: Foram incluídas 289 lesões, sendo 105 carcinomas basocelulares (CBC), 35 carcinomas espinocelulares (CEC), 24 melanomas, 2 carcinomas de células de Merkel, 15 tumores malignos mais raros e metástases cutâneas de outros tumores e 108 lesões benignas. Todas as lesões malignas foram visualizadas à US com 24 MHz. As principais características ultrassonográficas dos tumores malignos foram: hipoecogenicidade (98.9%) e ecotextura homogênea (90.6%). A presença de vascularização foi mais frequente nas lesões malignas e este foi o parâmetro mais relevante para diferenciação entre malignas e benignas. Conclusão: A USAF demonstrou ser uma ferramenta bastante útil na avaliação dos tumores cutâneos, apoiando no diagnóstico, estadiamento, avaliação de resposta e seguimento. O diagnóstico diferencial entre os principais tumores cutâneos ainda não é possível utilizandose somente a USAF, sendo, portanto, a correlação com dermatoscopia e histopatologia fundamental para o diagnóstico definitivo. Não foram observadas características ultrassonográficas capazes de diferenciar com segurança nevos de melanomas em nossa amostra.

Ultrasound (US) is a diagnostic tool that has been widely used in medicine for over 50 years. In dermatology, the use of this technology is increasing, mainly after the development of higher frequency devices (above 20 MHz), which allow a more detailed assessment of the skin and its annexes. Despite the wide range of information that can be obtained through this exam and the need for non-invasive methods that help in the diagnosis, therapeutic planning and follow-up of skin tumors, the lack of uniformity in the assessment criteria and ultrasound parameters still hinders the routine use of this imaging technique in cutaneous oncology. Objectives: To describe the sonographic characteristics of cutaneous tumors in B-mode and Doppler and to compare them with dermoscopy and histology; and to evaluate morphological characteristics in B-mode and Doppler ultrasound that allow differentiating nevi and melanomas. Material and methods: Prospective analysis of patients with skin lesions suspicious for cancer from the A.C.Camargo Cancer Center. The lesions were assessed by dermoscopy, high-frequency ultrasound (24 MHz) and Doppler and then correlated with histology. Results: 289 lesions were included, of which 105 were basal cell carcinomas, 35 squamous cell carcinomas, 24 melanomas, 2 Merkel cell carcinomas, 15 less common malignant tumors and cutaneous metastases from other tumors and 108 benign lesions. All malignant lesions were visualized at 24 MHZ ultrasonography. The main sonographic characteristics of malignant tumors were: hypoechogenicity (98.9%) and homogeneous echotexture (90.6%) Vascularization was more frequent in malignant lesions and this was the most relevant parameter for differentiating malignant and benign lesions. Conclusion: High-frequency ultrasound proved to be a very useful tool in the assessment of skin tumors, supporting diagnosis, staging, assessment of therapeutic response and follow-up. It's still not possible differentiate the main skin tumors based only on sonographic criteria, therefore, the correlation with dermoscopy and histology is necessary for the definitive diagnosis. No sonographic features that could reliably differentiate nevi and melanomas were found in our sample

Ganglio centinela positivo en melanoma: factores predictivos y resultados del vaciamiento ganglionar / Positive sentinel lymph node in melanoma: predictive factors and results of completion lymph node dissection

RESUMEN Antecedentes: la biopsia del ganglio centinela (GC) es la técnica aceptada para determinar el pronóstico en estadios iniciales de melanoma cutáneo. La ventaja del vaciamiento ganglionar (VG) cuando el GC resulta positivo ha sido recientemente cuestionada. Objetivo: describir los porcentajes y factores asociados a metástasis en el GC, y en los ganglios no centinela (GnC) en los VG de pacientes con GC positivo. Material y métodos: se llevó a cabo un estudio retrospectivo de los registros clínicos y patológicos de 139 pacientes operados por melanoma cutáneo entre enero de 2012 y diciembre de 2019. Resultados: a 96 (69%) pacientes se les realizó biopsia de GC. El promedio de edad fue 61,7 años ± 17,5 (19-93); 53 (55,2%) fueron hombres. La lesión primaria estuvo ubicada en: extremidades 47 (49%), tronco 39 (40,6%), cabeza y cuello 10 (10,4%). El promedio de espesor de Breslow fue 5,01 mm (1,05- 50 mm) y se encontró ulceración en 35 casos (36,4%). El GC fue identificado en todas las oportunidades y en 39 (40,6%) fue positivo. Hubo asociación con el espesor ≥ 3 mm (p = 0,000017) y con la ulceración (p = 0,0011). A los pacientes con GC positivo se les efectuó el VG del territorio afectado: 23 axilar, 10 inguinal y 6 cervical. Veintitrés (59%) presentaron metástasis en GnC. Se asoció con el espesor (p = 0,022) y la ulceración (p = 0,019). Conclusión: existió un alto porcentaje de GnC positivos en la población estudiada, vinculado al espesor y la ulceración. Estas características, así como la dificultad de un estricto seguimiento, inducen a no abandonar el VG en pacientes con GC positivo.

