- Email: [email protected]
Halting metastasis through CXCR4 inhibition
Bioorganic & Medicinal Chemistry Letters 23 (2013) 20–25
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl
BMCL Digest
Halting metastasis through CXCR4 inhibition Deborah M. Ramsey ⇑, Shelli R. McAlpine ⇑ Department of Chemistry, University of New South Wales, Sydney NSW 2052, Australia
a r t i c l e
i n f o
Article history: Received 2 October 2012 Accepted 31 October 2012 Available online 12 November 2012 Keywords: CXCR4 SDF-1 Metastasis Cancer Heat shock protein 90
a b s t r a c t Metastasis occurs when cancer cells leave the primary tumor site and migrate to distant parts of the body. The CXCR4–SDF-1 pathway facilitates this migration, and its expression has become the hallmark of several metastatic cancers. Targeted approaches are currently being developed to inhibit this pathway, and several candidates are now in clinical trials. Continued exploration of CXCR4 inhibitors will generate compounds that have improved activity over current candidates. Ó 2012 Elsevier Ltd. All rights reserved.
Cells do not stand still. Cell migration is a primordial characteristic observed from the beginning of embryonic development, and directed cell movement continues throughout the life of an organism.1 During normal mammalian embryogenesis, embryonic stem cells (ESCs) traffic to numerous sites under the control of several chemokines, including stromal cell-derived factor-1 (SDF-1; also known as CXCL12) and its corresponding adhesive molecule CXCR4 (CD184).2,3 Adult pluripotent cells maintain a ‘homing’ capacity to specific tissues using the SDF-1/CXCR4 chemosensory system, leading to life-affirming effects such as wound repair and tissue regeneration.4–6 Cancer cells exploit the same migratory pathways utilized by embryonic and adult stem cells.7,8 Metastatic tumors express ESC transcription factors and spread beyond the primary tumor site through SDF-1/CXCR4-mediated migration.9 Expression of CXCR4 has been tied to tumor infiltration and poor prognosis in colon, breast, and gallbladder cancers.10–13 In addition, SDF-1 expression is correlated to reduced survival in ovarian cancer patients.11 A growing body of research points to the CXCR4 pathway as an emerging arena for cancer drug discovery, with the goal of specifically disrupting the receptor-ligand (CXCR4–SDF-1) interaction that drives the spread of tumors. In addition, a deeper understanding of the pathways involved in CXCR4 folding and expression points to the major role that molecular chaperones play in facilitating metastasis.14 Current therapies being tested in clinical trials are utilizing a combinatorial approach for inhibiting the spread of CXCR4-expressing cancer cells, targeting both the receptor-ligand interaction and other pro-oncogenic pathways (Table 1). ⇑ Corresponding authors. Tel.: +61 4 1672 8896; fax: +61 2 9385 6141. E-mail addresses: [email protected] (D.M. Ramsey), [email protected] (S.R. McAlpine). 0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.bmcl.2012.10.138
CXCR4 is a member of the G protein-coupled (GPC) chemokine receptor family, utilizing seven trans-membrane alpha helices to integrate into the cell membrane (Fig. 1). CXCR4 gene expression is downregulated in normal breast cells and non-invasive cancer cell lines, but invasive neuroblastoma and prostate cancer cell lines show high levels of CXCR4 transcription.15,16 Several factors may contribute to the observed increase in gene expression, such as DNA methylation or TNF signaling cascades.17,18 In addition, upregulation of the transcriptional activator HIF-1a leads to a direct increase in CXCR4 gene expression.19 Following post-translational modification, CXCR4 relies on the molecular chaperone heat shock protein 90 (Hsp90) for proper folding and delivery to the lipid bilayer (Fig. 2).14 Hsp90 is also the molecular chaperone for several members of the CXCR4/SDF-1 phosphorylation cascade, including the protein tyrosine kinase Src, human epidermal growth factor receptor 2 (HER2), and the serine/threonine kinase Akt.21,22 Inhibitors of Hsp90 have been studied in clinical trials for efficacy against metastatic cancers,14 and expression levels of several proteins in the CXCR4 pathway are reduced during treatment.23 However, toxicity issues limit the use of current Hsp90 inhibitors as a monotherapy in the clinic.23 The crystal structure of CXCR4 shows that the seven transmembrane alpha helices bundle together to form an extracellular binding pocket (Fig. 124). The extracellular loops (ECLs) and amino acids within the alpha helices contribute to binding interactions with potential ligands. Two disulfide bonds form between key cysteine residues on the extracellular face of the receptor, and these bonds are critical for shaping the binding pocket and for ligand binding: Cys28 of the N-terminal segment with Cys274 of helix VII (H7), and Cys109 with Cys186 in ECL2.20,24 CXCR4 can also homo- or heterodimerize.20,25 This ability could facilitate metastasis in cancer cells expressing high levels of CXCR4, since
21
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25 Table 1 Selected CXCR4 and SDF-1 Inhibitors in clinical triala Sponsor
Drug name
Secondary names
Combination therapy?
Phase
Cancer
MDX-1338, Medarex
Lenalidomide, Dexamethasone, or Bortezomib
IA
Multiple myeloma
Ablynx
BMS936564 ALX-0651
I
Healthy volunteers (safety study)
CXCR4 antagonists Biokine Therapeutics Ltd
BKT-140
I/IIA
Multiple myeloma
Cardiff University
Plerixafor
Daunorubicin, Clofarabine
I
Washington University School of Medicine Weill Medical College (Cornell University) Metastatix, Inc.
Plerixafor
4F-benzoyl-TN14003, BL-8040 Mozobil, Genzyme, AMD3100 Mozobil, Genzyme, AMD3100 Mozobil, Genzyme, AMD3100
Mitoxantrone, Etoposide, Cytarabine Decitabine
I/II
Acute myeloid leukemia; high risk myelodysplastic syndrome Acute myeloid leukemia
I
Acute myeloid leukemia
MSX-122
I
Refractory metastatic or locally advanced solid tumors
NOX-A12
II
Multiple myeloma
Anti-CXCR4 antibodies Bristol–Myers Squib
SDF-1 antagonists Noxxon Pharma AG a
Plerixafor
Information obtained from www.clinicaltrials.gov.
Figure 1. CXCR4 receptor structure and binding pocket. CXCR4 consists of seven trans-membrane helices (H1–7) and three extracellular loops (ECL1-3). The receptor folds into a bundle, creating a binding pocket for its ligand SDF-1.21
Figure 2. CXCR4/SDF-1 signaling cascade. CXCR4 is folded and delivered to the cell surface by Hsp90. Once the chemokine SDF-1 is bound, a phospho-relay transfers the signal to Akt, leading to actin reorganization and subsequent cell migration.
