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The CDKN2A gene remains understudied in melanoma compared to BRAF alterations. Inactivation of this tumor suppressor gene through homozygous deletions in the 9p21 chromosomal region leads to cellular proliferation and disrupts pro-apoptotic pathways. Genetic changes in CDKN2A are linked to multiple primary melanomas (MPM), with patients diagnosed with melanoma facing an elevated risk of developing additional primaries. We present the rare case of a 72-year-old Caucasian woman with nine metastasizing melanomas across diverse anatomical sites, posing a diagnostic challenge. Initial diagnosis in 2022 revealed ulcerated superficial spreading melanomas, progressing to intradermal and papillary dermal populations with neurotropism and angiotropism by early 2023. Lymph node metastases were identified, classifying the condition as pT3b N3b. Subsequent assessments in April 2023 revealed clinically suspicious melanocytic lesions diagnosed as intradermal and traumatized junctional nevi. In late 2023, cutaneous pigmented lesions and subcutaneous metastases were confirmed as nodular nevoid low-CSD multiple melanomas. Fluorescence in situ hybridization testing revealed homozygous CDKN2A deletion, necessitating close multidisciplinary collaboration for an optimized care plan for effective monitoring and intervention in this intricate clinical scenario. In summary, this case report highlights the diagnostic challenges of MPM in a single patient. Stressing the importance of immuno-histochemistry and CDKN2A genetic testing, our findings underscore the crucial role of these tools in accurately distinguishing malignant melanocytic proliferations from nevi and characterizing MPM cases.
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Inibidor p16 de Quinase Dependente de Ciclina , Melanoma , Neoplasias Cutâneas , Humanos , Melanoma/genética , Melanoma/diagnóstico , Feminino , Idoso , Inibidor p16 de Quinase Dependente de Ciclina/genética , Neoplasias Cutâneas/genética , Mutação , Neoplasias Primárias Múltiplas/genéticaRESUMO
Melanoma, a malignant neuroectodermic tumor originating from the neural crest, presents a growing global public health challenge and is anticipated to become the second most prevalent malignancy in the USA by 2040. The CDKN2A gene, particularly p16INK4a, plays a pivotal role in inhibiting the cell cycle via the cyclin D/CDK2-pRb pathway in certain tumors. In familial melanomas (FM), 40% exhibit CDKN2A mutations affecting p16INK4a, impacting checkpoint G1, and stabilizing p53 expression. This study aims to establish a scoring system using immunohistochemical antibodies, providing a cost-saving approach to classify multiple primary melanomas (MPM) and FM patients based on their mutational status, thus mitigating genetic testing expenses. This retrospective study included 23 patients with MPM and FM, assessing the p16, CD8, and Ki67 immunohistochemical status. Analyses of each parameter and associations between their value intervals and genetic CDKN2A status were conducted. A total score of at least 9 out of 10 points per tumor defined melanomas with homozygous CDKN2A deletions, exhibiting a sensitivity of 100% and specificity of 94.11%. In conclusion, p16, CD8, and Ki67 individually serve as valuable indicators for predicting melanoma evolution. The algorithm, comprising these three immunohistochemical parameters based on their prognostic and evolutionary significance, proves to be a valuable auxiliary diagnostic tool for cost-effective prediction of mutational status in detecting multiple and familial primary melanomas with CDKN2A homozygous deletion.
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Our study highlighted the immune changes by pro-inflammatory biomarkers in the gut-liver-axis-linked ROS-cell death mechanisms in chronic and acute inflammations when gut cells are exposed to endotoxins in patients with hepatic cirrhosis or steatosis. In duodenal tissue samples, gut immune barrier dysfunction was analyzed by pro-inflammatory biomarker expressions, oxidative stress, and cell death by flow cytometry methods. A significant innate and adaptative immune system reaction was observed as result of persistent endotoxin action in gut cells in chronic inflammation tissue samples recovered from hepatic cirrhosis with the A-B child stage. Instead, in patients with C child stage of HC, the endotoxin tolerance was installed in cells, characterized by T lymphocyte silent activation and increased Th1 cytokines expression. Interesting mechanisms of ROS-cell death were observed in chronic and acute inflammation samples when gut cells were exposed to endotoxins and immune changes in the gut-liver axis. Late apoptosis represents the chronic response to injury induction by the gut immune barrier dysfunction, oxidative stress, and liver-dysregulated barrier. Meanwhile, necrosis represents an acute and severe reply to endotoxin action on gut cells when the immune system reacts to pro-inflammatory Th1 and Th2 cytokines releasing, offering protection against PAMPs/DAMPs by monocytes and T lymphocyte activation. Flow cytometric analysis of pro-inflammatory biomarkers linked to oxidative stress-cell death mechanisms shown in our study recommends laboratory techniques in diagnostic fields.
