ETHNOPHARMACOLOGICAL RELEVANCE:Nerium oleander is used to treat liver-associated chronic metabolic diseases in traditional medicinal systems across the globe. The hepatoprotective effects of oleander are mentioned in Indian and Chinese traditional medicinal literature. AIM OF THE STUDY:The present study aimed to investigate the cellular mechanisms behind the hepatoprotective effects of a non-toxic dose of oleander (NO). MATERIALS AND METHODS:The hepatoprotective effects of NO were tested against lipopolysaccharide (LPS)-treated HepG2 cells. Oxidative stress response was studied using cellular enzymatic assays, and gene expression was analyzed using qRT-PCR. HepG2 cells were pretreated with TAK-242 (pharmacological inhibitor of TLR4) to decipher the anti-inflammatory mechanisms of NO. Cell-free metabolites were analyzed using GCMS and were subjected to pathway enrichment analysis. RESULTS:NO reduced systemic inflammation, serum lipid peroxidation byproducts, and glucose without affecting serum transaminase levels and hepatic histopathological features. NO attenuated the inflammation-induced loss of antioxidant enzyme activities and mRNA expressions of toll-like receptor-4 (TLR4)/nuclear factor κβ (NFκβ)-dependent inflammatory genes. In TAK-242 pretreated cells, LPS was unable to induce inflammatory and oxidative responses. However, NO treatment in TAK-242 pretreated cells with LPS stimulation further reduced the signs of inflammation and improved hepatoprotective activities. A comparative analysis of the intracellular global metabolome from HepG2 cells with and without NO treatment indicated NO-mediated favorable modulation of intracellular metabolic pathways that support cytoprotective activities. CONCLUSION:NO protects HepG2 cells from LPS-induced oxidative and inflammatory injury. The hepatoprotective effects of NO are mediated by a TLR4-independent process and through a favorable modulation of the intracellular global metabolome that supports cytoprotection.

OBJECTIVE:To perform testing for cytokines involved in dermal inflammatory reactions and to document and compare the effects of an oleander extract (OE), oleandrin, and oclacitinib on biomarkers relevant to allergic reactions. The effects of these compounds under inflamed culture conditions are of direct importance to the treatment of canine atopic dermatitis. METHODS:Testing involved primary canine dermal fibroblasts and the canine DH82 macrophage cell line; both cell types are important for initiating, regulating, and resolving dermal allergic reactions via cytokine communication. RESULTS:Under inflamed conditions, OE and oleandrin downregulated key cytokines secreted by canine dermal fibroblasts and the DH82 macrophage cell line; all of which are treatment targets in dermatitis. In the DH82 macrophage cultures, the most noteworthy reductions involved IL-6, IL-12/IL-23p40, interferon-γ, tumor necrosis factor-α, VEGF, and nerve growth factor-β. Oclacitinib triggered reductions of some cytokines involved in allergic reactions, including TGF-β1, IL-12/IL-23p40, and tumor necrosis factor-α; however, these reductions were less robust than the reductions triggered by OE and oleandrin and accompanied by increases in other cytokines involved in dermal inflammation, including IL-6, interferon-γ, and nerve growth factor-β. In cultures of primary dermal fibroblasts, OE and oleandrin reduced the levels of IL-8 and monocyte chemoattractant protein-1, whereas oclacitinib had little or no effect. CONCLUSIONS:Oleander extract and oleandrin directly modulate immune responses under inflamed conditions. Moreover, OE and oleandrin appear to provide a more beneficial overall cytokine regulation than oclacitinib under inflamed culture conditions. CLINICAL RELEVANCE:These results suggest that OE and oleandrin are efficacious agents to treat canine atopic dermatitis. Future studies should evaluate the efficacy of these compounds in dogs affected by atopic dermatitis.

