ResearchPad - kaposi-sarcoma https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[DLX1008 (brolucizumab), a single-chain anti-VEGF-A antibody fragment with low picomolar affinity, leads to tumor involution in an <i>in vivo</i> model of Kaposi Sarcoma]]> https://www.researchpad.co/article/elastic_article_14618 Kaposi Sarcoma (KS) is among the most angiogenic cancers in humans and an AIDS-defining condition. KS-associated herpesvirus (KSHV) is necessary for KS development, as is vascular endothelial growth factor (VEGF-A). DLX1008 is a novel anti-VEGF-A antibody single-chain variable fragment (scFv) with low picomolar affinity for VEGF-A. In vivo imaging techniques were used to establish the efficacy of DLX1008 and to establish the mechanism of action; this included non-invasive imaging by ultrasound and optical fluorescence, verified by post-mortem histochemistry. The results showed that DLX1008 was efficacious in a KS mouse model. The NSG mouse xenografts suffered massive internal necrosis or involution, consistent with a lack of blood supply. We found that imaging by ultrasound was superior to external caliper measurements in the validation of the angiogenesis inhibitor DLX1008. Further development of DLX1008 against VEGF-dependent sarcomas is warranted.

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<![CDATA[Conservation of Complex Nuclear Localization Signals Utilizing Classical and Non-Classical Nuclear Import Pathways in LANA Homologs of KSHV and RFHV]]> https://www.researchpad.co/article/5989daabab0ee8fa60ba93ff

ORF73 latency-associated nuclear antigen (LANA) of the Kaposi's sarcoma-associated herpesvirus (KSHV) is targeted to the nucleus of infected cells where it binds to chromatin and mediates viral episome persistence, interacts with cellular proteins and plays a role in latency and tumorigenesis. A structurally related LANA homolog has been identified in the retroperitoneal fibromatosis herpesvirus (RFHV), the macaque homolog of KSHV. Here, we report the evolutionary and functional conservation of a novel bi-functional nuclear localization signal (NLS) in KSHV and RFHV LANA. N-terminal peptides from both proteins were fused to EGFP or double EGFP fusions to examine their ability to induce nuclear transport of a heterologous protein. In addition, GST-pull down experiments were used to analyze the ability of LANA peptides to interact with members of the karyopherin family of nuclear transport receptors. Our studies revealed that both LANA proteins contain an N-terminal arginine/glycine (RG)-rich domain spanning a conserved chromatin-binding motif, which binds directly to importin β1 in a RanGTP-sensitive manner and serves as an NLS in the importin β1-mediated non-classical nuclear import pathway. Embedded within this domain is a conserved lysine/arginine-(KR)-rich bipartite motif that binds directly to multiple members of the importin α family of nuclear import adaptors in a RanGTP-insensitive manner and serves as an NLS in the classical importin α/β-mediated nuclear import pathway. The positioning of a classical bipartite kr-NLS embedded within a non-classical rg-NLS is a unique arrangement in these viral proteins, whose nuclear localization is critical to their functionality and to the virus life cycle. The ability to interact with multiple import receptors provides alternate pathways for nuclear localization of LANA. Since different import receptors can import cargo to distinct subnuclear compartments, a multifunctional NLS may provide LANA with an increased ability to interact with different nuclear components in its multifunctional role to maintain viral latency.

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<![CDATA[Clinical Factors Associated with Long-Term Complete Remission versus Poor Response to Chemotherapy in HIV-Infected Children and Adolescents with Kaposi Sarcoma Receiving Bleomycin and Vincristine: A Retrospective Observational Study]]> https://www.researchpad.co/article/5989dad9ab0ee8fa60bb9426

