Maternal HBV infection induces HBVeAg/PD-L1 dependent tolerance in offspring

Hepatitis B virus (HBV) can be vertically transmitted from mother to babies. Unlike exposure at adulthood, immunological consequences of such exposure to HBV in newborns is "tolerance" to chronic HBV infection.

Ordinarily mice are not naturally infected with HBV. New paper in Immunity reported development of mouse model to study immunology of vertical transmission of HBV.

This model is a combination of two independent processes: 1st, use of HBV transgenic mice and 2nd, hydrodynamic injection of plasmid that contained 1.3-mer HBV genomic DNA. Initially, the authors showed mice born to HBV Tg females (referred here as TGD mouse), but not controls, developed "chronic HBV infection" when exposed to HBV plasmid. 

  

Analysis of HBV-specific CD8 T cells from HBV-DNA injected TGD mice liver showed up-regulation of checkpoint inhibitor PD-1 on T cells.

Treatment of TGD mice with anti-PDL1 antibody enabled them to control HBV-DNA infection.

Moreover, the authors found that clodronate-liposome depletion of macrophages from TGD mice also enabled them to control HBV-DNA infection.

Finally, the authors found that HBVeAg played important role in viral persistence and tolerance by observing that (a) mice born to HBV-mut Tg females (HBV lacking eAg) do not develop persistent HVB-DNA infection, and (b) TGD mice exposed to HBV-DNA lacking eAg also were able to eliminate virus.

In summary, this study suggests that HBVeAg controls newborn's tolerance to HBV via inhibitory PD-L1 signaling.

Few additional thoughts: first, the authors reported that after HBV-DNA injection, around 15% of liver macrophages, called Kupffer cells, expressed viral antigen. However, they also found that >75% of Kupffer cells expressed PD-L1 upon HBV-DNA injection. Second, since HBV virus from HBV Tg pregnant females cannot directly infect newborns, it is not clear how HBVeAg is able to modulate macrophages and tolerize newborn's CD8 T cells (the authors also acknowledged this inconsistency).

David Usharauli

CD47 and PD-L1 work in tandem with MYK to keep T cells at bay

This week Science published a short study examining the role of MYK in tumor immune evasion mechanisms [MYC is a transcription factor that regulates the expression of a multitude of gene products involved in cell proliferation, growth, differentiation, and apoptosis"].The authors found that MYK selectively directs expression of two immune-inhibitory molecules, CD47 and PD-L1, in several mouse and human tumor cells lines.

Using Tet-off transgenic mouse model to control MYK expression in T cell acute lymphoblastic leukemia (MYC T-ALL), the authors showed that both "in vitro or in vivo MYC inactivation resulted in a rapid downregulation of CD47 and PD-L1 [but not other surface markers], both at the mRNA level, as detected by quantitative real-time PCR (qPCR), and at the protein level, as detected by flow cytometry".

Since previous results demonstrated "complete tumor clearance following the inactivation of oncogenes, including MYC", the authors wanted to examine whether MYK effect on tumor regression involved CD47/PD-L1 tandem. Indeed, forced expression of either CD47 or PD-L1 in MYC T-ALL reduced anti-tumor effect of MYK inactivation.

In summary, this study suggests that MYK oncogene provides tumor cells with defenses to repel immune cells. 

David Usharauli

Selective PD-L1/2 expression in lymphoid tissues allows tumor escape during allo-HSCT

The goal of allogeneic hematopoietic stem cell transplantation (allo-HSCT) is to achieve graft-versus-leukemia (GVL) effect to eliminate residual tumor cells in host [left after irradiation]. Since most allo-HSCs are derived from HLA-compatible donors, GVL reaction is mediated by donor T cells reacting against host minor histocompatibility antigens (miHAg; self or tumor-specific Ags). However, even reaction to self-miHAg could still induce graft-versus-host disease (GVHD) because self-miHAgs are broadly expressed in different tissues. Interestingly, GVHD preferentially affects peripheral tissues such as liver or gut. But why?

A new [and very good] paper in journal Immunity may have an answer for this question. In this study, the authors found that selective expression of PD-1 ligands, PD-L1 and PD-L2, in allo-HSCT recipients' lymphoid tissues [but not in peripheral tissues] provide a "cover" against cytotoxic activity exerted by donor CD8 T cells.