OBJECTIVE The purpose of the study reported herein was to determine the dose of oleander extract and oleandrin (the key pharmacologically active constituent) that could be safely administered PO to dogs. ANIMALS 42 purebred Beagle dogs were used to study an extract of Nerium oleander . METHODS 3 studies were performed in 42 purebred young adult (ages 12 months or older) Beagle dogs using a supercritical fluid extract of N oleander leaves. The first study was an 8-day initial dose-ranging study in 2 dogs, a second 7-day repeat-dosing study was performed in 4 dogs, and the final study was performed in 32 dogs where test subjects were given extract or placebo once daily for 28 consecutive days via oral (gavage) administration followed by a 14-day recovery period. RESULTS At 2.3 µg/kg of oleandrin, there were no observable adverse effects during the duration of the study. Adverse effects were not seen until doses exceeded 6.9 µg/kg of oleandrin, at which time mild, reversible clinical signs were noted. However, a dose > 460 µg of oleandrin/kg was fatal in 1 of 2 dogs in this study. CLINICAL RELEVANCE The studies reported here, taken in totality, suggest that doses exceeding 6.9 µg/kg of oleandrin may be associated with cardiac abnormalities. An estimated no treatment effective adverse event oral dose of oleandrin appears to be 4.6 µg of oleandrin/kg. Higher doses may be tolerable but should be used with appropriate monitoring.

Successful treatment of glioblastoma multiforme (GBM), an aggressive form of primary brain neoplasm, mandates the need to develop new therapeutic strategies. In this study, we investigated the potential of PBI-05204 in targeting GBM stem cells (GSCs) and the underlying mechanisms. Treatment with PBI-05204 significantly reduced both the number and size of tumor spheres derived from patient-derived GSCs (GBM9, GSC28 and TS543), and suppressed the tumorigenesis of GBM9 xenografts. Moreover, PBI-05204 treatment led to a significant decrease in the expression of CD44 and NANOG, crucial markers of progenitor stem cells, in GBM9 and GSC28 GSCs. This treatment also down-regulated GRP78 expression in both GSC types. Knocking down GRP78 expression through GRP78 siRNA transfection in GBM9 and GSC28 GSCs also resulted in reduced spheroid size and CD44 expression. Combining PBI-05204 with GRP78 siRNA further decreased spheroid numbers compared to GRP78 siRNA treatment alone. PBI-05204 treatment led to increased expression of pRIP1K and pRIP3K, along with enhanced binding of RIPK1/RIPK3 in GBM9 and GSC28 cells, resembling the effects observed in GRP78-silenced GSCs, suggesting that PBI-05204 induced necroptosis in these cells. Furthermore, oleandrin, a principle active cardiac glycoside component of PBI-05204, showed the ability to inhibit the self-renewal capacity in GSCs. These findings highlight the potential of PBI-05204 as a promising candidate for the development of novel therapies that target GBM stem cells.

Supplementary Figure 1 from Profiling Lipoxygenase Metabolism in Specific Steps of Colorectal Tumorigenesis

Perspective from Regulation of Prostaglandin Transporters in Colorectal Neoplasia

Supplementary Table 1 from Biological Activity of Celecoxib in the Bronchial Epithelium of Current and Former Smokers

The Nerium oleander extract PBI 05204 (PBI) and its cardiac glycoside constituent oleandrin have direct anti-viral properties. Their effect on the immune system, however, is largely unknown. We used an in vitro model of human peripheral blood mononuclear cells to document effects under three different culture conditions: normal, challenged with the viral mimetic polyinosinic:polycytidylic acid Poly I:C, and inflamed by lipopolysaccharide (LPS). Cells were evaluated for immune activation marks CD69, CD25, and CD107a, and culture supernatants were tested for cytokines. Both PBI and oleandrin directly activated Natural Killer (NK) cells and monocytes and triggered increased production of cytokines. Under viral mimetic challenge, PBI and oleandrin enhanced the Poly I:C-mediated immune activation of monocytes and NK cells and enhanced production of IFN-γ. Under inflammatory conditions, many cytokines were controlled at similar levels as in cultures treated with PBI and oleandrin without inflammation. PBI triggered higher levels of some cytokines than oleandrin. Both products increased T cell cytotoxic attack on malignant target cells, strongest by PBI. The results show that PBI and oleandrin directly activate innate immune cells, enhance anti-viral immune responses through NK cell activation and IFN-γ levels, and modulate immune responses under inflamed conditions. The potential clinical impact of these activities is discussed.