dimerization would amplify the signaling cascade upon ligand binding and lead to enhanced downstream effects. Crystal structures of CXCR4 homodimers indicate that ligand-induced confor-
mational changes to one receptor can actually increase the ligand binding affinity of a second receptor.20 The converse is also true, suggesting that the biological efficacy of CXCR4 antagonists may
22
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25
be tied to whether these modulators induce positive or negative binding cooperativity. The small chemokine SDF-1 binds CXCR4 and other cognate receptors (Fig. 1), triggering multiple signaling cascades that promote cell survival and metastasis.16,26 The source of SDF-1 can be normal tissue (including blood vascular endothelial cells and bone marrow), stromal cells within solid tumors (for paracrine signaling), or cancer cells themselves (for autocrine signaling).27–29 The crystal structure of SDF-1a consists of two monomers that form an asymmetrical dimer.30 Each monomer is composed of three beta strands that form an anti-parallel beta sheet, which is bordered on either side by an alpha helix. The N-terminal amino acids of SDF-1 are thought to be responsible for its ligand binding activity.30 Charged residues within the first alpha helix and the beta sheet, as well as an adjacent hydrophobic region within the beta sheet, may facilitate interactions with the trans-membrane helices of CXCR4. Once SDF-1 is bound, CXCR4, HER2, and possibly extracellular Hsp90, co-localize to lipid rafts, which are cell membrane clusters enriched in cholesterol.31 Not only do lipid rafts facilitate transfer of the signal into the cell interior, but recent evidence suggests that extracellular Hsp90 stabilizes HER2 and promotes this signaling event (Fig. 2).22,31 Once bound to SDF-1, activated CXCR4 increases intracellular calcium levels and induces a phosphorylation cascade.32 CXCR4 is then internalized and can be recycled back to the cell surface or degraded by lysosomal enzymes.32 Cell motility is regulated at the intracellular level by several phosphorylation cascades controlled by Src or Akt (also called protein kinase B). Akt serves as the central node in the CXCR4/SDF-1 signaling cascade (Fig. 2). Association with Hsp90 protects Akt from dephosphorylation and subsequent degradation. Akt phosphorylates the G protein Girdin and other downstream targets that modulate reorganization of actin fibers in the cell.33,34 Actin polymerization is further stabilized by Hsp90, leading to the formation of plasma membrane protrusions known as filopodia.35,36 Thus, tumor cells utilize the CXCR4/SDF-1 signaling cascade to induce actin polymerization and directed cell migration, and CXCR4-positive cancer cells take advantage of autocrine or paracrine-generated SDF-1 gradients to spread to other tissues. The strategic approach to blocking the CXCR4/SDF-1 pathway has centered largely on inhibitors of the receptor CXCR4 or the chemokine SDF-1. CXCR4 inhibitors fall into two categories: antiCXCR4 antibodies and CXCR4 antagonists (Table 1). Since CXCR4 is also the co-receptor for human immunodeficiency virus (HIV) and is required for viral entry into T cells, significant drug design advancements on CXCR4 inhibitors were made before they were utilized as anti-cancer therapies. Chemokine inhibitors are generally well tolerated by patients, and antagonists of SDF-1 are now exhibiting some success in clinical trials.37,38 In addition, targeting other extracellular proteins in the CXCR4 pathway (namely HER2) has also shown some therapeutic promise in inhibiting metastasis and could be combined in future studies with CXCR4/SDF-1 antagonists.39 Anti-CXCR4 antibodies. Amino acids within the CXCR4 binding pocket form specific interactions with SDF-1, but the receptor as a whole is quite flexible and can dislodge the binding of large molecules such as monoclonal antibodies that target its surface.40,41 As a result, anti-CXCR4 antibodies must form strong binding interactions with CXCR4 to form a physical barrier to chemokines attempting to enter the binding pocket (Fig. 3). A recent study examining CXCR4/SDF-1 interactions found that sulfonated tyrosine residues in one of the N-terminal extracellular loops of CXCR4 are absolutely critical for binding to SDF-1.42 Targeting these specific sulfonated tyrosine residues has proven an effective strategy for neutralizing the effects of chemokine-receptor interactions and avoiding the issues associated with monoclonal antibodies tar-
Figure 3. Anti-CXCR4 antibodies and nanobodies bind to the CXCR4 receptor and block SDF-1 from entering the binding pocket.
geted to flexible regions on CXCR4. The current anti-CXCR4 antibody under clinical investigation is the drug candidate MDX1338 (BMS-936564).40 MDX-1338 directly blocks the binding interaction between one of the N-terminal extracellular loops of CXCR4 and SDF-1, causing inhibition of cell migration.43 MDX1338 is potent in the low nanomolar range and reduces AML tumors in murine xenograft models.43 Clinical studies are ongoing, where MDX-1338 is used in combination with standard chemotherapeutic treatments to reduce multiple myeloma (MM; Table 1). One of the drawbacks of monoclonal antibody therapy is that multiple antibodies must be used to target multiple epitopes on a single receptor. Advancements in nanotechnology have led to the development of a unique solution: the nanobody. Nanobodies are antibody fragments with a single monomeric variable domain (monovalent), or two different variable domains held together by a flexible linker (biparatopic).44 Only four nanobodies are currently being studied in clinical trials, and one of those nanobodies (ALX0651) targets CXCR4. Recent data suggests that monovalent CXCR4 nanobodies act as neutral antagonists, which compete with SDF-1 for binding but fail to neutralize multiple CXCR4 receptors at once.44 On the other hand, biparatopic CXCR4 nanobodies are designed to act as inverse agonists, not only by blocking SDF-1, but also by inhibiting multiple CXCR4 receptors through the bivalent construction of the molecule (Fig. 3). This cumulative effect counteracts overexpression and homodimerization of CXCR4 receptors on cancer cell surfaces and leads to downstream effects such as inhibition of cell migration.44 ALX-0651 is a biparatopic nanobody currently in Phase I clinical trials (Table 1), where there is evidence suggesting that this molecule significantly reduces cell migration. Although clinical studies using ALX-0651 are just beginning, there is considerable promise for the use of this new technology in the development of novel anti-metastatic agents. CXCR4 antagonists. A well-known small molecule inhibitor of CXCR4 is AMD3100 (xylyl–bicyclam), which is currently in Phase I/II clinical trials (Fig. 4). AMD3100 was initially studied for its antiviral properties, and the observation that this molecule induced stem cell mobilization was a surprising side effect that led to the examination of its anti-cancer activity.41 The ability of AMD3100 to act as a stem cell mobilizer has been frequently exploited in the clinic, although its mechanism of action is under debate.6 Hematopoetic stem cells (HSCs) in the bone marrow maintain the balance between proliferation and differentiation, replenishing both blood and immune cells in the bloodstream to keep the system at a steady state. CXCR4-positive HSCs are retained in the bone
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25
23
Figure 4. Small molecule antagonists of CXCR4.
marrow by stromal cells producing SDF-1 gradients (paracrine signaling), and SDF-1 binds to CXCR4 and prevents HSC migration outside of the niche. Clinicians can manipulate this storehouse of stem cells for a variety of purposes, including tissue regeneration, or by inducing cancerous stem cells to leave the marrow during chemotherapy. For HSCs to leave the bone marrow, it is hypothesized that CXCR4 antagonists such as AMD3100 work by binding to CXCR4 and blocking the binding of endogenous SDF-1.45 Without the receptor’s ability to respond to its retention signal (i.e. SDF-1 in the bone marrow niche), CXCR4-positive HSCs will leave the marrow and enter the bloodstream. AMD3100 is a metal-chelating, bicyclic, reversible inhibitor that binds in a pocket between the transmembrane helices and extracellular loop (ECL) 2 of CXCR4, leading to allosteric modulation and effective blockade of SDF-1 binding (Fig. 5).46 This antagonist is specific for the CXCR4 receptor and does not cross-react with other GPC-receptors. AMD3100’s selectivity may be due in part to the key hydrophobic contacts formed between the bicyclam rings and two tryptophan residues within the binding pocket of CXCR4, Trp 195 and Trp283.47 Several Phase I/II clinical trials are utilizing this molecule in a combinatorial therapeutic approach to treat acute myeloid leukemia (AML)45. Interestingly, the affin-
ity of these bicyclam complexes for CXCR4 is enhanced when the macrocycle binds to copper, nickel or zinc ions.47,48 Further research is underway to study how metal complexes can strengthen the CXCR4 receptor binding interactions with bicyclam macrocycles. New developments in small molecule inhibitors of CXCR4 have led to some interesting leads that exhibit anti-metastatic properties. MSX-122, currently in Phase I clinical trials, is a small molecule antagonist of CXCR4 that inhibits metastasis of breast, melanoma, and head and neck carcinoma in murine models (Fig. 4).49 There have also been recent leads in the development of small molecules that target sites on CXCR4 other than its binding pocket. One such small molecule inhibitor is the natural product celastrol (Fig. 4), which prevents cell migration by inhibiting translation of CXCR4 mRNA.50 Other compounds have targeted a post-translational modification of CXCR4, where a recent in silico screen identified inhibitors that bound to a sulfotyrosine recognition site within one of the N-terminal extracellular loops of CXCR4.43 One inhibitor identified in this screen reduced SDF-1mediated calcium flux in treated THP-1 cells.42 Peptide inhibitors of CXCR4 are beginning to show promise in the clinic. BKT140 is an analog of T140 (sequence of T140: H-
Figure 5. CXCR4 antagonists compete for binding to the receptor binding pocket. The binding pocket (BP) of CXCR4 is the site for SDF-1 binding, as well as the target of CXCR4 antagonists.
