Recent advances of small extracellular vesicle biomarkers in breast cancer diagnosis and prognosis.

Current clinical tools for breast cancer (BC) diagnosis are insufficient but liquid biopsy of different bodily fluids has recently emerged as a minimally invasive strategy that provides a real-time snapshot of tumour biomarkers for early diagnosis, active surveillance of progression, and post-treatment recurrence. Extracellular vesicles (EVs) are nano-sized membranous structures 50-1000 nm in diameter that are released by cells into biological fluids. EVs contain proteins, nucleic acids, and lipids which play pivotal roles in tumourigenesis and metastasis through cell-to-cell communication. Proteins and miRNAs from small EVs (sEV), which range in size from 50-150 nm, are being investigated as a potential source for novel BC biomarkers using mass spectrometry-based proteomics and next-generation sequencing. This review covers recent developments in sEV isolation and single sEV analysis technologies and summarises the sEV protein and miRNA biomarkers identified for BC diagnosis, prognosis, and chemoresistance. The limitations of current sEV biomarker research are discussed along with future perspective applications.

CD44 variant 6 is associated with prostate cancer growth and chemo-/radiotherapy response in vivo.

We have previously demonstrated that CD44 variant 6 (CD44v6) is associated with prostate cancer (CaP) growth and therapeutic resistance in vitro, however, the role of CD44v6 in CaP in vivo is not fully understood. The purpose of this study is to investigate the effect of CD44v6 on CaP growth and chemo-/radiotherapy response in NOD/SCID mouse models in vivo and to validate its role as a therapeutic target for CaP therapy. CD44v6 was knocked down in PC-3M CaP cell line using short hairpin RNA. Subcutaneous (s.c.) and orthotopic CaP mouse xenografts were established. The effect of CD44v6 knockdown (KD) on tumour growth was evaluated in both s.c. and orthotopic models. Chemo-/radiotherapy response was evaluated in the s.c. model. Association of CD44v6 with PI3K/Akt pathway was validated using immunohistochemistry staining. We found that KD of CD44v6 significantly reduced tumour growth in both models, and enhanced the sensitivity of tumours to chemotherapy and radiotherapy in the s.c. model. In addition, we demonstrated that KD of CD44v6 is associated with downregulation of the PI3K/Akt/mTOR pathway. Our data confirm that CaP growth and chemo-/radiosensitivity in vivo is associated with CD44v6, which holds great promises as a therapeutic target in the treatment of CaP.

Inhibition of PI3K/Akt/mTOR signaling pathway alleviates ovarian cancer chemoresistance through reversing epithelial-mesenchymal transition and decreasing cancer stem cell marker expression.

BACKGROUND: Ovarian cancer is the most common malignant tumor of the female reproductive tract. Chemoresistance is a major challenge for current ovarian cancer therapy. However, the mechanism underlying epithelial ovarian cancer (EOC) chemoresistance is not completely uncovered. The phosphatidylinositol-3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling is an important intracellular pathway in regulating cell cycle, quiescence, and proliferation. The aim of this study is to investigate the role of PI3K/Akt/mTOR signaling pathway and its association with epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) marker expression in EOC chemoresistance. METHODS: The expressions of EMT and CSC markers were detected by immunofluorescence, western blot, and quantitative real-time PCR. BEZ235, a dual PI3K/mTOR inhibitor, was employed to investigate the role of PI3K/Akt/ mTOR signaling in regulating EMT and CSC marker expression. Students' t test and one-way ANOVA with Tukey's post-hoc test were used to compare the data from different groups. RESULTS: We found that EMT and CSC marker expression were significantly enhanced in chemoresistant EOC cells, which was accompanied by the activation of PI3K/Akt/mTOR signaling. Compared with single cisplatin treatment, combined treatment with BEZ235 and cisplatin significantly disrupted the colony formation ability, induced higher ROS level and more apoptosis in chemoresistant EOC cells. Furthermore, the combination approach effectively inhibited PI3K/Akt/mTOR signaling pathway, reversed EMT, and decreased CSC marker expression in chemoresistant EOC cells compared with cisplatin mono-treatment. CONCLUSIONS: Our results first demonstrate that EMT and enhanced CSC marker expression triggered by activated PI3K/Akt/mTOR signaling are involved in the chemoresistance of EOC, and BEZ235 in combination with cisplatin might be a promising treatment option to reverse EOC chemoresistance.