BACKGROUND:Bovine viral diarrhea virus (BVDV), bovine respiratory syncytial virus (BRSV). and bovine coronavirus (BCV) threaten the productivity of cattle worldwide. Development of therapeutics that can control the spread of these viruses is an unmet need. The present research was designed to explore the in vitro antiviral activity of the Nerium oleander derived cardiac glycoside oleandrin and a defined N. oleander plant extract (PBI-05204) containing oleandrin.METHODS:Madin Darby Bovine Kidney (MDBK) cells, Bovine Turbinate (BT) cells, and Human Rectal Tumor-18 (HRT-18) cells were used as in vitro culture systems for BVDV, BRSV and BCV, respectively. Cytotoxicity was established using serial dilutions of oleandrin or PBI-05204. Noncytotoxic concentrations of each drug were used either prior to or at 12 h and 24 h following virus exposure to corresponding viruses. Infectious virus titers were determined following each treatment.RESULTS:Both oleandrin as well as PBI-05204 demonstrated strong antiviral activity against BVDV, BRSV, and BCV, in a dose-dependent manner, when added prior to or following infection of host cells. Determination of viral loads by PCR demonstrated a concentration dependent decline in virus replication. Importantly, the relative ability of virus produced from treated cultures to infect new host cells was reduced by as much as 10,000-fold at noncytotoxic concentrations of oleandrin or PBI-05204.CONCLUSIONS:The research demonstrates the potency of oleandrin and PBI-05204 to inhibit infectivity of three important enveloped bovine viruses in vitro. These data showing non-toxic concentrations of oleandrin inhibiting infectivity of three bovine viruses support further investigation of in vivo antiviral efficacy.

Glioblastoma multiforme (GBM) is the most common as well as one of the most malignant types of brain cancer. Despite progress in development of novel therapies for the treatment of GBM, it remains largely incurable with a poor prognosis and a very low life expectancy. Recent studies have shown that oleandrin, a unique cardiac glycoside from Nerium oleander, as well as a defined extract (PBI-05204) that contains this molecule, inhibit growth of human glioblastoma, and modulate glioblastoma patient-derived stem cell-renewal properties. Here we demonstrate that PBI-05204 treatment leads to an increase in vitro in the sensitivity of GBM cells to radiation in which the main mechanisms are the transition from autophagy to apoptosis, enhanced DNA damage and reduced DNA repair after radiotherapy (RT) administration. The combination of PBI-05204 with RT was associated with reduced tumor progression evidenced by both subcutaneous as well as orthotopic implanted GBM tumors. Collectively, these results reveal that PBI-05204 enhances antitumor activity of RT in preclinical/murine models of human GBM. Given the fact that PBI-05204 has already been examined in Phase I and II clinical trials for cancer patients, its efficacy when combined with standard-of-care radiotherapy regimens in GBM should be explored.

Glioblastoma multiforme (GBM) is the most common as well as one of the most malignant types of brain cancer. Despite progress in development of novel therapies for the treatment of GBM, it remains largely incurable with a poor prognosis and a very low life expectancy. Recent studies have shown that oleandrin, a unique cardiac glycoside from Nerium oleander, as well as defined extract (PBI-05204) that contains this molecule, inhibit growth of human glioblastoma, and modulate glioblastoma patient-derived stem cell-renewal properties. The present study aimed to investigate the radiosensitization of PBI-05204 in glioblastoma using both in vitro and in vivo cancer models as well as to explore the potential mechanism of actions in GBM. The radiosensitizing effect of PBI-05204 was assessed against human GBM U87MG, U251, T98G and A172 cell lines as well as their relevant xenograft and orthotopic models. The induction of apoptosis, DNA damage and repair of DNA double strand breaks were assessed with determination of caspase 3 and 9 protein expression, DNA laddering, protein expression of rH2AX, Ku70, DNA-PKcs, and RAD51 as well as a Comet Assay. PBI-05204 treatment leads to an increased in vitro sensitivity of GBM cells, including U87MG, U251, T98G and A172 cells, to radiotherapy (RT) in which the main mechanisms are the transition from autophagy to apoptosis and enhanced DNA damage evidenced by increased expression of γH2AX. Additionally, relative increased expression of Ku70, DNA-PKcs and RAD51 due to RT were reduced by PBI-05204 in U87MG and U251 cells, suggesting PBI-05204 lessened RT mediated DNA repair. PBI-05204 significantly enhanced the RT mediated inhibition of tumor growth by 4.7-, 2.1- and 2.2-fold in U87MG, U251 and T98G xenograft models, respectively. The combination of RT and PBI-05204 showed a significantly enhancement of disease-free survival to 103.0 ± 63.2 days compared to the control group (p < 0.001) which was 3-fold longer than that of RT only group. Collectively, these results reveal that PBI-05204 enhances antitumor activity of RT in preclinical/murine models of human GBM. Given the fact that PBI-05204 has already been examined in Phase I and II clinical trials for cancer patients, its efficacy when combined with standard-of-care radiotherapy regimens in GBM should be explored in future clinical trials of this difficult to treat brain cancer. Citation Format: Alessandro Colapietro, Peiying Yang, Alessandra Rosetti, Andrea Mancini, Flora Vital, Stefano Martellucci, Francesco Marampon, Vincenzo Mattei, Giovanni Luca Gravina, Robert Newman, Claudio Festuccia. The botanical drug PBI-05204, a supercritical CO₂ extract of Nerium oleander, augments the antitumor efficacy of radiotherapy in treatment of human glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6307.