24
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25
Arg-Arg-Nal-Cys-Tyr-Arg-Lys-D-Lys-Pro-Tyr-Arg-Cit-Cys-Arg-OH), a 14-residue peptide that is a potent inverse agonist and CXCR4 antagonist and inhibits metastatic breast cancer in murine models.51 With its potency in the low nanomolar range, BKT140 is now being studied in Phase I/II clinical trials against MM (Table 1). Other peptide analogs have used virally-derived CXCR4 antagonists as the basis for lead compound development. Using 21 amino acids from the human herpesvirus 8 viral macrophage inflammatory protein II (vMIP-II), recent research indicates that D-amino acid analogs of this peptide bind CXCR4 more strongly than the original peptide and serve as better antagonists.40 These discoveries will contribute significantly to the design of new peptide inhibitors. SDF-1 antagonists. SDF-1 antagonists have been studied for their efficacy in both cancer and diabetes research, and promising results have been observed.38 The common SDF-1 antagonist is a 45 base pair L-RNA oligonucleotide (or Spiegelmer) known as NOX-A12.38 This aptamer is currently in Phase II clinical trials as a monotherapy against MM (Table 1). One question that remains to be explored in the clinic is whether CXCR4 antagonism alone can halt metastasis, or whether SDF-1 antagonism is also required for full therapeutic impact. Future clinical studies that combine CXCR4 and SDF-1 antagonists may determine if there is an additive benefit, or if one antagonist is better at inhibiting metastatic disease and preventing relapse. Inhibitors of other proteins in the CXCR4 pathway. The CXCR4/ SDF-1 pathway plays a major role in metastatic breast cancer, where the overexpression of CXCR4 is one of several biomarkers (including HER2/neu) used to diagnose aggressive disease.52 Thus, inhibitors of other proteins in the CXCR4 pathway may be required for optimal therapeutic effect. The role of CXCR4/SDF-1 pathway in metastatic breast cancer has become clearer in recent years. SDF-1 can induce the transactivation of HER2/neu in breast cancer cells,53 and CXCR4 can facilitate homing of metastatic breast cancer cells to the bone.52 More research is needed in metastatic breast cancer clinical trials to determine the efficacy of CXCR4/SDF-1 antagonists in combination with other chemotherapeutic agents. Anti-HER2 monoclonal antibodies, either alone or conjugated to other anticancer drugs (such as Trastuzumab emtansine or T-DM1), may be needed in addition to CXCR4/SDF-1 antagonists for successful treatment of metastatic breast cancer.54 The GPC-receptor CXCR4 and its chemokine SDF-1 play a critical role in cell migration and metastatic disease. Benefiting from early anti-viral research, the development of CXCR4 inhibitors has led to several successful drug candidates that are currently undergoing testing in Phase I/II clinical trials. Future development of carefully designed agonists and antagonists, as well as mining structures from natural product sources, will expand the number of new anti-cancer agents with the potential to halt CXCR4-mediated cell migration. The success of these compounds may require a combination of therapies targeted at multiple proteins involved in this metastatic pathway. ⁄ Dr. Deborah M. Ramsey and Associate Professor Shelli R. McAlpine, School of Chemistry, University of New South Wales, Sydney NSW 2052, Australia. Tel.: +61 2 9385 5505. Email: [email protected] and [email protected]. Acknowledgments We thank the University of New South Wales (UNSW), as well as NIH 1R01CA137873 for support of D.M.R. and S.R.M. References and notes 1. Hunger, C.; Odemis, V.; Engele, J. Exp. Cell Res. 2012, 318, 2178. 2. Cha, Y. R.; Fujita, M.; Butler, M.; Isogai, S.; Kochhan, E.; Siekmann, A. F.; Weinstein, B. M. Dev. Cell 2012, 22, 824.
3. Ciriza, J.; Hall, D.; Lu, A.; De Sena, J. R.; Al-Kuhlani, M.; García-Ojeda, M. E. PLoS One 2012, 7, e30542. 4. Ding, J.; Hori, K.; Zhang, R.; Marcoux, Y.; Honardoust, D.; Shankowsky, H. A.; Tredget, E. E. Wound Repair Regen. 2011, 19, 568. 5. Xia, W. H.; Li, J.; Su, C.; Yang, Z.; Chen, L.; Wu, F.; Zhang, Y. Y.; Yu, B. B.; Qiu, Y. X.; Wang, S. M.; Tao, J. Aging Cell 2012, 11, 111. 6. Brzoska, E.; Kowalewska, M.; Markowska, A.; Kowalski, K.; Archacka, K.; Zimowska, M.; Grabowska, I.; Czerwin´ska, A. M.; Góra, M.; Stremin´ska, W.; Jan´czyk-Ilach, K.; Ciemerych, M. A. Biol. Cell. 2012, Sept 14, Accepted for publication (Epub ahead of print). 7. Kleeberger, W.; Bova, G. S.; Nielsen, M. E.; Herawi, M.; Chuang, A. Y.; Epstein, J. I.; Berman, D. M. Cancer Res. 2007, 67, 9199. 8. Nian, W. Q.; Chen, F. L.; Ao, X. J.; Chen, Z. T. Mol. Cell. Biochem. 2011, 355, 241. 9. Belton, A.; Gabrovsky, A.; Bae, Y. K.; Reeves, R.; Iacobuzio-Donahue, C.; Huso, D. L.; Resar, L. M. PLoS one 2012, 7, e30034. 10. Zhang, N. H.; Li, J.; Li, Y.; Zhang, X. T.; Liao, W. T.; Zhang, J. Y.; Li, R.; Luo, R. C. Exp. Ther. Med. 2012, 3, 973. 11. Popple, A.; Durrant, L. G.; Spendlove, I.; Rolland, P.; Scott, I. V.; Deen, S.; Ramage, J. M. Br. J. Cancer 2012, 106, 1306. 12. Yao, X.; Zhou, L.; Han, S.; Chen, Y. J. Int. Med. Res. 2011, 39, 1253. 13. Hiller, D. J.; Meschonat, C.; Kim, R.; Li, B. D.; Chu, Q. D. Sugery 2011, 150, 459. 14. Mandawat, A.; Fiskus, W.; Buckley, K. M.; Robbins, K.; Rao, R.; Balusu, R.; Navenot, J.; Wang, Z.; Ustun, C.; Chong, D. G.; Atadja, P.; Fujii, N.; Peiper, S. C.; Bhalla, K. Blood 2010, 116, 5306. 15. Singh, S.; Singh, U. P.; Grizzle, W. E.; Lillard, J. W. J. Lab Invest. 2004, 84, 1666. 16. Müller, A.; Homey, B.; Soto, H.; Ge, N.; Catron, D.; Buchanan, M. E.; Mcclanahan, T.; Murphy, E.; Yuan, W.; Wagner, S. N.; Barrera, J. L.; Mohar, A.; Verástegui, E.; Zlotnik, A. Nature 2001, 410, 50. 