ABSTRACT Background: Sentinel lymph node (SLN) biopsy is the technique accepted to determine the prognosis of early cutaneous melanomas. The advantage of lymph node dissection (LND) when SLN biopsy is positive has recently been questioned. Objective: The aim of this study is to describe the percentages and factors associated with SLN and non-sentinel node (NSN) metastases in LNDs of SLN-positive patients. Material and methods: The clinical records and pathology reports of 139 patients undergoing surgery for cutaneous melanoma between January 2012 and December 2019 were retrospectively reviewed. Results: Ninety-six (69%) patients underwent SLN biopsy. Mean age was 61.7 ± 17.5 years (19-93) and 53 (55.2%) were men. The primary lesion was located in the extremities in 47 (49%) cases, in the trunk in 39 (40.6%), and in the head and neck in 10 (10.4%). Mean Breslow thickness was 5.01 mm (1.05-50 mm) and ulceration was found in 35 cases (36.4%). The SLN was identified in all the cases and was positive in 39 (40.6%). There was an association with thickness ≥ 3 mm (p = 0.000017) and ulceration (p = 0.0011). Those patients with positive SLN biopsy underwent LND of the territory involved: axillary in 23, inguinal in 10 and cervical in 6. Twenty-three (59%) presented NSLN metastases and were associated with thickness (p = 0.022) and ulceration (p = 0.019). Conclusion: There was a high percentage of positive NSLN in the population studied which was associated with thickness and ulceration. These characteristics and the difficulty to achieve strict follow-up are the reasons for completion LND in SLN-positive patients.

Aspectos clínicos do câncer durante o período gestacional: desafios diagnósticos e terapêuticos / Clinical aspects of cancer during the gestational period: diagnostic and therapeutic challenges

O câncer é uma das maiores causas de morte em mulheres na idade reprodutiva e ocorre em aproximadamente 0,05% a 0,1% das gestações. Os cânceres ginecológicos, de mama, tireoide e hematológicos são os mais comuns na gravidez. O obstetra é o principal médico para investigar sintomas que podem estar relacionados à malignidade. O diagnóstico pode ser dificultado devido à sobreposição de sintomas da gravidez, como náusea, vômitos, aumento do útero e das mamas, dor abdominal, além da limitação para uso de exames de imagem e alterações comuns em exames laboratoriais. O risco e o benefício do diagnóstico e o tratamento para o bem-estar materno e fetal devem ser avaliados com cuidado pelos profissionais envolvidos. Este artigo tem como objetivo realizar uma revisão sobre quando suspeitar e como investigar os principais cânceres na gestação.(AU)

Cancer is the major cause of death in women on reproductive age and occurs in approximately 0.05% to 0.1% of pregnancies. Gynecological, breast, thyroid and hema- tological cancers are the most common in pregnancy. The obstetrician is the primary physician to investigate symptoms that may be related to malignancy. The diagnosis can be difficult due to the overlap of pregnancy symptoms, such as nausea, vomiting, enlargement of the uterus and breasts, abdominal pain, in addition to the limitation for the use of imaging tests and common changes in laboratory tests. The risk and be- nefit of diagnosis and treatment for maternal and fetal well-being should be carefully assessed by the professionals involved. This article aims to conduct a review on when to suspect and how to investigate the main cancers in pregnancy.(AU)

Retrospective study of cutaneous neoplasms in horses treated at the Livestock Development Center of the Federal University of Bahia / Estudo retrospectivo das neoplasias cutâneas em equinos atendidos no Centro de Desenvolvimento da Pecuária da Universidade Federal da Bahia

The aim of this study was to determine the prevalence of cutaneous neoplasms in horses treated at the Center for the Development of Livestock at the Federal University of Bahia, as well as to correlate it with the coat color, breed, and age of the animal. For that, the attendance records for the last ten years were reviewed. When evaluating the files, 13 cases of cutaneous tumor in horses confirmed by histopathology and cytology were observed. The most prevalent skin tumors were sarcoid (38.5%), melanoma (23%), and fibrosarcoma (15.4%). Regarding the equine coat color, gray and sorrel horses were the most frequent with 30.7% and 23.1% of cases, respectively. As for the equine breed, the mangalarga marchador was the most prevalent (38.4%). Regarding age, 38.46% of the horses were up to 5 years old, 30.77% of the animals were between 4 and 10 years old, and 30.76% were between 11 and 16 years old. In the end, it can be concluded that sarcoid and melanoma were the most prevalent neoplasms.

Objetivou-se com este trabalho determinar a prevalência de neoplasias cutâneas em equinos atendidos no Centro de Desenvolvimento da Pecuária da Universidade Federal da Bahia, bem como correlaciona-la com a pelagem, raça e idade do animal. Para tanto revisou-se as fichas de atendimento dos últimos dez anos. Ao avaliar as fichas, observou-se 13 casos de tumor cutâneo em equinos confirmado por histopatologia ou citologia. Os tumores cutâneos mais prevalentes foram sarcoide (38,5%), melanoma (23%) e fibrossarcoma (15,4%). Com relação a pelagem, equinos tordilhos e alazões foram os mais frequentes com 30,7% e 23,1% dos casos, respectivamente. Quanto as raças, a mangalarga marchador foi a mais prevalente (38,4%). Em relação a idade, 38,46% dos equinos possuíam até 5 anos de idade, 30,77% dos animais apresentavam idade entre 4 e 10 anos e, 30,76% apresentavam idade entre 11 e 16 anos. Ao fim, pode-se concluir que o sarcoide e o melanoma foram as neoplasias mais prevalentes.