Epithelial cell adhesion molecule (EpCAM) is involved in prostate cancer chemotherapy/radiotherapy response in vivo.

BACKGROUND: Development of chemo-/radioresistance is a major challenge for the current prostate cancer (CaP) therapy. We have previously demonstrated that epithelial cell adhesion molecule (EpCAM) is associated with CaP growth and therapeutic resistance in vitro, however, the role of EpCAM in CaP in vivo is not fully elucidated. Here, we aimed to investigate how expression of EpCAM is involved in CaP growth and chemo-/radiotherapy response in NOD/SCID mouse models in vivo and to validate its role as a therapeutic target for CaP therapy. METHODS: EpCAM was knocked down in PC-3 CaP cell line using short hairpin RNA (shRNA). The effect of EpCAM-knockdown (KD) on tumour growth, chemo-/radiotherapy response and animal survival was evaluated on subcutaneous (s.c) and orthotopic mouse models. RESULTS: We found that KD of EpCAM significantly inhibited tumour growth, increased xenograft sensitivity to chemotherapy/radiotherapy, and prolonged the survival of tumour-bearing mice. In addition, we demonstrated that KD of EpCAM is associated with downregulation of the PI3K/Akt/mTOR pathway. CONCLUSIONS: In conclusion, our data confirms that CaP growth and chemo-/radioresistance in vivo is associated with over-expression of EpCAM, which serves both a functional biomarker and promising therapeutic target.

Proteomics discovery of chemoresistant biomarkers for ovarian cancer therapy.

INTRODUCTION: Chemoresistance is a major challenge to current ovarian cancer chemotherapy. It is important to identify biomarkers to distinguish chemosensitive and chemoresistant patients. Areas covered: We review the medical literature, discuss MS-based technologies with respect to chemoresistant ovarian cancer and summarize the promising chemoresistant biomarkers identified. In addition, the challenges and future perspectives of biomarker discovery research are explored. With the employment of mass spectrometry-based (MS-based) proteomics, biomarker discovery of ovarian cancer has made great progress in the last decade. Many potential biomarkers were identified by MS-based proteomics technologies, some of which have been validated for further extensive studies in clinical settings. Expert commentary: The discovery of chemoresistant biomarkers is a newly developing area and may provide a clue for predicting chemotherapeutic response and discover therapeutic targets for paving the way of personalized medicine. Multiple complementary MS-based proteomics approaches hold promise for finding novel therapeutic targets in ovarian cancer treatment.

CD44 variant 6 is associated with prostate cancer metastasis and chemo-/radioresistance.

INTRODUCTION: Prostate cancer (CaP) is the second leading malignancy in older men in Western countries. The role of CD44 variant 6 (CD44v6) in CaP progression and therapeutic resistance is still uncertain. Here, we investigated the roles of CD44v6 in CaP metastasis and chemo/radioresistance. Expression of CD44v6 in metastatic CaP cell lines, human primary CaP tissues and lymph node metastases was assessed using immunofluorescence and immunohistochemistry, respectively. METHODS: Knock down (KD) of CD44v6 was performed in PC-3M, DU145, and LNCaP cells using small interfering RNA (siRNA), and confirmed by confocal microscope, Western blot and quantitative real time polymerase chain reaction (qRT-PCR). Cell growth was evaluated by proliferation and colony formation assays. The adhesive ability and invasive potential were assessed using a hyaluronic acid (HA) adhesion and a matrigel chamber assay, respectively. Tumorigenesis potential and chemo-/radiosensitivity were measured by a sphere formation assay and a colony assay, respectively. RESULTS: Over-expression of CD44v6 was found in primary CaP tissues and lymph node metastases including cancer cells and surrounding stromal cells. KD of CD44v6 suppressed CaP proliferative, invasive and adhesive abilities, reduced sphere formation, enhanced chemo-/radiosensitivity, and down-regulated epithelial-mesenchymal transition (EMT), PI3K/Akt/mTOR, and Wnt/ß-catenin signaling pathway proteins in vitro. CONCLUSIONS: Our findings demonstrate that CD44v6 is an important cancer stem cell-like marker associated with CaP proliferation, invasion, adhesion, metastasis, chemo-/radioresistance, and the induction of EMT as well as the activation PI3K/Akt/mTOR and Wnt signaling pathways, suggesting that CD44v6 is a novel therapeutic target to sensitize CaP cells to chemo/radiotherapy.