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Endotoxinas , Inflamação , Criança , Humanos , Endotoxinas/metabolismo , Espécies Reativas de Oxigênio , Cirrose Hepática , Apoptose , Citocinas , BiomarcadoresRESUMO
Cutaneous melanoma is a public health problem. Efforts to reduce its incidence have failed, as it continues to increase. In recent years, many risk factors have been identified. Numerous diagnostic systems exist that greatly assist in early clinical diagnosis. The histopathological aspect illustrates the grim nature of these cancers. Currently, pathogenic pathways and the tumor microclimate are key to the development of therapeutic methods. Revolutionary therapies like targeted therapy and immune checkpoint inhibitors are starting to replace traditional therapeutic methods. Targeted therapy aims at a specific molecule in the pathogenic chain to block it, stopping cell growth and dissemination. The main function of immune checkpoint inhibitors is to boost cellular immunity in order to combat cancer cells. Unfortunately, these therapies have different rates of effectiveness and side effects, and cannot be applied to all patients. These shortcomings are the basis of increased incidence and mortality rates. This study covers all stages of the evolutionary sequence of melanoma. With all these data in front of us, we see the need for new research efforts directed at therapies that will bring greater benefits in terms of patient survival and prognosis, with fewer adverse effects.
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Melanoma represents an aggressive malignant tumor, encapsulating frequent loss of differentiation markers, with familial melanoma constituting a relatively commonly encountered entity, in direct relationship with cyclin-dependent kinase inhibitor 2A (CDKN2A). The present study aims to identify the association between the immunohistochemical p14-p16 profile, the molecular CDKN2A findings and clinically diagnosed familial or multiple primary melanomas (MPM). We conducted a 5-year retrospective cross-sectional study, on patients diagnosed with familial or MPM. P14 and p16 immunohistochemical staining has been applied on the selected surgical specimens simultaneously with the performance of fluorescence in situ hybridization (FISH) CDKN2A testing. 13 out of the 23 included cases displayed p14 and/or p16 immunohistochemical absence and the main positive relationships were encountered between CDKN2A homozygous deletion and p14â ±â p16 negative immunoreactions. Cases with exclusive p16 absent reaction (nâ =â 7) were more frequently associated with the presence of distant metastases (85.71%), while samples with exclusive p14 immunohistochemical loss exhibited more favorable histopathological prognostic markers. The average percentage of p16-stained nuclei in the superficial dermis and the deep dermis were equal (29.54% for each), therefore infirming its potential predictive and/or prognostic utility. The present study is the first of its type to approach the clinical, evolutionary and immunophenotypic correlations between p14-p16 immunohistochemical testing, CDKN2A molecular biology pattern, familial melanoma and spontaneous MPM in a cohort of Romanian patients. This analysis highlighted the value of singular p16 immunohistochemical absence as a predictor for aggressive biological behavior and unfavorable prognosis in familial melanoma and/or MPM, in comparison with the exclusive loss of p14, indifferent to the histopathological subtype. The present study emphasizes the utility of immunohistochemistry as a less expensive method of complementing the current testing arsenal and could represent the starting point for the elaboration of tailored diagnostic and therapeutic algorithms, based on the discovered p14-p16-CDKN2A significant correlation.
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Melanoma , Neoplasias Cutâneas , Humanos , Melanoma/genética , Estudos Retrospectivos , Hibridização in Situ Fluorescente , Melanoma Maligno Cutâneo , Homozigoto , Estudos Transversais , Deleção de Sequência , Neoplasias Cutâneas/genética , Inibidor p16 de Quinase Dependente de Ciclina/genéticaRESUMO
Background and Objectives: In the realm of the rising incidence of cutaneous and mucous melanoma, CDKN2A mutations characterize familial and multiple primary melanoma cases. The involvement of tumor-infiltrating lymphocytes (TILs) is interconnected with survival rates, but may extend even further. The aim of this study is to verify the accuracy of the classical "naked eye" count of CD8-positive T cells comprised within the tumoral population and peritumoral infiltrate versus that obtained via a special software run by the aid of artificial intelligence (AI), used to determine the percentage of CD8-positive TILs. Materials and Methods: The present retrospective cross-sectional study conducted over a period of 5 years (2018-2022) focused on patients diagnosed with mucous and/or cutaneous melanoma, with a positive family history for melanoma, or personal antecedents of primary malignant melanocytic lesions. The 23 selected cases were diagnosed histopathologically, tested for CDKN2A mutations through fluorescent hybridization in situ, and CD8 immunohistochemistry was performed. The included slides were evaluated both manually (naked-eye examination) and automatically (via QuPath platform) for quantifying the CD8-positive TILs. Results: The number of CD8-positive TILs in melanoma samples has been more accurately identified through the use of an AI-mediated software as compared to the human-eye evaluation performed by experimental pathologists. A higher percentage of CD8-positive intratumoral lymphocytes versus stromal lymphocytes was positively associated with more numerous metastatic sites. Conclusions: The CD8 lymphocytic phenotype harbors major significance in the context of familial and multiple primary melanoma and may comprise a cost-effective investigation meant to help in the establishment of melanoma prognosis and response to immunotherapy.
