Identification of a New Functional Domain in Angiopoietin-like 3 (ANGPTL3) and Angiopoietin-like 4 (ANGPTL4) Involved in Binding and Inhibition of Lipoprotein Lipase (LPL)

Angiopoietin-like 3 (ANGPTL3) and angiopoietin-like 4 (ANGPTL4) are secreted proteins that regulate triglyceride (TG) metabolism in part by inhibiting lipoprotein lipase (LPL). Recently, we showed that treatment of wild-type mice with monoclonal antibody (mAb) 14D12, specific for ANGPTL4, recapitulated the Angptl4 knock-out (-/-) mouse phenotype of reduced serum TG levels. In the present study, we mapped the region of mouse ANGPTL4 recognized by mAb 14D12 to amino acids Gln29–His53, which we designate as specific epitope 1 (SE1). The 14D12 mAb prevented binding of ANGPTL4 with LPL, consistent with its ability to neutralize the LPL-inhibitory activity of ANGPTL4. Alignment of all angiopoietin family members revealed that a sequence similar to ANGPTL4 SE1 was present only in ANGPTL3, corresponding to amino acids Glu32–His55. We produced a mouse mAb against this SE1-like region in ANGPTL3. This mAb, designated 5.50.3, inhibited the binding of ANGPTL3 to LPL and neutralized ANGPTL3-mediated inhibition of LPL activity in vitro. Treatment of wild-type as well as hyperlipidemic mice with mAb 5.50.3 resulted in reduced serum TG levels, recapitulating the lipid phenotype found in Angptl3-/- mice. These results show that the SE1 region of ANGPTL3 and ANGPTL4 functions as a domain important for binding LPL and inhibiting its activity in vitro and in vivo. Moreover, these results demonstrate that therapeutic antibodies that neutralize ANGPTL4 and ANGPTL3 may be useful for treatment of some forms of hyperlipidemia. Angiopoietin-like 3 (ANGPTL3) and angiopoietin-like 4 (ANGPTL4) are secreted proteins that regulate triglyceride (TG) metabolism in part by inhibiting lipoprotein lipase (LPL). Recently, we showed that treatment of wild-type mice with monoclonal antibody (mAb) 14D12, specific for ANGPTL4, recapitulated the Angptl4 knock-out (-/-) mouse phenotype of reduced serum TG levels. In the present study, we mapped the region of mouse ANGPTL4 recognized by mAb 14D12 to amino acids Gln29–His53, which we designate as specific epitope 1 (SE1). The 14D12 mAb prevented binding of ANGPTL4 with LPL, consistent with its ability to neutralize the LPL-inhibitory activity of ANGPTL4. Alignment of all angiopoietin family members revealed that a sequence similar to ANGPTL4 SE1 was present only in ANGPTL3, corresponding to amino acids Glu32–His55. We produced a mouse mAb against this SE1-like region in ANGPTL3. This mAb, designated 5.50.3, inhibited the binding of ANGPTL3 to LPL and neutralized ANGPTL3-mediated inhibition of LPL activity in vitro. Treatment of wild-type as well as hyperlipidemic mice with mAb 5.50.3 resulted in reduced serum TG levels, recapitulating the lipid phenotype found in Angptl3-/- mice. These results show that the SE1 region of ANGPTL3 and ANGPTL4 functions as a domain important for binding LPL and inhibiting its activity in vitro and in vivo. Moreover, these results demonstrate that therapeutic antibodies that neutralize ANGPTL4 and ANGPTL3 may be useful for treatment of some forms of hyperlipidemia. Lipoprotein lipase (LPL) 5The abbreviations used are: LPL, lipoprotein lipase; ANGPTL3, angiopoietin-like 3; ANGPTL4, angiopoietin-like 4; hANGPTL3, recombinant human angiopoietin-like 3 protein (full-length); N′-hANGPTL3, recombinant human angiopoietin-like 3 amino acids Asp20–Pro243; N′-mANGPTL4, mouse ANGPTL4 amino acids Gly25–Leu185; KLH, keyhole limpet hemocyanin; mAb KLH, isotype-matched control mAb against KLH; ORF, open reading frame; LDLr, low density lipoprotein receptor; ApoE, apolipoprotein E; -/-, homozygous null; +/-, heterozygous null; +/+, wild-type littermate; Angptl4K/K, homozygous for the Angptl4 E40K variant allele; Angptl4E/K, heterozygous for the Angptl4 E40K variant allele; Angptl4E/E, Angptl4 wild-type littermate from parents heterozygous for Angptl4 E40K variant allele; TG, triglyceride; mAb, monoclonal antibody; SE1, specific epitope 1; BSA, bovine serum albumin; Ni-NTA, nickel-nitrilotriacetic acid; ELISA, enzyme-linked immunosorbent assay; HRP, horseradish peroxidase; DGGR, 1,2-O-dilauryl-rac-glycero-3-glutaric acid-(6′-methylresorufin) ester. plays a pivotal role in lipid metabolism by catalyzing the hydrolysis of plasma triglycerides (TGs). LPL is likely to be regulated by mechanisms that depend on nutritional status and on the tissue in which it is expressed (1Goldberg I.J. Merkel M. Front. Biosci. 