Acromion and Distal Clavicle Grafts for Arthroscopic Glenoid Reconstruction.

BACKGROUND: We intended to determine if an acromion or distal clavicle bone graft could restore large glenoid defects using two novel, screw-free graft fixation techniques. METHODS: Twenty-four sawbone shoulder models were divided into four groups (n = 6 per group) according to fixation technique and bone graft: (1) modified buckle-down technique with clavicle graft, (2) modified buckle-down technique with acromion graft, (3) cross-link technique with acromion graft, (4) cross-link technique with clavicle graft. Testing was performed sequentially in (1) intact models, (2) after creation of a 30% by-width glenoid defect and (3) after repair. The shoulder joint was translated anteriorly, and glenohumeral contact pressures and load were measured to quantify the biomechanical stability. RESULTS: Maximum contact pressures were restored to 42-56% of intact glenoid using acromion and clavicle grafts with novel fixation techniques. Acromion grafts attained higher maximum contact pressures than clavicle grafts in all groups. Peak translational forces increased by 171-368% after all repairs. CONCLUSIONS: This controlled laboratory study on sawbone models found that both the acromion and distal clavicle are suitable autologous bone graft options for treating large anterior glenoid defects, having appropriate dimensions and contours for reconstructing the glenoid arc. The modified buckle-down and cross-link techniques are two graft fixation techniques that restore stability to the shoulder joint upon repairing a large glenoid defect and are advantageous in being screw-free and simple to execute.

Superior and Anterior Glenoid Labral Tears Are Associated With Increased Neurofilament Concentration.

BACKGROUND: Pain is a common presentation after glenohumeral labral injuries. However, the source of that pain is undetermined. PURPOSE/HYPOTHESIS: We aimed to determine if there is a differential expression of nerve fibers around the glenoid labrum and if torn labra have increased neuronal expression compared with untorn labra (rotator cuff repair labra). We hypothesized that the superior labrum would have a higher concentration of neurofilament than would the rest of the labrum and that the concentration of neurofilament would increase at the site of a labral tear. STUDY DESIGN: Descriptive laboratory study. METHODS: Seven labra were sampled at the 3-, 5-, 9-, and 12-o'clock positions during total shoulder arthroplasty. Samples were also collected at the 3-, 5-, and 12-o'clock positions during rotator cuff repair (16 labra), anterior labral repair (6 labra), type II superior labral anterior to posterior (SLAP) repair (4 labra), and capsular release for idiopathic capsulitis (5 labra). Sections were immunostained with antibodies to neurofilament, a specific neuronal marker that is used to identify central and peripheral nerve fibers, and the concentration and intensity of immunostained-positive cells assessed. RESULTS: The concentration of neurofilament staining was similar in the superior, anterior, posterior, and inferior glenoid labrum in untorn labra (8 neurofilament expressing cells per square millimeter; P = .3). Torn labra exhibited a 3- to 4-fold increase in neuronal expression, which was isolated to the location of the tear in SLAP (P = .09) and anterior labral tears (P = .02). The concentration of neurofilament expressing cells in torn glenoid labrum samples was comparable that in with the glenoid labrum of adhesive capsulitis samples (P = .7). CONCLUSION: This study supports the hypothesis that after a tear of the anterior or superior labrum the labrum in that region becomes populated with new nerves fibers and that these fibers may be responsible for the pain noted by patients with superior (SLAP) and/or anterior labral (Bankart) tears. CLINICAL RELEVANCE: This study suggests that neural infiltration contributes to the pain experienced by patients with labral tears. It may help with patient education and direct future management of labral lesions.