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Melanoma , Neoplasias Primárias Múltiplas , Neoplasias Cutâneas , Humanos , Melanoma/genética , Melanoma/patologia , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/patologia , Estudos Retrospectivos , Melanoma Maligno Cutâneo , Inteligência Artificial , Estudos Transversais , Linfócitos T CD8-Positivos/patologia , Linfócitos do Interstício Tumoral , Prognóstico , Fenótipo , Neoplasias Primárias Múltiplas/patologia , Inibidor p16 de Quinase Dependente de Ciclina/genéticaRESUMO
Lipomatous tumors are the most frequent soft tissue neoplasms. Sometimes their differential diagnosis is difficult to perform only by microscopic analysis. This study aims to create a histopathological scoring system and highlight the impact of intratumoral microvascular density. This study was conducted over 10 years. We analyzed the main pathogenic pathways (MDM2 and CDK4), as well as the tumor microvascularization (CD31 and CD34) by immunohistochemical tests. We also analyzed the status of the MDM2 gene by CISH. These data, together with the clinical and histopathological information, were statistically analyzed by appropriate tests. We identified 112 eligible cases, with most of the patients being in their sixth decade of life, with a slight predominance of the female sex. We found important associations like tumor location linked to nuclear pleomorphism severity and microvascularization density correlated with atypia severity. Also, we observed that a maximum diameter of a tumor of at least 69 mm is associated with the presence of tumor necrosis. The score designed in this study shows an increased sensitivity and specificity for the diagnosis of lipomas (100%, respectively, 97%), atypical lipomatous tumors (93.8%, respectively, 82.3%), and liposarcomas (100%, respectively, 90.5%). This present study enhances the present data by bringing to attention the histopathological score with a role in differential diagnosis, as well as in the prediction of immunohistochemical and genetic tests. Also, we highlighted the importance of microvascular density, especially in the diagnosis of liposarcomas.
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(1) Background: Human cytomegalovirus (CMV) infection is one of the most frequent opportunistic infections in immunosuppressed patients. Romania has one of the highest incidences of patients living with human immunodeficiency virus (HIV) which determines an immunosuppressive state. The aim of this study was to establish the prevalence of CMV infection among women living with HIV in Southeastern Romania and also to evaluate and correlate antiretroviral therapy (ART) with CD4 level and CMV disease evolution. (2) Methods: Seventy women living with HIV from Southeastern Romania were screened for CMV infection using antigen quantification. Of these, 50 were included in the study. First, the patients filled out a questionnaire regarding social conditions and other associated diseases. Then, we explored the statistical correlations between the data and HIV status, CD4+ cell counts, viral load, and antiretroviral therapy (ART). (3) Results: Median age of the patients was 33 years. Twenty-nine cases were diagnosed with HIV after sexual life beginning and 21 before. Most of the patients had a CD4 level over 200 cells/µL. ART duration in the CD4 under 200 cells/µL group was a bit longer than that in the CD4 over 200 cells/µL group. Forty-one patients had undetectable viremia. CD4 average value in the lot of patients with undetectable viremia was 704.71 cells/µL and in the lot with detectable viremia was 452.44 cells/µL. Viremia values correlated negatively with CD4 level. A positive correlation between IgG CMV values and ART therapy length was identified. A negative significant correlation between values of IgG CMV and values of CD4 was identified. CD4 value correlated negatively with IgG CMV values and with CMV avidity. (4) Conclusions: IgG CMV values had a weak positive correlation with ART therapy length, and a negative statistically significant correlation with values of CD4. CMV avidity has a negative correlation with CD4 value.