Kaposi sarcoma (KS) is the most common HIV-associated malignancy in children and adolescents in Africa. Pediatric KS is distinct from adult disease. We evaluated the clinical characteristics associated with long-term outcomes. We performed a retrospective observational analysis of 70 HIV-infected children and adolescents with KS less than 18 years of age diagnosed between 8/2010 and 6/2013 in Lilongwe, Malawi. Local first-line treatment included bleomycin and vincristine plus nevirapine-based highly active anti-retroviral therapy (HAART). Median age was 8.6 years (range 1.7–17.9); there were 35 females (50%). Most common sites of presentation were: lymph node (74%), skin (59%), subcutaneous nodules (33%), oral (27%), woody edema (24%), and visceral (16%). Eighteen (26%) presented with lymphadenopathy only. Severe CD4 suppression occurred in 28%. At time of KS diagnosis, 49% were already on HAART. Overall, 28% presented with a platelet count < 100 x 109/L and 37% with hemoglobin < 8 g/dL. The 2-year event-free (EFS) and overall survival (OS) were 46% and 58% respectively (median follow-up 29 months, range 15–50). Multivariable analysis of risk of death and failure to achieve EFS demonstrated that visceral disease (odds ratios [OR] 19.08 and 11.61, 95% CI 2.22–163.90 and 1.60–83.95 respectively) and presenting with more than 20 skin/oral lesions (OR 9.57 and 22.90, 95% CI 1.01–90.99 and 1.00–524.13 respectively) were independent risk factors for both. Woody edema was associated with failure to achieve EFS (OR 7.80, 95% CI 1.84–33.08) but not death. Univariable analysis revealed that lymph node involvement was favorable for EFS (OR 0.28, 95% CI 0.08–0.99), while T1 TIS staging criteria, presence of cytopenias, and severe immune suppression were not associated with increased mortality. Long-term complete remission is achievable in pediatric KS, however outcomes vary according to clinical presentation. Based on clinical heterogeneity, treatment according to risk-stratification is necessary to improve overall outcomes.

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<![CDATA[Opposing Regulation of PROX1 by Interleukin-3 Receptor and NOTCH Directs Differential Host Cell Fate Reprogramming by Kaposi Sarcoma Herpes Virus]]> https://www.researchpad.co/article/5989db10ab0ee8fa60bcc007

Lymphatic endothelial cells (LECs) are differentiated from blood vascular endothelial cells (BECs) during embryogenesis and this physiological cell fate specification is controlled by PROX1, the master regulator for lymphatic development. When Kaposi sarcoma herpes virus (KSHV) infects host cells, it activates the otherwise silenced embryonic endothelial differentiation program and reprograms their cell fates. Interestingly, previous studies demonstrated that KSHV drives BECs to acquire a partial lymphatic phenotype by upregulating PROX1 (forward reprogramming), but stimulates LECs to regain some BEC-signature genes by downregulating PROX1 (reverse reprogramming). Despite the significance of this KSHV-induced bidirectional cell fate reprogramming in KS pathogenesis, its underlying molecular mechanism remains undefined. Here, we report that IL3 receptor alpha (IL3Rα) and NOTCH play integral roles in the host cell type-specific regulation of PROX1 by KSHV. In BECs, KSHV upregulates IL3Rα and phosphorylates STAT5, which binds and activates the PROX1 promoter. In LECs, however, PROX1 was rather downregulated by KSHV-induced NOTCH signal via HEY1, which binds and represses the PROX1 promoter. Moreover, PROX1 was found to be required to maintain HEY1 expression in LECs, establishing a reciprocal regulation between PROX1 and HEY1. Upon co-activation of IL3Rα and NOTCH, PROX1 was upregulated in BECs, but downregulated in LECs. Together, our study provides the molecular mechanism underlying the cell type-specific endothelial fate reprogramming by KSHV.

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<![CDATA[Amplification of the Angiogenic Signal through the Activation of the TSC/mTOR/HIF Axis by the KSHV vGPCR in Kaposi's Sarcoma]]> https://www.researchpad.co/article/5989dad6ab0ee8fa60bb8207

Background

Kaposi's sarcoma (KS) is a vascular neoplasm characterized by the dysregulated expression of angiogenic and inflammatory cytokines. The driving force of the KS lesion, the KSHV-infected spindle cell, secretes elevated levels of vascular endothelial growth factor (VEGF), essential for KS development. However, the origin of VEGF in this tumor remains unclear.

