Thomas Albers

Multi-kinase inhibitors can associate with heat shock proteins through their NH2-termini by which they suppress chaperone function

We performed proteomic studies using the GRP78 chaperone-inhibitor drug AR-12 (OSU-03012) as bait. Multiple additional chaperone and chaperone-associated proteins were shown to interact with AR-12, including: GRP75, HSP75, BAG2; HSP27; ULK-1; and thioredoxin. AR-12 down-regulated in situ immuno-fluorescence detection of ATP binding chaperones using antibodies directed against the NH2-termini of the proteins but only weakly reduced detection using antibodies directed against the central and COOH portions of the proteins. Traditional SDS-PAGE and western blotting assessment methods did not exhibit any alterations in chaperone detection. AR-12 altered the sub-cellular distribution of chaperone proteins, abolishing their punctate speckled patterning concomitant with changes in protein co-localization. AR-12 inhibited chaperone ATPase activity, which was enhanced by sildenafil; inhibited chaperone – chaperone and chaperone – client interactions; and docked in silico with the ATPase domains of HSP90 and of HSP70. AR-12 combined with sildenafil in a GRP78 plus HSP27 –dependent fashion to profoundly activate an eIF2α/ATF4/CHOP/Beclin1 pathway in parallel with inactivating mTOR and increasing ATG13 phosphorylation, collectively resulting in formation of punctate toxic autophagosomes. Over-expression of [GRP78 and HSP27] prevented: AR-12 –induced activation of ER stress signaling and maintained mTOR activity; AR-12 –mediated down-regulation of thioredoxin, MCL-1 and c-FLIP-s; and preserved tumor cell viability. Thus the inhibition of chaperone protein functions by AR-12 and by multi-kinase inhibitors very likely explains why these agents have anti-tumor effects in multiple genetically diverse tumor cell types.

Blocking Myristoylation of Src Inhibits Its Kinase Activity and Suppresses Prostate Cancer Progression

Protein N-myristoylation enables localization to membranes and helps maintain protein conformation and function. N-myristoyltransferases (NMT) catalyze co- or posttranslational myristoylation of Src family kinases and other oncogenic proteins, thereby regulating their function. In this study, we provide genetic and pharmacologic evidence that inhibiting the N-myristoyltransferase NMT1 suppresses cell-cycle progression, proliferation, and malignant growth of prostate cancer cells. Loss of myristoylation abolished the tumorigenic potential of Src and its synergy with androgen receptor in mediating tumor invasion. We identified the myristoyl-CoA analogue B13 as a small-molecule inhibitor of NMT1 enzymatic activity. B13 exposure blocked Src myristoylation and Src localization to the cytoplasmic membrane, attenuating Src-mediated oncogenic signaling. B13 exerted its anti-invasive and antitumor effects against prostate cancer cells, with minimal toxic side-effects in vivo Structural optimization based on structure-activity relationships enabled the chemical synthesis of LCL204, with enhanced inhibitory potency against NMT1. Collectively, our results offer a preclinical proof of concept for the use of protein myristoylation inhibitors as a strategy to block prostate cancer progression. Cancer Res; 77(24); 6950-62. ©2017 AACR.

Diazapentacene derivatives: synthesis, properties, and structures

Two new diazapentacene derivatives were synthesized and studied photochemically and electrochemically. Both compounds indicate potential as ambipolar semiconductors. A single crystal X-ray structure has been obtained for one of the compounds.

Multi-kinase inhibitors interact with sildenafil and ERBB1/2/4 inhibitors to kill tumor cells in vitro and in vivo

We have recently demonstrated that multi-kinase inhibitors such as sorafenib and pazopanib can suppress the detection of chaperones by in situ immuno-fluorescence, which is further enhanced by phosphodiesterase 5 inhibitors. Sorafenib and pazopanib inhibited the HSP90 ATPase activity with IC50 values of ~1.0 μM and ~75 nM, respectively. Pazopanib docked in silico with two possible poses into the HSP90 ATP binding pocket. Pazopanib and sildenafil combined to reduce the total protein levels of HSP1H/p105 and c-MYC and to reduce their co-localization. Sorafenib/pazopanib combined with sildenafil in a [GRP78+HSP27] –dependent fashion to: (i) profoundly activate an eIF2α/Beclin1 pathway; (ii) profoundly inactivate mTOR and increase ATG13 phosphorylation, collectively resulting in the formation of toxic autophagosomes. In a fresh PDX isolate of NSCLC combined knock down of [ERBB1+ERBB3] or use of the ERBB1/2/4 inhibitor afatinib altered cell morphology, enhanced ATG13 phosphorylation, inactivated NFκB, and further enhanced [sorafenib/pazopanib + sildenafil] lethality. Identical data to that with afatinib were obtained knocking down PI3K p110α/β or using buparlisib, copanlisib or the specific p110α inhibitor BYL719. Afatinib adapted NSCLC clones were resistant to buparlisib or copanlisib but were more sensitive than control clones to [sorafenib + sildenafil] or [pazopanib + sildenafil]. Lapatinib significantly enhanced the anti-tumor effect of [regorafenib + sildenafil] in vivo; afatinib and BYL719 enhanced the anti-tumor effects of [sorafenib + sildenafil] and [pazopanib] in vivo, respectively.

Abstract A070: Targeting N-myristoyltransferase 1 inhibits prostate cancer progression

N-myristoyltransferase (NMT) facilitates co/post-translational myristoylation of several oncogenic proteins, regulating their functions in tumor progression. N-myristoyltransferase (NMT), which has been shown to be upregulated in a variety of cancers, is overexpressed in androgen-independent prostate cancer cells. We demonstrate that genetically, ablation of NMT1 inhibited proliferation of prostate cancer cells, tumor growth, and suppressed myristoylation profile of prostate cancer cells. Screening a panel of myristoyl-CoA analogs against purified human NMT1 protein leads to identifying the B13 as an inhibitor for NMT1. B13 significantly suppressed prostate cancer cell proliferation, migration, and invasion by cell cycle arrest, and inhibited the growth of prostate xenograft tumors with minimal pathologic effect on major organs in vivo. Structure activity relationship based optimization of B13 led to LCL204, which showed better inhibitory properties towards NMT1. We demonstrate that targeting protein myristoylation is a potential therapeutic approach to inhibit prostate tumor progression. Citation Format: Omar Alsaidan, Sungjin Kim, Octavia Goodwin, Qianjin Li, Aiping Bai, Alicja Bielawska, George Zheng, Iryna Lebedyeva, Thomas Albers, James S. Norris, Scott D. Pegan, Houjian Cai. Targeting N-myristoyltransferase 1 inhibits prostate cancer progression [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A070.

Benzotriazole-Based Strategies Toward Peptidomimetics, Conjugates, and Other Peptide Derivatives

Benzotriazole-mediated routes to peptidomimetics and peptide conjugates have been discussed in detail. The Katritzky group developed a benzotriazole methodology toward the activation of carbonyl groups of amino acids and modified analogs, which allowed the synthesis of cyclic peptides, azapeptides, azidopeptides, aminoxypeptides, oxyazapeptides, depsipeptides, and isopeptides. High-yielding reactions of N-, O-, S-, and C-acylated nucleophiles with activated aminoacyl or peptidoyl benzotriazole derivatives have also been reported. Benzotriazole methodology enabled the efficient incorporation of bioactive moieties into peptides, peptidomimetics, amino acids, and other carbonyl-containing compounds. Predominant number of reported products retained chiral purity. Some of the products displayed promising biological activities such as anticancer and antibacterial activity along with the improved stability under physiological conditions.