Actualización en fotoprotección / Update in photoprotection

Hoy en día entre las principales patologías por las que muchos pacientes acuden a servicios de dermatología suelen encontrarse las fotodermatosis o enfermedades de la piel causadas por exposición aguda o crónica a la radiación ultravioleta. Tal vez uno de los motivos de la elevada incidencia de estas enfermedades se debe al desarrollo de diferentes actividades cotidianas bajo el sol, sin la protección adecuada, ya que gran parte de la población todavía no ha tomado conciencia del daño que puede ocasionar el sol en la piel; en su papel como exacerbante de entidades como el lupus o diferentes cuadros de alergia; así como desencadenante del cáncer de piel. Al respecto cada año se producen más de 130.000 casos de melanoma en todo el mundo (estimaciones de la Organización Mundial de la Salud); y entre 2 a 3 millones de cánceres de piel que no son tipo melanoma. Mientras los cánceres de piel no-melanomas tienen fácil tratamiento quirúrgico y raramente son malignos; los melanomas (cánceres de piel por excesiva exposición a la radiación ultravioleta) son muy agresivos, los tratamientos no son muy eficaces a largo plazo, y la mortalidad es relativamente elevada. A lo anteriormente mencionado se suma la situación de confinamiento por la pandemia de COVID-19 debido a la cuál hemos estado en un periodo prolongado de baja exposición solar más larga de lo habitual, teniendo como consecuencia fisiopatológica bajos niveles de síntesis de vitamina D corporal, con un incremento en el riesgo de infecciones, incluida la del coronavirus. De igual manera procesos de protección solar como la melanogénesis dependientes del impacto de la radiación ultravioleta (UV) en la piel frenada por el mismo confinamiento, así como la reducción en la hiperqueratosis e hiperplasia epidérmica, como respuesta gradual de aclimatación lumínica de la piel, han traído como consecuencia un mayor daño por la mayor sensibilidad frente a las irradiancias solares más energéticas y agresivas (UV), con una piel sin período de adaptación, lo que podría repercutir a largo plazo en una mayor incidencia de cáncer de piel. De esta forma tenemos que considerar que en el periodo de confinamiento por la COVID-19 las medidas de foto protección habituales cobran especial importancia, por lo que es recomendable exposiciones solares directas, diarias, breves, para aumentar los niveles séricos de vitamina D3, además de lograr los beneficios que aportan la exposición solar a nuestro estado de ánimo, a través de la estimulación de neurotransmisores involucrados en los mecanismos de bienestar emocional(afectado también por el confinamiento). Por tales motivos el objetivo de la presente revisión será brindar una actualización en lo referente a tipos de fotoprotectores, su uso; y sobre las recomendaciones en foto prevención que deben inculcarse en nuestra población.

Diagnostic and prognostic significance of long noncoding RNA LINC00173 in patients with melanoma

SUMMARY OBJECTIVE: A growing volume of literature has suggested long noncoding RNAs (lncRNAs) as important players in tumor progression. In this study, we aimed to investigate the expression and prognostic value of lncRNA LINC00173 (LINC00173) in melanoma. METHODS: LINC00173 expression was measured in 163 paired cancerous and noncancerous specimen samples by real-time polymerase chain reaction. The correlations between LINC00173 expression with clinicopathological characteristics and prognosis were analyzed by chi-square test, log-rank test, and multivariate survival analysis. Receiver-operating characteristic curves were used for the assessment of the diagnostic value of LINC00173 for melanoma patients. RESULTS: The expression level of LINC00173 in melanoma specimens was distinctly higher than that in adjacent non-neoplasm specimens (p<0.01). Besides, LINC00173 was expressed more frequently in patients with advanced melanoma than in patients with early melanoma. Multivariate assays confirmed that LINC00173 expression level was an independent prognostic predictor of melanoma patients (p<0.05). CONCLUSION: Our data indicated that LINC00173 expression could serve as an unfavorable prognostic biomarker for melanoma patients.

Fotoenvejecimiento cutáneo y su relación con el cáncer de piel: revisión sistemática / Cutaneous photoaging and its association with skin cancer: a systematic review

Introducción. Un factor fundamental en el envejecimiento de la piel es la exposición acumulativa a la luz solar. Al avanzar el proceso de fotoenvejecimiento, el colágeno y las fibras elásticas experimentan mayores daños, generando patologías asociadas a cáncer. Se plantea como objetivo de esta revisión, analizar la evidencia que existe en relación a las diferentes situaciones del fotoenvejecimiento cutáneo y su asociación con el cáncer de piel. Metodología. Se elaboró una revisión sistemática, exploratoria, a través de una búsqueda bibliográfica en gestores de bases de datos en salud. Se seleccionaron 18 artículos completos entre los años 2019 y 2020. Resultados. Se agruparon los hallazgos de los artículos en dos matrices de información, la primera que indica los temas principales del fotoenvejecimiento y la relación con los factores asociados a cáncer de piel, y una segunda matriz que presenta la interrelación de la asociación con la conclusión más relevante del artículo. Conclusiones. El fotoenvejecimiento es un problema que crece día a día impactando en la vida del ser humano, creando repercusiones a nivel físico y mental que afectan la calidad de vida. Es necesario crear conciencia acerca de los mecanismos de fotoprotección, como el uso de bloqueadores solares, antioxidantes, evitar la luz solar y uso de lentes solares, entre otros; y, en los casos que ya exista algún daño por fotoenvejecimiento, elegir la mejor alternativa e individualizar cada paciente según su tipo de piel y enfermedades asociadas con la edad, para un mejor enfoque de tratamiento, y así impactar de manera positiva en el crecimiento exponencial que existe hoy en día en diversos tipos de cáncer de piel y lesiones precancerosas

