Targeting prion-like protein doppel selectively suppresses tumor angiogenesis

Al-Hilal, Taslim A.,Chung, Seung Woo,Choi, Jeong Uk,Alam, Farzana,Park, Jooho,Kim, Seong Who,Kim, Sang Yoon,Ahsan, Fakhrul,Kim, In-San,Byun, Youngro American Society for Clinical Investigation 2016 The Journal of clinical investigation Vol.126 No.4

<P>Controlled and site-specific regulation of growth factor signaling remains a major challenge for current antiangiogenic therapies, as these antiangiogenic agents target normal vasculature as well tumor vasculature. In this article, we identified the prion-like protein doppel as a potential therapeutic target for tumor angiogenesis. We investigated the interactions between doppel and VEGFR2 and evaluated whether blocking the doppel/VEGFR2 axis suppresses the process of angiogenesis. We discovered that tumor endothelial cells (TECs), but not normal ECs, express doppel; tumors from patients and mouse xenografts expressed doppel in their vasculatures. Induced doppel overexpression in ECs enhanced vascularization, whereas doppel constitutively colocalized and complexed with VEGFR2 in TECs. Doppel inhibition depleted VEGFR2 from the cell membrane, subsequently inducing the internalization and degradation of VEGFR2 and thereby attenuating VEGFR2 signaling. We also synthesized an orally active glycosaminoglycan (LHbisD4) that specifically binds with doppel. We determined that LHbisD4 concentrates over the tumor site and that genetic loss of doppel in TECs decreases LHbisD4 binding and targeting both in vitro and in vivo. Moreover, LHbisD4 eliminated VEGFR2 from the cell membrane, prevented VEGF binding in TECs, and suppressed tumor growth. Together, our results demonstrate that blocking doppel can control VEGF signaling in TECs and selectively inhibit tumor angiogenesis.</P>

Design, Synthesis, and Therapeutic Evaluation of Poly(acrylic acid)–tetraDOCA Conjugate as a Bile Acid Transporter Inhibitor

Park, Jooho,Al-Hilal, Taslim A.,Jeong, Jee-Heon,Choi, Jeong uk,Byun, Youngro American Chemical Society 2015 Bioconjugate chemistry Vol.26 No.8

<P>Regulation of cholesterol and bile acid homeostasis has been attracting attention as a pharmaceutical target for the treatment of diseases, such as hypercholesterolaemia and type 2 diabetes. In recent years, small bile acid analogues have been developed for the purpose of apical sodium-dependent bile acid transporter (ASBT) inhibition. Here, we designed a novel hydrophilic ASBT inhibitor using oligomeric bile acid with a high affinity with ASBT. Polyacrylic acid-tetraDOCA conjugates (PATD) have the ability to bind to ASBT in order to induce hypocholesterolemic effects. Both the viability and the functionality of PATD were evaluated in vitro, showing that PATDs were effective in inhibiting the increases of cholesterol in the blood and oil in the liver induced by high fat diet (HFD). The results indicated that the newly developed biomaterials with oligomeric bile acids and a hydrophilic polymer are potent therapeutic agents for hyperlipidemia.</P>

Absorption Mechanism of a Physical Complex of Monomeric Insulin and Deoxycholyl-<small>l</small>-lysyl-methylester in the Small Intestine

Mahmud, Foyez,Jeon, Ok-Cheol,Al-Hilal, Taslim A.,Kweon, Seho,Yang, Victor C.,Lee, Dong Soo,Byun, Youngro American Chemical Society 2015 MOLECULAR PHARMACEUTICS Vol.12 No.6

