Cytoplasm Of Tumor Cells Research Articles

Expression of CD68 and CD163 in malignant epithelial cells of oral squamous cell carcinoma: phenotypic shift or mere cell fusion

Background / purposeThe progression of epithelial to mesenchymal tissue (EMT) is a highly intricate process that facilitates the transformation of cancer cells, allowing them to changeover their characteristic epithelial properties to mesenchymal attributes. This notable change empowers the cells with enhanced mobility and the ability to migrate to distant locations. Furthermore, it is imperative to adopt the idea of macrophage tumor cell fusion to achieve comprehensive considerate of this phenomenon. Our primary objective was to conduct a thorough investigation of macrophage-restricted antigens expression, specifically CD68 and CD163, in malignant epithelial cells of oral cavity squamous cell carcinoma (OSCC) to elucidate aforementioned perceptions. Materials and methodsCD68 and CD163 immunohistochemical expression were assessed in oral squamous cell carcinoma (OSCC), encompassing both the neoplastic cells and the tumor-associated macrophages (TAMs). ResultsBoth CD68 and CD163 antigens were revealed in OSCC malignant epithelial cells in a granular cell pattern, localized in membrane and cytoplasm of tumor cells respectively as well as in the infiltrating TAMs. ConclusionThe macrophage antigens were not limited to the infiltrating tumor-associated macrophages (TAMs), but were also observed in a substantial proportion of OSCC malignant epithelial cells within the tumor parenchyma. This particular expression pattern may represent a subset of tumor cells that have undergone an epithelial to a mesenchymal phenotypic transition. In addition, fusion of macrophages with tumor cells cannot be excluded; both might be associated with increased metastatic activity of OSCC.

Correlation of MMP-9 (Matrix Metalloproteinase 9) and SMA (Smooth Muscle Actin) expression with Basal Cell Carcinoma’s risk of recurrence

Basal Cell Carcinoma (BCC) is the most common cancer in many countries, with its incidence steadily rising. Data from the Indonesian Pathologists Association show that among 1,530 skin cancer cases, BCC was the most frequent (39.93%), followed by Squamous Cell Carcinoma (23%) and Malignant Melanoma (7.9%). According to the 2018 WHO classification, BCC is divided into Low-Risk and High-Risk groups based on the risk of recurrence. BCC invades surrounding tissues through extracellular matrix degradation and enhanced cell motility, driven by the expression of MMP-9 (Matrix Metalloproteinase 9) and SMA (Smooth Muscle Actin). This study aims to evaluate the expression of MMP-9 and SMA in BCC using immunohistochemistry and analyze their correlation with the risk of recurrence. A total of 40 paraffin blocks from BCC patients diagnosed between 2019 and 2021 at Dr. Saiful Anwar General Hospital in Malang, Indonesia, were sampled. The blocks were sectioned, stained with MMP-9 and SMA antibodies, and examined for positive expression indicated by brown staining in the cytoplasm of tumor cell and peritumoral stroma. Spearman correlation showed r = 0.811 for MMP-9 and r = 0.857 for SMA (p < 0.05, CI: 95%), indicating a significant relationship between these markers and the risk of recurrence. Higher MMP-9 and SMA expression corresponded to a higher risk of BCC recurrence, suggesting their potential as prognostic biomarkers. However, further research is needed to assess recurrence risk more accurately and determine the most appropriate therapy.

Mutations of the Cx43 Gene in Non-Small Cell Lung Cancer: Association with Aberrant Localization of Cx43 Protein Expression and Tumor Progression

Background and Objectives: The Connexin43 (Cx43) gene is a suspected tumor suppressor gene, as re-expressed wild-type Cx43 genes reduce the malignancy potential of tumor cells. However, the role of Cx43 gene expression in human lung tumorigenesis remains unclear. Materials and Methods: Tumor tissues from 165 primary lung cancer patients were collected to study Cx43 protein expression and gene mutations using immunohistochemistry and direct DNA sequencing. In addition, Cx43 genes with or without mutations were transfected to CL-3 human lung cancer cells to confirm the function of these mutant forms of the Cx43 gene. Results: Aberrant localization of Cx43 protein in the nucleus and cytoplasm of tumor cells was detected in 14 out of 165 non-small cell lung cancer (NSCLC) patients. Mutations in the Cx43 gene were also found in patients with aberrant Cx43 localization, and transfection of these mutant genes into lung cancer cells enhanced their proliferation. Conclusions: To our knowledge, this is the first study to demonstrate Cx43 gene mutations in human lung neoplasm, supporting the hypothesis that Cx43 may function as a tumor suppressor in some lung cancer patients. Additionally, the findings suggest an association between aberrant localization of Cx43 protein expression and tumor progression.

Responsive Supramolecular Nanomicelles Formed through Self-Assembly of Acyclic Cucurbit[n]uril for Targeted Drug Delivery to Cancer Cells.