17. Küry, P.; Köller, H.; Hamacher, M.; Cornely, C.; Hasse, B.; Müller, H. W. Mol. Cell. Neurosci. 2003, 24, 1. 18. Ramos, E. A.; Grochoski, M.; Braun-Prado, K.; Seniski, G. G.; Cavalli, I. J.; Ribeiro, E. M.; Camargo, A. A.; Costa, F. F.; Klassen, G. PLoS One 2011, 6, e29461. 19. Rahimi, M.; George, J.; Tang, C. Int. J. Cancer 1850, 2010, 126. 20. Wu, B.; Chien, E. Y.; Mol, C. D.; Fenalti, G.; Liu, W.; Katritch, V.; Abagyan, R.; Brooun, A.; Wells, P.; Bi, F. C.; Hamel, D. J.; Kuhn, P.; Handel, T. M.; Cherezov, V.; Stevens, R. C. Science 2010, 330, 1066. 21. Hawle, P.; Siepmann, M.; Harst, A.; Siderius, M.; Reusch, H. P.; Obermann, W. M. J. Mol. Cell. Biol. 2006, 26, 8385. 22. Sidera, K.; Gaitanou, M.; Stellas, D.; Matsas, R.; Patsavoudi, E. J. Biol. Chem. 2008, 283, 2031. 23. Gartner, E. M.; Silverman, P.; Simon, M.; Flaherty, L.; Abrams, J.; Ivy, P.; Lorusso, P. M. Breast Cancer Res. Treat. 2012, 131, 933. 24. Zhou, H.; Tai, H. H. Arch. Biochem. Biophys. 2000, 373, 211. 25. Rodríguez, D.; Gutiérrez-De-Terán, H. Proteins 1919, 2012, 80. 26. Duda, D. G.; Kozin, S. V.; Kirkpatrick, N. D.; Xu, L.; Fukumura, D.; Jain, R. K. Clin. Cancer Res. 2011, 17, 2074. 27. Orimo, A.; Gupta, P. B.; Sgroi, D. C.; Arenzana-Seisdedos, F.; Delaunay, T.; Naeem, R.; Carey, V. J.; Richardson, A. L.; Weinberg, R. A. Dev. Cell 2005, 121, 335. 28. Zabel, B. A.; Wang, Y.; Lewén, S.; Berahovich, R. D.; Penfold, M. E.; Zhang, P.; Powers, J.; Summers, B. C.; Miao, Z.; Zhao, B.; Jalili, A.; Janowska-Wieczorek, A.; Jaen, J. C.; Schall, T. J. J. Immunol. 2009, 183, 3204. 29. Jin, D. K.; Shido, K.; Kopp, H. G.; Petit, I.; Shmelkov, S. V.; Young, L. M.; Hooper, A. T.; Amano, H.; Avecilla, S. T.; Heissig, B.; Hattori, K.; Zhang, F.; Hicklin, D. J.; Wu, Y.; Zhu, Z.; Dunn, A.; Salari, H.; Werb, Z.; Hackett, N. R.; Crystal, R. G.; Lyden, D.; Rafii, S. Nat. Med. 2006, 12, 557. 30. Dealwis, C.; Fernandez, E. J.; Thompson, D. A.; Simon, R. J.; Siani, M. A.; Lolis, E. Proc. Natl. Acad. Sci. U.S.A. 1998, 95, 6941. 31. Chinni, S. R.; Yamamoto, H.; Dong, Z.; Sabbota, A.; Bonfil, R. D.; Cher, M. L. Mol. Cancer Res. 2008, 6, 446. 32. Ho, T. K.; Shiwen, X.; Abraham, D.; Tsui, J.; Baker, D. Cardiol. Res. Pract. 2012, 2012, 143. 33. Liao, Y.; Hung, M. Am. J. Transl. Res. 2010, 2, 19. 34. Enomoto, A.; Murakami, H.; Asai, N.; Morone, N.; Watanabe, T.; Kawai, K.; Murakumo, Y.; Usukura, J.; Kaibuchi, K.; Takahashi, M. Dev. Cell 2005, 9, 389. 35. Taiyab, A.; Rao, C. M. Biochem. Biophys. Acta 2011, 1813, 213. 36. Arjonen, A.; Kaukonen, R.; Ivaska, J. Cell Adhes. Migr. 2011, 5, 421. 37. Madhusudan, S.; Foster, M.; Muthuramalingam, S. R.; Braybrooke, J. P.; Wilner, S.; Kaur, K.; Han, C.; Hoare, S.; Balkwill, F.; Talbot, D. C.; Ganesan, T. S.; Harris, A. L. Clin. Cancer Res. 2004, 10, 6528. 38. Darisipudi, M. N.; Kulkarni, O. P.; Sayyed, S. G.; Ryu, M.; Migliorini, A.; Sagrinati, C.; Parente, E.; Vater, A.; Eulberg, D.; Klussmann, S.; Romagnani, P.; Anders, H. J. Am. J. Pathol. 2011, 179, 116. 39. Sachdev, J. C.; Jahanzeb, M. Clin. Breast Cancer 2012, 12, 19. 40. Zhou, N.; Luo, Z.; Luo, J.; Fan, X.; Cayabyab, M.; Hiraoka, M.; Liu, D.; Han, X.; Pesavento, J.; Dong, C. Z.; Wang, Y.; An, J.; Kaji, H.; Sodroski, J. G.; Huang, Z. J. Biol. Chem. 2002, 277, 17476. 41. Peled, A.; Wald, O.; Burger, J. Expert Opin. Investig. Drugs 2012, 21, 341. 42. Veldkamp, C. T.; Ziarek, J. J.; Peterson, F. C.; Chen, Y.; Volkman, B. F. J. Am. Chem. Soc. 2010, 132, 7242. 43. Kuhne, M.; Mulvey, T.; Belanger, B.; Chen, S.; Pan, C.; Chong, C.; Niekro, W.; Kempe, T.; Henning, K. A.; Cohen, L.; Korman, A.; Cardarelli, P. M. Presented at: 100th American Association for Cancer Research Annual Meeting; Denver, CO., USA. 2009; 2009, Apr 18–22, Abstract LB. 44. Jähnichen, S.; Blanchetot, C.; Maussang, D.; Gonzalez-Pajuelo, M.; Chow, K. Y.; Bosch, L.; De Vrieze, S.; Serruys, B.; Ulrichts, H.; Vandevelde, W.; Saunders, M.;
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25
45.
46. 47. 48.
49.
De Haard, H. J.; Schols, D.; Leurs, R.; Vanlandschoot, P.; Verrips, T.; Smit, M. J. Proc. Natl. Acad. Sci. U.S.A. 2010, 107, 20565. Fricker, S. P.; Anastassov, V.; Cox, J.; Darkes, M. C.; Grujic, O.; Idzan, S. R.; Labrecque, J.; Lau, G.; Mosi, R. M.; Nelson, K. L.; Qin, L.; Santucci, Z.; Wong, R. S. Biochem. Pharmacol. 2006, 72, 588. Wong, R. S.; Bodart, V.; Metz, M.; Labrecque, J.; Bridger, G.; Fricker, S. P. Mol. Pharmacol. 2008, 74, 1485. Hunter, T. M.; Mcnae, I. W.; Liang, X.; Bella, J.; Parsons, S.; Walkinshaw, M. D.; Sadler, P. J. Proc. Natl. Acad. Sci. U.S.A. 2005, 102, 2288. Ross, A.; Soares, D. C.; Covelli, D.; Pannecouque, C.; Budd, L.; Collins, A.; Robertson, N.; Parsons, S.; De Clercq, E.; Kennepohl, P.; Sadler, P. J. Inorg. Chem 2010, 49, 1122. Liang, Z.; Zhan, W.; Zhu, A.; Yoon, Y.; Lin, S.; Sasaki, M.; Klapproth, J. M.; Yang, H.; Grossniklaus, H. E.; Xu, J.; Rojas, M.; Voll, R. J.; Goodman, M. M.; Arrendale,
50. 51.
52.
53. 54.
25
R. F.; Liu, J.; Yun, C. C.; Snyder, J. P.; Liotta, D. C.; Shim, H. PLoS One 2012, 7, e34038. Yadav, V. R.; Sung, B.; Prasad, S.; Kannappan, R.; Cho, S. G.; Liu, M.; Chaturvedi, M. M.; Aggarwal, B. J. Mol. Med. (Berl) 2010, 88, 1243. Tamamura, H.; Hori, A.; Kanzak, I. N.; Hiramatsu, K.; Mizumoto, M.; Nakashima, H.; Yamamoto, N.; Otaka, A.; Fujii, N. FEBS Lett. 2003, 550, 79. Cabioglu, N.; Sahin, A. A.; Morandi, P.; Meric-Bernstam, F.; Islam, R.; Lin, H. Y.; Bucana, C. D.; Gonzalez-Angulo, A. M.; Hortobagyi, G. N.; Cristofanilli, M. Ann. Oncol. 2009, 20, 1013. Cabioglu, N.; Summy, J.; Miller, C.; Parikh, N. U.; Sahin, A. A.; Tuzlali, S.; Pumiglia, K.; Gallick, G. E.; Price, J. E. Cancer Res. 2005, 65, 6493. Chudasama, V. L.; Stark, F.; Harrold, J. M.; Tibbitts, J.; Girish, S. R.; Gupta, M.; Frey, N.; Mager, D. E. Clin. Pharmacol. Ther. 2012, 92, 520.