Methodology/Principal Findings

Here we report that the KSHV G protein-coupled receptor (vGPCR) upregulates VEGF in KS through an intricate paracrine mechanism. The cytokines secreted by the few vGPCR-expressing tumor cells activate in neighboring cells multiple pathways (including AKT, ERK, p38 and IKKβ) that, in turn, converge on TSC1/2, promoting mTOR activation, HIF upregulation, and VEGF secretion. Conditioned media from vGPCR-expressing cells lead to an mTOR-dependent increase in HIF-1α and HIF-2α protein levels and VEGF upregulation. In a mouse allograft model for KS, specific inhibition of the paracrine activation of mTOR in non-vGPCR-expressing cells was sufficient to inhibit HIF upregulation in these cells, and abolished the ability of the vGPCR-expressing cells to promote tumor formation in vivo. Similarly, pharmacologic inhibition of HIF in this model blocked VEGF secretion and also lead to tumor regression.

Conclusions/Significance

Our findings provide a compelling explanation for how the few tumor cells expressing vGPCR can contribute to the dramatic amplification of VEGF secretion in KS, and further provide a molecular mechanism for how cytokine dysregulation in KS fuels angiogenesis and tumor development. These data further suggest that activation of HIF by vGPCR may be a vulnerable target for the treatment of patients with KS.

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<![CDATA[Non-AIDS Associated Kaposi's Sarcoma: Clinical Features and Treatment Outcome]]> https://www.researchpad.co/article/5989da1aab0ee8fa60b7c978

Background

Kaposi's sarcoma (KS) in HIV negative patients is rare and has to be distinguished from AIDS associated KS. Two groups are at risk to develop non-AIDS related KS: elderly men mainly of Mediterranean origin and persons with iatrogenic immunosuppression.

Patients and Methods

In order to define risk-groups and major clinical features we retrospectively evaluated clinical data of all patients with non-AIDS associated KS presenting to the Department of Dermatology, University Hospital Tuebingen between 1987 and 2009. Data were extracted from the tumor registry of the Comprehensive Cancer Center Tuebingen and from patient records.

Results

20 patients with non-AIDS KS have been identified. The average age at KS onset was 66.6 years; the male-to-female-ratio was 3∶1. Most of the patients were immigrants from Mediterranean or Eastern European countries (60%). 15 cases of classic KS versus 5 cases of iatrogenic KS were observed. In 95% of the cases, KS was limited to the skin, without mucosal, lymph node or visceral manifestation. KS lesions were in all cases multiple and mostly bilateral, the most common localization was the skin of the lower extremities. Tumor control was achieved in nearly all cases by the use of local or systemic therapy. No patient died from KS.

Conclusions

Unlike KS in AIDS patients, non-AIDS associated KS is a rather localized process which rarely involves lymph nodes or organs. It is mostly seen in elderly males from Mediterranean or Eastern European countries and in most cases responsive on local or systemic therapeutic strategies.

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<![CDATA[Assessing and Responding to Palliative Care Needs in Rural Sub-Saharan Africa: Results from a Model Intervention and Situation Analysis in Malawi]]> https://www.researchpad.co/article/5989da8bab0ee8fa60b9e243

Introduction

Palliative care is rarely accessible in rural sub-Saharan Africa. Partners In Health and the Malawi government established the Neno Palliative Care Program (NPCP) to provide palliative care in rural Neno district. We conducted a situation analysis to evaluate early NPCP outcomes and better understand palliative care needs, knowledge, and preferences.

Methods

Employing rapid evaluation methodology, we collected data from 3 sources: 1) chart review of all adult patients from the NPCP’s first 9 months; 2) structured interviews with patients and caregivers; 3) semi-structured interviews with key stakeholders.

Results

The NPCP enrolled 63 patients in its first 9 months. Frequent diagnoses were cancer (n = 50, 79%) and HIV/AIDS (n = 37 of 61, 61%). Nearly all (n = 31, 84%) patients with HIV/AIDS were on antiretroviral therapy. Providers registered 112 patient encounters, including 22 (20%) home visits. Most (n = 43, 68%) patients had documented pain at baseline, of whom 23 (53%) were treated with morphine. A majority (n = 35, 56%) had ≥1 follow-up encounter. Mean African Palliative Outcome Scale pain score decreased non-significantly between baseline and follow-up (3.0 vs. 2.7, p = 0.5) for patients with baseline pain and complete pain assessment documentation. Providers referred 48 (76%) patients for psychosocial services, including community health worker support, socioeconomic assistance, or both. We interviewed 36 patients referred to the NPCP after the chart review period. Most had cancer (n = 19, 53%) or HIV/AIDS (n = 10, 28%). Patients frequently reported needing income (n = 24, 67%) or food (n = 22, 61%). Stakeholders cited a need to make integrated palliative care widely available.