The supramolecular drug delivery systems (SDDSs) based on host-guest recognition through noncovalent interactions, capable of responsive behavior and dynamic switching to external stimuli, have attracted considerable attention in cancer therapy. In this study, a targeted dual-functional drug delivery system was designed and synthesized. A hydrophilic macrocyclic host molecule (acyclic cucurbit[n]uril ACB) was modified with folic acid (FA) as a targeting ligand. The guest molecule consists of a disulfide bond attached to adamantane (DA) and cannabidiol (CBD) at both ends of the response element of glutathione. Recognition and self-assembly of host and guest molecules successfully functionalize supramolecular nanomicelles (SNMs), targeting cancer cells and releasing drugs in a high glutathione environment. The interactions between host and guest molecules were investigated by using nuclear magnetic resonance (NMR), fluorescence titration, Fourier-transform infrared spectroscopy (FT-IR), and thermal analysis (TGA). Transmission electron microscopy (TEM) and dynamic light scattering (DLS) confirmed the nanostructure of the SNMs. Experimentation with 5,5'-dithiobis (2-nitrobenzoic acid) (DTNB) demonstrated the responsiveness of SNMs to glutathione (GSH). In vitro cytotoxicity assays demonstrated that SNMs had a greater targeting efficacy for four types of cancer cells (HeLa, HCT-116, A549, and HepG2) compared to normal 293T cells. Cellular uptake studies revealed that HeLa cells more readily absorbed SNMs, leading to their accumulation in the tumor cell cytoplasm. Fluorescence colocalization assays verified that SNMs efficiently accumulated in organelles related to energy metabolism and signaling, including mitochondria and the endoplasmic reticulum, affecting cellular metabolic death. Both flow cytometry and confocal nuclear staining assays confirmed that SNMs effectively induced apoptosis over time, ultimately resulting in the death of cancer cells. These findings demonstrate that SNMs exhibit excellent targeting ability, responsiveness, high bioavailability, and stability, suggesting significant potential in drug delivery applications.

UCHL1 Overexpression Is Related to the Aggressive Phenotype of Non-small Cell Lung Cancer.

Ubiquitin C-terminal hydrolase L1 (UCHL1), which encodes thiol protease that hydrolyzes a peptide bond at the C-terminal glycine residue of ubiquitin, regulates cell differentiation, proliferation, transcriptional regulation, and numerous other biological processes and may be involved in lung cancer progression. UCHL1 is mainly expressed in the brain and plays a tumor-promoting role in a few cancer types; however, there are limited reports regarding its role in lung cancer. Single-cell RNA (scRNA) sequencing using 10X chromium v3 was performed on a paired normal-appearing and tumor tissue from surgical specimens of a patient who showed unusually rapid progression. To validate clinical implication of the identified biomarkers, immunohistochemical (IHC) analysis was performed on 48 non-small cell lung cancer (NSCLC) tissue specimens, and the correlation with clinical parameters was evaluated. We identified 500 genes overexpressed in tumor tissue compared to those in normal tissue. Among them, UCHL1, brain expressed X-linked 3 (BEX3), and midkine (MDK), which are associated with tumor growth and progression, exhibited a 1.5-fold increase in expression compared to that in normal tissue. IHC analysis of NSCLC tissues showed that only UCHL1 was specifically overexpressed. Additionally, in 48 NSCLC specimens, UCHL1 was specifically upregulated in the cytoplasm and nuclear membrane of tumor cells. Multivariable logistic analysis identified several factors, including smoking, tumor size, and high-grade dysplasia, to be typically associated with UCHL1 overexpression. Survival analyses using The Cancer Genome Atlas (TCGA) datasets revealed that UCHL1 overexpression is substantially associated with poor survival outcomes. Furthermore, a strong association was observed between UCHL1 expression and the clinicopathological features of patients with NSCLC. UCHL1 overexpression was associated with smoking, tumor size, and high-grade dysplasia, which are typically associated with a poor prognosis and survival outcome. These findings suggest that UCHL1 may serve as an effective biomarker of NSCLC.

Exosome-decorated bio-heterojunctions reduce heat and ROS transfer distance for boosted antibacterial and tumor therapy

Photothermal and photodynamic therapies represent effective modalities for combatting bacteria and tumor cells. However, therapeutic outcomes are constrained by limitations related to the heat and reactive oxygen species (ROS) transfer distance from photosensitizers to targets. To address this issue, we have devised and developed exosome-decorated bio-heterojunctions (E-bioHJ) consisted of MXene (Ti3C2), liquid metal (LM) and exosomes sourced from CT26 cells to enhance the phototherapeutic consequences. Engineering E-bioHJ enhances phototherapeutic effect in antibacterial and anti-tumor treatment, which is ascribed to reducing transfer distance of the heat and ROS. When adorned with exosomes, E-bioHJ is targetedly delivered into the cytoplasm of tumor cells to generate amount heat and ROS under 808 nm near-infrared radiation, which further induces mitochondrial dysfunction and apoptosis/necroptosis. As envisaged, this study presents a novel tactic to enhance the antibacterial and anti-tumor efficacy of biomaterials through reducing the heat and ROS delivery travel distance.

