Phosphorylated Annexin A2 at Tyrosine 23 Regulates Exosome Release and Biogenesis in Triple Negative Breast Cancer

dc.creatorDesai, Priyanka P.
dc.creatorTripathi, Amit K.
dc.creatorDonkor, Michael
dc.creatorThyagarajan, Srikantha
dc.creatorJones, Harlan
dc.creatorVan Treuren, Timothy
dc.creatorLampe, Jana B.
dc.creatorChaudhary, Panka J.
dc.creatorVishwanatha, Jamboor
dc.creator.orcid0000-0002-3784-8799 (Tripathi, Amit K.)
dc.creator.orcid0000-0002-5488-0328 (Donkor, Michael)
dc.description.abstractPurpose: Exosomes are highly involved in the progression of diverse diseases. Targeting exosome biogenesis and release is a potential strategy for the treatment of the disease like cancer which urges an improved understanding of the process. During the exosomes biogenesis, invagination of the plasma membranes forms early endosomes which mature into late endosomes and multivesicular bodies. Annexin A2 (AnxA2), a calcium dependent phospholipid binding protein, is one of the cargo proteins which gets uploading into the exosomes and impart aggressive phenotype in triple negative breast cancer (TNBC). The mechanism how AnxA2 uploads the exosomal cargo into the exosomes and releases exosomes in the tumor microenvironment remains to be unidentified. In this study, we have explored the potential mechanism for exosome biogenesis and release to target it in TNBC, which lacks the targeted based therapies. Methods: Plasmids expressing constitutive phosphomimetic (AnxA2-Y23E) and non-phosphomimetic AnxA2 (AnxA2-Y23F) mutant gene were transfected in MDA-MB-231 cells. Exosomes isolated from AnxA2-Y23E and AnxA2-Y23F mutant cells were analyzed for expression of the exosomal cargo proteins and RNAs by Western blot and RT-PCR. The number of exosomes released were analyzed by Nanotrack analysis (NTA). Mutant cells treated with Rapamycin, mTORC1(Mammalian Target of Rapamycin Complex 1) inhibitor, were analyzed for the cargo and exosomal secretion. Mutant cells were injected in nude mice to generate tumors. Serum exosomes were isolated and analyzed for cargo and number of exosome release by NTA. Results: In this study, we found that phosphorylated Annexin A2 at tyrosine 23 increases exosome secretion. It loads proteins like AnxA2, CD9 (Cluster of Differentiation 9), LC3B, and Tsg101(Tumor susceptibility gene 101), and AnxA2 and mTOR mRNA into the exosomes. Moreover, secretion and loading of cargo into the exosomes is regulated by increased phosphorylation of AnxA2 and reduced downstream mTORC1 activity. Conclusions: Phosphorylation of AnxA2 at tyrosine 23 regulates exosome secretion and cargo loading into the exosomes in TNBC.
dc.description.sponsorshipResearch reported in this abstract was supported by National Institutes of Health under Award Numbers R01CA220273 (JKV) and S21MD012472 (JKV)
dc.titlePhosphorylated Annexin A2 at Tyrosine 23 Regulates Exosome Release and Biogenesis in Triple Negative Breast Cancer