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The spectrum of major and minor salivary gland disorders varies widely. Epidemiological data on some injury categories are rare and often not up-to-date. This study aims to analyze epidemiological data using clinical, paraclinical, and histopathological parameters. Study was carried out for 5 years on the nonneoplastic and tumoral pathology of the salivary glands. Data were statistically analyzed using the appropriate parameters. Data analysis according to the biological behavior of the lesions revealed great heterogeneity. Statistically significant correlations were observed between the type of injury, age (Pâ =â .002) and gender (0.033). The environment of origin of the patients as well as the comorbidities reflected in most cases the nature of the process. Associations were also observed between the biological behavior of the lesions and the hemicranial topography (Pâ =â .019), the type of salivary gland (Pâ =â .024), and the surgical technique used (Pâ <â .001). Most cases were identified in the major salivary glands, often in the parotid. The most common diseases are represented by nonspecific chronic sialadenitis (nonneoplastic lesion), pleomorphic adenoma and Warthin tumor (benign tumors), mucoepidermoid carcinoma (malignant tumor), and squamous carcinoma (secondary tumor). They presented axial diameters between 2 to 95 mm. The most used curative technique was subtotal excision with facial nerve preservation. In conclusion, the study highlighted the main epidemiological aspects of salivary gland disorders. Some data agree with the specialty literature, and particular aspects are also observed. Therefore, this research is useful both in the medical and research fields.
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Adenoma Pleomorfo , Carcinoma Mucoepidermoide , Doenças das Glândulas Salivares , Neoplasias das Glândulas Salivares , Humanos , Estudos Retrospectivos , Neoplasias das Glândulas Salivares/patologia , Glândulas Salivares/patologia , Adenoma Pleomorfo/patologia , Doenças das Glândulas Salivares/epidemiologia , Carcinoma Mucoepidermoide/patologiaRESUMO
BACKGROUND AND AIMS: High-grade gastrointestinal neuroendocrine neoplasms (GI-NENs) are divided into well-differentiated G3 neuroendocrine tumors (NETs G3) and neuroendocrine carcinomas (NECs), having identical cut-offs of proliferation, but different biomolecular origins. This translates in distinct treatment choices. Our aim was to establish if p53/Rb1 immunohistochemical status in GI-NENs with Ki67 index >20% can predict the histopathological diagnosis. METHODS: p53/Rb1 immunolabelling was performed on 42 cases of high-grade GI-NENs, diagnosed as NET G3, NEC and mixed neuroendocrine-non-neuroendocrine neoplasms (MiNEN) with NEC component. Immunolabeled slides were digitally scanned, with automatic quantification of p53 and Rb1, blind to the diagnosis. RESULTS: The p53 positive percentage was stratified; two cut-offs were selected, naming the intervals as N (null, <1%), T (tumor, 1%-20%) and C (carcinoma, >20%). The Rb1 expression loss in >90% of neoplastic cells was considered mutational. NETs G3 mainly showed the T status (14/16, 87.5%), followed by N (1/16, 6.25%) and C (1/16, 6.25%); NECs and NEC components in MiNENs predominantly expressed the C status (19/26, 73.08%), followed by N (5/26, 19.23%) and T (2/26, 7.69%) (p<0.001, χ 2 =27.017). NET G3s showed positive expression for Rb1; 73.08% of NECs expressed negative Rb1 (p<0.001, χ 2 =21.351). NECs and NEC components in MiNENs showed Rb1 mutational status in 13 C cases (13/19, 68.42%), 4 N cases (4/5, 80%) and in both the T cases (p=0.002, χ 2 =11.187). CONCLUSIONS: Our results highlight the correlations between the p53/Rb1 immunostainings and the histopathological diagnosis of high-grade GI-NENs. NECs and NEC components in MiNENs showed a p53 mutational status (0% or 21-100%) and predominantly negative Rb1 expression. NETs G3 showed a p53 wild-type status (1-20%) and retained Rb1 expression. These findings suggest that the differential diagnosis of high-grade GI-NENs may benefit from p53/Rb1 immunohistochemical tests in everyday practice.