Features of Vascular Endothelial Growth Factor А (Vegf-А) Expression in Atypical Teratoid-Rhabdoid Tumor of the Central Nervous System in Children

Atypical teratoid rhabdoid tumors (AT/RT) are highly malignant neoplasms composed of poorly differentiated germline cells and a variable number of rhabdoid cells that can differentiate into different cell types. A key diagnostic factor for AT/RT is the absence of nuclear expression of the INI1 protein (SMARCB1 gene product) when tested with an anti-INI1 antibody in an immunohistochemical study. Currently, there is no standard treatment for AT/RT, but treatment approaches have become more aggressive and multimodal. In one study, AT/RT was found to be positive for VEGF. The purpose of our study was to examine the expression of VEGF-A in AT/RT in the central nervous system of children. We analyzed biopsy samples from 6 male patients aged 3 months to 8 years (average age 4.5 years). The level of VEGF-A expression was measured by counting the number of brown pixels in the cytoplasm of tumor cells using the PhotoM program in 1 mm2 of tumor tissue. Immunohistochemistry (IHC) revealed varying levels of cytoplasmic staining with an antibody to VEGF-A in 5 out of 6 patients, with the youngest patient (No. 6) showing staining only in endothelial cells. In conclusion, children with AT/RT exhibit different patterns of VEGF-A expression, potentially dependent on the molecular subgroup. Identifying this marker in these highly aggressive tumors could aid in developing new treatment protocols and determining which patients would benefit most from treatment.

Development of a deep-learning model tailored for HER2 detection in breast cancer to aid pathologists in interpreting HER2-low cases.

Over 50% of breast cancer cases are "Human epidermal growth factor receptor 2 (HER2) low breast cancer (BC)", characterized by HER2 immunohistochemistry (IHC) scores of 1+ or 2+ alongside no amplification on fluorescence in situ hybridization (FISH) testing. The development of new anti-HER2 antibody-drug conjugates (ADCs) for treating HER2-low breast cancers illustrates the importance of accurately assessing HER2 status, particularly HER2-low breast cancer. In this study we evaluated the performance of a deep-learning (DL) model for the assessment of HER2, including an assessment of the causes of discordances of HER2-Null between a pathologist and the DL model. We specifically focussed on aligning the DL model rules with the ASCO/CAP guidelines, including stained cells' staining intensity and completeness of membrane staining. We trained a DL model on a multicentric cohort of breast cancer cases with HER2-IHC scores (n = 299). The model was validated on two independent multicentric validation cohorts (n = 369 and n = 92), with all cases reviewed by three senior breast pathologists. All cases underwent a thorough review by three senior breast pathologists, with the ground truth determined by a majority consensus on the final HER2 score among the pathologists. In total, 760 breast cancer cases were utilized throughout the training and validation phases of the study. The model's concordance with the ground truth (ICC = 0.77 [0.68-0.83]; Fisher P = 1.32e-10) is higher than the average agreement among the three senior pathologists (ICC = 0.45 [0.17-0.65]; Fisher P = 2e-3). In the two validation cohorts, the DL model identifies 95% [93% - 98%] and 97% [91% - 100%] of HER2-low and HER2-positive tumours, respectively. Discordant results were characterized by morphological features such as extended fibrosis, a high number of tumour-infiltrating lymphocytes, and necrosis, whilst some artefacts such as nonspecific background cytoplasmic stain in the cytoplasm of tumour cells also cause discrepancy. Deep learning can support pathologists' interpretation of difficult HER2-low cases. Morphological variables and some specific artefacts can cause discrepant HER2-scores between the pathologist and the DL model.

Demonstration of Epstein-Barr Virus by In Situ Hybridization in Papillary Thyroid Carcinomas Developing on Background of Hashimoto's Thyroiditis.

