Scientists have identified a platelet-derived chemokine that naturally inhibits infection by a wide range of primary HIV-1 strains, and may point to new therapeutic strategies against the virus. The chemokine, CXCL4 (PF-4), acts similarly to other known anti-HIV chemokines in that it interferes with the earliest stages of infection at the point of viral entry into cells. However, while other known HIV-blocking chemokines prevent HIV gaining access to its target cells by binding to one of the two cell-surface chemokine receptors—CCR5 or CXCR4—which represent the co-receptors used by different strains of the virus to gain a foothold, CLXL4 binds directly to the major viral envelope glycoprotein gp120. This means the platelet-derived chemokine has broad-spectrum inhibitory activity that is unrestricted by viral dependence on the CCR5 or CXCR4 co-receptors.
Reporting in PNAS, the researchers at the National Institute of Allergy and Infectious Diseases (NIAID), and San Raffaele Scientific Institute in Milan, say their discovery could provide new insights into HIV pathogenesis and aid in the design of new therapeutic and preventive approaches. Paolo Lusso, Ph.D., and colleagues describe their findings in a paper titled “Identification of the platelet-derived chemokine CXCL4/PF-4 as a broad-spectrum HIV-1 inhibitor.”
The team initially demonstrated that treating CD4+ T cells with exogenous recombinant human CXCL4 blocked infection by a range of CXCR4- and CCR5-tropic primary HIV-1 isolates, but only if treatment was effected either before the cells were exposed to the virus, or at the point of exposure. If CXCL4 administration was delayed to an hour after infection, the antiviral efficacy was reduced, and administration at four hours after infection was completely ineffective. This indicated that CXCL4 blocks an early stage in viral replication, rather than later stages such as reverse transcription, gene transcription/translation, or maturation. Using an HIV-1 entry assay the researchers were then able to demonstrate that CXCL4 blocked attachment to and entry of the virus into CD4+ T cells.
They were able to rule out the possibility that CXCL4 might act by directly blocking CD4 or other co-receptors based on various lines of evidence. These included the observations that expression of such receptors in primary CD4+ T cells wasn’t downmodulated on treatment with CXCL4, while soluble CD4 didn’t bind to CXCL4 in enzyme immunoassays. Moreover, CXCL4 had no antagonistic effects on CR5- or CSCR4-mediated signaling. Interestingly, CXCR3B, which is the only high-affinity receptor currently known for CXCL4, isn’t highly expressed on HIV-1 target cells adnd hasn’t even been identified as a co-receptor for HIV-1.
In contrast the results from an HIV-1 virion-capture assays and co-immunoprecipitation studies showed that CXCL4 does directly bind to the HIV-1 envelope glycoprotein gp120. Further assays using anti-gp120 antibodies indicated that the chemokine binds to a region on the outer surface of the gp120 glycoprotein that is near to but doesn’t overlap the CD4 binding site.
CXCL4 is primarily produced by megakaryocytes and their platelet offspring, which release it on activation. “Thus, it is important to emphasize that the effective antiviral concentrations of CXCL4 against various primary HIV-1 isolates in physiologically relevant target cells such as CD4+ T cells and macrophages are fully compatible with the physiological levels of this chemokine,” the investigators write. Moreover, CXCL4 exerts immunomodulatory and proinflammatory effects, including monocyte activation and differentiation, which may have particular clinical relevance in terms of platelet activation that occurs during inflammatory processes such as HIV-1 infection. “Thus, CXCL4 may play a dual role in the pathogenesis of HIV-1 disease, on one side by suppressing HIV-1 replication but on the other by fostering immunologic activation, inflammation, and coagulation abnormalities.”
The authors admit that in vivo studies investigating the role of endogenous chemokines in complex diseases like AIDS are tricky, and can point to associations, but not pinpoint cause. However, they note studies do indicate that higher serum levels of CXCL4 in HIV-1 patients correlate with a less advanced clinical stage, “suggesting that this cytokine may play a protective role in vivo.”
Unfortunately, the team has also identified primary HIV-1 isolates that are insensitive to CXCL4-mediated inhibition at the doses used in their reported studies. It thus seems that some strains of the virus may be able to escape CXCL4-mediated protection. The NIAID researchers are currently investigating the influence of both endogenous CXCL4 production and polymorphisms in the CXCL4 gene to see how they affect the course of HIV-1 infection.