2001; 6: D388-D405Crossref PubMed Google Scholar, 2Merkel M. Eckel R.H. Goldberg I.J. J. Lipid Res. 2002; 43: 1997-2006Abstract Full Text Full Text PDF PubMed Scopus (455) Google Scholar, 3Preiss-Landl K. Zimmermann R. Hammerle G. Zechner R. Curr. Opin. Lipidol. 2002; 13: 471-481Crossref PubMed Scopus (198) Google Scholar). Two secreted proteins, angiopoietin-like 3 (ANGPTL3) and angiopoietin-like 4 (ANGPTL4), play important roles in the regulation of LPL activity (4Koishi R. Ando Y. Ono M. Shimamura M. Yasumo H. Fujiwara T. Horikoshi H. Furukawa H. Nat. Genet. 2002; 30: 151-157Crossref PubMed Scopus (331) Google Scholar, 5Koster A. Chao Y.B. Mosior M. Ford A. Gonzalez-DeWhitt P.A. Hale J.E. Li D. Qiu Y. Fraser C.C. Yang D.D. Heuer J.G. Jaskunas S.R. Eacho P. Endocrinology. 2005; 146: 4943-4950Crossref PubMed Scopus (354) Google Scholar). ANGPTL3 and ANGPTL4 consist of a signal peptide, an N-terminal segment containing coiled-coil domains, and a C-terminal fibrinogen-like domain. The N-terminal segment as well as full-length ANGPTL3 and ANGPTL4 have been shown to inhibit LPL activity, and deletion of the N-terminal segment of ANGPTL3 and ANGPTL4 resulted in total loss of LPL-inhibiting activity (6Ono M. Shimizugawa T. Shimamura M. Yoshida K. Noji-Sakikawa C. Ando Y. Koishi R. Furukawa H. J. Biol. Chem. 2003; 278: 41804-41809Abstract Full Text Full Text PDF PubMed Scopus (172) Google Scholar, 7Ge H. Cha J.Y. Gopal H. Harp C. Yu X. Repa J.J. Li C. J. Lipid Res. 2005; 46: 1484-1490Abstract Full Text Full Text PDF PubMed Scopus (121) Google Scholar). These observations clearly indicate that the N-terminal region of ANGPTL4 contains the functional domain that inhibits LPL and affects plasma lipid levels. The coiled-coil domains have been proposed to be responsible for oligomerization (8Ge H. Yang G. Yu X. Pourbahrami T. Li C. J. Lipid Res. 2004; 45: 2071-2079Abstract Full Text Full Text PDF PubMed Scopus (91) Google Scholar); however, it is not known whether the coiled-coil domains directly mediate the inhibition of LPL activity. To define the physiological role of ANGPTL4 more clearly, we characterized the pharmacological consequences of ANGPTL4 inhibition in mice treated with the ANGPTL4-neutralizing monoclonal antibody (mAb) 14D12 (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). Injection of mAb 14D12 significantly lowered fasting TG levels in C57BL/6J mice relative to levels in C57BL/6J mice treated with an isotype-matched anti-KLH control (KLH) mAb (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). These reduced TG values similar to in fasting plasma TG levels in Angptl4 knock-out (-/-) mice. This that mAb 14D12 is a ANGPTL4-neutralizing antibody that is to inhibit ANGPTL4 activity and the reduced lipid phenotype found in mice. The pharmacological of mAb 14D12 the epitope recognized by mAb 14D12 and this binding ANGPTL4 as an LPL ANGPTL4 is to directly with LPL A. T. G. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google it is not which amino acids ANGPTL4 mediate this we show that amino acids of the epitope for mAb This designated specific epitope 1 a domain that the ANGPTL4 and LPL and the of this we present that ANGPTL3 contains an SE1 and with antibodies with ANGPTL3 SE1 we whether the ANGPTL3 SE1 region is in LPL binding and We whether treatment of mice with an SE1 mAb the phenotype of serum TG and levels found in Angptl3-/- mice. we the therapeutic of an SE1 mAb for treatment of in apolipoprotein low density lipoprotein mice. bovine serum and from and from 1,2-O-dilauryl-rac-glycero-3-glutaric acid-(6′-methylresorufin) was from the of was from by the of from was from was from of Angptl3-/- and for the Angptl4 E40K Angptl3-/- mouse was by and Angptl4 knock-out mice produced in and to the of secreted and with a with the S. M. M. PubMed Scopus (91) Google as shown in and mice The with Angptl3-/- mice to knock-out mice. mice with Angptl3-/- mice to mice. The of mice been (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). The for mice is shown in and is The for Angptl4 in mice is and in with and in with and and the of on for the and of The Scholar). on a and to and mice and mice on mice and mice a