This study aimed to demonstrate the role of Epstein-Barr Virus (EBV) in papillary thyroid carcinomas (PTC) developing on the background of Hashimoto's thyroiditis (HT). The presence of EBV in tumoral tissue, lymphocytes, and peritumoral normal thyroid tissue was investigated using the in situ hybridization method in paraffin blocks. The subtypes of PTC, tumor diameter, TNM stage, multifocality, invasion of thyroid capsule, perineural invasion, and muscular tissue invasion were identified and compared according to EBV involvement. Eighty-one patients with HT diagnosis, with 93.8% (n=76) female and 6.2% (n=5) male, were included in the study. Papillary microcarcinoma was the pathological diagnosis in 24.2% (n=15) of the cases. EBV was identified in 58.06% (n=36) of the tumor cells nuclei, 58.06% (n=36) in the tumor cell cytoplasm, 16.12% (n=10) in tumor infiltrative lymphocytes, and 53.2% (n=33) in normal parenchymal follicle epithelial cells (NPFEC). In the T2 stage, the rate of EBV nuclear positivity in patients was significantly higher (p=0.034). The classic variant of papillary carcinoma was accompanied by a significantly higher rate of EBV-negative NPFEC (67.6%, p=0.049). In multifocal tumors, EBV positivity was found to be significantly higher in lymphocytes in the surrounding tissues (58.3%, p=0.034). A significant increase in EBV positivity in the surrounding tissue lymphocytes was observed in multifocal PTC developing on a background of HT. This suggests a possible association between HT and EBV.

Utilizing a Pathomics Biomarker to Predict the Effectiveness of Bevacizumab in Ovarian Cancer Treatment.

The purpose of this investigation is to develop and initially assess a quantitative image analysis scheme that utilizes histopathological images to predict the treatment effectiveness of bevacizumab therapy in ovarian cancer patients. As a widely accessible diagnostic tool, histopathological slides contain copious information regarding underlying tumor progression that is associated with tumor prognosis. However, this information cannot be readily identified by conventional visual examination. This study utilizes novel pathomics technology to quantify this meaningful information for treatment effectiveness prediction. Accordingly, a total of 9828 features were extracted from segmented tumor tissue, cell nuclei, and cell cytoplasm, which were categorized into geometric, intensity, texture, and subcellular structure features. Next, the best performing features were selected as the input for SVM (support vector machine)-based prediction models. These models were evaluated on an open dataset containing a total of 78 patients and 288 whole slides images. The results indicated that the sufficiently optimized, best-performing model yielded an area under the receiver operating characteristic (ROC) curve of 0.8312. When examining the best model's confusion matrix, 37 and 25 cases were correctly predicted as responders and non-responders, respectively, achieving an overall accuracy of 0.7848. This investigation initially validated the feasibility of utilizing pathomics techniques to predict tumor responses to chemotherapy at an early stage.

High Expression of Nicotinamide Adenine Dinucleotide Phosphate Oxidase 4 Correlates with High Grade of Clear Cell Renal Cell Carcinoma

Background Clear cell renal cell carcinoma (ccRCC) is the most common histological subtype in renal cell tumors (RCT). This is a very aggressive cancer, often diagnosed at advanced stage leading to increase morbidity and mortality rates. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 4 (NOX-4) expression plays role in the regulation of several cancer types. The expression of NOX-4 is associated with a poorer degree of differentiation and a shorter survival in RCT. Therefore, NOX-4 expression is predictor of prognosis and potential therapeutic target. Research on the correlation of NOX-4 expression in RCTs is still very limited, while Correlation of NOX-4 expression in ccRCC does not exist yet. Objective To assess the relationship between NOX-4 expression and ISUP grading in ccRCC. Method In this study, 30 slide samples and ccRCC paraffin blocks were diagnosed histopathologically at Department of Anatomic Pathology of H. Adam Malik General Hospital from January 2012 to April 2022. All clinicopathological data were taken from medical records/pathology archives. The expression of NOX-4 immunohistochemical was assessed in nucleus and cytoplasm of tumor cells. The assessment was carried out semiquantitatively based on intensity and percentage of expressed tumor cells. The relationship between NOX-4 expression and ISUP grading was assessed using Fisher's exact test. Results Of the 30 ccRCC samples, high grade ccRCC was found in 21 cases (70%) and strong expression of NOX-4 in cytoplasm and nucleus in 20 cases (66.7%). Strong expression of NOX-4 in high grade ccRCC was found in 17 cases (81.0%). A significant relationship was observed between NOX-4 expression and ISUP grading on ccRCC with a p-value of 0.030 (p<0.05) using Fisher's exact test. Conclusion There is a significant relationship between NOX-4 expression and ISUP grading in ccRCC. Therefore, NOX-4 expression can be used for the prognosis of ccRCC and therapy development.

The effects of enoxaparin treatment in a xenograft mouse model of oral squamous cell carcinoma: A pilot study.