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl
BMCL Digest
Halting metastasis through CXCR4 inhibition Deborah M. Ramsey ⇑, Shelli R. McAlpine ⇑ Department of Chemistry, University of New South Wales, Sydney NSW 2052, Australia
a r t i c l e
i n f o
Article history: Received 2 October 2012 Accepted 31 October 2012 Available online 12 November 2012 Keywords: CXCR4 SDF-1 Metastasis Cancer Heat shock protein 90
a b s t r a c t Metastasis occurs when cancer cells leave the primary tumor site and migrate to distant parts of the body. The CXCR4–SDF-1 pathway facilitates this migration, and its expression has become the hallmark of several metastatic cancers. Targeted approaches are currently being developed to inhibit this pathway, and several candidates are now in clinical trials. Continued exploration of CXCR4 inhibitors will generate compounds that have improved activity over current candidates. Ó 2012 Elsevier Ltd. All rights reserved.
Cells do not stand still. Cell migration is a primordial characteristic observed from the beginning of embryonic development, and directed cell movement continues throughout the life of an organism.1 During normal mammalian embryogenesis, embryonic stem cells (ESCs) traffic to numerous sites under the control of several chemokines, including stromal cell-derived factor-1 (SDF-1; also known as CXCL12) and its corresponding adhesive molecule CXCR4 (CD184).2,3 Adult pluripotent cells maintain a ‘homing’ capacity to specific tissues using the SDF-1/CXCR4 chemosensory system, leading to life-affirming effects such as wound repair and tissue regeneration.4–6 Cancer cells exploit the same migratory pathways utilized by embryonic and adult stem cells.7,8 Metastatic tumors express ESC transcription factors and spread beyond the primary tumor site through SDF-1/CXCR4-mediated migration.9 Expression of CXCR4 has been tied to tumor infiltration and poor prognosis in colon, breast, and gallbladder cancers.10–13 In addition, SDF-1 expression is correlated to reduced survival in ovarian cancer patients.11 A growing body of research points to the CXCR4 pathway as an emerging arena for cancer drug discovery, with the goal of specifically disrupting the receptor-ligand (CXCR4–SDF-1) interaction that drives the spread of tumors. In addition, a deeper understanding of the pathways involved in CXCR4 folding and expression points to the major role that molecular chaperones play in facilitating metastasis.14 Current therapies being tested in clinical trials are utilizing a combinatorial approach for inhibiting the spread of CXCR4-expressing cancer cells, targeting both the receptor-ligand interaction and other pro-oncogenic pathways (Table 1). ⇑ Corresponding authors. Tel.: +61 4 1672 8896; fax: +61 2 9385 6141. E-mail addresses: [email protected] (D.M. Ramsey), [email protected] (S.R. McAlpine). 0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.bmcl.2012.10.138
CXCR4 is a member of the G protein-coupled (GPC) chemokine receptor family, utilizing seven trans-membrane alpha helices to integrate into the cell membrane (Fig. 1). CXCR4 gene expression is downregulated in normal breast cells and non-invasive cancer cell lines, but invasive neuroblastoma and prostate cancer cell lines show high levels of CXCR4 transcription.15,16 Several factors may contribute to the observed increase in gene expression, such as DNA methylation or TNF signaling cascades.17,18 In addition, upregulation of the transcriptional activator HIF-1a leads to a direct increase in CXCR4 gene expression.19 Following post-translational modification, CXCR4 relies on the molecular chaperone heat shock protein 90 (Hsp90) for proper folding and delivery to the lipid bilayer (Fig. 2).14 Hsp90 is also the molecular chaperone for several members of the CXCR4/SDF-1 phosphorylation cascade, including the protein tyrosine kinase Src, human epidermal growth factor receptor 2 (HER2), and the serine/threonine kinase Akt.21,22 Inhibitors of Hsp90 have been studied in clinical trials for efficacy against metastatic cancers,14 and expression levels of several proteins in the CXCR4 pathway are reduced during treatment.23 However, toxicity issues limit the use of current Hsp90 inhibitors as a monotherapy in the clinic.23 The crystal structure of CXCR4 shows that the seven transmembrane alpha helices bundle together to form an extracellular binding pocket (Fig. 124). The extracellular loops (ECLs) and amino acids within the alpha helices contribute to binding interactions with potential ligands. Two disulfide bonds form between key cysteine residues on the extracellular face of the receptor, and these bonds are critical for shaping the binding pocket and for ligand binding: Cys28 of the N-terminal segment with Cys274 of helix VII (H7), and Cys109 with Cys186 in ECL2.20,24 CXCR4 can also homo- or heterodimerize.20,25 This ability could facilitate metastasis in cancer cells expressing high levels of CXCR4, since
21
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25 Table 1 Selected CXCR4 and SDF-1 Inhibitors in clinical triala Sponsor
Drug name
Secondary names
Combination therapy?
Phase
Cancer
MDX-1338, Medarex
Lenalidomide, Dexamethasone, or Bortezomib
IA
Multiple myeloma
Ablynx
BMS936564 ALX-0651
I
Healthy volunteers (safety study)
CXCR4 antagonists Biokine Therapeutics Ltd
BKT-140
I/IIA
Multiple myeloma
Cardiff University
Plerixafor
Daunorubicin, Clofarabine
I
Washington University School of Medicine Weill Medical College (Cornell University) Metastatix, Inc.
Plerixafor
4F-benzoyl-TN14003, BL-8040 Mozobil, Genzyme, AMD3100 Mozobil, Genzyme, AMD3100 Mozobil, Genzyme, AMD3100
Mitoxantrone, Etoposide, Cytarabine Decitabine
I/II
Acute myeloid leukemia; high risk myelodysplastic syndrome Acute myeloid leukemia
I
Acute myeloid leukemia
MSX-122
I
Refractory metastatic or locally advanced solid tumors
NOX-A12
II
Multiple myeloma
Anti-CXCR4 antibodies Bristol–Myers Squib
SDF-1 antagonists Noxxon Pharma AG a
Plerixafor
Information obtained from www.clinicaltrials.gov.
Figure 1. CXCR4 receptor structure and binding pocket. CXCR4 consists of seven trans-membrane helices (H1–7) and three extracellular loops (ECL1-3). The receptor folds into a bundle, creating a binding pocket for its ligand SDF-1.21
Figure 2. CXCR4/SDF-1 signaling cascade. CXCR4 is folded and delivered to the cell surface by Hsp90. Once the chemokine SDF-1 is bound, a phospho-relay transfers the signal to Akt, leading to actin reorganization and subsequent cell migration.
