Total Osteopontin and Its Isoform OPN4 Are Differently Expressed in Respiratory Samples during Influenza A(H1N1)pdm09 Infection and Progression.

Influenza A virus (IAV) infection affects the human respiratory tract, causing an acute and highly contagious disease. Individuals with comorbidities and in the extremes of age are classified as risk groups for serious clinical outcomes. However, part of the severe infections and fatalities are observed among young healthy individuals. Noteworthy, influenza infections lack specific prognostic biomarkers that would predict the disease severity. Osteopontin (OPN) has been proposed as a biomarker in a few human malignancies and its differential modulation has been observed during viral infections. However, OPN expression levels in the primary site of IAV infection have not been previously investigated. Therefore, we evaluated the transcriptional expression patterns of total OPN (tOPN) and its splicing isoforms (OPNa, OPNb, OPNc, OPN4, and OPN5) in 176 respiratory secretion samples collected from human influenza A(H1N1)pdm09 cases and a group of 65 IAV-negative controls. IAV samples were differentially classified according to their disease severity. tOPN was more frequently detected in IAV samples (34.1%) when compared with the negative controls (18.5%) (p < 0.05), as well as in fatal (59.1%) versus non-fatal IAV samples (30.5%) (p < 0.01). OPN4 splice variant transcript was more prevalent in IAV cases (78.4%) than in the negative controls (66.1%) (p = 0.05) and in severe cases (85.7%) in relation to the non-severe ones (69.2%) (p < 0.01). OPN4 detection was also associated with severity symptoms such as dyspnea (p < 0.05), respiratory failure (p < 0.05), and oxygen saturation < 95% (p < 0.05). In addition, the OPN4 expression level was increased in the fatal cases of respiratory samples. Our data indicated that tOPN and OPN4 had a more pronounced expression pattern in IAV respiratory samples, pointing to the potential use of these molecules as biomarkers to evaluate disease outcomes.

The Intracellular and Secreted Sides of Osteopontin and Their Putative Physiopathological Roles.

Classically, osteopontin (OPN) has been described as a secreted glycophosprotein. Indeed, most data concerning its physiological and pathological roles are mainly related to the secreted OPN (sOPN). However, there are several instances in which intracellular OPN (iOPN) has been described, presenting some specific roles in distinct experimental models, such as in the immune system, cancer cells, and neurological disorders. We herein aimed to highlight and discuss some of these secreted and intracellular roles of OPN and their putative clinical and biological impacts. Moreover, by consolidating data from the OPN protein database, we also analyzed the occurrence of signal peptide (SP) sequences and putative subcellular localization, especially concerning currently known OPN splicing variants (OPN-SV). Comprehending the roles of OPN in its distinct cellular and tissue environments may provide data regarding the additional applications of this protein as biomarkers and targets for therapeutic purposes, besides further describing its pleiotropic roles.

Multiparametric flow cytometry directing the evaluation of CRLF2 rearrangements and JAK2 status in pediatric B cell precursor acute lymphoblastic leukemia.

INTRODUCTION: This study aimed to determine whether cytokine receptor-like factor 2 (CRLF2) antigen expression evaluated using multiparametric flow cytometry (MFC) could predict the genotype of CRLF2 and Janus kinase 2 (JAK2) status for application in the diagnosis of pediatric B-cell precursor acute lymphoblastic leukemia (BCP-ALL). METHODS: A total of 321 BCP-ALL bone marrow samples were collected, 291 at diagnosis and 13 at first relapse, while 17 samples were excluded due to low cellular viability. The CRLF2 antigen expression was evaluated using flow cytometry (percentage of positivity and median fluorescence intensity [MFI]). The CRLF2 transcript levels were assessed via quantitative reverse transcription polymerase chain reaction using SYBR Green. The CRLF2 rearrangements (CRLF2-r) were identified using the CRLF2 break-apart probe via fluorescence in situ hybridization. Sanger sequencing was performed to identify the JAK2 exon 16 mutations. RESULTS: We observed that 60 of the 291 cases (20.6%) presented CRLF2 antigen positivity, whereas the CRLF2 transcript overexpression was found in 19 of 113 cases (16.8%). The JAK2 mutation was found in four out of 116 cases (3.4%), all of which had CRLF2 ≥10% of positive cells and intermediate or high MFI (p < 0.0001). In addition, in the 13 cases with the CRLF2-r, a positive correlation was found with the CRLF2 antigen intermediate (61.5%) MFI (p = 0.017). Finally, the CRLF2-positive antigen was identified in the BCP-ALL subclones. CONCLUSION: The identification of the CRLF2 antigen using the MFC, based on the percentage of positivity and MFI values, is a useful tool for predicting JAK2 mutations and CRLF2-r.