Recent studies suggest that enoxaparin may have therapeutic effects on oral squamous cell carcinoma. We aimed to assess this effect utilizing xenograft mouse model through evaluations of proliferation and angiogenesis markers at the RNA and protein levels. Mice were divided into enoxaparin treatment (n = 4), positive control (n = 4) and negative control (n = 3) groups. Immunohistochemical analyses were performed utilizing Bcl-2, Bax and Ki-67 antibodies. Expression levels of proliferation and apoptosis related genes were calculated utilizing qRT-PCR. Time-dependent proliferation assays were performed in OSC-19 and HEK293 cell-lines. Bax antibody showed positive staining in the cytoplasm and nuclei of tumor cells, while Bcl-2 antibody displayed staining only in the cytoplasm. A proliferation index of 15%-20% was found in all groups with the Ki-67 marker indicating no metastasis. Enoxaparin treatment caused decrease in BCL2, BAX and CCNB1 genes' expressions. Compared to HEK293, proliferation assays demonstrated higher division rates in OSC-19 with a significant decrease in viability after 96 h. Reduced BCL-2 expression indicates a regression of tumor growth, but reduced BAX expression is not correlated with increased apoptosis. Despite the aggressive nature of OSC-19, our results showed a low cell viability with a high division rate when compared with the control HEK293. This paralleled our in vivo findings that showed absence of lymph node metastasis across all mice groups. This discrepancy with the literature suggests that further investigations of the underlying mechanisms and protein-level analyses are needed to draw definitive conclusions about the effect of enoxaparin on OSC-19 behavior.

Evaluation of Paxillin Expression in Odontogenic Cysts and Tumors.

Paxillin (PXN) is one of the proteins involved in cell adhesion. PXN and integrins constitute a key site for the focal adhesion between the cell and extracellular matrix. Several studies have shown that PXN is a factor in tumor formation, progression, invasion, and metastasis. This study evaluated PXN expression in four types of odontogenic lesions with different aggressive behaviors. In this retrospective cross-sectional study, PXN expression was immunohistochemically assessed in 68 paraffin-embedded tissue samples from patients with the confirmed diagnosis of four types of odontogenic lesions, including 14 dentigerous cysts (DC), 20 odontogenic keratocyst (OKC), 16 unicystic ameloblastoma, and 18 solid ameloblastoma. The PXN expression in these samples were scored based on the percentage and intensity of immunoreactivity, and compared among the groups by Chi-square test. The PXN marker was detected in the cytoplasm of tumor cells (unicystic and solid ameloblastoma) and the epithelial layer of cysts (DC and OKC). The intensively stained marker of PXN was observed in 9 cases (64.3%) of the DC, 14 cases (70%) of OKC, 12 cases (75%) of unicystic ameloblastoma, and 13 cases (72.2%) of solid ameloblastoma. However, there was not statistical difference of PXN protein expression between DC and OKC (p Value = 0.51) and unicystic and solid ameloblastoma (p = 0.58), also the same was true for cysts and tumors (p = 0.37). The expression of PXN is not related to the biological behaviors of odontogenic lesions.

Cytoplasmic PPARγ Significantly Correlates With P53 Immunohistochemical Expression and Tumor Size in Localized Tenosynovial Giant Cell Tumor.

Tenosynovial giant cell tumor (TGCT)is a monoarticular fibrohistiocytic benign or locally aggressive soft tissue tumor that originates from the synovium of joints, bursae, and tendon sheaths.It has an inflammatory neoplastic nature, with a clinical presentationranging from pain, swelling, stiffness, and limited range of movement to joint instability and blockage. Its uncommon incidence leads to a poorly understood pathogenesis. Localized formsof TGCT (LTGCT) can cause significant morbidity, interfere with daily patient activities, and decrease the patient's quality of life in challenging cases. This study aimed to investigate the immunohistochemical expression of PPARγ (peroxisome proliferator-activated receptor gamma) and P53 in LTGCT to understand the disease better and offer potential therapeutic targets. The study is cross-sectional, in which 27 LTGCT cases were collectedfrom the Pathology Department, Faculty of Medicine, Cairo University, Cairo, Egypt.Solitary and multiple LTGCT cases retrieved between January 2018 and December 2022 were included, and immunohistochemically stained with anti-PPARγ and P53 antibodies. The TGCT samples were excluded if they were insufficient for sectioning, processing, and interpretation, over-fixed, had process artifacts, or were of the diffuse TGCT type. Scoring of stain expression was performed by ImageJ (National Institutes of Health, Bethesda, MD) analysis using the threshold method and was expressed in percent area/high power field. Clinicopathological correlations were analyzed. All the 27 collected LTGCT cases were located in the small joints of patients' hands. Cases with solitary LGTCTs constituted 55.6%(n = 15), while 44.4% (n = 12) had multipleLTGCTs related to one affected site/case (e.g., multiple tumors in one finger). PPARγ was expressed in the cytoplasm of mononuclear and multinucleated tumor cells and foamy histiocytes, while P53 expression was mainly in mononuclear cells' nuclei. PPARγ significantly correlated with P53 expression (r = 0.9 and P = 0.000). PPARγ (r = 0.4 and P = 0.02) and P53 (r = 0.5 and P = 0.01) were positively correlated with tumor size. Only P53 expression was positively correlated with tumor multiplicity (r = 0.4 and P = 0.03). Using the receiver operating characteristic curve test, the P53 cutoff score detecting the multiplicity of TGCTs was ≥20.5%, with a 75% sensitivity and 80% specificity. PPARγ and P53 have a significant role in LTGCT growth, while P53 plays a role in tumor multiplicity. They can be possible targets in LTGCTs unfit for excision.