Introduction. A fundamental factor in skin aging is cumulative exposure to sunlight. As the photoaging process progresses, collagen and elastic fibers experience greater damage, generating pathologies associated with cancer. The aim of this review is to analyze the evidence that exists in relation to the different situations of cutaneous photoaging and its association with skin cancer. Methodology. A systematic, exploratory review was prepared through a bibliographic search in database managers in health. A total of 18 articles were selected between the years 2019 and 2020. Results. The articles information were grouped into two matrix, the first one that indicates the main themes of photoaging and the relationship with the factors associated with skin cancer, and a second matrix that presents the interrelationship of these factors with the most relevant conclusion of the study. Conclusions. Photoaging is a problem that grows day by day impacting on human life, creating repercussions on a physical and mental level affecting the quality of life. It is necessary to raise awareness about photoprotection mechanisms such as the use of sunscreen, antioxidants , avoid sunlight, use of solar lenses, among others; and in cases where there is already some damage caused by photoaging, choose the best alternative and individualize each patient according to their skin type and diseases associated with age, for a better treatment approach, and thus positively impact the exponential growth that exists today in various types of skin cancer and precancerous lesions

Leucemia linfocítica crônica associada a melanoma metastático em homem de 53 anos / Metastatic melanoma in a 53-year-old male with relapsed chronic lymphocytic leukemia

Introdução: A associação de leucemia linfocítica crônica (LLC) e melanoma tem sido estudada nos últimos anos. Acredita-se que a imunossupressão causada pelo tratamento da doença seja o fator de risco mais importante para o aumento da susceptibilidade ao desenvolvimento e disseminação do câncer de pele. Relato de Caso: Este relato de caso descreve homem de 53 anos, em tratamento de leucemia linfocítica crônica desde 2018, já submetido a diversos ciclos de quimioterapia com fludarabina. Apresentou histórico de exérese de melanoma nodular epitelioide no couro cabeludo em 2019, removido com margens livres. Um ano após a cirurgia, paciente evoluiu com piora do estado geral com necessidade de hospitalização. Investigação adicional revelou focos de metástase em pulmões, fígado, rins, estômago, sistema nervoso central e linfonodos. Análise histopatológica foi positiva para melanoma. A possibilidade de tratamento foi descartada pela equipe de oncologia, que sugeriu cuidados paliativos. Discussão: Um dos mecanismos mais discutidos para explicar esta associação de neoplasias é a imunossupressão resultante do tratamento da LLC, que deixa o paciente suscetível ao desenvolvimento e à disseminação do melanoma. Além disso, a fludarabina, quimioterápico geralmente usado para remissão da LLC, é conhecida por depletar células T-helper e tem sido descrita como cofator deste processo. A associação de leucemia e melanoma cutâneo têm sido descrita nos últimos anos, porém não há nenhum protocolo de tratamento para esta condição.

BACKGROUND: The association of Chronic Lymphocytic Leukemia (CLL) and melanoma have been studied in the last years. The immunosuppression caused by the treatment of CLL seems to be the major factor of increasing patients' susceptibility to the development and spread of skin cancer. CASE REPORT: This case report describes a 53-year-old male patient, in CLL treatment since 2018, already submitted to many cycles of chemotherapy with fludarabine. History of an exeresis of epithelioid nodular melanoma of the scalp in 2019, which was removed with a clear margin. One year later, he presented with a poor general condition with hospitalization indication. Additional investigation revealed metastatic lesions in lungs, liver, kidneys, stomach, central nervous system, and lymph nodes. Histopathologic analysis positive for melanoma. The possibility of treatment was discarded by the Oncology team, which suggested palliative care. DISCUSSION: One of the most discussed mechanisms to explain this cancer association is the immunosuppression developed during the treatment of CLL, increasing patients' susceptibility to the development and spread of melanoma. In addition, the use of fludarabine, a chemotherapy commonly used in relapsed CLL, is known to deplete T-helper cells and has been described as a cofactor of this process. The association of leukemia and cutaneous melanoma has been reported in the last years, yet there is no surveillance protocol.

Application of CRISPR-Cas9 to interrogate novel gene functions in cutaneous melanoma

Melanoma accounts for 3% of skin neoplasms and is the leading cause of death from skin disorders worldwide. The high mortality rate associated with this disease stems from the high capacity of melanoma patients to develop metastases and treatment relapse with inhibitors of the MAPK signaling pathway (such as BRAF inhibitors), commonly used in melanoma therapy. Thus, the investigation of genes involved in the mechanisms of melanoma development is essential for new and more effective therapeutic strategies. Hence, we describe in this thesis two projects involving the genes SIN3B and IRF4 as possible biomarkers for cutaneous melanoma. Initially, through bioinformatics analyses performed by our group, an upregulation of SIN3B was found in metastatic melanomas. This result together with the understanding of SIN3B role in regulating gene expression and oncogenic transformation, prompted us to describe in this thesis some mechanisms by which SIN3B may influence melanoma development. We then sought to characterize the gene function using SIN3B-deleted cells, generated by the CRISPR-Cas9 methodology. Initially, we observed increased SIN3B expression in BRAF-mutant metastatic melanomas, where we noted that the long splicing variant of the gene (NM_001297595.1) was effectively prevalent in melanomas. Subsequently, we designed gRNAs between the exons 2 and 3 of the human SIN3B gene and engineered three knockout clones and three control clones (containing empty lentiCRISPRv2 plasmid) from different melanoma cell lines (SKMEL28, A2058, and A375). Through functional analyses, it was observed that the absence of the gene did not interfere in the proliferation of tumor cells; however, it led to a decrease in invasive properties. These results were verified by Boyden chamber assays and transcriptome analysis (total RNA sequencing of deleted cells), where a decrease in migration and motility pathways was observed. Additionally, a screening of synthetically lethal genes with SIN3B was performed with a genome wide CRISPR library. These results showed that USP7 and STK11 genes, which belong to the FoxO signaling pathway, were essential in SIN3B-depleted melanoma cells. Finally, through a collaborative project with the Wellcome Trust Sanger Institute, previous large-scale sequencing analyses demonstrated that deletion of the IRF4 gene was lethal for melanoma cells. Accordingly, we performed IRF4 silencing in vitro and noticed that the lack of IRF4 promotes cell death and apoptosis, independently of MYC and MITF, known in the literature to be downstream targets of this gene. Therefore, these data suggest that IRF4 plays a vital role in melanoma cell survival. Taken together, both works herein described in this thesis demonstrate how CRISPR-Cas9 can be applied to study the functions and mechanisms of genes involved in melanoma progression, collectively helping in the development of more effective therapeutic strategies for this tumor