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Carcinoma Neuroendócrino , Neoplasias Gastrointestinais , Tumores Neuroendócrinos , Neoplasias Pancreáticas , Humanos , Proteína Supressora de Tumor p53/genética , Diagnóstico Diferencial , Neoplasias Pancreáticas/patologia , Tumores Neuroendócrinos/diagnóstico , Tumores Neuroendócrinos/genética , Tumores Neuroendócrinos/metabolismo , Neoplasias Gastrointestinais/diagnóstico , Neoplasias Gastrointestinais/genética , Carcinoma Neuroendócrino/diagnóstico , Carcinoma Neuroendócrino/genética , Carcinoma Neuroendócrino/patologiaRESUMO
Myocytic tumors of the uterus present vast morphological heterogeneity, which makes differential diagnosis between the different entities necessary. This study aims to enrich the existing data and highlight new potential therapeutic targets regarding aspects related to the pathogenic process and the tumor microenvironment in order to improve the quality of life of women. We performed a 5-year retrospective study, including particular cases of uterine myocyte tumors. Immunohistochemical analyses of pathogenic pathways (p53, RB1, and PTEN) and tumor microclimate using markers (CD8, PD-L1, and CD105), as well as genetic testing of the PTEN gene, were performed. The data were statistically analyzed using the appropriate parameters. In cases of atypical leiomyoma, a significant association was observed between PTEN deletion and an increased number of PD-L1+ T lymphocytes. For malignant lesions and STUMP, PTEN deletion was associated with the advanced disease stage. Advanced cases were also associated with an increased mean CD8+ T cell count. An increased number of lymphocytes was associated with an increased percentage of RB1+ nuclei. The study corroborated clinical and histogenetic data, highlighting the importance of the differential diagnosis of these tumors to improve the management of patients and increase their quality of life.
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BACKGROUND AND AIMS: MicroRNAs (miR) have altered expression in multiple autoimmune disorders including inflammatory bowel disease. The aim of the study was to assess the tissue and circulating miR-31, miR-200b, and miR-200c expression levels as potential biomarkers for intestinal disease activity in patients with Crohn's disease (CD). METHODS: The study included 45 patients with histopathological confirmed CD and active disease (defined as fecal calprotectin >50 µg/g and Simple Endoscopic Score (SES) of CD >3), and 21 subjects as controls for the validation cohort. Demographic and clinical data, biomarkers (fecal calprotectin), endoscopy data, the expression levels of miR-31, miR-200b, and miR-200c in tissue and serum were assessed (by RT-PCR). Receiver operating characteristic analysis was performed to assess the miR-31, miR-200b, and miR-200c expression levels as potential biomarkers for active CD. RESULTS: Mean fecal calprotectin was 1540±890 µg/g. Mean SES-CD was 8.9±4.2. Tissue and circulating miR- 31 were significantly correlated with fecal calprotectin (r=0.81, r=0.83, p<0.01) and with SES-CD (r=0.82, r=0.79, p<0.01). The expression level of miR-31 was significantly upregulated in CD tissue cases compared to the control tissue samples (6.24±1.57 vs. 3.70±1.44; p <0.01). Similarly, serum miR-31 expression levels in CD patients were significantly upregulated compared to the control serum samples (0.78±0.42 vs. -2.07±1.00; p<0.01). The expression levels of tissue miR-200b and miR-200c were significantly upregulated in CD tissue cases compared to the control tissue samples (-5.25±0.93 vs. -4.69±0.80, p=0.03 for miR-200b, and -0.86±0.96 vs. 0.39±0.66, p<0.01 for miR-200c). Similarly, serum miR-200b and miR-200c expression levels in CD patients were significantly upregulated compared to the control serum samples (p < 0.05). Receiver operating characteristic analysis revealed that the expression levels of the selected miRNAs could help to discriminate active CD patients from healthy controls with very good specificity and sensitivity. CONCLUSIONS: Tissue and circulating miR-31, miR-200b, and miR-200c reflect disease activity in CD patients and can be used as biomarkers for active disease.
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MicroRNA Circulante , Doença de Crohn , MicroRNAs , Humanos , MicroRNA Circulante/genética , Doença de Crohn/diagnóstico , Doença de Crohn/genética , MicroRNAs/genética , Biomarcadores , Complexo Antígeno L1 LeucocitárioRESUMO
RATIONALE: Dedifferentiated liposarcoma is defined as a malignant tumor that changes its shape from a well-differentiated liposarcoma to a non-liposarcomatous form. Most paratesticular liposarcomas manifest as an inguinal, painless, slow-growing mass. The standard treatment is extensive surgical excision, radiotherapy being proposed for cases with positive margins, those with recurrence, or in cases of the existence of unfavorable prognostic factors. PATIENT CONCERNS: We present the case of a young patient diagnosed initially with left hydrocele, which after 2 years proved to mask a differentiated liposarcoma of the spermatic cord. The initial clinical manifestations were represented by the increase in volume of the left groin-scrotal region and pain at this level. DIAGNOSIS: Microscopic examination in hematoxylin-eosin staining highlighted the presence of lipoblasts and fibroblasts in association with areas of hemorrhage and tumor necrosis. The performed immunohistochemical tests confirmed the diagnosis of dedifferentiated liposarcoma. To support and confirm the presence of the mouse double minute 2 homolog gene mutation, chromogenic in situ hybridization analysis was performed. INTERVENTIONS: The initial treatment was the surgical one. After 2 weeks, the patient received zolendronic acid for hypercalcemia which was caused by the osseous metastasis. OUTCOMES: The patient died secondary to acute renal failure caused by hypercalcemia despite the treatment received. LESSONS: This case underlines the importance of both the correct management of oncological patients, as well as immunohistochemical and genetic tests in the identification of prognostic factors, with the ultimate goal of administering an appropriate oncological treatment.