Conclusions

We identified a high prevalence of pain and psychosocial needs among patients with serious chronic illnesses in rural Malawi. Early NPCP results suggest that comprehensive palliative care can be provided in rural Africa by integrating disease-modifying treatment and palliative care, linking hospital, clinic, and home-based services, and providing psychosocial support that includes socioeconomic assistance.

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<![CDATA[An APE1 inhibitor reveals critical roles of the redox function of APE1 in KSHV replication and pathogenic phenotypes]]> https://www.researchpad.co/article/5989db5aab0ee8fa60bdf349

APE1 is a multifunctional protein with a DNA base excision repair function in its C-terminal domain and a redox activity in its N-terminal domain. The redox function of APE1 converts certain transcription factors from inactive oxidized to active reduced forms. Given that among the APE1-regulated transcription factors many are critical for KSHV replication and pathogenesis, we investigated whether inhibition of APE1 redox function blocks KSHV replication and Kaposi’s sarcoma (KS) phenotypes. With an shRNA-mediated silencing approach and a known APE-1 redox inhibitor, we demonstrated that APE1 redox function is indeed required for KSHV replication as well as KSHV-induced angiogenesis, validating APE1 as a therapeutic target for KSHV-associated diseases. A ligand-based virtual screening yielded a small molecular compound, C10, which is proven to bind to APE1. C10 exhibits low cytotoxicity but efficiently inhibits KSHV lytic replication (EC50 of 0.16 μM and selective index of 165) and KSHV-mediated pathogenic phenotypes including cytokine production, angiogenesis and cell invasion, demonstrating its potential to become an effective drug for treatment of KS.

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<![CDATA[A Computational Profiling of Changes in Gene Expression and Transcription Factors Induced by vFLIP K13 in Primary Effusion Lymphoma]]> https://www.researchpad.co/article/5989da22ab0ee8fa60b7f48f

Infection with Kaposi's sarcoma associated herpesvirus (KSHV) has been linked to the development of primary effusion lymphoma (PEL), a rare lymphoproliferative disorder that is characterized by loss of expression of most B cell markers and effusions in the body cavities. This unique clinical presentation of PEL has been attributed to their distinctive plasmablastic gene expression profile that shows overexpression of genes involved in inflammation, adhesion and invasion. KSHV-encoded latent protein vFLIP K13 has been previously shown to promote the survival and proliferation of PEL cells. In this study, we employed gene array analysis to characterize the effect of K13 on global gene expression in PEL-derived BCBL1 cells, which express negligible K13 endogenously. We demonstrate that K13 upregulates the expression of a number of NF-κB responsive genes involved in cytokine signaling, cell death, adhesion, inflammation and immune response, including two NF-κB subunits involved in the alternate NF-κB pathway, RELB and NFKB2. In contrast, CD19, a B cell marker, was one of the genes downregulated by K13. A comparison with K13-induced genes in human vascular endothelial cells revealed that although there was a considerable overlap among the genes induced by K13 in the two cell types, chemokines genes were preferentially induced in HUVEC with few exceptions, such as RANTES/CCL5, which was induced in both cell types. Functional studies confirmed that K13 activated the RANTES/CCL5 promoter through the NF-κB pathway. Taken collectively, our results suggest that K13 may contribute to the unique gene expression profile, immunophenotype and clinical presentation that are characteristics of KSHV-associated PEL.