O melanoma representa 3% dos tipos de neoplasias cutâneas e é a maior causa das mortes por distúrbios de pele no mundo. A alta taxa de mortalidade associada à essa doença advém da alta capacidade de pacientes com melanoma desenvolverem metástases, e apresentarem recidiva após tratamento com inibidores da via de sinalização MAPK (como da proteína BRAF), comumente utilizados no tratamento de pacientes metastáticos. Assim, a investigação de genes envolvidos nos mecanismos de desenvolvimento do melanoma é primordial para novas estratégias terapêuticas mais efetivas. Dessa forma, descrevemos no presente trabalho dois projetos envolvendo os genes SIN3B e IRF4 como possíveis biomarcadores para melanoma cutâneo. Em análises prévias de bioinformática realizados pelo nosso grupo, SIN3B foi identificado tendo maior expressão em melanomas metastáticos. Além disso, diversos estudos mostraram que o gene está envolvido na regulação da expressão gênica e transformação oncogênica. Dessa forma, descrevemos nessa tese alguns mecanismos pelos quais SIN3B pode influenciar no desenvolvimento do melanoma, através da caracterização funcional de células SIN3B-deletadas pela metodologia CRISPR-Cas9. Inicialmente, observamos aumento na expressão de SIN3B em melanomas metastáticos BRAF-mutados, onde notamos que a variante de splicing longa do gene (NM_001297595.1), era efetivamente prevalente em melanomas. Assim, desenhamos sequências de RNA guias entre os éxons 2 e 3 do gene SIN3B humano e, obtivemos três clones knockout e outros três clones controle (contendo plasmídeo vazio) em diferentes linhagens de melanoma (SKMEL28, A2058 e A375), para caracterização funcional. Observou-se que a ausência do gene não interferiu na proliferação das células tumorais, contudo, acarretou na diminuição de processos invasivos. Esses resultados foram averiguados através de ensaios em câmara de Boyden e análises de transcriptoma (sequenciamento de RNA total das células deletadas), onde notou-se diminuição das vias de migração e motilidade. Adicionalmente, um rastreamento de genes sinteticamente letais com SIN3B foi realizado com uma biblioteca de CRISPR capaz de silenciar todo o genoma. Esses resultados mostraram que os genes USP7 e STK11, ambos pertencentes à via de sinalização de FoxO, são essenciais nas células SIN3B deletadas. Por fim, através de um projeto colaborativo com o Wellcome Trust Sanger Institute, análises prévias de sequenciamento de larga escala demonstraram que a deleção do gene IRF4 era letal para células de melanoma. Dessa forma, realizamos o silenciamento de IRF4 in vitro e notamos que a ausência do gene promove morte celular e apoptose, independentemente de MYC e MITF, conhecidos na literatura por serem alvos downstream do gene. Portanto, esses dados sugerem que IRF4 tem um papel importante na sobrevivência de células de melanoma. Em conjunto, ambos trabalhos descritos nessa tese, demonstram como a metodologia CRISPR-Cas9 pode auxiliar no entendimento de processos importantes para a malignidade do melanoma e contribuir para estratégias terapêuticas mais efetivas para esse tumor

Estudo do composto STX140, um análogo do estradiol, em linhagens de melanoma humano / STX140 compound study in human melanoma

O câncer de pele pode ser classificado como não melanoma e melanoma. O melanoma apresenta baixa incidência entre os cânceres de pele, porém é a forma mais letal e é considerado um dos tipos mais resistentes ao tratamento. Devido à infiltração de células malignas nos tecidos, vasos linfáticos e vasos sanguíneos, o melanoma invade e se espalha rapidamente. Suas metástases são frequentemente localizadas em linfonodos, cérebro, fígado e outros órgãos. Melanomas metastáticos abrigam múltiplas mutações gênicas e muitos tumores apresentam resistência aos tratamentos, como por exemplo com inibidores BRAF, devido à mutações e ativação de vias paralelas. Ou seja, existe uma necessidade clara da busca de novas opções de tratamento. Em trabalho realizado por nosso grupo, Massaro et al mostraram que o derivado de estradiol 2- Metoxiestradiol induz apoptose em células de melanoma e senescência. Neste sentido, o composto STX140, (um análogo do estradiol com biodisponibilidade superior), que já se mostrou eficaz no combate ao câncer de mama em diversos estudos in vitro e in vivo, será então avaliado para sua ação no melanoma de forma inédita. Este trabalho teve como principal objetivo explorar a ação antitumoral em células de melanoma do composto STX140, especialmente a indução de senescência. Utilizando a cultura de células de melanoma foram realizados os ensaios de: viabilidade celular - IC50, formação de colônias, análise do ciclo celular e caracterização de morte celular por citometria de fluxo, ensaio In vitro scratch, coloração para ß-galactosidase, PCR quantitativo e ELISA. Os resultados mostraram que o composto STX140: diminui a viabilidade celular, inibe a proliferação, formação de colônias e migração em linhagens de melanoma (não resistentes e resistentes ao vemurafenibe, inibidor de BRAF). Além do mais, o composto atuou diminuindo a secreção da interleucina pró-tumoral IL-8 em células resistentes. O STX140 induziu senescência nas células de melanoma que foram positivas para ß-galactosidase, também havendo aumento da expressão de genes chave de vias de senescência (CDKN1A e GADD45A) nas células de melanoma resistentes tratadas com o composto. Em conclusão, o STX140 mostrou ter um potencial antitumoral contra o melanoma, diminuindo sua viabilidade celular, inibindo sua proliferação e migração, induzindo senescência, diminuindo a secreção de interleucina pró- tumoral, com efeito mais acentuado nas linhagens de melanoma resistente