Treatment of rhabdomyosarcoma (RMS), the most common a soft tissue sarcoma in childhood, provides intensive multimodal therapy, with radiotherapy (RT) playing a critical role for local tumor control. However, since RMS efficiently activates mechanisms of resistance to therapies, despite improvements, the prognosis remains still largely unsatisfactory, mainly in RMS expressing chimeric oncoproteins PAX3/PAX7-FOXO1, and fusion-positive (FP)-RMS. Cardiac glycosides (CGs), plant-derived steroid-like compounds with a selective inhibitory activity of the Na+/K+-ATPase pump (NKA), have shown antitumor and radio-sensitizing properties. Herein, the therapeutic properties of PBI-05204, an extract from Nerium oleander containing the CG oleandrin already studied in phase I and II clinical trials for cancer patients, were investigated, in vitro and in vivo, against FN- and FP-RMS cancer models. PBI-05204 induced growth arrest in a concentration dependent manner, with FP-RMS being more sensitive than FN-RMS, by differently regulating cell cycle regulators and commonly upregulating cell cycle inhibitors p21(Waf1/Cip1) and p27(Cip1/Kip1). Furthermore, PBI-05204 concomitantly induced cell death on both RMS types and senescence in FN-RMS. Notably, PBI-05204 counteracted in vitro migration and invasion abilities and suppressed the formation of spheroids enriched in CD133(+) cancer stem cells (CSCs). PBI-05204 sensitized both cell types to RT by improving the ability of RT to induce G2 growth arrest and counteracting the RT-induced activation of both Non-Homologous End-Joining and homologous recombination DSBs repair pathways. Finally, the antitumor and radio-sensitizing proprieties of PBI-05204 were confirmed in vivo. Notably, both in vitro and in vivo evidence confirmed the higher sensitivity to PBI-05204 of FP-RMS. Thus, PBI-05204 represents a valid radio-sensitizing agent for the treatment of RMS, including the intrinsically radio-resistant FP-RMS.

Stress can alter immunological, neurochemical and endocrinological functions, but its role in cancer progression is not well understood. Here, we show that chronic behavioral stress results in higher levels of tissue catecholamines, greater tumor burden and more invasive growth of ovarian carcinoma cells in an orthotopic mouse model. These effects are mediated primarily through activation of the tumor cell cyclic AMP (cAMP)–protein kinase A (PKA) signaling pathway by the β 2 adrenergic receptor (encoded by ADRB2 ). Tumors in stressed animals showed markedly increased vascularization and enhanced expression of VEGF, MMP2 and MMP9, and we found that angiogenic processes mediated the effects of stress on tumor growth in vivo . These data identify β-adrenergic activation of the cAMP–PKA signaling pathway as a major mechanism by which behavioral stress can enhance tumor angiogenesis in vivo and thereby promote malignant cell growth. These data also suggest that blocking ADRB-mediated angiogenesis could have therapeutic implications for the management of ovarian cancer.