A review of bone grafting techniques for glenoid reconstruction.

Background: Traumatic anterior shoulder dislocations can cause bony defects of the anterior glenoid rim and are often associated with recurrent shoulder instability. For large glenoid defects of 20-30% without a mobile bony fragment, glenoid reconstruction with bone grafts is often recommended. This review describes two broad categories of glenoid reconstruction procedures found in literature: coracoid transfers involving the Bristow and Latarjet procedures, and free bone grafting techniques. Methods: An electronic search of MEDLINE and PubMed was conducted to find original articles that described glenoid reconstruction techniques or modifications to existing techniques. Results: Coracoid transfers involve the Bristow and Latarjet procedures. Modifications to these procedures such as arthroscopic execution, method of graft attachment and orientation have been described. Free bone grafts have been obtained from the iliac crest, distal tibia, acromion, distal clavicle and femoral condyle. Conclusion: Both coracoid transfers and free bone grafting procedures are options for reconstructing large bony defects of the anterior glenoid rim and have had similar clinical outcomes. Free bone grafts may offer greater flexibility in graft shaping and choice of graft size depending on the bone stock chosen. Novel developments tend towards minimising invasiveness using arthroscopic approaches and examining alternative non-rigid graft fixation techniques.

Survival prediction in triple negative breast cancer using multiple instance learning of histopathological images.

Computational pathology is a rapidly expanding area for research due to the current global transformation of histopathology through the adoption of digital workflows. Survival prediction of breast cancer patients is an important task that currently depends on histopathology assessment of cancer morphological features, immunohistochemical biomarker expression and patient clinical findings. To facilitate the manual process of survival risk prediction, we developed a computational pathology framework for survival prediction using digitally scanned haematoxylin and eosin-stained tissue microarray images of clinically aggressive triple negative breast cancer. Our results show that the model can produce an average concordance index of 0.616. Our model predictions are analysed for independent prognostic significance in univariate analysis (hazard ratio = 3.12, 95% confidence interval [1.69,5.75], p < 0.005) and multivariate analysis using clinicopathological data (hazard ratio = 2.68, 95% confidence interval [1.44,4.99], p < 0.005). Through qualitative analysis of heatmaps generated from our model, an expert pathologist is able to associate tissue features highlighted in the attention heatmaps of high-risk predictions with morphological features associated with more aggressive behaviour such as low levels of tumour infiltrating lymphocytes, stroma rich tissues and high-grade invasive carcinoma, providing explainability of our method for triple negative breast cancer.

Fecal DNA Virome Is Associated with the Development of Colorectal Neoplasia in a Murine Model of Colorectal Cancer.

Alteration of the gut virome has been associated with colorectal cancer (CRC); however, when and how the alteration takes place has not been studied. Here, we employ a longitudinal study in mice to characterize the gut virome alteration in azoxymethane (AOM)-induced colorectal neoplasia and identify important viruses associated with tumor growth. The number and size of the tumors increased as the mice aged in the AOM treated group, as compared to the control group. Tumors were first observed in the AOM group at week 12. We observed a significantly lower alpha diversity and shift in viral profile when tumors first appeared. In addition, we identified novel viruses from the genera Brunovirus, Hpunavirus that are positively associated with tumor growth and enriched at a late time point in AOM group, whereas members from Lubbockvirus show a negative correlation with tumor growth. Moreover, network analysis revealed two clusters of viruses in the AOM virome, a group that is positively correlated with tumor growth and another that is negatively correlated with tumor growth, all of which are bacteriophages. Our findings suggest that the gut virome changes along with tumor formation and provides strong evidence of a potential role for bacteriophage in the development of colorectal neoplasia.