Multiparametric flow cytometry directing the evaluation of CRLF2 rearrangements and JAK2 status in pediatric B cell precursor acute lymphoblastic leukemia

Asbtract Introduction This study aimed to determine whether cytokine receptor-like factor 2 (CRLF2) antigen expression evaluated using multiparametric flow cytometry (MFC) could predict the genotype of CRLF2 and Janus kinase 2 (JAK2) status for application in the diagnosis of pediatric B-cell precursor acute lymphoblastic leukemia (BCP-ALL). Methods A total of 321 BCP-ALL bone marrow samples were collected, 291 at diagnosis and 13 at first relapse, while 17 samples were excluded due to low cellular viability. The CRLF2 antigen expression was evaluated using flow cytometry (percentage of positivity and median fluorescence intensity [MFI]). The CRLF2 transcript levels were assessed via quantitative reverse transcription polymerase chain reaction using SYBR Green. The CRLF2 rearrangements (CRLF2-r) were identified using the CRLF2 break-apart probe via fluorescence in situ hybridization. Sanger sequencing was performed to identify the JAK2 exon 16 mutations. Results We observed that 60 of the 291 cases (20.6%) presented CRLF2 antigen positivity, whereas the CRLF2 transcript overexpression was found in 19 of 113 cases (16.8%). The JAK2 mutation was found in four out of 116 cases (3.4%), all of which had CRLF2 ≥10% of positive cells and intermediate or high MFI (p < 0.0001). In addition, in the 13 cases with the CRLF2-r, a positive correlation was found with the CRLF2 antigen intermediate (61.5%) MFI (p= 0.017). Finally, the CRLF2-positive antigen was identified in the BCP-ALL subclones. Conclusion The identification of the CRLF2 antigen using the MFC, based on the percentage of positivity and MFI values, is a useful tool for predicting JAK2 mutations and CRLF2-r.

Bovine Lactoferrin Induces Cell Death in Human Prostate Cancer Cells.

Bovine lactoferrin (bLf) is a multifunctional protein widely associated with anticancer activity. Prostate cancer is the second most frequent type of cancer worldwide. This study was aimed at evaluating the influence of bLf on cell viability, cell cycle progression, reactive oxygen species (ROS) production, and rate of apoptosis in the human prostate cancer cell line (DU-145). MTT assay and trypan blue exclusion were used to analyze cell viability. Morphological changes were analyzed through optical microscopy after 24 h and 48 h of bLf treatment. FITC-bLf internalization and cellular damage were observed within 24 h by confocal fluorescence microscopy. Cell cycle analyses were performed by flow cytometry and propidium iodide. For caspases 3/7 activation and reactive oxygen species production evaluation, cells were live-imaged using the high-throughput system Operetta. The cell viability assays demonstrated that bLf induces cell death and morphological changes after 24 h and 48 h of treatment compared to control on DU-145 cells. The bLf internalization was detected in DU-145 cells, G1-phase arrest of the cell cycle, caspase 3/7 activation, and increased oxidative stress on bLf-treated cells. Our data support that bLf has an important anticancer activity, thus offering new perspectives in preventing and treating prostate cancer.