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Neoplasias dos Genitais Masculinos , Hipercalcemia , Lipossarcoma , Cordão Espermático , Humanos , Masculino , Animais , Camundongos , Neoplasias dos Genitais Masculinos/diagnóstico , Neoplasias dos Genitais Masculinos/cirurgia , Neoplasias dos Genitais Masculinos/patologia , Lipossarcoma/diagnóstico , Lipossarcoma/cirurgia , Lipossarcoma/patologia , Escroto/patologia , Cordão Espermático/patologiaRESUMO
(1) Background: A malignant gastrointestinal neuroectodermal tumor (GNET) is an ultra-rare primary neoplasm with a distinctive histopathological, immunohistochemical, molecular, and ultramicroscopic profile, synonymous terminology with clear cell sarcoma-like tumor of the gastrointestinal tract. This case report aims to describe a case of GNET with challenging mesenchymal, lymphoid, and melanic tumor differential diagnosis. (2) Case presentation: We discuss the case of a 67-year-old male patient who presented with diffuse abdominal pain, intermittent lack of intestinal transit, and frequent episodes of nausea, followed by segmental resection of the jejunum and sigmoid colon. The patient had no relevant medical history. The surgical specimen underwent immunohistochemical staining and morphological evaluation. (3) Results: Histopathological analysis reveals a moderately homogeneous polyhedral-epithelioid and spindle cell neoplastic proliferation with a zonal discohesive pattern and extensive and focal fasciculated architecture. Twenty monoclonal antibodies were used for immunostaining, which allowed GNET to be diagnosed on the basis of the tumoral immune profile, characterized by positive reactivity of S100, SOX10, and CD 56. (4) Conclusions: The poor prognosis of GNET is highlighted in the present study, along with the vital importance of differential diagnosis issues with mesenchymal, lymphoid, and melanic tumors, which make the diagnosis difficult for both pathologists and clinicians.
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(1) Background: Because melanoma is an aggressive tumor with an unfavorable prognosis, we aimed to characterize the PD-L1 expression in melanomas in association with T cell infiltrates because PD-1/PD-L1 blockade represents the target in treating melanoma strategy. (2) Methods: The immunohistochemical manual quantitative methods of PD-L1, CD4, and CD8 TILs were performed in melanoma tumor microenvironment cells. (3) Results: Most of the PD-L1 positive, expressing tumors, have a moderate score of CD4+ TILs and CD8+TILs (5-50% of tumor area) in tumoral melanoma environment cells. The PD-L1 expression in TILs was correlated with different degrees of lymphocytic infiltration described by the Clark system (X2 = 8.383, p = 0.020). PD-L1 expression was observed often in melanoma cases, with more than 2-4 mm of Breslow tumor thickness being the associated parameters (X2 = 9.933, p = 0.014). (4) Conclusions: PD-L1 expression represents a predictive biomarker with very good accuracy for discriminating the presence or absence of malign tumoral melanoma cells. PD-L1 expression was an independent predictor of good prognosis in patients with melanomas.
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Oxidative stress is associated with aging, cancers, and numerous metabolic and chronic disorders, and phenolic compounds are well known for their health-promoting role due to their free-radical scavenging activity. These phytochemicals could also exhibit pro-oxidant effects. Due to its bioactive phenolic secondary metabolites, Usnea barbata (L.) Weber ex. F.H. Wigg (U. barbata) displays anticancer and antioxidant activities and has been used as a phytomedicine for thousands of years. The present work aims to analyze the properties of U. barbata extract in canola oil (UBO). The UBO cytotoxicity on oral squamous cell carcinoma (OSCC) CLS-354 cell line and blood cell cultures was explored through complex flow cytometry analyses regarding apoptosis, reactive oxygen species (ROS) levels, the enzymatic activity of caspase 3/7, cell cycle, nuclear shrinkage (NS), autophagy (A), and synthesis of deoxyribonucleic acid (DNA). All these studies were concomitantly performed on canola oil (CNO) to evidence the interaction of lichen metabolites with the constituents of this green solvent used for extraction. The obtained data evidenced that UBO inhibited CLS-354 oral cancer cell proliferation through ROS generation (316.67 × 104), determining higher levels of nuclear shrinkage (40.12%), cell cycle arrest in G0/G1 (92.51%; G0 is the differentiation phase, while during G1 phase occurs preparation for cell division), DNA fragmentation (2.97%), and autophagy (62.98%) than in blood cells. At a substantially higher ROS level in blood cells (5250.00 × 104), the processes that lead to cell death-NS (30.05%), cell cycle arrest in G0/G1 (86.30%), DNA fragmentation (0.72%), and autophagy (39.37%)-are considerably lower than in CLS-354 oral cancer cells. Our work reveals the ROS-mediated anticancer potential of UBO through DNA damage and autophagy. Moreover, the present study suggests that UBO pharmacological potential could result from the synergism between lichen secondary metabolites and canola oil phytoconstituents.