<P>Currently, oral administration of insulin still remains the best option to avoid the burden of repeated subcutaneous injections and to improve its pharmacokinetics. The objective of the present investigation was to demonstrate the absorption mechanism of insulin in the physical complexation of deoxycholyl-<SMALL>l</SMALL>-lysyl-methylester (DCK) for oral delivery. The oral insulin/DCK complex was prepared by making a physical complex of insulin aspart with DCK through ion-pair interaction in water. For the cellular uptake study, fluorescein-labeled insulin or DCK were prepared according to a standard protocol and applied to Caco-2 or MDCK cell lines. For the PK/PD studies, we performed intrajejunal administration of different formulation of insulin/DCK complex to diabetic rats. The resulting insulin and DCK complex demonstrated greatly enhanced lipophilicity as well as increased permeation across Caco-2 monolayers. The immunofluorescence study revealed the distribution of the complex in the cytoplasm of Caco-2 cells. Moreover, in the apical sodium bile acid transporter (ASBT) transfected MDCK, the insulin/DCK complex showed interaction with ASBT, and also demonstrated absorption through passive diffusion. We could not find that any evidence of endocytosis in relation to the uptake of insulin complex in vitro. In the rat intestine model, the highest absorption of insulin complex was observed in the jejunum at 1 h and then in the ileum at 2–4 h. In PK/PD study, the complex showed a similar PK profile to that of SC insulin. Overall, the study showed that the effect of DCK on enhancing the absorption of insulin resulted from transcellular processes as well as bile acid transporter activity.</P><P><B>Graphic Abstract</B> <IMG SRC='http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/mpohbp/2015/mpohbp.2015.12.issue-6/mp500626a/production/images/medium/mp-2014-00626a_0009.gif'></P><P><A href='http://pubs.acs.org/doi/suppl/10.1021/mp500626a'>ACS Electronic Supporting Info</A></P>

Transplantation of Heterospheroids of Islet Cells and Mesenchymal Stem Cells for Effective Angiogenesis and Antiapoptosis

Shin, Jung-Youn,Jeong, Jee-Heon,Han, Jin,Bhang, Suk Ho,Jeong, Gun-Jae,Haque, Muhammad R.,Al-Hilal, Taslim A.,Noh, Myungkyung,Byun, Youngro,Kim, Byung-Soo Mary Ann Liebert 2015 Tissue engineering. Part A Vol.21 No.5

<P>Although islet transplantation has been suggested as an alternative therapy for type 1 diabetes, there are efficiency concerns that are attributed to poor engraftment of transplanted islets. Hypoxic condition and delayed vasculogenesis induce necrosis and apoptosis of the transplanted islets. To overcome these limitations in islet transplantation, heterospheroids (HSs), which consist of rat islet cells (ICs) and human bone marrow-derived mesenchymal stem cells (hMSCs), were transplanted to the kidney and liver. The HSs cultured under the hypoxic condition system exhibited a significant increase in antiapoptotic gene expression in ICs. hMSCs in the HSs secreted angiogenic and antiapoptotic proteins. With the HS system, ICs and hMSCs were successfully located in the same area of the liver after transplantation of HSs through the portal vein, whereas the transplantation of islets and the dissociated hMSCs did not result in localization of transplanted ICs and hMSCs in the same area. HS transplantation resulted in an increase in angiogenesis at the transplantation area and a decrease in the apoptosis of transplanted ICs after transplantation into the kidney subcapsule compared with transplantation of islet cell clusters (ICCs). Insulin production levels of ICs were higher in the HS transplantation group compared with the ICC transplantation group. The HS system may be a more efficient transplantation method than the conventional methods for the treatment of type 1 diabetes.</P>

A heparin conjugate, LHbisD4, inhibits lymphangiogenesis and attenuates lymph node metastasis by blocking VEGF-C signaling pathway

Choi, Jeong Uk,Chung, Seung Woo,Al-Hilal, Taslim A.,Alam, Farzana,Park, Jooho,Mahmud, Foyez,Jeong, Jee-Heon,Kim, Sang Yoon,Byun, Youngro IPC Science and Technology Press 2017 Biomaterials Vol.139 No.-

<P><B>Abstract</B></P> <P>Clinical studies have found that the incidence of cancer metastasis through the lymphatic vessels are 3–5 times higher than that through the blood vessels. These findings suggest the potency of anti-lymphangiogenic therapy in reducing the incidence of cancer metastasis. Previously, we reported LHbisD4, which is the conjugate of low molecular weight heparin (LMWH) and four bis-deoxycholates as a potent anti-angiogenic drug with less toxicity and orally active property. Here, we show that LHbisD4 could also suppress the formation of new lymphatic vessels and attenuate the incidence of metastasis by blocking VEGF-C signaling pathway. LHbisD4 significantly enhanced binding affinity with VEGF-C when compared with LMWH, which enables LHbisD4 to suppress the proliferation, migration and formation of tubular structures of human dermal lymphatic endothelial cells(HDLECs) in <I>in vitro</I> condition even in the presence of excessive amounts of VEGF-C. Similarly, we found that the density of lymphatic vessels in the primary tumor tissue in breast cancer bearing mice was significantly diminished when LHbisD4 was administered compared with the control group. Also, the incidence of axillary lymph nodes and distant organ metastasis was significantly reduced in the LHbisD4 administered group, which demonstrates that LHbisD4 could successfully lower the incidence of metastasis through blocking VEGF-C induced lymphangiogenesis. Based on these results, we propose LHbisD4 as a potent anti-cancer drug that can reduce the incidence of metastasis by suppressing lymphangiogenesis through blocking VEGF-C signaling pathway.</P>