Abstract PO4-25-08: Secretory breast carcinoma (SBC) in an 8-year-old male: Report of an exceptionally rare case with review of clinicoradiologic and pathomolecular findings of SBC in the male pediatric population

Abstract Secretory breast carcinoma (SBC) is a rare and distinct disease, comprising approximately 0.01% of all breast carcinomas. This is a case of an 8-year-old boy who presented with a 1.5-year history of a non-tender, slowly enlarging left subareolar mass, not associated with nipple discharge. Serologic workup revealed minimally elevated prolactin with normal-range FSH, LH, estradiol and testosterone, as well as normal-range beta-hCG, LDH and AFP. Ultrasound showed a circumscribed solid hypoechoic lesion with internal arterial vascularity, favoured to represent a sebaceous cyst. Excisional biopsy was performed, and histologic examination revealed the classic intra- and extracellular secretory material typically seen in SBC. Immunohistochemical (IHC) stains for mammaglobin, S-100 and high molecular weight keratin (CK5) were positive. ER was positive, though not strong staining, and PR and HER2 were negative. A screen for NTRK fusions with pan-TRK IHC was positive. No skin, perineural or lymphovascular invasion was identified. RNA-based NGS demonstrated ETV6-NTRK3 fusion and confirmed the diagnosis of SBC. The patient underwent mastectomy and SLNB, which showed no residual tumor and no positive lymph nodes. A full metastatic work-up was not performed and the patient did not receive neoadjuvant therapy. First described as “juvenile breast carcinoma,” SBC has since been recognized to occur across a wide age range, with most cases occurring in adults. SBC in male pediatric patients is exceptionally rare, with fewer than 20 cases reported in the literature. In this demographic, SBC is usually detected clinically as a subareolar mass not associated with nipple discharge, and appears as a well-circumscribed mass on imaging, mimicking benign entities. SBC is usually detected at a low stage, treated surgically, and follows a clinically indolent course with excellent outcomes. Positive axillary nodes have been identified in a few male children, who subsequently underwent chemo- and/or radiotherapy, with no recorded recurrences in these cases. Distant metastases are an extremely uncommon feature of SBC and have not been reported in the pediatric male population; however, due to the rarity of this diagnosis, particularly in this demographic, robust long-term follow-up data are still lacking. The name “secretory” derives from the histologic appearance of the dense eosinophilic secretory material seen in tumor lumina and bubbly material in the cytoplasm of tumor cells. Similar to the cytologic characteristics of other translocation-driven neoplasms, SBC shows minimal pleomorphism. Mitoses are absent or rare. IHC stains are typically positive for S-100 and high molecular weight keratins and negative for ER, PR, and HER2. Despite the immunophenotypic similarities to triple negative/basal-like carcinomas (TNBC/BLC) the molecular relationship between SBC and TNBC/BLC is unclear. Though initially defined by its distinctive secretions on histology, SBC is now chiefly characterized by a balanced translocation, t(12;15)(p13;q25), resulting in oncogenic ETV6-NTRK3 fusion. Recent cases have demonstrated this using a variety of molecular techniques, including FISH, RT-PCR, and in our case, RNA-based NGS. This translocation also characterizes non-breast secretory carcinomas and has been identified in several other neoplasms, including pediatric mesenchymal tumours and adult acute myeloid leukemia. The rarity of SBC, particularly in the male pediatric population, underlines the necessity of this case report. Additionally, this case highlights the importance of considering SBC as a potential diagnosis in male children presenting with a breast mass, even in the presence of benign imaging characteristics. Table 1: Clinicoradiologic and pathomolecular characteristics of SBC in pediatric males AND, axillary node dissection; CT, chemotherapy; ER, estrogen receptor; FISH, fluorescence in situ hybridization; LE, local excision; MRM, modified radical mastectomy; nd, no data; NED, no evidence of disease; NGS, next-generation sequencing; NS, nodal sampling; PA, periareolar; PR, progesterone receptor; RA, retroareolar; RM, radical mastectomy; PCR, polymerase chain reaction; RT, radiotherapy; SA, subareolar; SLNB, sentinel lymph node biopsy; SM, simple mastectomy; WLE, wide local excision *Not reported as strong staining Table 2: Immunohistochemical phenotype of secretory breast carcinoma EMA, epithelial membrane antigen; ER, estrogen receptor; PR, progesterone receptor *Not reported as strong staining Table 3: Laboratory investigations AFP, alpha fetoprotein; FSH, follicle stimulating hormone; hCG, human chorionic gonadotropin; LDH, lactate dehydrogenase; LH, luteinizing hormone Citation Format: Rebecca Brassington, Svetlana Silverman, Ioana Bratu. Secretory breast carcinoma (SBC) in an 8-year-old male: Report of an exceptionally rare case with review of clinicoradiologic and pathomolecular findings of SBC in the male pediatric population [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-25-08.