Skin cancer can be classified as non-melanoma and melanoma. Melanoma has a low incidence among skin cancers, but it is the most lethal form and is considered one of the most resistant to treatment. Due to the infiltration of malignant cells into tissues, lymphatic vessels and blood vessels, melanoma invades and spreads rapidly. Its metastases are often located in lymph nodes, brain, liver and other organs. Metastatic melanomas presents multiple gene mutations and many tumors are resistant to treatments, such as with BRAF inhibitors, due to mutations and activation of parallel pathways. In other words, there is a clear need to search for new treatment options. In work carried out by our group, Massaro et al showed that the estradiol derivative 2- Methoxyestradiol induces apoptosis in melanoma cells and senescence. In this sense, the compound STX140, (an estradiol analogue with superior bioavailability), which has already been shown to be effective against breast cancer in vitro and in vivo studies will be then evaluated for its action on melanoma. The main objective of this work is to explore the antitumor action of the compound STX140 in melanoma cells, especially the induction of senescence. Using the melanoma cell culture the following assays were performed: cell viability - IC50, clonogenic, cell cycle analysis and cell death characterization by flow cytometry, wound assay, staining for ß-galactosidase, quantitative PCR and ELISA. Preliminary data from this work showed that the compound STX140: decreases cell viability, inhibits proliferation, colony formation and migration in melanoma cell lines (non-resistant and resistant to vemurafenib, BRAF inhibitor). It also decreased the secretion of pro-tumor interleukin IL-8 in resistant cells. STX140 induced senescence in melanoma cells, that were positive for ß-galactosidase, and there was also increased expression of key genes of senescence pathways (CDKN1A and GADD45A) in resistant melanoma cells treated with the compound. In conclusion, STX140 has been shown to have antitumor potential against melanoma, decreasing its cell viability, inhibiting its proliferation and migration, inducing senescence, decreasing pro-tumor interleukin secretion, with a more pronounced effect on resistant melanoma cell lines

Role and mechanism of Vγ4 T cells in impaired wound healing of rapamycin-induced full-thickness skin defects in mice / 中华烧伤杂志