Osteopontin Expression in Thyroid Cancer: Deciphering EMT-Related Molecular Mechanisms.

Thyroid cancer is the most common tumor arising from the endocrine system and generally presents good prognosis. However, its aggressive subtypes are related to therapeutic resistance and early metastasis. Epithelial-mesenchymal transition (EMT) and its reverse process, the mesenchymal-epithelial transition (MET), are key events mediating cancer progression, including in thyroid cancer. The matricellular protein osteopontin (OPN) has been reported as a master regulator of EMT in many tumor types. Although high OPN expression has been described and associated with important aspects of thyroid cancer progression, there is no clear evidence regarding OPN as a regulator of EMT in thyroid cancer. Thus, taking together the known roles of OPN in the modulation of EMT in cancer and the information reporting the expression of OPN in thyroid tumor progression, this review aims at summarizing and discussing data related to EMT in thyroid cancer and its putative relation to the roles of OPN in the development of thyroid cancer. These data provide new insights into the molecular mechanisms by which OPN could potentially modulate EMT in thyroid tumors, generating evidence for future studies that may contribute to new therapeutic, prognostic and/or diagnostic tools.

LRP1B Expression as a Putative Predictor of Response to Pegylated Liposomal Doxorubicin Treatment in Ovarian Cancer.

BACKGROUND: Pegylated liposomal doxorubicin (PLD) is among the most active therapies for recurrent/progressive ovarian cancer (OC). Low-density lipoprotein receptor-related protein 1B (LRP1B) is one of the 10 most significantly deleted genes in human cancers. It mediates endocytosis of several factors from the cellular environment including liposomes. Although the LRP1B role in cancer has not been fully disclosed, its contribution to resistance to liposomal therapies has been hypothesized. This study aimed to evaluate the impact of LRP1B protein as a possible marker of response to PLD in patients with OC. METHODS: LRP1B expression and response to PLD were analyzed in OC cell lines by qRT-PCR and PrestoBlue viability assay, respectively. LRP1B protein expression was evaluated for the first time, in tumor samples from PLD-treated patients and controls (other chemotherapies) by immunohistochemistry. Association of LRP1B staining score (determined based on intensity and percentage of positively stained cells) with clinicopathological features, response to therapy and survival outcomes was evaluated. RESULTS: OC cells with increased expression of LRP1B were more sensitive to PLD. LRP1B staining score was associated with clinicopathological features, response to therapy, and survival outcomes. Higher LRP1B levels were associated with prolonged progression-free survival. This association was more evident in patients treated with PLD and in responders to PLD. CONCLUSION: Our results support a possible role of LRP1B as a predictor of response to PLD in patients with OC.

Susceptibility of lung cancer patients to COVID-19: A review of the pandemic data from multiple nationalities.

Several studies have highlighted that cancer patients tend to be more susceptible to develop severe infection and to die from COVID-19. Certain medical conditions such as immunosuppression, presence of comorbidities, and underlying pulmonary damage are possible determinants of disease severity, especially in lung cancer patients. While recent studies have shown that lung cancer is one of the most prevalent tumor types among COVID-19 cancer patients, we still have an incomplete view of how data from several countries work as a whole. The aim of this review was to investigate COVID-19 prevalence in lung cancer patient cohorts and their probability to develop severe illness and death when compared to nonlung cancer patients from multiple nationalities, including countries that have been the epicenters of the pandemic. We also focus on some intrinsic lung cancer features that might influence COVID-19 outcomes. An integrative view of the susceptibility of lung cancer patients might be especially relevant to assist physicians in evaluating the risks of COVID-19 in these patients, and to foster better decisions on treatment delay.

(3,3'-Methylene)bis-2-hydroxy-1,4-naphthoquinones induce cytotoxicity against DU145 and PC3 cancer cells by inhibiting cell viability and promoting cell cycle arrest.

