Remote strikes and defense: activated T cell-derived exosomal PD-1 attenuates PD-L1-induced immune dysfunction in triple-negative breast cancer

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Triple-negative breast cancer (TNBC) is a group of heterogeneous tumors lacking effective shared therapeutic targets. Among numerous potential markers for predicting the biological behavior and clinical outcome of TNBC, a high proportion of the tumor-infiltrating lymphocytes (TILs), currently proposed as a marker of the immunomodulatory signature, is strongly associated with a favorable prognosis despite the ununified classification of TNBC genomic subtypes.

As broadly accepted, PD-1 expressed on the effector T lymphocytes transduces suppressive signaling by the ligation with PD-L1 expressed on cancer cell surface. Therefore, in the combat between cytotoxic T lymphocytes (CTLs) and cancer cells, PD-1 seems to serve as a “traitor” and assists cancer cells to survive by escaping CTL-killing based on its conventional biofunction as a co-inhibitory immune checkpoint. However, the clinical observation of PD-1 in TNBC tumors tells another story. Interestingly, PD-1 expression is significantly associated with longer disease-free survival and overall survival in patients with TNBC. Especially, high numbers of PD-1-positive immune infiltrates are associated with significantly increased disease-free survival. These clinical findings raised an interesting question, from which our project was inspired: whether PD-1 on T lymphocytes might somehow harbor biofunctions in restricting the immune evasion of cancer cells aside from its immunosuppressive activity? 

At that time, our lab just published a paper on Cell Research describing how “cunning” the TNBC cells are to release exosomes carrying PD-L1 to remotely devitalize CTLs, thereby escaping being “executed”. Based on the special characteristic of exosomes, my mentor Dr. Hung and I had a discussion about whether it would be possible that tumor-specific T lymphocytes also release exosomes carrying PD-1 to defend against cancerous PD-L1. Soon after the assumption, with the generous assistance of Dr. Yi Yang, Dr. Riyao Yang and Dr. Linlin Sun, who were also working on projects in cancer immunology and exosomes at that time, we quickly performed the first several qualitative experiments and validated the PD-1 expression in exosomes isolated from stimuli-activated peripheral blood mononuclear cell-derived T lymphocytes and Jurkat model T lymphocytes. From here, our project started to take shape.

After validating the sub-localization of PD-1 on exosome surface by transmission electron microscopy and in-vitro binding of PD-L1 and exosomal PD-1, we were more confident of our hypothesis. Yet still, we needed to make sure the role of exosomal PD-1 and how it impacts on cancer immune surveillance during the confrontation between CTLs and cancer cells. By establishing in-vitro tumor-specific T cell-killing system and in-vivo orthotopic TNBC tumor models, we validated our hypothesis that exosomal PD-1 facilitates CTLs to attenuate the immune evasion of PD-L1-expressing TNBC cells via both occupying cell surface PD-L1 loci and neutralizing exosomal PD-L1 particles.

In summary, our previous study showed that TNBC cell-derived exosomal PD-L1 acts like “PD-1-targeting missiles” to impair CTL function, and our current study shows that this sort of “airstrike” is not launched merely by cancer cells but also by activated T lymphocytes within the tumor microenvironment via releasing exosomal PD-1 as protective “PD-L1-targeting missiles”. Based on our findings, we provide a potential strategy of surface modification for therapeutic tumor-targeting exosomes. It would be rational in the future studies to discover novel approaches to transfer these surface inhibitory immune checkpoints, such as PD-1, Tim-3 and LAG-3, etc., which are currently well-documented as immune suppressors, into releasable exosomal forms to remotely block their ligands expressed on cancer cells. Furthermore, in combination with chemotherapy agents or nucleic acid cargos, exosomes loaded with surface inhibitory immune checkpoint receptors might be able to attenuate the immune suppressive microenvironment extracellularly while simultaneously attacking cancer cells intracellularly. 

As a Ph.D. student, who just finished M.D. training and was quite junior in basic research at the start of this project, I was given many helpful advices from my mentors and all the experienced senior lab members in daily discussions, and they all helped me grow up fast during the exploration in science. This also thanks to Dr. Hung for running the lab in a not only productive but also harmonious way. Even Dr. Hung assumed the President position in China Medical University at Taiwan since February 2019, he organized the project through monthly visit to Houston before the COVID-19 pandemic, and continues the input via Skype, WeChat, etc.  Indeed, working during the COVID-19 pandemic was really a tough time since we were doing strictly time-restricted shifts and everyone was only allowed to work for half a day, which significantly delayed the progress. However, we are glad that things are getting back to normal steadily with all the timely regulations and efforts made by everyone in UT MD. Anderson Cancer Center. Have to say, the scientific training in the Hung Lab would definitely be a memorable experience in my life. 

Yufan Qiu

Ph.D. Candidate, UT MD Anderson Cancer Center