Objective: To investigate the role and mechanism of Vγ4 T cells in impaired wound healing of rapamycin-induced full-thickness skin defects in mice. Methods: The experimental research methods were applied. Eighty-six C57BL/6J male mice (hereinafter briefly referred to as wild-type mice) aged 8-12 weeks were selected for the following experiments. Vγ4 T cells were isolated from axillary lymph nodes of five wild-type mice for the following experiments. Intraperitoneal injection of rapamycin for 42 mice was performed to establish rapamycin-treated mice model for the following experiments. Eighteen wild-type mice were divided into normal control group without any treatment, trauma only group, and trauma+CC chemokine ligand 20 (CCL20) inhibitor group according to the random number table (the same grouping method below), with 6 mice in each group. The full-thickness skin defect wound was made on the back of mice in the latter two groups (the same wound model below), and mice in trauma+CCL20 inhibitor group were continuously injected subcutaneously with CCL20 inhibitor at the wound edge for 3 days after injury. Another 6 rapamycin-treated mice were used to establish wound model as rapamycin+trauma group. On post injury day (PID) 3, the epidermal cells of the skin tissue around the wound of each trauma mice were extracted by enzyme digestion, and the percentage of Vγ4 T cells in the epidermal cells was detected by flow cytometry. In normal control group, the epidermal cells of the normal skin tissue in the back of mice were taken at the appropriate time point for detection as above. Five wild-type mice were used to establish wound models. On PID 3, the epidermal cells were extracted from the skin tissue around the wound. The cell populations were divided into Vγ4 T cells, Vγ3 T cells, and γδ negative cells by fluorescence-activated cell sorter, which were set as Vγ4 T cell group, Vγ3 T cell group, and γδ negative cell group (with cells in each group being mixed with B16 mouse melanoma cells), respectively. B16 mouse melanoma cells were used as melanoma cell control group. The expression of interleukin-22 (IL-22) mRNA in cells of each group was detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction (RT-PCR), with the number of samples being 6. Thirty rapamycin-treated mice were used to establish wound models, which were divided into Vγ4 T cell only group and Vγ4 T cell+IL-22 inhibitor group performed with corresponding injections and rapamycin control group injected with phosphate buffer solution (PBS) immediately after injury, with 10 mice in each group. Another 10 wild-type mice were taken to establish wound models and injected with PBS as wild-type control group. Mice in each group were injected continuously for 6 days. The percentage of wound area of mice in the four groups was calculated on PID 1, 2, 3, 4, 5, and 6 after injection on the same day. Six wild-type mice and 6 rapamycin-treated mice were taken respectively to establish wound models as wild-type group and rapamycin group. On PID 3, the mRNA and protein expressions of IL-22 and CCL20 in the peri-wound epidermis tissue of mice in the two groups were detected by real-time fluorescence quantitative RT-PCR and Western blotting, respectively. The Vγ4 T cells were divided into normal control group without any treatment and rapamycin-treated rapamycin group. After being cultured for 24 hours, the mRNA and protein expressions of IL-22 of cells in the two groups were detected by real-time fluorescence quantitative RT-PCR and Western blotting, respectively, with the number of samples being 6. Data were statistically analyzed with independent sample t test, analysis of variance for repeated measurement, one-way analysis of variance, Bonferroni method, Kruskal-Wallis H test, and Wilcoxon rank sum test. Results: The percentage of Vγ4 T cells in the epidermal cells of the skin tissue around the wound of mice in trauma only group on PID 3 was 0.66% (0.52%, 0.81%), which was significantly higher than 0.09% (0.04%, 0.14%) in the epidermal cells of the normal skin tissue of mice in normal control group (Z=4.31, P<0.01). The percentages of Vγ4 T cells in the epidermal cells of the skin tissue around the wound of mice in rapamycin+trauma group and trauma+CCL20 inhibitor group on PID 3 were 0.25% (0.16%, 0.37%) and 0.24% (0.17%, 0.35%), respectively, which were significantly lower than that in trauma only group (with Z values of 2.27 and 2.25, respectively, P<0.05). The mRNA expression level of IL-22 of cells in Vγ4 T cell group was significantly higher than that in Vγ3 T cell group, γδ negative cell group, and melanoma cell control group (with Z values of 2.96, 2.45, and 3.41, respectively, P<0.05 or P<0.01). Compared with that in wild-type control group, the percentage of wound area of mice in rapamycin control group increased significantly on PID 1-6 (P<0.01), the percentage of wound area of mice in Vγ4 T cell+IL-22 inhibitor group increased significantly on PID 1 and PID 3-6 (P<0.05 or P<0.01). Compared with that in rapamycin control group, the percentage of wound area of mice in Vγ4 T cell only group decreased significantly on PID 1-6 (P<0.05 or P<0.01). Compared with that in Vγ4 T cell only group, the percentage of wound area of mice in Vγ4 T cell+IL-22 inhibitor group increased significantly on PID 3-6 (P<0.05 or P<0.01). On PID 3, compared with those in wild-type group, the expression levels of IL-22 protein and mRNA (with t values of -7.82 and -5.04, respectively, P<0.01) and CCL20 protein and mRNA (with t values of -7.12 and -5.73, respectively, P<0.01) were decreased significantly in the peri-wound epidermis tissue of mice in rapamycin group. After being cultured for 24 hours, the expression levels of IL-22 protein and mRNA in Vγ4 T cells in rapamycin group were significantly lower than those in normal control group (with t values of -7.75 and -6.04, respectively, P<0.01). Conclusions: In mice with full-thickness skin defects, rapamycin may impair the CCL20 chemotactic system by inhibiting the expression of CCL20, leading to a decrease in the recruitment of Vγ4 T cells to the epidermis, and at the same time inhibit the secretion of IL-22 by Vγ4 T cells, thereby slowing the wound healing rate.

Ultrasonographic features and clinical pathological of liver metastasis in patients with melanoma / 中华肿瘤杂志

Objective: To investigate the ultrasonographic features and clinical pathological of liver metastasis in patients with melanoma. Methods: Thirteen patients with liver metastasis from melanoma treated in Tianjin Medical University Cancer Institute and Hospital from 2013 to 2019 were selected, and their ultrasonographic and clinicopathological characteristics were analyzed retrospectively. Results: Eleven of the 13 patients had multiple liver lesions. The maximum diameter of the lesions was (5.89±2.73) cm. Five cases of lesions were mixed echo (3 cases with high melanin content), 4 cases of lesions were hyperechoic (3 cases with low melanin content), 3 cases of lesions were hypoechoic (all with high melanin content), 1 case of lesions were equal echo (with high melanin content). The lesions in 11 patients had clear boundaries, while other 2 patients lacked the clear borders. Cystic areas were present in the lesions of 3 patients. Six cases had irregular lesions (lobulated), and 7 cases had regular lesions (round, oval). There were acoustic halos around the lesion in 9 cases and smooth and uneven acoustic halos in 5 cases. The results of immunohistochemistry showed that 11 cases were positive for S-100, HMB45 and Melan-A. One patient was not tested for HMB45, while S-100 and Melan-A were positive. One patient did not undergo Melan-A test, while S-100 and HMB45 were positive. Conclusion: Most of the liver metastases of melanoma are mixed echo or hyperechoic, most of them are nodular with clear boundaries combined with vocal halo, and a few of the lesions have cystic areas.