EXPRESSION PATTERN OF THE MRPS18 FAMILY PROTEINS IN CHORDOMA

Chordomas are rare tumors of the axial skeleton that arise from the notochord; they are observed usually at the base of the skull and rarer along the spine. Chordomas are resistant to conventional chemotherapy, that is why it is important, to find the new diagnostic and/or prognostic markers. We have shown recently, that genes of the mitochondrial ribosomal protein S18 (MRPS18) family are differentially expressed in gliomas. The MRPS18-2 oncoprotein is expressed at higher levels in cancerous cells compared to their normal counterpart. Aim. We wanted to investigate the peculiarities of the expression pattern of the MRPS18 family genes in chordoma to better understand their role in cancerogenesis, using an immunohistochemical analysis on chordoma clinical samples. Methods. 15 specimens of chordoma of the base of the skull and 5 samples of the spine localization were studied retrospectively. The specific antibodies against MRPS18-1, MRPS18-2, MRPS18-3, and RB was used for an immunohistochemical analysis. All cases were stained in parallel with appropriate negative control. Results. We found that MRPS18 family proteins are differentially expressed in chordoma tissues. MRPS18-1 showed the strongest signal in all tumor samples. The lowest level of the signal intensity was recorded for the MRPS18-3 protein. In several samples the RB signal was detected in cytoplasm of tumor cells. Importantly, in these samples the strong staining for MRPS18-2 was observed as well. The fine mechanism of sequestering of RB in cytoplasm of cancer cells should be further elucidated. Conclusion, These preliminary results should lead to a larger work on a role of RB-MRPS18 family proteins interaction for chordoma development.

Dual miRNA-Triggered DNA Walker Assisted by APE1 for Specific Recognition of Tumor Cells.

The identification of a specific tumor cell is crucial for the early diagnosis and treatment of cancer. However, it remains a challenge due to the limited sensitivity and accuracy, long response time, and low contrast of the recent approaches. In this study, we develop a dual miRNA-triggered DNA walker (DMTDW) assisted by APE1 for the specific recognition of tumor cells. miR-10b and miR-155 were selected as the research models. Without miR-10b and miR-155 presence, the DNA walker remains inactive as its walking strand of W is locked by L1 and L2. After miR-10b and miR-155 are input, the DNA walker is triggered as miR-10b and miR-155 bind to L1 and L2 of W-L1-L2, respectively, unlocking W. The DNA walker is driven by endogenous APE1 that is highly catalytic and is highly expressed in the cytoplasm of tumor cells but barely expressed in normal cells, ensuring high contrast and reaction efficiency for specific recognition of tumor cells. Dual miRNA input is required to trigger the DNA walker, making this strategy with a high accuracy. The DMTDW strategy exhibited high sensitivity for miRNA analysis with a detection limit of 44.05 pM. Living cell-imaging experiments confirmed that the DMTDW could effectively respond to the fluctuation of miRNA and specifically identified MDA-MB-231 cells from different cell lines. The proposed DMTDW is sensitive, rapid, and accurate for specific tumor cell recognition. We believe that the DMTDW strategy can become a powerful diagnostic tool for the specific recognition of tumor cells.

Co-encapsulation of granzyme B and perforin in nanocapsules for tumour therapy: biomimicking immune cells

Granzyme B (GrB)-based immunotherapy is of interest for cancer treatment. However, insufficient cellular uptake and a lack of targeting remain challenges to make use of GrB for solid tumour therapy. As GrB induced cell death requires the help of perforin (PFN), we designed a system (nGPM) for the co-delivery of GrB and PFN. Therefore, GrB and PFN were loaded in a porous polymeric nanocapsule rich in acetylcholine analogues and matrix metalloproteinase-2 (MMP-2) responsive peptides. The neutrally charged nGPM nanocapsules showed as long circulating time and accumulated at the tumour sites. Once in the tumour the outside shell of nanocapsules became degraded by overexpressed MMP-2 proteases, resulting in the release of GrB and PFN. We found that the PFN complex formed small pores on the surface of tumour cells which allow GrB to enter the cytoplasm of tumour cells inducing cell apoptosis and tumour suppression significantly.