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Carcinoma de Células Escamosas , Neoplasias de Cabeça e Pescoço , Neoplasias Bucais , Usnea , Humanos , Neoplasias Bucais/tratamento farmacológico , Neoplasias Bucais/metabolismo , Usnea/química , Usnea/metabolismo , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas de Cabeça e Pescoço , Óleo de Brassica napus/farmacologia , Autofagia , Dano ao DNA , Espécies Reativas de Oxigênio/metabolismo , Apoptose , Extratos Vegetais/farmacologia , Fenóis/farmacologia , DNA/farmacologia , Linhagem Celular TumoralRESUMO
Necrosis and increased microvascular density in glioblastoma IDH-wild-type are the consequence of both hypoxia and cellular immaturity. Our study aimed to identify the main clinical-imaging and morphogenetic risk factors associated with tumor necrosis and microvascular in the prognosis of patient survival. We performed a retrospective study (10 years) in which we identified 39 cases. We used IDH1, Ki-67 and Nestin immunomarkers, as well as CDKN2A by FISH. The data were analyzed using SPSS Statistics. The clinical characterization identified only age over 50 years as a risk factor (HR = 3.127). The presence of the tumor residue, as well as the absence of any therapeutic element from the trimodal treatment, were predictive factors of mortality (HR = 1.024, respectively HR = 7.460). Cellular immaturity quantified by Nestin was associated with reduced overall survival (p = 0.007). Increased microvascular density was associated with an increased proliferative index (p = 0.009) as well as alterations of the CDKN2A gene (p < 0.001). CDKN2A deletions and cellular immaturity were associated with an increased percentage of necrosis (p < 0.001, respectively, p = 0.017). The main risk factors involved in the unfavorable prognosis are moderate and increased Nestin immunointensity, as well as the association of increased microvascular density with age over 50 years. Necrosis was not a risk factor.
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Oral squamous cell carcinoma (OSCC) is the most frequent oral malignancy, with a high death rate and an inadequate response to conventional chemotherapeutic drugs. Medical research explores plant extracts' properties to obtain potential nanomaterial-based anticancer drugs. The present study aims to formulate, develop, and characterize mucoadhesive oral films loaded with Usnea barbata (L.) dry acetone extract (F-UBA) and to investigate their anticancer potential for possible use in oral cancer therapy. U. barbata dry acetone extract (UBA) was solubilized in ethanol: isopropanol mixture and loaded in a formulation containing hydroxypropyl methylcellulose (HPMC) K100 and polyethylene glycol 400 (PEG 400). The UBA influence on the F-UBA pharmaceutical characteristics was evidenced compared with the references, i.e., mucoadhesive oral films containing suitable excipients but no active ingredient loaded. Both films were subjected to a complex analysis using standard methods to evaluate their suitability for topical administration on the oral mucosa. Physico-chemical and structural characterization was achieved by Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), thermogravimetric analysis (TGA), scanning electron microscopy (SEM), and atomic force microscopy (AFM). Pharmacotechnical evaluation (consisting of the measurement of specific parameters: weight uniformity, thickness, folding endurance, tensile strength, elongation, moisture content, pH, disintegration time, swelling rate, and ex vivo mucoadhesion time) proved that F-UBAs are suitable for oral mucosal administration. The brine shrimp lethality (BSL) assay was the F-UBA cytotoxicity prescreen. Cellular oxidative stress, caspase 3/7 activity, nuclear condensation, lysosomal activity, and DNA synthesis induced by F-UBA in blood cell cultures and oral epithelial squamous cell carcinoma (CLS-354) cell line were investigated through complex flow cytometry analyses. Moreover, F-UBA influence on both cell type division and proliferation was determined. Finally, using the resazurin-based 96-well plate microdilution method, the F-UBA antimicrobial potential was explored against Staphylococcus aureus ATCC 25923, Pseudomonas aeruginosa ATCC 27353, Candida albicans ATCC 10231, and Candida parapsilosis ATCC 22019. The results revealed that each UBA-loaded film contains 175 µg dry extract with a usnic acid (UA) content of 42.32 µg. F-UBAs are very thin (0.060 ± 0.002 mm), report a neutral pH (7.01 ± 0.01), a disintegration time of 146 ± 5.09 s, and an ex vivo mucoadhesion time of 85 ± 2.33 min, and they show a swelling ratio after 6 h of 211 ± 4.31%. They are suitable for topical administration on the oral mucosa. Like UA, they act on CLS-354 tumor cells, considerably increasing cellular oxidative stress, nuclear condensation, and autophagy and inducing cell cycle arrest in G0/G1. The F-UBAs inhibited the bacterial and fungal strains in a dose-dependent manner; they showed similar effects on both Candida sp. and higher inhibitory activity against P. aeruginosa than S. aureus. All these properties lead to considering the UBA-loaded mucoadhesive oral films suitable for potential application as a complementary therapy in OSCC.