We developed a novel method for the synthesis of bis-naphthoquinones (BNQ), which are hybrids of lawsone (2-hydroxy-1,4-naphthoquinone) and 3-hydroxy-juglone (3,5-dihydroxy-1,4-naphthoquinone). The anticancer activity of three synthesized compounds, named 4 (RC10), 5 (RCDFC), and 6 (RCDOH) was evaluated in vitro against two metastatic prostate cancer (PCa) cell lines, DU145 and PC3, using MTT assays. We found that 4 (RC10) and 5 (RCDFC) induced cytotoxicity against DU145 and PC3 cells. Flow cytometry analysis revealed that these two compounds promoted cell cycle arrest in G1/S and G2/M phases, increased Sub-G1 peak and induced inhibition in cell viability. We also showed that these effects are cell-type context dependent and more selective for these tested PCa cells than for HUVEC non-tumor cells. The two BNQ compounds 4 (RC10) and 5 (RCDFC) displayed promising anticancer activity against the two tested metastatic PCa cell lines, DU145 and PC3. Their effects are mainly associated with inhibition of cell viability, possibly through apoptotic cell death, besides altering the SubG1, G1/S and G2/M phases of cell cycle. 5 (RCDFC) compound was found to be more selective than 4 (RC10), when comparing their cytotoxic effects in relation to HUVEC non-tumoral cells. Future work should also test these compounds in combination with other chemotherapeutic drugs to evaluate their effects on further sensitizing drug-resistant metastatic PCa cells.

Bioactive compounds, antioxidant activity and antiproliferative effects in prostate cancer cells of green and roasted coffee extracts obtained by microwave-assisted extraction (MAE).

Coffee consumption has been investigated as a protective factor against prostate cancer. Coffee may be related to prostate cancer risk reduction due to its phytochemical compounds, such as caffeine, chlorogenic acids, and trigonelline. The roasting process affects the content of the phytochemicals and undesired compounds can be formed. Microwave-assisted extraction is an alternative to conventional extraction techniques since it preserves more bioactive compounds. Therefore, this study aimed to evaluate the phytochemical composition and the putative preventive effects in prostate cancer development of coffee beans submitted to four different coffee-roasting degrees extracted using microwave-assisted extraction. Coffea arabica green beans (1) were roasted into light (2), medium (3) and dark (4) and these four coffee samples were submitted to microwave-assisted extraction. The antioxidant capacity of these samples was evaluated by five different methods. Caffeine, chlorogenic acid and caffeic acid were measured through HPLC. Samples were tested against PC-3 and DU-145 metastatic prostate cancer cell lines regarding their effects on cell viability, cell cycle progression and apoptotic cell death. We found that green and light roasted coffee extracts had the highest antioxidant activity. Caffeine content was not affected by roasting, chlorogenic acid was degraded due to the temperature, and caffeic acid increased in light roasted and decreased in medium and dark roasted. Green and light roasted coffee extracts promoted higher inhibition of cell viability, caused greater cell cycle arrest in S and G2/M and induced apoptosis more compared to medium and dark roasted coffee extracts and the control samples. Coffee extracts were more effective against DU-145 than in PC-3 cells. Our data provide initial evidence that among the four tested samples, the consumption of green and light coffee extracts contributes to inhibit prostate cancer tumor progression features, potentially preventing aspects related to advanced prostate cancer subtypes.

Osteopontin­c isoform inhibition modulates ovarian cancer cell cisplatin resistance, viability and plasticity.