EGCG and ECG induce apoptosis and decrease autophagy via the AMPK/mTOR and PI3K/AKT/mTOR pathway in human melanoma cells / 中国天然药物

Catechins have been proven to exert antitumor effects in different kinds of cancers. However, the underlying mechanisms have not been completely clarified yet. This study aimed to assess the effects and mechanisms of (-)-epigallocatechin-3-gallate (EGCG) and (-)-epicatechin-3-gallate (ECG) on human melanoma skin A375 cells. Results showed that EGCG and ECG inhibited the proliferation of A375 cells and ECG showed better inhibitory effect. Flow cytometry analysis had shown that EGCG and ECG induced apoptosis and led to cell cycle arrest. EGCG and ECG decreased Bcl-2 expression and upregulated Caspase-3 protein level, indicating the development of apoptosis. Furthermore, EGCG and ECG could decreased mitochondrial membrane potential of A375 cells. In addition, the expression of Beclin-1, LC3 and Sirt3 were downregulated at protein levels, which known to be associated with autophagy. After autophagy was increased by rapamycin, the apoptotic trend was not change, indicating that apoptosis and autophagy are independent. Mechanistically, EGCG and ECG treatments decreased phosphorylated-AMPK (p-AMPK) and increased the ratios of p-PI3K, p-AKT and p-mTOR in melanoma cells. Conclusively, EGCG and ECG induced apoptosis via mitochondrial signaling pathway, downregulated autophagy through modulating the AMPK/mTOR and PI3K/AKT/mTOR signaling pathway. It indicated that EGCG and ECG may be utilized in human melanoma treatment.

Multi-target combinatory strategy to overcome tumor immune escape / 医学前沿

Immune therapy has become the fourth approach after surgery, chemotherapy, and radiotherapy in cancer treatment. Many immune checkpoints were identified in the last decade since ipilimumab, which is the first immune checkpoint inhibitor to cytotoxic T-lymphocyte associated protein 4, had been approved by the US Food and Drug Administration (FDA) for the treatment of unresectable or metastatic melanoma in 2011. The use of several antibody drugs that target PD1/PD-L1 for various cancer treatments has been approved by the FDA. However, fewer people are benefitting from immune checkpoint inhibitor treatment in solid cancers. Approximately 80% of patients do not respond appropriately because of primary or acquired therapeutic resistance. Along with the characterization of more immune checkpoints, the combinatory treatment of multiimmune checkpoint inhibitors becomes a new option when monotherapy could not receive a good response. In this work, the author focuses on the combination therapy of multiple immune checkpoints (does not include targeted therapy of oncogenes or chemotherapy), introduces the current progression of multiple immune checkpoints and their related inhibitors, and discusses the advantages of combination therapy, as well as the risk of immune-related adverse events.

Interleukins in the treatment of melanoma / 中华医学杂志(英文版)

Interleukins (ILs) and associated cytokines serve as the means of communication for immune cells and non-immune cells. The use of ILs in harnessing the immune system to cancer treatment has been a promising approach. ILs not only nurture an environment enabling cancer growth but also simultaneously trigger a productive tumor-directed immune response. These properties of ILs are increasingly being explored as a strategy to improve the outcomes of cancer. Here, we describe recently innovative technological approaches that have been developed to improve the pharmacokinetics, safety, and efficacies of IL-2, 15, 10, and 18 in the treatment of melanoma. Furthermore, the combination of ILs and immune checkpoint inhibition may synergize to reshape the tumor environment, thus yielding better clinical benefits in the future.

Linfoma con morfología en anillo de sello: patología con distintivos diagnósticos diferenciales / Lymphoma with signet ring morphology: pathology with distinctive differential diagnoses

Introducción: el linfoma con células en anillo de sello es una entidad poco frecuente y simuladora de neoplasias epiteliales, sarcomas y condiciones reactivas de histiocitos. Representa una variante morfológica de distintos linfomas no Hodgkin, por lo que su diagnóstico puede representar un desafío y debe ser considerado al realizar estudios complementarios. Objetivo: mostrar un caso con una morfología muy poco frecuente y recalcar la importancia de conocer esta entidad para no cometer errores en su diagnóstico. Caso clínico: se documenta el caso de un hombre de 67 años que desarrolló crecimientos ganglionares en axila derecha, cuello, ingle derecha, y región submandibular, con aparente afección en pulmones y bazo, que fue diagnosticado como linfoma B difuso de células grandes con morfología en anillo de sello, originado en el centro germinal. Se realizó estudio de microscopía electrónica de transmisión para una mejor caracterización de la morfología. Desafortunadamente el paciente no regresó a consulta de seguimiento, por lo que no inició tratamiento, falleció a los 6 meses posteriores al diagnóstico. Conclusiones: el linfoma con fenotipo en anillo de sello es poco frecuente, y puede presentarse en cualquier tipo de linfomas no Hodgkin; sin embargo, esta morfología es más comúnmente asociada a carcinomas y, en menor frecuencia, a sarcomas, melanomas o histiocitos reactivos, por lo que el considerar esta entidad junto con el uso adecuado de estudios complementarios es de gran importancia para su adecuado diagnóstico.

Background: Signet-ring cell lymphoma is a rare entity that simulates epithelial neoplasms, sarcomas and reactive histiocytes conditions. It represents a morphological variant of non-Hodgkin's lymphomas, its diagnosis can represent a challenge, therefore it should be considered in complementary studies. Objective: The aim of this work is to show a case with a very rare morphology and to emphasize the importance of awareness this entity and avoid mistakes in its diagnosis. Clinical case: We present a case of a 67-year-old man, who developed lymph node growths in the right armpit, neck, right groin, and submandibular region, with apparent involvement of the lungs and spleen; was diagnosed as diffuse large B cell lymphoma with signet-ring morphology, originated in the germinal center. Transmission electron microscopy study was carried out for a more precise characterization of the morphology. Unfortunately, the patient did not return for a follow-up consultation, so he did not start treatment and died 6 months after diagnosis. Conclusions: Lymphoma with the signet-ring phenotype is rare, and can occur in any type of non-Hodgkin lymphoma; however, this morphology is more commonly associated with carcinomas and, less frequently, with sarcomas, melanomas or reactive histiocytes conditions, therefore should be considered this entity together with the appropriate use of complementary studies for proper diagnosis.