dimerization would amplify the signaling cascade upon ligand binding and lead to enhanced downstream effects. Crystal structures of CXCR4 homodimers indicate that ligand-induced confor-
mational changes to one receptor can actually increase the ligand binding affinity of a second receptor.20 The converse is also true, suggesting that the biological efficacy of CXCR4 antagonists may
22
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25
be tied to whether these modulators induce positive or negative binding cooperativity. The small chemokine SDF-1 binds CXCR4 and other cognate receptors (Fig. 1), triggering multiple signaling cascades that promote cell survival and metastasis.16,26 The source of SDF-1 can be normal tissue (including blood vascular endothelial cells and bone marrow), stromal cells within solid tumors (for paracrine signaling), or cancer cells themselves (for autocrine signaling).27–29 The crystal structure of SDF-1a consists of two monomers that form an asymmetrical dimer.30 Each monomer is composed of three beta strands that form an anti-parallel beta sheet, which is bordered on either side by an alpha helix. The N-terminal amino acids of SDF-1 are thought to be responsible for its ligand binding activity.30 Charged residues within the first alpha helix and the beta sheet, as well as an adjacent hydrophobic region within the beta sheet, may facilitate interactions with the trans-membrane helices of CXCR4. Once SDF-1 is bound, CXCR4, HER2, and possibly extracellular Hsp90, co-localize to lipid rafts, which are cell membrane clusters enriched in cholesterol.31 Not only do lipid rafts facilitate transfer of the signal into the cell interior, but recent evidence suggests that extracellular Hsp90 stabilizes HER2 and promotes this signaling event (Fig. 2).22,31 Once bound to SDF-1, activated CXCR4 increases intracellular calcium levels and induces a phosphorylation cascade.32 CXCR4 is then internalized and can be recycled back to the cell surface or degraded by lysosomal enzymes.32 Cell motility is regulated at the intracellular level by several phosphorylation cascades controlled by Src or Akt (also called protein kinase B). Akt serves as the central node in the CXCR4/SDF-1 signaling cascade (Fig. 2). Association with Hsp90 protects Akt from dephosphorylation and subsequent degradation. Akt phosphorylates the G protein Girdin and other downstream targets that modulate reorganization of actin fibers in the cell.33,34 Actin polymerization is further stabilized by Hsp90, leading to the formation of plasma membrane protrusions known as filopodia.35,36 Thus, tumor cells utilize the CXCR4/SDF-1 signaling cascade to induce actin polymerization and directed cell migration, and CXCR4-positive cancer cells take advantage of autocrine or paracrine-generated SDF-1 gradients to spread to other tissues. The strategic approach to blocking the CXCR4/SDF-1 pathway has centered largely on inhibitors of the receptor CXCR4 or the chemokine SDF-1. CXCR4 inhibitors fall into two categories: antiCXCR4 antibodies and CXCR4 antagonists (Table 1). Since CXCR4 is also the co-receptor for human immunodeficiency virus (HIV) and is required for viral entry into T cells, significant drug design advancements on CXCR4 inhibitors were made before they were utilized as anti-cancer therapies. Chemokine inhibitors are generally well tolerated by patients, and antagonists of SDF-1 are now exhibiting some success in clinical trials.37,38 In addition, targeting other extracellular proteins in the CXCR4 pathway (namely HER2) has also shown some therapeutic promise in inhibiting metastasis and could be combined in future studies with CXCR4/SDF-1 antagonists.39 Anti-CXCR4 antibodies. Amino acids within the CXCR4 binding pocket form specific interactions with SDF-1, but the receptor as a whole is quite flexible and can dislodge the binding of large molecules such as monoclonal antibodies that target its surface.40,41 As a result, anti-CXCR4 antibodies must form strong binding interactions with CXCR4 to form a physical barrier to chemokines attempting to enter the binding pocket (Fig. 3). A recent study examining CXCR4/SDF-1 interactions found that sulfonated tyrosine residues in one of the N-terminal extracellular loops of CXCR4 are absolutely critical for binding to SDF-1.42 Targeting these specific sulfonated tyrosine residues has proven an effective strategy for neutralizing the effects of chemokine-receptor interactions and avoiding the issues associated with monoclonal antibodies tar-
Figure 3. Anti-CXCR4 antibodies and nanobodies bind to the CXCR4 receptor and block SDF-1 from entering the binding pocket.
geted to flexible regions on CXCR4. The current anti-CXCR4 antibody under clinical investigation is the drug candidate MDX1338 (BMS-936564).40 MDX-1338 directly blocks the binding interaction between one of the N-terminal extracellular loops of CXCR4 and SDF-1, causing inhibition of cell migration.43 MDX1338 is potent in the low nanomolar range and reduces AML tumors in murine xenograft models.43 Clinical studies are ongoing, where MDX-1338 is used in combination with standard chemotherapeutic treatments to reduce multiple myeloma (MM; Table 1). One of the drawbacks of monoclonal antibody therapy is that multiple antibodies must be used to target multiple epitopes on a single receptor. Advancements in nanotechnology have led to the development of a unique solution: the nanobody. Nanobodies are antibody fragments with a single monomeric variable domain (monovalent), or two different variable domains held together by a flexible linker (biparatopic).44 Only four nanobodies are currently being studied in clinical trials, and one of those nanobodies (ALX0651) targets CXCR4. Recent data suggests that monovalent CXCR4 nanobodies act as neutral antagonists, which compete with SDF-1 for binding but fail to neutralize multiple CXCR4 receptors at once.44 On the other hand, biparatopic CXCR4 nanobodies are designed to act as inverse agonists, not only by blocking SDF-1, but also by inhibiting multiple CXCR4 receptors through the bivalent construction of the molecule (Fig. 3). This cumulative effect counteracts overexpression and homodimerization of CXCR4 receptors on cancer cell surfaces and leads to downstream effects such as inhibition of cell migration.44 ALX-0651 is a biparatopic nanobody currently in Phase I clinical trials (Table 1), where there is evidence suggesting that this molecule significantly reduces cell migration. Although clinical studies using ALX-0651 are just beginning, there is considerable promise for the use of this new technology in the development of novel anti-metastatic agents. CXCR4 antagonists. A well-known small molecule inhibitor of CXCR4 is AMD3100 (xylyl–bicyclam), which is currently in Phase I/II clinical trials (Fig. 4). AMD3100 was initially studied for its antiviral properties, and the observation that this molecule induced stem cell mobilization was a surprising side effect that led to the examination of its anti-cancer activity.41 The ability of AMD3100 to act as a stem cell mobilizer has been frequently exploited in the clinic, although its mechanism of action is under debate.6 Hematopoetic stem cells (HSCs) in the bone marrow maintain the balance between proliferation and differentiation, replenishing both blood and immune cells in the bloodstream to keep the system at a steady state. CXCR4-positive HSCs are retained in the bone
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25
23
Figure 4. Small molecule antagonists of CXCR4.
marrow by stromal cells producing SDF-1 gradients (paracrine signaling), and SDF-1 binds to CXCR4 and prevents HSC migration outside of the niche. Clinicians can manipulate this storehouse of stem cells for a variety of purposes, including tissue regeneration, or by inducing cancerous stem cells to leave the marrow during chemotherapy. For HSCs to leave the bone marrow, it is hypothesized that CXCR4 antagonists such as AMD3100 work by binding to CXCR4 and blocking the binding of endogenous SDF-1.45 Without the receptor’s ability to respond to its retention signal (i.e. SDF-1 in the bone marrow niche), CXCR4-positive HSCs will leave the marrow and enter the bloodstream. AMD3100 is a metal-chelating, bicyclic, reversible inhibitor that binds in a pocket between the transmembrane helices and extracellular loop (ECL) 2 of CXCR4, leading to allosteric modulation and effective blockade of SDF-1 binding (Fig. 5).46 This antagonist is specific for the CXCR4 receptor and does not cross-react with other GPC-receptors. AMD3100’s selectivity may be due in part to the key hydrophobic contacts formed between the bicyclam rings and two tryptophan residues within the binding pocket of CXCR4, Trp 195 and Trp283.47 Several Phase I/II clinical trials are utilizing this molecule in a combinatorial therapeutic approach to treat acute myeloid leukemia (AML)45. Interestingly, the affin-
ity of these bicyclam complexes for CXCR4 is enhanced when the macrocycle binds to copper, nickel or zinc ions.47,48 Further research is underway to study how metal complexes can strengthen the CXCR4 receptor binding interactions with bicyclam macrocycles. New developments in small molecule inhibitors of CXCR4 have led to some interesting leads that exhibit anti-metastatic properties. MSX-122, currently in Phase I clinical trials, is a small molecule antagonist of CXCR4 that inhibits metastasis of breast, melanoma, and head and neck carcinoma in murine models (Fig. 4).49 There have also been recent leads in the development of small molecules that target sites on CXCR4 other than its binding pocket. One such small molecule inhibitor is the natural product celastrol (Fig. 4), which prevents cell migration by inhibiting translation of CXCR4 mRNA.50 Other compounds have targeted a post-translational modification of CXCR4, where a recent in silico screen identified inhibitors that bound to a sulfotyrosine recognition site within one of the N-terminal extracellular loops of CXCR4.43 One inhibitor identified in this screen reduced SDF-1mediated calcium flux in treated THP-1 cells.42 Peptide inhibitors of CXCR4 are beginning to show promise in the clinic. BKT140 is an analog of T140 (sequence of T140: H-
Figure 5. CXCR4 antagonists compete for binding to the receptor binding pocket. The binding pocket (BP) of CXCR4 is the site for SDF-1 binding, as well as the target of CXCR4 antagonists.