With continued expansion of the coronavirus disease (COVID-19) pandemic, caused by severe acute respiratory syndrome 2 (SARS-CoV-2), both antiviral drugs as well as effective vaccines are desperately needed to treat patients at high risk of life-threatening disease. Here, we present in vitro evidence for significant inhibition of SARS-CoV-2 by oleandrin and a defined extract of N. oleander (designated as PBI-06150). Using Vero cells, we found that prophylactic (pre-infection) oleandrin (as either the pure compound or as the active principal ingredient in PBI-06150) administration at concentrations as low as 0.05 µg/ml exhibited potent antiviral activity against SARS-CoV-2, with an 800-fold reduction in virus production, and a 0.1 µg/ml concentration resulted in a greater than 3000-fold reduction in infectious virus production. The half maximal effective concentration (EC50) values were 11.98 ng/ml when virus output was measured at 24 h post-infection, and 7.07 ng/ml measured at 48 h post-infection. Therapeutic (post-infection) treatment up to 24 h after SARS-CoV-2 infection of Vero cells also reduced viral titers, with 0.1 µg/ml and 0.05 µg/ml concentrations causing greater than 100-fold reduction as measured at 48 h, and the 0.05 µg/ml concentration resulting in a 78-fold reduction. Concentrations of oleandrin up to 10 µg/ml were well tolerated in Vero cells. We also present in vivo evidence of the safety and efficacy of defined N. oleander extract (PBI-06150), which was administered to golden Syrian hamsters in a preparation containing as high as 130 µg/ml of oleandrin. In comparison to administration of control vehicle, PBI-06150 provided a statistically significant reduction of the viral titer in the nasal turbinates (nasal conchae). The potent prophylactic and therapeutic antiviral activities demonstrated here, together with initial evidence of its safety and efficacy in a relevant hamster model of COVID-19, support the further development of oleandrin and/or defined extracts containing this molecule for the treatment of SARS-CoV-2 and associated COVID-19 disease and potentially also for reduction of virus spread by persons diagnosed early after infection.

More importantly, our published studies demonstrated the safety of the plant extract in the Food and Drug Administration (FDA)-approved Phase I and II clinical trials in cancer patients through the use of our novel drug PBI-05204. [...]that is our current aim for PBI-05204. With respect to oleandrin and extracts containing this molecule, we point out that although toxicity can result in the use of high doses of this molecule, when used appropriately either for cancer or, as suggested, for treatment of certain viral-mediated diseases (including envelope viruses), the compound can be used with both safety and, we hope to prove, clinical efficacy.

Glioblastoma multiform (GBM) is the most common primary glial tumor resulting in very low patient survival despite current extensive therapeutic efforts. Emerging evidence suggests that more effective treatments are required to overcome tumor heterogeneity, drug resistance and a complex tumor-supporting microenvironment. PBI-05204 is a specifically formulated botanical drug consisting of a modified supercritical C02 extract of Nerium oleander that has undergone both phase I and phase II clinical trials in the United States for treatment of patients with a variety of advanced cancers. The present study was designed to investigate the antitumor efficacy of this botanical drug against glioblastoma using both in vitro and in vivo cancer models as well as exploring efficacy against glioblastoma stem cells. All three human GBM cell lines, U87MG, U251, and T98G, were inhibited by PBI-05204 in a concentration dependent manner that was characterized by induction of apoptosis as evidenced by increased ANNEXIN V staining and caspase activities. The expression of proteins associated with both Akt and mTOR pathway was suppressed by PBI-05240 in all treated human GBM cell lines. PBI-05204 significantly suppressed U87 spheroid formation and the expression of important stem cell markers such as SOX2, CD44, and CXCR4. Oral administration of PBI-05204 resulted in a dose-dependent inhibition of U87MG, U251, and T98G xenograft growth. Additionally, PBI-05204–treated mice carrying U87-Luc cells as an orthotropic model exhibited significantly delayed onset of tumor proliferation and significantly increased overall survival. Immunohistochemical staining of xenograft derived tumor sections revealed dose-dependent declines in expression of Ki67 and CD31 positive stained cells but increased TUNEL staining. PBI-05204 represents a novel therapeutic botanical drug approach for treatment of glioblastoma as demonstrated by significant responses with in vivo tumor models. Both in vitro cell culture and immunohistochemical studies of tumor tissue suggest drug induction of tumor cell apoptosis and inhibition of PI3k/mTOR pathways as well as cancer stemness. Given the fact that PBI-05204 has already been examined in phase I and II clinical trials for cancer patients, its efficacy when combined with standard of care chemotherapy and radiotherapy should be explored in future clinical trials of this difficult to treat brain cancer.

