Harr Michael, Lavik Andrew, McColl Karen, Zhong Fei, Haberer Ben, Aldabbagh Khadijah, Yee Vivien, Distelhorst Clark W
Case Western Reserve University.
Res Sq. 2023 Jan 9:rs.3.rs-2436910. doi: 10.21203/rs.3.rs-2436910/v1.
There is increasing evidence that the T-cell protein, Lck, is involved in the pathogenesis of chronic lymphocytic leukemia (CLL) as well as other leukemias and lymphomas. We previously discovered that Lck binds to domain 5 of inositol 1,4,5-trisphosphate receptors (IP3R) to regulate Ca2+ homeostasis. Using bioinformatics, we targeted a region within domain 5 of IP3R-1 predicted to facilitate protein-protein interactions (PPIs). We generated a synthetic 21 amino acid peptide, KKRMDLVLELKNNASKLLLAI, which constitutes a domain 5 sub-domain (D5SD) of IP3R-1 that specifically binds Lck via its SH2 domain. With the addition of an HIV-TAT sequence to enable cell permeability of D5SD peptide, we observed wide-spread, Ca2+-dependent, cell killing of hematological cancer cells when the Lck-IP3R PPI was disrupted by TAT-D5SD. All cell lines and primary cells were sensitive to D5SD peptide, but malignant T-cells were less sensitive compared with B-cell or myeloid malignancies. Mining of RNA-seq data showed that LCK was expressed in primary diffuse large B-cell lymphoma (DLBCL) as well as acute myeloid leukemia (AML). In fact, LCK shows a similar pattern of expression as many well-characterized AML oncogenes and is part of a protein interactome that includes FLT3-ITD, Notch-1, and Kit. Consistent with these findings, our data suggest that the Lck-IP3R PPI may protect malignant hematopoietic cells from death. Importantly, TAT-D5SD showed no cytotoxicity in three different non-hematopoietic cell lines; thus its ability to induce cell death appears specific to hematopoietic cells. Together, these data show that a peptide designed to disrupt the Lck-IP3R PPI has a wide range of pre-clinical activity in leukemia and lymphoma.
越来越多的证据表明,T细胞蛋白Lck参与了慢性淋巴细胞白血病(CLL)以及其他白血病和淋巴瘤的发病机制。我们之前发现Lck与肌醇1,4,5-三磷酸受体(IP3R)的结构域5结合,以调节Ca2+稳态。利用生物信息学,我们靶向IP3R-1结构域5内一个预测可促进蛋白质-蛋白质相互作用(PPI)的区域。我们生成了一种合成的21个氨基酸的肽,KKRMDLVLELKNNASKLLLAI,它构成了IP3R-1的结构域5子结构域(D5SD),该子结构域通过其SH2结构域特异性结合Lck。通过添加HIV-TAT序列以使D5SD肽具有细胞通透性,当Lck-IP3R PPI被TAT-D5SD破坏时,我们观察到血液系统癌细胞出现广泛的、Ca2+依赖性的细胞杀伤。所有细胞系和原代细胞对D5SD肽敏感,但恶性T细胞与B细胞或髓系恶性肿瘤相比敏感性较低。对RNA-seq数据的挖掘显示,LCK在原发性弥漫性大B细胞淋巴瘤(DLBCL)以及急性髓系白血病(AML)中表达。事实上,LCK显示出与许多已充分表征的AML致癌基因相似的表达模式,并且是包括FLT3-ITD、Notch-1和Kit在内的蛋白质相互作用组的一部分。与这些发现一致,我们的数据表明Lck-IP3R PPI可能保护恶性造血细胞免于死亡。重要的是,TAT-D5SD在三种不同的非造血细胞系中没有显示出细胞毒性;因此其诱导细胞死亡的能力似乎对造血细胞具有特异性。总之,这些数据表明,一种设计用于破坏Lck-IP3R PPI的肽在白血病和淋巴瘤中具有广泛的临床前活性。