Multi-stage inhibition in breast cancer metastasis by orally active triple conjugate, LHTD4 (low molecular weight heparin-taurocholate-tetrameric deoxycholate)

( Farzana Alam ),( Taslim A. Al Hilal ),( Jooho Park ),( Jeong Uk Choi ),( Foyez Mahmud ),( Jee Heon Jeong ),( In San Kim ),( Sang Yoon Kim ),( Seung Rim Hwang ),( Youngro Byun ) 영남대학교 약품개발연구소 2016 영남대학교 약품개발연구소 연구업적집 Vol.26 No.-

Targeting multiple stages in metastatic breast cancer is one of the effective ways to inhibit metastatic progression. To target human metastatic breast cancer as well as improving patient compliance, we developed an orally active low molecular weight heparin (LMWH)-taurocholate conjugated with tetra-meric deoxycholic acid, namely LHTD4, which followed by physical complexation with a synthetic bile acid enhancer, DCK. In breast cancer, both transforming growth factor-β1(TGF-β1) and CXCL12 exhibit enhanced metastatic activity during the initiation and progression stages of breast cancer, thus we direct the focus on investigating the antimetastatic effect of LHTD4/DCK complex by targeting TGF- βl and CXCL12. Computer simulation study and SPR analysis were performed for the binding confirmation of LHTD4 with TGF- βl and CXCL12. We carried out in vitro phosphorylation assays of the consecutive re-ceptors of TGF-βl and CXCL12 (TGF-βl R1 and CXCR4, respectively). Effects of LHTD4 on in vitro cell migration (induced by TGF- βl) and chemotaxis (mediated by CXCL12) were investigated. The in vivo anti-metastatic effect of LHTD4 was evaluated in an accelerated metastasis model and an orthotopic MDA-MB-231 breast cancer model. The obtained KD values ofTGF- β1 and CXCL12 with LHTD4 were 0.85 and 0.019 ,βM respectively. The simulation study showed that binding affinities of LHTD4 fragment with either TGF-βl or CXCL 12 through additional electrostatic interaction was more stable than that of LMWH fragment. In vitro phosphorylation assays of TGF-βl R1 and CXCR4 showed that the effective inhibition of receptor phosphorylation was observed with the treatment of LHTD4. The expressions of epithelial to mesenchymal transition (EMT) marker proteins such as vimentin and Snail were prevented by LTHD4 treatment in In vitro studies with TGF-~l treated MDA-MB-231 cells. Moreover, we observed that LHTD4 negatively regulated the functions of TGF-βl and CXCL 12 on migration and invasion of breast cancer cell. In several advanced orthotopic and experimental breast cancer metastasis murine models. the treatment with LHTD4 (5 mgjkg daily. p.o.) significantly inhibited metastasis compared to the control. Overall, LHTD4 exhibited anti-metastatic effects by inhibiting TGF-βl and CXCL12. and the clinically relevant dose of orally active LHTD4 was found to be effective in preclinical studies without any apparent toxicity.

Medicinal Chemistry : Article ; Size Controlled Heparin Fragment.Deoxycholic Acid Conjugate Showed Anticancer Property by Inhibiting VEGF165

( Joo Ho Park ),( Jee Heon Jeong ),( Taslim A. Al-hilal ),( Ji Young Kim ),( Young Ro Byun ) 영남대학교 약품개발연구소 2015 영남대학교 약품개발연구소 연구업적집 Vol.25 No.-

Heparin is a highly sulfated, long, and linear polysaccharide, which can inhibit tumor growth by interacting with growth factors such as bFGF and VEGF. Several researchers have shown the anti-angiogenic effect of heparin and its conjugates in relation to growth factor inhibition. For drug development and inhibition of growth factors using heparin conjugates, the molecular size of heparin may be crucial considering the size of the heparin binding site of growth factors. In this study, we synthesized heparin fragments and deoxycholic acid conjugated heparin fragments (HFD) to search for the optimal size-controlled conjugate that will inhibit the angiogenic effect of VEGF165. We have also shown that the HFDs could have an enhanced therapeutic effect in vitro and in vivo consequent to the molecular size control. HFDs have significant anti-angiogenic effects by blocking the angiogenic activity of VEGF165 depending on its molecular size. Among them, HFD2 was a promising candidate for oral angiogenesis inhibitor. These results suggest that size-controlled synthesis is necessary for heparin-based drug development.