Over the past 15 years, investigators have reported on the utility and safety of cardiac glycosides for numerous health benefits including those as treatments for malignant disease, stroke-mediated ischemic injury and certain neurodegenerative diseases. In addition to those, there is a growing body of evidence for novel antiviral effects of selected cardiac glycoside molecules. One unique cardiac glycoside, oleandrin derived from Nerium oleander, has been reported to have antiviral activity specifically against 'enveloped' viruses including HIV and HTLV-1. Importantly, a recent publication has presented in vitro evidence for oleandrin's ability to inhibit production of infectious virus particles when used for treatment prior to, as well as after infection by SARS-CoV-2/COVID-19. This review will highlight the known in vitro antiviral effects of oleandrin as well as present previously unpublished effects of this novel cardiac glycoside against Ebola virus, Cytomegalovirus, and Herpes simplex viruses.

With continued expansion of the COVID-19 pandemic, antiviral drugs are desperately needed to treat patients at high risk of life-threatening disease and even to limit spread if administered early during infection. Typically, the fastest route to identifying and licensing a safe and effective antiviral drug is to test those already shown safe in early clinical trials for other infections or diseases. Here, we tested in vitro oleandrin, derived from the Nerium oleander plant and shown previously to have inhibitory activity against several viruses. Using Vero cells, we found that prophylactic oleandrin administration at concentrations down to 0.05 μg/ml exhibited potent antiviral activity against SARS-CoV-2, with an 800-fold reduction in virus production, and a 0.1 μg/ml dose resulted in a greater than 3,000-fold reduction in infectious virus production. The EC 50 values were 11.98ng/ml when virus output was measured at 24 hours post-infection, and 7.07ng/ml measured at 48 hours post-infection. Therapeutic (post-infection) treatment up to 24 hours after infection of Vero cells also reduced viral titers, with the 0.1 μg/ml dose causing greater than 100-fold reductions as measured at 48 hours, and the 0.05 μg/ml dose resulting in a 78-fold reduction. The potent prophylactic and therapeutic antiviral activities demonstrated here strongly support the further development of oleandrin to reduce the severity of COVID-19 and potentially also to reduce spread by persons diagnosed early after infection.IMPORTANCE:COVID-19, a pandemic disease caused by infection with SARS-CoV-2, has swept around the world to cause millions of infections and hundreds-of-thousands of deaths due to the lack of vaccines and effective therapeutics. We tested oleandrin, derived from the Nerium oleander plant and shown previously to reduce the replication of several viruses, against SARS-CoV-2 infection of Vero cells. When administered both before and after virus infection, nanogram doses of oleandrin significantly inhibited replication by up to 3,000-fold, indicating the potential to prevent disease and virus spread in persons recently exposed to SARS-CoV-2, as well as to prevent severe disease in persons at high risk. These results indicate that oleandrin should be tested in animal models and in humans exposed to infection to determine its medical usefulness in controlling the pandemic.

At present, there are no antiretroviral drugs that inhibit incorporation of the envelope glycoprotein into newly-synthesized virus particles. The botanical glycoside, oleandrin, derived from extracts of Nerium oleander, has previously been shown to reduce the levels of the gp120 envelope glycoprotein on human immunodeficiency virus type-1 (HIV-1) particles and inhibit HIV-1 infectivity in vitro. We therefore tested whether oleandrin or an extract from N. oleander could also inhibit the infectivity of the human T-cell leukemia virus type-1 (HTLV-1): A related enveloped retrovirus and emerging tropical infectious agent. The treatment of HTLV-1+ lymphoma T-cells with either oleandrin or a N. oleander extract did not significantly inhibit viral replication or the release of p19Gag-containing particles into the culture supernatants. However, the collected virus particles from treated cells exhibited reduced infectivity on primary human peripheral blood mononuclear cells (huPBMCs). Unlike HIV-1, extracellular HTLV-1 particles are poorly infectious and viral transmission typically occurs via direct intercellular interactions across a virological synapse. We therefore investigated whether oleandrin or a N. oleander extract could inhibit virus transmission from a GFP-expressing HTLV-1+ lymphoma T-cell-line to huPBMCs in co-culture assays. These results demonstrated that both oleandrin and the crude phytoextract inhibited the formation of virological synapses and the transmission of HTLV-1 in vitro. Importantly, these findings suggest oleandrin may have broad antiviral activity against enveloped viruses by reducing the incorporation of the envelope glycoprotein into mature particles, a stage of the infection cycle not targeted by modern HAART.