BSA conjugates bearing multiple copies of the basic domain of HIV-1 Tat: Prototype for the development of multitarget inhibitors of extracellular Tat

Abstract

a b s t r a c tThe transactivating factor (Tat) of HIV-1 is involved in AIDS progression and associated pathologies. Tat possesses a basic amino acid sequence implicated in heparan sulfate proteoglycan (HSPG)-mediated internalization, nuclear localization and transactivation by Tat and in the interaction of Tat with integrins and with the vascular endothelial growth factor receptor 2 (KDR) (kinase insert domain receptor). A BSA conjugate bearing an average of four copies of a peptide representing the basic domain/nuclear localization signal of Tat (BSA-Tat-NLS) inhibits transactivation by Tat exogenously added to cells but not by Tat endogenously produced after cell transfection with a tat cDNA, indicating that BSA-Tat-NLS does not interfere with Tat at an intracellular level. Surface plasmon resonance (SPR) experiments revealed that BSA-Tat-NLS binds to the HSPG analogue heparin. Accordingly, BSA-Tat-NLS binds to HSPGs of HL3T1 cell surface and inhibits HSPG-dependent Tat internalization. BSA-Tat-NLS retains its inhibitory potential when pre-incubated with HL3T1 cells before Tat administration, possibly by masking cell-surface HSPGs thus preventing Tat binding and internalization. SPR experiments revealed that BSA-Tat-NLS binds also to integrin ␣ v ␤ 3 and KDR. Accordingly, it inhibits pro-angiogenic endothelial cell adhesion to Tat and motogenesis. In conclusion, BSA-Tat-NLS binds/masks three different cell-surface receptors of Tat inhibiting different biological activities. These data point to BSA-Tat-NLS as a prototype for the development of Tat-antagonists endowed with a multitargeted mechanism of action.

Identification of a Dendrimeric Heparan Sulfate-Binding Peptide That Inhibits Infectivity of Genital Types of Human Papillomaviruses

“…The EC 50 and saturation binding values obtained for the SB105-A10 peptide with both wild-type CHO and HL3T1 cells configure a low-affinity, high-capacity binding pattern, consistent with previously reported interactions of viral peptides (11) and proteins (12,49,47) with cell surface HSPGs. Taken together, these data demonstrate that the binding of SB105-A10 to the surfaces of two different epithelial cell types occurs mainly via HSPGs.…”

“…5) comparable to the K d of its binding to heparin (15.6 to 87.6 nM) (Table 4). Also, these values are comparable to those already reported for the binding of viral proteins/peptides to heparin or HSPGs (11,12,49). (iii) SB105-A10 retained its capacity to inhibit HPV infection in the preattachment assay, demonstrating that its anti-HPV-16 potential is dependent on its interaction with the cell surface.…”

The AGMA1 poly(amidoamine) inhibits the infectivity of herpes simplex virus in cell lines, in human cervicovaginal histocultures, and in vaginally infected mice

“…Interestingly are also the observation that the binding of HSV-1 and 2 glycoproteins gD to nectin-1 depends on several basic amino acids, including L25, R36, R134 and R222 [65] and that HSV-2 infection can be mediated by a v b 3 integrin [66] that is well known to bind its physiological or pathological ligand via basic domains [67e69]. Taken together, these data suggest that the high positive charge of AGMA1 may mediate its binding to receptors different from HSPGs, conferring to the polymer a "multitarget" mechanism of action, as already demonstrated for cationic dendrimer-like compounds [70].…”

Heparin/Heparan Sulfate Proteoglycans Glycomic Interactome in Angiogenesis: Biological Implications and Therapeutical Use

“…HIV-1 Tat is a HIV-1 encoded pro-angiogenic peptide [ 181 ] that binds heparin with a K d equal to 5–64 nM [ 182 , 183 ]. It contains a basic domain composed by a linear stretch of positively charged amino acids ( Table 3 ) mainly responsible for its interaction with heparin that, in turn, requires sulfation of the 2-O -, N-O -, and 6-O -positions ( Table 2 ).…”