ALTEN: A High-Fidelity Primary Tissue-Engineering Platform to Assess Cellular Responses Ex Vivo.

To fully investigate cellular responses to stimuli and perturbations within tissues, it is essential to replicate the complex molecular interactions within the local microenvironment of cellular niches. Here, the authors introduce Alginate-based tissue engineering (ALTEN), a biomimetic tissue platform that allows ex vivo analysis of explanted tissue biopsies. This method preserves the original characteristics of the source tissue's cellular milieu, allowing multiple and diverse cell types to be maintained over an extended period of time. As a result, ALTEN enables rapid and faithful characterization of perturbations across specific cell types within a tissue. Importantly, using single-cell genomics, this approach provides integrated cellular responses at the resolution of individual cells. ALTEN is a powerful tool for the analysis of cellular responses upon exposure to cytotoxic agents and immunomodulators. Additionally, ALTEN's scalability using automated microfluidic devices for tissue encapsulation and subsequent transport, to enable centralized high-throughput analysis of samples gathered by large-scale multicenter studies, is shown.

Triple-negative breast cancer therapeutic resistance: Where is the Achilles' heel?

Triple-negative breast cancer (TNBC) shows a higher response rate to systemic therapy compared with other breast cancer subtypes. However, the tumor differentiation of TNBC is poorer, with an early tendency to metastasis and a higher recurrence rate. Relapsed and metastatic TNBCs usually progress more rapidly, showing strong resistance to chemotherapy and radiotherapy. Due to the lack of combinatorial targeted drugs, alternative treatments fail to improve these patient's prognosis and the quality of life. Finding the Achilles' heel of TNBC is critical for patients with advanced TNBC. Here, we summarize the latest advances in the mechanisms underlying TNBC therapeutic resistance, consider how these mechanisms may affect the development and utilization of TNBC targeted drugs, and discuss the rationale of relevant signals as therapeutic targets. Also, we review the clinical trials registered in ClinicalTrial.gov for TNBC patients, which comprehensively reveals current research and development of novel TNBC targeted drugs and future trends.

Immunotherapy for triple-negative breast cancer: A molecular insight into the microenvironment, treatment, and resistance.

Clinicians have very limited options to treat triple-negative breast cancer (TNBC) due to the lack of effective targeted drugs. Recently, the findings of the mechanism underlying tumor-intrinsic immune escape have fueled a wave of studies into immunotherapy in breast cancer (BC). Compared with other BC subtypes, TNBC shows a better response to immunotherapy due to the higher level of tumor mutation burden and lymphocyte infiltration. Thereinto, immune checkpoint inhibitors (ICIs) achieved the first success of immunotherapy for TNBC and are widely utilized with conventional treatments in the neoadjuvant/adjuvant and advanced stages. However, a large number of TNBC patients fail to demonstrate a good response to ICIs, and the acquired resistance to ICI-based therapies is clinically emerging, which is a major challenge for immunotherapy in TNBC. Here we review the latest advances in TNBC immune microenvironment, immunotherapy, and immunotherapeutic resistance and discuss the challenges and potential approaches to improve the clinical benefit of immunotherapy against TNBC.

THOC2 and THOC5 Regulate Stemness and Radioresistance in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer. Radioresistance and stemness are substantial obstacles to TNBC treatment. The THO complex (THOC) is a subunit of the TRanscription-EXport complex that functions in the coupling of transcription to nascent RNA splicing, elongation, and export. However, its role in regulating TNBC therapeutic resistance is not reported yet. In this study, the authors demonstrate that cancer stem cells are enriched in radioresistant TNBC cells and describe the role of the THOC in regulating TNBC radioresistance and stemness. The authors find that THOC2 and THOC5 are upregulated in radioresistant TNBC cells and associated with a poor prognosis in TNBC patients. Further investigation reveals that THOC2 promotes the stem-like properties and radioresistance of TNBC cells in a THOC5-dependent manner by facilitating the release of sex-determining region Y (SRY)-box transcription factor 2 (SOX2) and homeobox transcription factor (NANOG) transcripts from the nucleus. Silencing THOC2 or THOC5 expression decreases the protein expression of SOX2 and NANOG, depletes the stem-like properties, and causes radiosensitization in these TNBC cells. Moreover, THOC2 or THOC5 depletion blocks the xenograft tumorigenesis and growth of radioresistant TNBC in vivo. These findings uncover the novel correlations of THOC with TNBC stemness and therapeutic resistance, proposing alternative therapeutic strategies against relapsed TNBC.