To mimic allo-HSCTs, the authors have used female-to-male BM stem cell transplantation model. In this model, self-miHAg is represented by male antigen, HY. To induce GVHD, female BM cells were transplanted alongside with HY-specific transgenic MataHari CD8 T cells. As expected, only male recipients of female HSCs + MataHari CD8 T cells developed GVHD (in liver, gut and skin tissue).

To address the question why only those peripheral tissues were affected by GVHD, the authors examined the hypothesis that HY specific CD8 T cell cytotoxic activity were differentially affected by different tissues. Indeed, co-transfer of labeled male and female targets revealed that recipients of allo-HSCs + MataHari CD8 T cells showed selective reduction of cytotoxic activity against male targets in lymphoid tissues (but not in liver).

This observation was supported by the fact that in contrast to peripheral tissues, MataHari CD8 T cells obtained from lymphoid tissues expressed low level of granzyme B (molecule involved in cytotoxic activity).

To understand why it is the case, the authors examined expression of inhibitory molecules on CD8 T cells. This revealed that while PD-1 on CD8 T cells were similarly expressed irrespective of tissue origin, its ligands, PD-L1 and PD-L2 were selectively up-regulated in lymphoid tissues.

The role of PD-L1 and PD-L2 in inhibition of CD8 T cell cytotoxicity in lymphoid tissues were confirmed in  experiment with anti-PD1 antibody.

Finally, using anti-PD-1 antibody injection, the authors showed that B cell leukemia cells that were hiding in lymphoid tissues of allo-HSCs male recipients could be now eliminated by MataHari CD8 T cells.

In summary, this study showed that (a) during GVHD donor CD8 T cell activity is differentially regulated by different tissues based on availability of inhibitory PD-1 signaling and that (b) this is exploited by leukemia cells to hide in lymphoid tissues but it could be overcome by anti-PD1 antibody injection.

David Usharauli

Gene signature of tumors under immune pressure

Tumor is a tissue undergoing homeostasis-resistance turnover. Animal studies suggested that immune system can detect and sometimes eliminate tumor. In humans, however, the data are more complex and there is no clear answer, especially for solid tumors.

Since experimentation on human subjects ad libitum is obviously not an option, the scientists are constantly developing indirect, ex vivo approaches to study human tumor immunology. Advances in omics science made it possible to analyze large data set to find a clinically-relevant correlations between tumor-specific signatures.

This is exactly what this new paper published in journal Cell tried to do. This group led by Nir Hacohen at the Massachusetts General Hospital, has analyzed human tumor samples for gene signatures looking for evidence for their adaptation to local anti-tumor immune response. Such knowledge, if proven reliable, could be helpful  in determining the effectiveness of tumor immunotherapies. 

The authors has decided to focus on cytolytic gene signature (granzyme A and perforin) as a readout for tumors sensitivity to local immune response. Initial analyses showed that tumors varied in their expression of cytolytic signature. 

The authors reasoned that one reason for difference in cytolytic gene signature could be due to presence of tissue-specific viruses, frequently associating with tumors, like HPV virus for cervical cancer tissues. 

Additionally, the authors found there was evidence for depletion (reduction) of neo-epitope expression from predicted number of HLA-binding non-silent mutations, implying these tumors were under immune pressure.

Correspondingly, the authors identified several mutated genes that were enriched in tumors under immune pressure. One of these genes was caspase 8, a well-known member of cell death pathway.

Finally, the authors found there was positive or negative correlation between high cytolytic gene signature and amplification of gene regions containing immuno-regulatory genes such as PD-L1, PD-L2 and IDO1, IDO2.

In summary, these results provided indirect evidence suggesting that human immune system can detect tumor neo-epitopes and mount cytolytic response to eliminate it, thus exerting pressure on tumor cells to develop inhibitory counter-measures. Targeting these pathways could benefit the development of effective immunotherapeutic approaches.

Now there are few limitation to this study. First, cytolytic signature molecules, granzyme A and perforin, can be evidence of immune suppression rather than immune activity. Second, since the authors do not provide tumor patients survival data, it makes hard to argue for usefulness of these results for clinical application.

David Usharauli