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<![CDATA[Epigenetic Landscape of Kaposi's Sarcoma-Associated Herpesvirus Genome in Classic Kaposi's Sarcoma Tissues]]> https://www.researchpad.co/article/5989db53ab0ee8fa60bdcdee

Kaposi's sarcoma-associated herpesvirus (KSHV) is etiologically related to Kaposi's sarcoma (KS), primary effusion lymphoma (PEL) and multicentric Castleman's disease (MCD). It typically displays two different phases in its life cycle, the default latency and occasional lytic replication. The epigenetic modifications are thought to determine the fate of KSHV infection. Previous studies elegantly depicted epigenetic landscape of latent viral genome in in vitro cell culture systems. However, the physiologically relevant scenario in clinical KS tissue samples is unclear. In the present study, we established a protocol of ChIP-Seq for clinical KS tissue samples and mapped out the epigenetic landscape of KSHV genome in classic KS tissues. We examined AcH3 and H3K27me3 histone modifications on KSHV genome, as well as the genome-wide binding sites of latency associated nuclear antigen (LANA). Our results demonstrated that the enriched AcH3 was mainly restricted at latent locus while H3K27me3 was widespread on KSHV genome in classic KS tissues. The epigenetic landscape at the region of vIRF3 gene confirmed its silenced state in KS tissues. Meanwhile, the abundant enrichment of LANA at the terminal repeat (TR) region was also validated in the classic KS tissues, however, different LANA binding sites were observed on the host genome. Furthermore, we verified the histone modifications by ChIP-qPCR and found the dominant repressive H3K27me3 at the promoter region of replication and transcription activator (RTA) in classic KS tissues. Intriguingly, we found that the TR region in classic KS tissues was lacking in AcH3 histone modifications. These data now established the epigenetic landscape of KSHV genome in classic KS tissues, which provides new insights for understanding KSHV epigenetics and pathogenesis.

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<![CDATA[KS-Detect – Validation of Solar Thermal PCR for the Diagnosis of Kaposi’s Sarcoma Using Pseudo-Biopsy Samples]]> https://www.researchpad.co/article/5989dad5ab0ee8fa60bb7d25

Resource-limited settings present unique engineering challenges for medical diagnostics. Diagnosis is often needed for those unable to reach central healthcare systems, making portability and independence from traditional energy infrastructure essential device parameters. In 2014, our group presented a microfluidic device that performed a solar-powered variant of the polymerase chain reaction, which we called solar thermal PCR. In this work, we expand on our previous effort by presenting an integrated, portable, solar thermal PCR system targeted towards the diagnosis of Kaposi’s sarcoma. We call this system KS-Detect, and we now report the system’s performance as a diagnostic tool using pseudo-biopsy samples made from varying concentrations of human lymphoma cell lines positive for the KS herpesvirus (KSHV). KS-Detect achieved 83% sensitivity and 70% specificity at high (≥10%) KSHV+ cell concentrations when diagnosing pseudo-biopsy samples by smartphone image. Using histology, we confirm that our prepared pseudo-biopsies contain similar KSHV+ cell concentrations as human biopsies positive for KS. Through our testing of samples derived from human cell lines, we validate KS-Detect as a viable, portable KS diagnostic tool, and we identify critical engineering considerations for future solar-thermal PCR devices.

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<![CDATA[Rapid Multiplexed Immunoassay for Detection of Antibodies to Kaposi’s Sarcoma-Associated Herpesvirus]]> https://www.researchpad.co/article/5989dadaab0ee8fa60bb9511

Diagnosis of KSHV-infected individuals remains a challenge. KSHV prevalence is high in several populations with high prevalence of HIV, leading to increased risk of development of Kaposi’s sarcoma (KS). While current assays are reliable for detecting antibodies to KSHV, none are routinely utilized to identify individuals with KSHV infection and thus at increased risk for KS due to assay complexity, lack of access to testing, and cost, particularly in resource-limited settings. Here we describe the addition of KSHV proteins LANA and K8.1 to a previously evaluated HIV/co-infection multiplexed fluorescence immunoassay system. This study demonstrates assay performance by measuring antibody reactivity for KSHV and HIV-1 in a collection of clinical specimens from patients with biopsy-proven KS and sourced negative controls. The KSHV assay correctly identified 155 of 164 plasma samples from patients with biopsy-proven KS and 85 of 93 KSHV antibody (Ab)-negative samples for a sensitivity of 95.1% and specificity of 91.4%. Assay performance for HIV-1 detection was also assessed with 100% agreement with independently verified HIV-1 Ab-positive and Ab-negative samples. These results demonstrate good sensitivity and specificity for detection of antibody to KSHV antigens, and demonstrate the potential for multiplexed co-infection testing in resource-limited settings to identify those at increased risk for HIV-1-related complications.