Activation of the eIF2α/ATF4 axis drives triple-negative breast cancer radioresistance by promoting glutathione biosynthesis.

Triple-negative breast cancer (TNBC) is the most aggressive breast cancer subtype. Radiotherapy is an effective option for the treatment of TNBC; however, acquired radioresistance is a major challenge to the modality. In this study, we show that the integrated stress response (ISR) is the most activated signaling pathway in radioresistant TNBC cells. The constitutive phosphorylation of eIF2α in radioresistant TNBC cells promotes the activation of ATF4 and elicits the transcription of genes implicated in glutathione biosynthesis, including GCLC, SLC7A11, and CTH, which increases the intracellular level of reduced glutathione (GSH) and the scavenging of reactive oxygen species (ROS) after irradiation (IR), leading to a radioresistant phenotype. The cascade is significantly up-regulated in human TNBC tissues and is associated with unfavorable survival in patients. Dephosphorylation of eIF2α increases IR-induced ROS accumulation in radioresistant TNBC cells by disrupting ATF4-mediated GSH biosynthesis and sensitizes them to IR in vitro and in vivo. These findings reveal ISR as a vital mechanism underlying TNBC radioresistance and propose the eIF2α/ATF4 axis as a novel therapeutic target for TNBC treatment.

Multiplexed immunofluorescence identifies high stromal CD68+PD-L1+ macrophages as a predictor of improved survival in triple negative breast cancer.

Triple negative breast cancer (TNBC) comprises 10-15% of all breast cancers and has a poor prognosis with a high risk of recurrence within 5 years. PD-L1 is an important biomarker for patient selection for immunotherapy but its cellular expression and co-localization within the tumour immune microenvironment and associated prognostic value is not well defined. We aimed to characterise the phenotypes of immune cells expressing PD-L1 and determine their association with overall survival (OS) and breast cancer-specific survival (BCSS). Using tissue microarrays from a retrospective cohort of TNBC patients from St George Hospital, Sydney (n = 244), multiplexed immunofluorescence (mIF) was used to assess staining for CD3, CD8, CD20, CD68, PD-1, PD-L1, FOXP3 and pan-cytokeratin on the Vectra Polaris™ platform and analysed using QuPath. Cox multivariate analyses showed high CD68+PD-L1+ stromal cell counts were associated with improved prognosis for OS (HR 0.56, 95% CI 0.33-0.95, p = 0.030) and BCSS (HR 0.47, 95% CI 0.25-0.88, p = 0.018) in the whole cohort and in patients receiving chemotherapy, improving incrementally upon the predictive value of PD-L1+ alone for BCSS. These data suggest that CD68+PD-L1+ status can provide clinically useful prognostic information to identify sub-groups of patients with good or poor prognosis and guide treatment decisions in TNBC.

Cryopreservation of human cancers conserves tumour heterogeneity for single-cell multi-omics analysis.