24
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25
Arg-Arg-Nal-Cys-Tyr-Arg-Lys-D-Lys-Pro-Tyr-Arg-Cit-Cys-Arg-OH), a 14-residue peptide that is a potent inverse agonist and CXCR4 antagonist and inhibits metastatic breast cancer in murine models.51 With its potency in the low nanomolar range, BKT140 is now being studied in Phase I/II clinical trials against MM (Table 1). Other peptide analogs have used virally-derived CXCR4 antagonists as the basis for lead compound development. Using 21 amino acids from the human herpesvirus 8 viral macrophage inflammatory protein II (vMIP-II), recent research indicates that D-amino acid analogs of this peptide bind CXCR4 more strongly than the original peptide and serve as better antagonists.40 These discoveries will contribute significantly to the design of new peptide inhibitors. SDF-1 antagonists. SDF-1 antagonists have been studied for their efficacy in both cancer and diabetes research, and promising results have been observed.38 The common SDF-1 antagonist is a 45 base pair L-RNA oligonucleotide (or Spiegelmer) known as NOX-A12.38 This aptamer is currently in Phase II clinical trials as a monotherapy against MM (Table 1). One question that remains to be explored in the clinic is whether CXCR4 antagonism alone can halt metastasis, or whether SDF-1 antagonism is also required for full therapeutic impact. Future clinical studies that combine CXCR4 and SDF-1 antagonists may determine if there is an additive benefit, or if one antagonist is better at inhibiting metastatic disease and preventing relapse. Inhibitors of other proteins in the CXCR4 pathway. The CXCR4/ SDF-1 pathway plays a major role in metastatic breast cancer, where the overexpression of CXCR4 is one of several biomarkers (including HER2/neu) used to diagnose aggressive disease.52 Thus, inhibitors of other proteins in the CXCR4 pathway may be required for optimal therapeutic effect. The role of CXCR4/SDF-1 pathway in metastatic breast cancer has become clearer in recent years. SDF-1 can induce the transactivation of HER2/neu in breast cancer cells,53 and CXCR4 can facilitate homing of metastatic breast cancer cells to the bone.52 More research is needed in metastatic breast cancer clinical trials to determine the efficacy of CXCR4/SDF-1 antagonists in combination with other chemotherapeutic agents. Anti-HER2 monoclonal antibodies, either alone or conjugated to other anticancer drugs (such as Trastuzumab emtansine or T-DM1), may be needed in addition to CXCR4/SDF-1 antagonists for successful treatment of metastatic breast cancer.54 The GPC-receptor CXCR4 and its chemokine SDF-1 play a critical role in cell migration and metastatic disease. Benefiting from early anti-viral research, the development of CXCR4 inhibitors has led to several successful drug candidates that are currently undergoing testing in Phase I/II clinical trials. Future development of carefully designed agonists and antagonists, as well as mining structures from natural product sources, will expand the number of new anti-cancer agents with the potential to halt CXCR4-mediated cell migration. The success of these compounds may require a combination of therapies targeted at multiple proteins involved in this metastatic pathway. ⁄ Dr. Deborah M. Ramsey and Associate Professor Shelli R. McAlpine, School of Chemistry, University of New South Wales, Sydney NSW 2052, Australia. Tel.: +61 2 9385 5505. Email: [email protected] and [email protected]. Acknowledgments We thank the University of New South Wales (UNSW), as well as NIH 1R01CA137873 for support of D.M.R. and S.R.M. References and notes 1. Hunger, C.; Odemis, V.; Engele, J. Exp. Cell Res. 2012, 318, 2178. 2. Cha, Y. R.; Fujita, M.; Butler, M.; Isogai, S.; Kochhan, E.; Siekmann, A. F.; Weinstein, B. M. Dev. Cell 2012, 22, 824.
3. Ciriza, J.; Hall, D.; Lu, A.; De Sena, J. R.; Al-Kuhlani, M.; García-Ojeda, M. E. PLoS One 2012, 7, e30542. 4. Ding, J.; Hori, K.; Zhang, R.; Marcoux, Y.; Honardoust, D.; Shankowsky, H. A.; Tredget, E. E. Wound Repair Regen. 2011, 19, 568. 5. Xia, W. H.; Li, J.; Su, C.; Yang, Z.; Chen, L.; Wu, F.; Zhang, Y. Y.; Yu, B. B.; Qiu, Y. X.; Wang, S. M.; Tao, J. Aging Cell 2012, 11, 111. 6. Brzoska, E.; Kowalewska, M.; Markowska, A.; Kowalski, K.; Archacka, K.; Zimowska, M.; Grabowska, I.; Czerwin´ska, A. M.; Góra, M.; Stremin´ska, W.; Jan´czyk-Ilach, K.; Ciemerych, M. A. Biol. Cell. 2012, Sept 14, Accepted for publication (Epub ahead of print). 7. Kleeberger, W.; Bova, G. S.; Nielsen, M. E.; Herawi, M.; Chuang, A. Y.; Epstein, J. I.; Berman, D. M. Cancer Res. 2007, 67, 9199. 8. Nian, W. Q.; Chen, F. L.; Ao, X. J.; Chen, Z. T. Mol. Cell. Biochem. 2011, 355, 241. 9. Belton, A.; Gabrovsky, A.; Bae, Y. K.; Reeves, R.; Iacobuzio-Donahue, C.; Huso, D. L.; Resar, L. M. PLoS one 2012, 7, e30034. 10. Zhang, N. H.; Li, J.; Li, Y.; Zhang, X. T.; Liao, W. T.; Zhang, J. Y.; Li, R.; Luo, R. C. Exp. Ther. Med. 2012, 3, 973. 11. Popple, A.; Durrant, L. G.; Spendlove, I.; Rolland, P.; Scott, I. V.; Deen, S.; Ramage, J. M. Br. J. Cancer 2012, 106, 1306. 12. Yao, X.; Zhou, L.; Han, S.; Chen, Y. J. Int. Med. Res. 2011, 39, 1253. 13. Hiller, D. J.; Meschonat, C.; Kim, R.; Li, B. D.; Chu, Q. D. Sugery 2011, 150, 459. 14. Mandawat, A.; Fiskus, W.; Buckley, K. M.; Robbins, K.; Rao, R.; Balusu, R.; Navenot, J.; Wang, Z.; Ustun, C.; Chong, D. G.; Atadja, P.; Fujii, N.; Peiper, S. C.; Bhalla, K. Blood 2010, 116, 5306. 15. Singh, S.; Singh, U. P.; Grizzle, W. E.; Lillard, J. W. J. Lab Invest. 2004, 84, 1666. 16. Müller, A.; Homey, B.; Soto, H.; Ge, N.; Catron, D.; Buchanan, M. E.; Mcclanahan, T.; Murphy, E.; Yuan, W.; Wagner, S. N.; Barrera, J. L.; Mohar, A.; Verástegui, E.; Zlotnik, A. Nature 2001, 410, 50. 