Osteopontin (OPN) is upregulated in several types of tumor and has been associated with chemoresistance. However, the contribution of OPN splicing isoforms (OPN­SIs) to chemoresistance requires further investigation. The present study aimed to evaluate the expression patterns of each tested OPN­SI in cisplatin (CDDP)­resistant ovarian carcinoma cell lines, focusing on the role of the OPN­c isoform (OPNc) in drug resistance. ACRP ovarian cancer cells resistant to CDDP, as well as their parental cell line A2780, were used. Analyses of the transcriptional expression of OPN­SIs, epithelial­mesenchymal transition (EMT) markers and EMT­related cytokines were performed using reverse transcription­quantitative PCR. OPNc was silenced in ACRP cells using anti­OPNc DNA oligomers and stably overexpressed by transfecting A2780 cells with a mammalian expression vector containing the full length OPNc cDNA. Functional assays were performed to determine cell proliferation, viability and colony formation. The results demonstrated that among the three tested OPN­SIs, OPNc was the most upregulated transcript in the ACRP cells compared with the parental A2780 cells. In addition, the expression levels of P­glycoprotein multidrug transporter were upregulated in CDDP­resistant ACRP cells compared with those in A2780 cells. OPNc knockdown sensitized ACRP cells to CDDP treatment and downregulated P­gp expression levels compared with those in the negative control group. Additionally, silencing of OPNc impaired cell proliferative and colony formation abilities, as well as reversed the expression levels of EMT markers and EMT­related cytokines compared with those in the negative control cells. Notably, although stable OPNc overexpression resulted in increased A2780 cell proliferation, it notably increased CDDP sensitivity compared with that in the cells transfected with a control vector. These results suggested that OPNc silencing may represent a putative approach to sensitize resistant ovarian cancer cells to chemotherapeutic agents.

Osteopontin-4 and Osteopontin-5 splice variants are expressed in several tumor cell lines.

Among osteopontin splice variants (OPN-SV), the expression profile of osteopontin-4 (OPN4) and osteopontin-5 (OPN5) has not been addressed in distinct cancer types. We herein aimed to investigate their expression in several cancer cell lines, besides comparing it in relation to the three previously described OPN-SV: OPNa, OPNb and OPNc. Total RNA from cancer cell lines, including prostate (PC3 and DU145), ovarian (A2780), breast (MCF-7 and MDA-MB-231), colorectal (Caco-2, HT-29 and HCT-116), thyroid (TT, TPC1 and 8505c) and lung (A549 and NCI-H460) was extracted, followed by cDNA synthesis. OPN-SV transcript analysis by RT-PCR or RT-qPCR were performed using OPN-SV specific oligonucleotides and gapdh and actin transcripts were used as housekeeping controls. OPN4 and OPN5 transcripts displayed co-expression in most tested cell lines. OPN4 was found expressed in similar or higher levels in relation to OPN5. Moreover, in most tested cell lines, OPN4 is also expressed in similar levels to OPNa or OPNb. The expression of OPN5 is also generally variable in relation to the other OPN-SV, but expressed in similar or higher levels in relation to OPNc, depending on each tested cell line. OPN4 and OPN5 seem to be co-expressed in several tumor types and OPN4 is one of the most overexpressed OPN-SV in distinct tumor cell lines. Once both OPN4 and OPN5 are differentially expressed and also evidence tumor-specific expression patterns, we hypothesize that similarly to the other OPN-SV, they also possibly contribute to key aspects of tumor progression, what should be further functionally investigated in distinct tumor models.

Molecular Aspects of Thyroid Calcification.

In thyroid cancer, calcification is mainly present in classical papillary thyroid carcinoma (PTC) and in medullary thyroid carcinoma (MTC), despite being described in benign lesions and in other subtypes of thyroid carcinomas. Thyroid calcifications are classified according to their diameter and location. At ultrasonography, microcalcifications appear as hyperechoic spots ≤ 1 mm in diameter and can be named as stromal calcification, bone formation, or psammoma bodies (PBs), whereas calcifications > 1 mm are macrocalcifications. The mechanism of their formation is still poorly understood. Microcalcifications are generally accepted as a reliable indicator of malignancy as they mostly represent PBs. In order to progress in terms of the understanding of the mechanisms behind calcification occurring in thyroid tumors in general, and in PTC in particular, we decided to use histopathology as the basis of the possible cellular and molecular mechanisms of calcification formation in thyroid cancer. We explored the involvement of molecules such as runt-related transcription factor-2 (Runx-2), osteonectin/secreted protein acidic and rich in cysteine (SPARC), alkaline phosphatase (ALP), bone sialoprotein (BSP), and osteopontin (OPN) in the formation of calcification. The present review offers a novel insight into the mechanisms underlying the development of calcification in thyroid cancer.