BACKGROUND: High throughput single-cell RNA sequencing (scRNA-Seq) has emerged as a powerful tool for exploring cellular heterogeneity among complex human cancers. scRNA-Seq studies using fresh human surgical tissue are logistically difficult, preclude histopathological triage of samples, and limit the ability to perform batch processing. This hindrance can often introduce technical biases when integrating patient datasets and increase experimental costs. Although tissue preservation methods have been previously explored to address such issues, it is yet to be examined on complex human tissues, such as solid cancers and on high throughput scRNA-Seq platforms. METHODS: Using the Chromium 10X platform, we sequenced a total of ~ 120,000 cells from fresh and cryopreserved replicates across three primary breast cancers, two primary prostate cancers and a cutaneous melanoma. We performed detailed analyses between cells from each condition to assess the effects of cryopreservation on cellular heterogeneity, cell quality, clustering and the identification of gene ontologies. In addition, we performed single-cell immunophenotyping using CITE-Seq on a single breast cancer sample cryopreserved as solid tissue fragments. RESULTS: Tumour heterogeneity identified from fresh tissues was largely conserved in cryopreserved replicates. We show that sequencing of single cells prepared from cryopreserved tissue fragments or from cryopreserved cell suspensions is comparable to sequenced cells prepared from fresh tissue, with cryopreserved cell suspensions displaying higher correlations with fresh tissue in gene expression. We showed that cryopreservation had minimal impacts on the results of downstream analyses such as biological pathway enrichment. For some tumours, cryopreservation modestly increased cell stress signatures compared to freshly analysed tissue. Further, we demonstrate the advantage of cryopreserving whole-cells for detecting cell-surface proteins using CITE-Seq, which is impossible using other preservation methods such as single nuclei-sequencing. CONCLUSIONS: We show that the viable cryopreservation of human cancers provides high-quality single-cells for multi-omics analysis. Our study guides new experimental designs for tissue biobanking for future clinical single-cell RNA sequencing studies.

Inhibition of micrometastatic prostate cancer cell spread in animal models by 213Bilabeled multiple targeted alpha radioimmunoconjugates.

PURPOSE: To investigate the therapeutic potential of 213Bilabeled multiple targeted alpha-radioimmunoconjugates for treating prostate cancer (CaP) micrometastases in mouse models. EXPERIMENTAL DESIGN: PC-3 CaP cells were implanted s.c., in the prostate, and intratibially in NODSCID mice. The expression of multiple tumor-associated antigens on tumor xenografts and micrometastases was detected by immunohistochemistry. Targeting vectors were two monoclonal antibodies, and a plasminogen activator inhibitor type 2 that binds to cell surface urokinase plasminogen activator, labeled with 213Bi using standard methodology. In vivo efficacy of multiple alpha conjugates (MTAT) at different activities was evaluated in these mouse models. Tumor growth was monitored during observations and local regional lymph node metastases were assessed at the end of experiments. RESULTS: The take rate of PC-3 cells was 100% for each route of injection. The tumor-associated antigens (MUC1, urokinase plasminogen activator, and BLCA-38) were heterogeneously expressed on primary tumors and metastatic cancer clusters at transit. A single i.p. injection of MTAT (test) at high and low doses caused regression of the growth of primary tumors and prevented local lymph node metastases in a concentration-dependent fashion; it also caused cancer cells to undergo necrosis and apoptosis. CONCLUSIONS: Our results suggest that MTAT can impede primary PC-3 CaP growth at three different sites in vivo through induction of apoptosis, and can prevent the spread of cancer cells and target lymph node micrometastases in a concentration-dependent manner. MTAT, by targeting multiple antigens, can overcome heterogeneous antigen expression to kill small CaP cell clusters, thus providing a potent therapy for micrometastases.

Exosomal microRNAs as liquid biopsy biomarkers in prostate cancer.

Prostate cancer (PCa) is the most commonly diagnosed solid-organ cancer in males. The PSA testing may cause overdiagnosis and overtreatment for PCa patients. There is an urgent need for new biomarkers with greater discriminative precision for diagnosis and risk-stratification, to select for prostate biopsy and treatment of PCa. Liquid biopsy is a promising field with the potential to provide comprehensive information on the genetic landscape at diagnosis and to track genomic evolution over time in order to tailor the therapeutic choices at all stages of PCa. Exosomes, containing RNAs, DNAs and proteins, have been shown to be involved in tumour progression and a rich potential source of tumour biomarkers, especially for profiling analysis of their miRNAs content. In this review, we summarise the exosomal miRNAs in PCa diagnosis, prognosis and management, and further discuss their possible technical challenges associated with isolating PCa-specific exosomes.