17. Küry, P.; Köller, H.; Hamacher, M.; Cornely, C.; Hasse, B.; Müller, H. W. Mol. Cell. Neurosci. 2003, 24, 1. 18. Ramos, E. A.; Grochoski, M.; Braun-Prado, K.; Seniski, G. G.; Cavalli, I. J.; Ribeiro, E. M.; Camargo, A. A.; Costa, F. F.; Klassen, G. PLoS One 2011, 6, e29461. 19. Rahimi, M.; George, J.; Tang, C. Int. J. Cancer 1850, 2010, 126. 20. Wu, B.; Chien, E. Y.; Mol, C. D.; Fenalti, G.; Liu, W.; Katritch, V.; Abagyan, R.; Brooun, A.; Wells, P.; Bi, F. C.; Hamel, D. J.; Kuhn, P.; Handel, T. M.; Cherezov, V.; Stevens, R. C. Science 2010, 330, 1066. 21. Hawle, P.; Siepmann, M.; Harst, A.; Siderius, M.; Reusch, H. P.; Obermann, W. M. J. Mol. Cell. Biol. 2006, 26, 8385. 22. Sidera, K.; Gaitanou, M.; Stellas, D.; Matsas, R.; Patsavoudi, E. J. Biol. Chem. 2008, 283, 2031. 23. Gartner, E. M.; Silverman, P.; Simon, M.; Flaherty, L.; Abrams, J.; Ivy, P.; Lorusso, P. M. Breast Cancer Res. Treat. 2012, 131, 933. 24. Zhou, H.; Tai, H. H. Arch. Biochem. Biophys. 2000, 373, 211. 25. Rodríguez, D.; Gutiérrez-De-Terán, H. Proteins 1919, 2012, 80. 26. Duda, D. G.; Kozin, S. V.; Kirkpatrick, N. D.; Xu, L.; Fukumura, D.; Jain, R. K. Clin. Cancer Res. 2011, 17, 2074. 27. Orimo, A.; Gupta, P. B.; Sgroi, D. C.; Arenzana-Seisdedos, F.; Delaunay, T.; Naeem, R.; Carey, V. J.; Richardson, A. L.; Weinberg, R. A. Dev. Cell 2005, 121, 335. 28. Zabel, B. A.; Wang, Y.; Lewén, S.; Berahovich, R. D.; Penfold, M. E.; Zhang, P.; Powers, J.; Summers, B. C.; Miao, Z.; Zhao, B.; Jalili, A.; Janowska-Wieczorek, A.; Jaen, J. C.; Schall, T. J. J. Immunol. 2009, 183, 3204. 29. Jin, D. K.; Shido, K.; Kopp, H. G.; Petit, I.; Shmelkov, S. V.; Young, L. M.; Hooper, A. T.; Amano, H.; Avecilla, S. T.; Heissig, B.; Hattori, K.; Zhang, F.; Hicklin, D. J.; Wu, Y.; Zhu, Z.; Dunn, A.; Salari, H.; Werb, Z.; Hackett, N. R.; Crystal, R. G.; Lyden, D.; Rafii, S. Nat. Med. 2006, 12, 557. 30. Dealwis, C.; Fernandez, E. J.; Thompson, D. A.; Simon, R. J.; Siani, M. A.; Lolis, E. Proc. Natl. Acad. Sci. U.S.A. 1998, 95, 6941. 31. Chinni, S. R.; Yamamoto, H.; Dong, Z.; Sabbota, A.; Bonfil, R. D.; Cher, M. L. Mol. Cancer Res. 2008, 6, 446. 32. Ho, T. K.; Shiwen, X.; Abraham, D.; Tsui, J.; Baker, D. Cardiol. Res. Pract. 2012, 2012, 143. 33. Liao, Y.; Hung, M. Am. J. Transl. Res. 2010, 2, 19. 34. Enomoto, A.; Murakami, H.; Asai, N.; Morone, N.; Watanabe, T.; Kawai, K.; Murakumo, Y.; Usukura, J.; Kaibuchi, K.; Takahashi, M. Dev. Cell 2005, 9, 389. 35. Taiyab, A.; Rao, C. M. Biochem. Biophys. Acta 2011, 1813, 213. 36. Arjonen, A.; Kaukonen, R.; Ivaska, J. Cell Adhes. Migr. 2011, 5, 421. 37. Madhusudan, S.; Foster, M.; Muthuramalingam, S. R.; Braybrooke, J. P.; Wilner, S.; Kaur, K.; Han, C.; Hoare, S.; Balkwill, F.; Talbot, D. C.; Ganesan, T. S.; Harris, A. L. Clin. Cancer Res. 2004, 10, 6528. 38. Darisipudi, M. N.; Kulkarni, O. P.; Sayyed, S. G.; Ryu, M.; Migliorini, A.; Sagrinati, C.; Parente, E.; Vater, A.; Eulberg, D.; Klussmann, S.; Romagnani, P.; Anders, H. J. Am. J. Pathol. 2011, 179, 116. 39. Sachdev, J. C.; Jahanzeb, M. Clin. Breast Cancer 2012, 12, 19. 40. Zhou, N.; Luo, Z.; Luo, J.; Fan, X.; Cayabyab, M.; Hiraoka, M.; Liu, D.; Han, X.; Pesavento, J.; Dong, C. Z.; Wang, Y.; An, J.; Kaji, H.; Sodroski, J. G.; Huang, Z. J. Biol. Chem. 2002, 277, 17476. 41. Peled, A.; Wald, O.; Burger, J. Expert Opin. Investig. Drugs 2012, 21, 341. 42. Veldkamp, C. T.; Ziarek, J. J.; Peterson, F. C.; Chen, Y.; Volkman, B. F. J. Am. Chem. Soc. 2010, 132, 7242. 43. Kuhne, M.; Mulvey, T.; Belanger, B.; Chen, S.; Pan, C.; Chong, C.; Niekro, W.; Kempe, T.; Henning, K. A.; Cohen, L.; Korman, A.; Cardarelli, P. M. Presented at: 100th American Association for Cancer Research Annual Meeting; Denver, CO., USA. 2009; 2009, Apr 18–22, Abstract LB. 44. Jähnichen, S.; Blanchetot, C.; Maussang, D.; Gonzalez-Pajuelo, M.; Chow, K. Y.; Bosch, L.; De Vrieze, S.; Serruys, B.; Ulrichts, H.; Vandevelde, W.; Saunders, M.;
D. M. Ramsey, S. R. McAlpine / Bioorg. Med. Chem. Lett. 23 (2013) 20–25
45.
46. 47. 48.
49.
De Haard, H. J.; Schols, D.; Leurs, R.; Vanlandschoot, P.; Verrips, T.; Smit, M. J. Proc. Natl. Acad. Sci. U.S.A. 2010, 107, 20565. Fricker, S. P.; Anastassov, V.; Cox, J.; Darkes, M. C.; Grujic, O.; Idzan, S. R.; Labrecque, J.; Lau, G.; Mosi, R. M.; Nelson, K. L.; Qin, L.; Santucci, Z.; Wong, R. S. Biochem. Pharmacol. 2006, 72, 588. Wong, R. S.; Bodart, V.; Metz, M.; Labrecque, J.; Bridger, G.; Fricker, S. P. Mol. Pharmacol. 2008, 74, 1485. Hunter, T. M.; Mcnae, I. W.; Liang, X.; Bella, J.; Parsons, S.; Walkinshaw, M. D.; Sadler, P. J. Proc. Natl. Acad. Sci. U.S.A. 2005, 102, 2288. Ross, A.; Soares, D. C.; Covelli, D.; Pannecouque, C.; Budd, L.; Collins, A.; Robertson, N.; Parsons, S.; De Clercq, E.; Kennepohl, P.; Sadler, P. J. Inorg. Chem 2010, 49, 1122. Liang, Z.; Zhan, W.; Zhu, A.; Yoon, Y.; Lin, S.; Sasaki, M.; Klapproth, J. M.; Yang, H.; Grossniklaus, H. E.; Xu, J.; Rojas, M.; Voll, R. J.; Goodman, M. M.; Arrendale,
50. 51.
52.
53. 54.
25
R. F.; Liu, J.; Yun, C. C.; Snyder, J. P.; Liotta, D. C.; Shim, H. PLoS One 2012, 7, e34038. Yadav, V. R.; Sung, B.; Prasad, S.; Kannappan, R.; Cho, S. G.; Liu, M.; Chaturvedi, M. M.; Aggarwal, B. J. Mol. Med. (Berl) 2010, 88, 1243. Tamamura, H.; Hori, A.; Kanzak, I. N.; Hiramatsu, K.; Mizumoto, M.; Nakashima, H.; Yamamoto, N.; Otaka, A.; Fujii, N. FEBS Lett. 2003, 550, 79. Cabioglu, N.; Sahin, A. A.; Morandi, P.; Meric-Bernstam, F.; Islam, R.; Lin, H. Y.; Bucana, C. D.; Gonzalez-Angulo, A. M.; Hortobagyi, G. N.; Cristofanilli, M. Ann. Oncol. 2009, 20, 1013. Cabioglu, N.; Summy, J.; Miller, C.; Parikh, N. U.; Sahin, A. A.; Tuzlali, S.; Pumiglia, K.; Gallick, G. E.; Price, J. E. Cancer Res. 2005, 65, 6493. Chudasama, V. L.; Stark, F.; Harrold, J. M.; Tibbitts, J.; Girish, S. R.; Gupta, M.; Frey, N.; Mager, D. E. Clin. Pharmacol. Ther. 2012, 92, 520.