Predictive Biomarkers and Patient Outcome in Platinum-Resistant (PLD-Treated) Ovarian Cancer.

Identification of predictive biomarkers for ovarian cancer (OC) treatment, particularly in the platinum-resistant/refractory setting, is highly relevant for clinical management. E-cadherin, vimentin, and osteopontin (OPN) are proteins associated with tumor microenvironment (TME) remodelling that play key roles in cancer. This study aimed to evaluate the association between the staining patterns of these proteins with survival outcomes in a series of OC patients, namely in patients with platinum-resistant/refractory disease. Low E-cadherin expression and high vimentin expression in all patient groups (as well as for E-cadherin in the platinum-resistant arm) were significantly associated with longer overall survival (OS). Low cytoplasmic OPN expression (and cytoplasmic and membrane OPN in the platinum-resistant arm) were significantly associated with longer OS. In patients that responded to treatment (pegylated liposomal doxorubicin (PLD) or other), low cytoplasmic OPN expression was also associated with longer progression-free survival (PFS). In the other hand, high nuclear OPN-c expression in patients that respond to treatment was associated with longer OS and longer PFS. Longer PFS was also associated with high expression of both nuclear and cytoplasm OPN-c, in platinum-resistant patients and in those that responded to PLD. Our study indicates that the expression of E-cadherin, vimentin, and OPN may have prognostic implications. Nuclear OPN-c and cytoplasm OPN expression are putative predictive markers in platinum-resistant (PLD treated) ovarian cancer patients.

Amazon Fruits Inhibit Growth and Promote Pro-apoptotic Effects on Human Ovarian Carcinoma Cell Lines.

Murici (Byrsonima crassifolia (L.) Kunth and B. verbascifolia (L.) DC.) and tapereba (Spondias mombin) are Amazonian fruits that contain bioactive compounds. Biochemical and molecular characterization of these fruits can reveal their potential use in preventing diseases, including cancer. The extracts were characterized regarding the presence and profile of carotenoids by high-performance liquid chromatography (HPLC), total phenolic content by the Folin-Ciocalteu assay, and antioxidant activity by antioxidant value 2,2-diphenyl-1-picrylhydrazyl (DPPH) content analysis, 22,20-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) content analysis, Ferric-Reducing Ability of Plasma (FRAP), and Oxygen Radical Absorbance Capacity (ORAC) analysis. The extracts of tapereba and murici studied were important sources of total carotenoids and lutein, respectively. The extracts were then tested for their effect on the viability of the A2780 ovarian cancer (OC) cell line and its cisplatin (CDDP)-resistant derived cell line, called ACRP, by using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays. Their influence on cell cycle and apoptosis were analyzed by using flow cytometry. Murici and tapereba cell extracts exhibited a strong bioactivity by inhibiting A2780 and ACRP cell viability by 76.37% and 78.37%, respectively, besides modulating the cell cycle and inducing apoptotic cell death. Our results open new perspectives for the development of innovative therapeutic strategies using these Amazon fruit extracts to sensitize ovarian cancer cells to current chemotherapeutic options.

The long non-coding RNA PCA3: an update of its functions and clinical applications as a biomarker in prostate cancer.