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<![CDATA[Predictive Clinical Factors in the Diagnosis of Gastrointestinal Kaposi's Sarcoma and Its Endoscopic Severity]]> https://www.researchpad.co/article/5989d9fcab0ee8fa60b7237d

Background

The diagnosis of gastrointestinal (GI) involvement in Kaposi's sarcoma (KS) is important to make because the need for treatment depends on the extent of the disease. Moreover, severe GI lesions can cause serious complications. Endoscopy with biopsy is an extremely useful method to diagnose GI-KS. However, determining the indications for endoscopy is difficult because KS can occur without GI symptoms or cutaneous KS. This study sought to clarify predictive clinical factors for GI-KS and its severity on endoscopy.

Methodology/Principal Findings

A total of 1,027 HIV-infected patients who underwent endoscopy were analyzed. Sexual behavior, CD4 count, HIV RNA, history of highly active antiretroviral therapy (HAART), GI symptoms, and cutaneous KS were assessed. Endoscopic severity including bulky tumor, ulceration, and number of lesions were evaluated. Thirty-three patients had GI-KS and 46 patients cutaneous KS. Among the GI-KS patients, 78.8% (26/33) had no GI symptoms and 24.2% (8/33) had no cutaneous KS. Univariate analysis identified men who have sex with men (MSM), CD4 <100 cells/µL, HIV RNA ≥10,000 copies/mL, no history of HAART, and cutaneous KS were significantly associated with GI-KS. Among these factors, cutaneous KS was closely related to GI-KS on multivariable analysis. Among patients without cutaneous KS, MSM and CD4 count <100 cells/µL were the only independent clinical factors related to GI-KS. Bulky tumor was significantly associated with CD4 <100 cells/µL and large number of lesions was significantly associated with HIV-RNA ≥10,000 copies/mL.

Conclusions

To diagnose GI-KS, clinical factors need to be considered before endoscopy. The presence of GI symptoms is not useful in predicting GI-KS. MSM and CD4 count <100 cells/µL are predictive factors among patients without cutaneous KS. Caution should be exercised especially in patients with low CD4 counts or high HIV viral loads as they are more likely to develop severe GI-KS lesions.

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<![CDATA[The SH3BGR/STAT3 Pathway Regulates Cell Migration and Angiogenesis Induced by a Gammaherpesvirus MicroRNA]]> https://www.researchpad.co/article/5989dae9ab0ee8fa60bbe755

Kaposi’s sarcoma (KS)-associated herpesvirus (KSHV) is a gammaherpesvirus etiologically associated with KS, a highly disseminated angiogenic tumor of hyperproliferative spindle endothelial cells. KSHV encodes 25 mature microRNAs but their roles in KSHV-induced tumor dissemination and angiogenesis remain unknown. Here, we investigated KSHV-encoded miR-K12-6-3p (miR-K6-3p) promotion of endothelial cell migration and angiogenesis, which are the underlying mechanisms of tumor dissemination and angiogenesis. We found that ectopic expression of miR-K6-3p promoted endothelial cell migration and angiogenesis. Mass spectrometry, bioinformatics and luciferase reporter analyses revealed that miR-K6-3p directly targeted sequence in the 3’ untranslated region (UTR) of SH3 domain binding glutamate-rich protein (SH3BGR). Overexpression of SH3BGR reversed miR-K6-3p induction of cell migration and angiogenesis. Mechanistically, miR-K6-3p downregulated SH3BGR, hence relieved STAT3 from SH3BGR direct binding and inhibition, which was required for miR-K6-3p maximum activation of STAT3 and induction of cell migration and angiogenesis. Finally, deletion of miR-K6 from the KSHV genome abrogated its effect on the SH3BGR/STAT3 pathway, and KSHV-induced migration and angiogenesis. Our results illustrated that, by inhibiting SH3BGR, miR-K6-3p enhances cell migration and angiogenesis by activating the STAT3 pathway, and thus contributes to the dissemination and angiogenesis of KSHV-induced malignancies.

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