Immunohistochemical expression of Beclin-1 and LC3B in benign and borderline ovarian tumors

Background Epithelial ovarian tumors account for ⁓90% of ovarian cancers. Recently conducted studies have demonstrated autophagy role in tumor development and progression. Autophagic markers include Beclin-1, essential for autophagosome formation and LC3B, required for the elongation step during autophagy. Aim To investigate Beclin-1 and LC3B expression in benign and borderline epithelial ovarian tumors along with its relation to clinicopathologic parameters. Materials and methods Thirty-six cases of epithelial ovarian neoplasms were subjected to hematoxylin and eosin staining and immunohistochemical staining with Beclin-1 and LC3B. Results Beclin-1 was expressed in the nucleus and/or the cytoplasm of tumor cells. The cytoplasmic expression of Beclin-1 was significantly more frequent in borderline cases compared with benign tumors. High Beclin-1 cytoplasmic expression was significantly associated with tumor size, gross appearance, and tumor histological type. Benign Brenner tumors were the only benign tumors expressing cytoplasmic Beclin-1 localization. Regarding LC3B expression, nuclear and cytoplasmic subcellular localizations were also detected. Cytoplasmic LC3B localization was significantly more frequent in the borderline groups. A significant relation was observed between high cytoplasmic LC3B expression and tumor size, tumor gross appearance, and histological type. The majority of benign Brenner tumors showed a high cytoplasmic LC3B expression. Conclusions The expression of Beclin-1 and LC3B varies in benign versus borderline epithelial ovarian tumors. Cytoplasmic expression is predominant in borderline tumors. Beclin-1 and LC3B cytoplasmic expression is significantly high in small-sized and solid benign tumors. Benign Brenner tumors are the only benign tumors showing cytoplasmic Beclin-1 and LC3B localization.

Abstract 6182: Artificial intelligence (AI)-powered quantification of glypican-3 (GPC3) expression facilitates patient selection for GPC3-targeted therapy in solid tumors

Abstract Introduction: GPC3 is a glycosyl-phosphatidylinositol-anchored cell surface glycoprotein that functions to control cell growth and differentiation in early development. Membrane GPC3 is highly expressed in hepatocellular carcinoma (HCC) and moderately in squamous non-small cell lung cancer (SQ-NSCLC) with limited expression in normal tissues which makes it an attractive target for immune cell engagers. This study aims to develop an AI-powered algorithm to evaluate GPC3 expression and compare it with manual immunohistochemistry (IHC) scoring in HCC, adeno-NSCLC and SQ-NSCLC. Methods: 147 NSCLC (94 SQ-NSCLC and 53 adeno-NSCLC) and 113 HCC FFPE tumor blocks were obtained from commercial sources. GPC3 protein level was quantified by a qualified IHC assay for tumor cell membrane and cytoplasm individually. PD-L1 expression is reported as tumor cell proportion score (TPS), combined positive score (CPS) and immune cell proportion score (IPS/IC) TPS, regardless of staining pattern and intensity. FFPE slides were stained for GPC3+ (GC33, Ventana), PD-L1+ (22C3, Dako) cells and then scanned at 20x using the Aperio Versa8 scanner. Machine learning (ML) models were developed using the digitized whole slide images to identify GPC3+ tumor area. Data-driven cutoffs were applied to model-generated human-interpretable features of GPC3+ percentage of tumor area to classify samples as positive and negative. As an orthogonal approach to the GPC3+ percentage cutoff method, tissue and cell annotations were used to train a convolutional neural network (CNN) to classify GPC3 positivity. Results: The epidemiology profiling results revealed that GPC3 protein is highly expressed in HCC (57.5% cases with > 1% positive cells), followed by SQ-NSCLC (52.1%) and adeno-NSCLC (5.7%). GPC3 expression was only detected in tumor cells with two staining patterns observed: membrane + cytoplasm and cytoplasm-only. There was no significant correlation between GPC3 and PD-L1 expression across tumor types, although a trend of lower level of PD-L1 was observed in the samples with GPC3 ≥50%. ML model quantification of GPC3+ and GPC3- showed high concordance with the consensus score of 2 independent pathologists on a test set of held-out samples not used in training. This concordance was comparable to that of individual annotators to consensus. Instead, GPC3 positivity results using both cutoff and CNN methods showed moderate concordance with 2-way pathologist consensus. Conclusions: Here we profiled GPC3 expression across tumor types and showed high level of expression in HCC followed by SQ-NSCLC. We have established an AI-powered digital pathology platform that can provide a standardized, scalable, and reproducible method of characterizing GPC3 positivity to support further patient selection in clinical study. Citation Format: Ellen Meng, Robin Meng, Ying Tong, Yuchen Li, Hong Wang, Paola Fiorentini, Elham Attieh, Qi Tang, Asma Kefsi, Serena Masciari, Giovanni Abbadessa, Cecile Combeau, Lei Tang, Benoit Pasquier, Rui Wang. Artificial intelligence (AI)-powered quantification of glypican-3 (GPC3) expression facilitates patient selection for GPC3-targeted therapy in solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6182.