Prostate cancer antigen 3 (PCA3) is an overexpressed prostate long non-coding RNA (lncRNA), transcribed from an intronic region at the long arm of human chromosome 9q21-22. It has been described that PCA3 modulates prostate cancer (PCa) cell survival through modulating androgen receptor (AR) signaling, besides controlling the expression of several androgen responsive and cancer-related genes, including epithelial-mesenchymal transition (EMT) markers and those regulating gene expression and cell signaling. Also, PCA3 urine levels have been successfully used as a PCa diagnostic biomarker. In this review, we have highlighted recent findings regarding PCA3, addressing its gene structure, putative applications as a biomarker, a proposed origin of this lncRNA, roles in PCa biology and expression patterns. We also updated data regarding PCA3 interactions with cancer-related miRNAs and expression in other tissues and diseases beyond the prostate. Altogether, literature data indicate aberrant expression and dysregulated activity of PCA3, suggesting PCA3 as a promising relevant target that should be even further evaluated on its applicability for PCa detection and management.

The LQB-223 Compound Modulates Antiapoptotic Proteins and Impairs Breast Cancer Cell Growth and Migration.

Drug resistance represents a major issue in treating breast cancer, despite the identification of novel therapeutic strategies, biomarkers, and subgroups. We have previously identified the LQB-223, 11a-N-Tosyl-5-deoxi-pterocarpan, as a promising compound in sensitizing doxorubicin-resistant breast cancer cells, with little toxicity to non-neoplastic cells. Here, we investigated the mechanisms underlying LQB-223 antitumor effects in 2D and 3D models of breast cancer. MCF-7 and MDA-MB-231 cells had migration and motility profile assessed by wound-healing and phagokinetic track motility assays, respectively. Cytotoxicity in 3D conformation was evaluated by measuring spheroid size and performing acid phosphatase and gelatin migration assays. Protein expression was analyzed by immunoblotting. Our results show that LQB-223, but not doxorubicin treatment, suppressed the migratory and motility capacity of breast cancer cells. In 3D conformation, LQB-223 remarkably decreased cell viability, as well as reduced 3D culture size and migration. Mechanistically, LQB-223-mediated anticancer effects involved decreased proteins levels of XIAP, c-IAP1, and Mcl-1 chemoresistance-related proteins, but not survivin. Survivin knockdown partially potentiated LQB-223-induced cytotoxicity. Additionally, cell treatment with LQB-223 resulted in changes in the mRNA levels of epithelial-mesenchymal transition markers, suggesting that it might modulate cell plasticity. Our data demonstrate that LQB-223 impairs 3D culture growth and migration in 2D and 3D models of breast cancer exhibiting different phenotypes.

Loss of the p53 transactivation domain results in high amyloid aggregation of the Δ40p53 isoform in endometrial carcinoma cells.

Dysfunctional p53 formation and activity can result from aberrant expression and subcellular localization of distinct p53 isoforms or aggregates. Endometrial carcinoma (EC) is a cancer type in which p53 status is correlated with prognosis, and TP53 mutations are a frequent genetic modification. Here we aimed to evaluate the expression patterns of different p53 isoforms and their contributions to the formation and subcellular localization of p53 amyloid aggregates in both EC and endometrial nontumor cell lines. We found that full-length (fl) p53 and a truncated p53 isoform, Δ40p53, resulting from alternative splicing of exon 2 or alternative initiation of translation at ATG-40, are the predominantly expressed p53 variants in EC cells. However, Δ40p53 was the major p53 isoform in endometrial nontumor cells. Immunofluorescence assays revealed that Δ40p53 is mainly localized to cytoplasmic punctate structures of EC cells, resembling solid-phase structures similar to those found in neurodegenerative pathologies. Using light-scattering kinetics, CD, and transmission EM, we noted that the p53 N-terminal transactivation domain significantly reduces aggregation of the WT p53 DNA-binding domain, confirming the higher aggregation tendency of Δ40p53, which lacks this domain. This is the first report of cytoplasmic Δ40p53 in EC cells being a major component of amyloid aggregates. The differential aggregation properties of p53 isoforms in EC cells may open up new avenues in the development of therapeutic strategies that preferentially target specific p53 isoforms to prevent p53 amyloid aggregate formation.