Size Controlled Heparin Fragment–Deoxycholic Acid Conjugate Showed Anticancer Property by Inhibiting VEGF₁₆₅

Park, Jooho,Jeong, Jee-Heon,Al-Hilal, Taslim A.,Kim, Ji-young,Byun, Youngro American Chemical Society 2015 Bioconjugate chemistry Vol.26 No.5

<P>Heparin is a highly sulfated, long, and linear polysaccharide, which can inhibit tumor growth by interacting with growth factors such as bFGF and VEGF. Several researchers have shown the anti-angiogenic effect of heparin and its conjugates in relation to growth factor inhibition. For drug development and inhibition of growth factors using heparin conjugates, the molecular size of heparin may be crucial considering the size of the heparin binding site of growth factors. In this study, we synthesized heparin fragments and deoxycholic acid conjugated heparin fragments (HFD) to search for the optimal Size-controlled conjugate that will inhibit the angiogenic effect of VEGF(165). We have also shown that the HFDs could have an enhanced therapeutic effect in vitro and in vivo consequent to the molecular size control. HFDs have significant anti-angiogenic:effects by blocking the angiogenic activity of VEGF(165) depending on its molecular size. Among them, HFD2 was a promising Candidate for oral angiogenesis inhibitor. These results suggest that size-controlled synthesis is necessary for heparin-based drug development.</P>

Oral delivery of ionic complex of ceftriaxone with bile acid derivative in non-human primates.

Jeon, Ok-Cheol,Hwang, Seung Rim,Al-Hilal, Taslim A,Park, Jin Woo,Moon, Hyun Tae,Lee, Seulki,Park, Jae Hyung,Byun, Youngro Kluwer Academic/Plenum Publishers 2013 Pharmaceutical research Vol.30 No.4

<P>Since the absorption of ceftriaxone (CTO) in the intestine is restricted by its natural physiological characteristics, we developed a series of small synthetic compounds derived from bile acids to promote the absorption of CTO in the gastrointestinal tract.</P>

Antithrombotic efficacy of an oral low molecular weight heparin conjugated with deoxycholic asset on microsurgical anastomosis in rats

Eom, Jin Sup,Koh, Kyung S.,Al-Hilal, Taslim A.,Park, Jin Woo,Jeon, Ok Cheol,Moon, Hyun Tae,Byun, Youngro Elsevier 2010 Thrombosis research Vol.126 No.3

<P><B>Abstract</B></P><P><B>Introduction</B></P><P>Thrombogenic occlusion, at the anastomotic microvessels, contributed impaired blood flow to flap failure. The effect of an orally active low molecular weight heparin (LMWH) derivative conjugated with deoxycholic acid (DOCA) on the patency of anastomosis of the crushed rat artery was investigated, expecting its antithrombogenic effect.</P><P><B>Materials and Methods</B></P><P>60 Femoral arteries of 30 rats, divided into three groups of 20 each, were reanastomosed. LMWH-DOCA conjugate was orally administered prior to and after operation for 5 consecutive days. On the sixth day of operation, the patency of the anastomosed artery was evaluated.</P><P><B>Results</B></P><P>The patency of oral LMWH-DOCA (10mg/kg) group was significantly enhanced from 15% to 45%, compared to non-treated control group. On the other hand, when the dosage of LMWH-DOCA was reduced to 1mg/kg, its efficacy on anastomosis was not as efficacious in terms of patency. The intima of crushed artery was impaired and thrombus formation was examined in the control group. In the drug treated group, the patency was only compromised by a thin layer of thrombus that covered the inner layer of the vessel without causing any damage to the internal elastic lamina.</P><P><B>Conclusion</B></P><P>The medication of oral LMWH-DOCA conjugate has been vetted in microvascular anastomosis of the crushed artery. LMWH-DOCA was potentially useful for improving the patency in compromised vessels after microsurgery.</P>