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Adult low-grade gliomas are a rare and aggressive pathology of the central nervous system. Some of their characteristics contribute to the patient's life expectancy and to their management. This study aimed to characterize and identify the main prognostic factors of low-grade gliomas. The six-year retrospective study statistically analyzed the demographic, imaging, and morphogenetic characteristics of the patient group through appropriate parameters. Immunohistochemical tests were performed: IDH1, Ki-67, p53, and Nestin, as well as FISH tests on the CDKN2A gene and 1p/19q codeletion. The pathology was prevalent in females, with patients having an average age of 56.31 years. The average tumor volume was 41.61 cm3, producing a midline shift with an average of 7.5 mm. Its displacement had a negative impact on survival. The presence of a residual tumor resulted in decreased survival and is an independent risk factor for mortality. Positivity for p53 identified a low survival rate. CDKN2A mutations were an independent risk factor for mortality. We identified that a negative prognosis is influenced by the association of epilepsy with headache, tumor volume, and immunoreactivity to IDH1 and p53. Independent factors associated with mortality were midline shift, presence of tumor residue, and CDKN2A gene deletions and amplifications.
Assuntos
Neoplasias Encefálicas , Glioma , Feminino , Humanos , Isocitrato Desidrogenase/genética , Nestina/genética , Prognóstico , Estudos Retrospectivos , Proteína Supressora de Tumor p53/genética , Antígeno Ki-67/genética , Mutação , Glioma/genética , Glioma/patologiaRESUMO
Somatic mutations in the oncogenes of the epidermal growth factor receptor signaling pathway play vital roles in colorectal carcinogenesis and have been closely linked with clinical resistance to monoclonal therapy. In this study, we have analyzed the mutation frequencies of 5 genes and compared the genetic findings with clinicopathological variables in order to determine diagnostically relevant alterations and compare these findings with those of other studies In our Sanger sequencings, KRAS (exons 2, 3, and 4), NRAS (exons 2, 3, and 4), PIK3CA (exons 9 and 20), BRAF (exon 15), AKT1 (exon 2) genes, and microsatellite instability (MSI) status were analyzed using an ABI 3500 analyzer in a cohort of 58 Romanian colorectal cancer (CRC) patients who underwent surgical resection at Emergency County Clinical Hospital in ConstanÈa, Romania. In our series, mutation rates of KRAS, BRAF, PIK3CA, and AKT1 genes were 39.63%, 8.62%, 6.88%, and 3.44%, respectively. By contrast, we did not find any tumor harboring mutation in the NRAS gene. Notably, the KRAS and PIK3CA mutations were not mutually exclusive, 1 patient harbored 2 mutations in exon2, codon 12 (Gly12Val) of KRAS and exon 20, codon 1047 (His1047Arg) of PIK3CA. The finding of our study are generally consistent with data found in the literature. Regarding to clinicopathological variables, mutation of KRAS was associated with distant metastasis at the time of diagnosis, while mutation of BRAF was significantly associated with MSI-H in contrast with MSI-L/MSS tumors. Moreover, PIK3CA mutation tends to be located in the proximal segment of the colon and to be well/moderately differentiated compared to wild-type tumors. In conclusion, the assessment of these mutations suggests that CRC patients from southeast Romania exhibit a mutation profile similar to other populations. These results could contribute to creating a better method of qualifying patients for molecularly targeted therapies and obtaining better screening strategies.