mice on of Lipid for lipid from from the TG levels with a serum with a TG levels with a serum with a and of mice by with protein and with protein with a containing SE1 of ANGPTL3 acids to for in for in to the serum by from the mice and with as a The the and tissue 1 was and for The was as of and was by on the protein and ANGPTL3 by and the for in and mAb for and mAb as (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). The 14D12 and anti-KLH have been (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). epitope for in vitro and of ANGPTL4 proteins by a corresponding to a of was with and This the the C-terminal of the open reading epitope of mouse ANGPTL4 amino acids the of of mouse ANGPTL4 that and The from mouse ANGPTL4 with containing the of the the of the The for in vitro and by the an Angptl4 and To the epitope of mAb 14D12, a of ANGPTL4 proteins amino acids The mouse ANGPTL4 amino acids that by amino acids was with the containing the sequence The for in vitro and by as proteins containing the mouse ANGPTL4 by in vitro and to the The of in vitro proteins was by with antibody was by of the proteins with mAb was used to the epitope of with ANGPTL3 not and of ANGPTL3 and the of human ANGPTL3 with an C-terminal epitope was with the in to the of recombinant a density of 1 and for in The by for and the containing secreted was and of from was as (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). human LPL with an C-terminal epitope was and and as LPL was by M. G. T. J. PubMed Scopus Google Scholar). 4 The was and with to a of and with to a of The was to a in of a of 1 The was with of and of The was with 1 the in The for LPL activity with the as and the and and of N-terminal of ANGPTL3 and human ANGPTL3 amino acids and human ANGPTL4 amino acids mouse ANGPTL4 amino acids was was by to a of 1 the an reading of for by for 4 and the proteins with to the on 4 and in of 1 was to the a of 1 and the was on for The was and for The was and a The was to 1 of and with for 1 4 The was by and the was The was in of and to a The was and the was with of The was with and in 1 The protein was from the with in by and the containing the recombinant protein against and was by of for ANGPTL3 SE1 and a as of LPL with activity was in vitro with the to a of the of and M. R. 2001; PubMed Scopus Google Scholar). was by in to and The was and a This is for containing LPL in LPL with the and by of to of was for with a with a and a The of was a and is expressed as the in relative of of ANGPTL4 with 4 recombinant human LPL was by to the of a with of in the was and the for 1 with of The with of The with of containing LPL 4 LPL with mAb, mAb, anti-KLH 1 the with in was to well and for The was and well was as was to well and was to LPL was by the with a LPL was by the LPL to from LPL to of ANGPTL3 with 4 of a with of in the was and the for 1 with of The with of The with of containing LPL LPL with mAb, mAb, anti-KLH 1 of antibodies to well and for LPL and antibodies from the by with was to well and was to LPL was by the with a LPL was by the LPL to from LPL to and and values by with a for are as the sequence the N-terminal domains of ANGPTL4 and ANGPTL3 by J. Res. PubMed Scopus Google and by by of by a was of a in ANGPTL4 in LPL a monoclonal 14D12, that with ANGPTL4 and its LPL-inhibiting activity in vitro and in (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). We that by the epitope in ANGPTL4 that with this we a similar region ANGPTL3 and the by which these proteins inhibit this we expressed a of amino acids as proteins and of these proteins with mAb results showed that mAb 14D12 only to the protein containing amino acids We the epitope for mAb 14D12 by of with protein and a of these proteins with mAb 14D12 This amino acids to the epitope amino acids with a of amino We found that mAb 14D12 only with amino acids and with amino acids Moreover, we found that mAb 14D12 not with the that amino acids and are in binding with mAb In antibody binding to the amino acids from to the binding of mAb 14D12 was for amino acids for amino acids These results indicate that the epitope for mAb 14D12 amino acids We define this region as ANGPTL4 ANGPTL4 been shown to directly with LPL A. T. G. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar); however, the domains in this have not been To whether mAb 14D12 is to inhibit binding of ANGPTL4 with LPL, we an binding this LPL with 14D12 to a 4 the and with to the of LPL to of LPL to was in the of mAb 14D12 in the of control mAb that to the of ANGPTL4 in C-terminal to the SE1 domain of these reduced serum in more relative to the control not The mAb showed a binding to that was from all was mAb 14D12 serum TG levels in serum mAb levels in and for in vitro and was to inhibit the binding of LPL to in vitro These results that the SE1 region of ANGPTL4 is with binding and inhibition of with mouse ANGPTL4 of for with a of corresponding to mouse ANGPTL4 amino acids and The to in a The ANGPTL4 E40K variant is with TG levels in S. Y. A. Nat. Genet. 2007; PubMed Scopus Google and the of the SE1 domain in and mice. To whether the E40K variant is with TG levels in mice as well as we mice only this of mice heterozygous for the Angptl4 E40K produced levels of the ANGPTL4 present in the and of wild-type and and sequence revealed that the E40K variant was the only ANGPTL4 present in mice not serum TG levels in Angptl4K/K, Angptl4E/K, and mice on a we found these levels to be in and mice in The of of the E40K on serum TG levels was similar to the in Angptl4 heterozygous and mice ANGPTL3 an SE1 epitope that the SE1 region of ANGPTL4 likely a region for inhibiting LPL activity. To whether angiopoietin angiopoietin-like family members this we the protein with amino acids the fibrinogen-like the J. Res. PubMed Scopus Google Scholar). The only angiopoietin angiopoietin-like protein ANGPTL4 with a was ANGPTL3. results the was with amino acids Moreover, results with mouse human ANGPTL3 as the Alignment of the N-terminal of mouse and human ANGPTL4 and ANGPTL3 protein are shown in the N-terminal of ANGPTL3 and ANGPTL4 are similar the SE1 of ANGPTL4 and its corresponding sequence in ANGPTL3 are and ANGPTL3 to an SE1 segment and the domain of ANGPTL4 and ANGPTL3 are we that the SE1 region in ANGPTL3 likely with LPL and inhibits its activity by a similar to that of ANGPTL4 A. T. G. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). this we produced a monoclonal 5.50.3, which was with the SE1 region mouse and human ANGPTL3 and for the SE1 of LPL by ANGPTL3 the SE1 To whether the SE1 region of ANGPTL3 is in inhibition of LPL, we recombinant human LPL with in the of of mAb 5.50.3 control mAb and the for LPL activity. In the of mAb, inhibited LPL by mAb as as on the LPL-inhibiting activity of ANGPTL3. mAb 5.50.3 prevented inhibition of LPL by with an of and an of We mAb 5.50.3 with and that C-terminal to and of the SE1 region for ability to neutralize ANGPTL3-mediated inhibition of In this LPL was with in the of mAb 5.50.3, mAb mAb control mAb and for LPL activity. mAb 5.50.3 neutralized inhibition of LPL by hANGPTL3, mAb mAb control mAb on the LPL-inhibiting activity of These results show that mAb 5.50.3 is of the LPL-inhibiting activity of ANGPTL3 in vitro and that the SE1 region is for inhibition of LPL by of with mouse ANGPTL3 of for with corresponding to mouse ANGPTL3 amino acids and The to in a The SE1 in of LPL with whether ANGPTL3, ANGPTL4, directly with LPL, we an binding In this LPL with mAb, mAb KLH, mAb 5.50.3 to a 4 the and with antibody to the of LPL binding to LPL binding was in not in of mAb 5.50.3 binding of LPL to by more of mAb on These results indicate that the SE1 region of ANGPTL3, that of ANGPTL4, is with the binding and inhibition of of LPL by ANGPTL3 in in of results of show that the SE1 region of ANGPTL3 is in inhibiting LPL activity. We that antibodies that with the SE1 region and inhibition of LPL activity by ANGPTL3 LPL activity and TG levels in vivo. To this we Angptl3-/- mice to as that in the of ANGPTL3 on serum lipid levels. produced and of Angptl3-/- mice. We treated C57BL/6J mice with control mAb SE1 mAb 5.50.3 and the TG levels of the mice with of Angptl3-/- mice and wild-type TG levels in Angptl3-/- mice in and in mice treated with mAb 5.50.3 in mice treated with control mAb These results that mAb 5.50.3 inhibition of LPL by ANGPTL3 in vivo. We whether inhibition of LPL activity by ANGPTL3 is a for serum lipid levels in mouse of hyperlipidemia. To this we treated mice with control mAb mAb 5.50.3 and the of treatment on serum TG and levels. The TG and levels of these mice to of mice mice. TG levels in mice in and in mice treated with mAb 5.50.3 in mice treated with mAb serum TG levels in mice in and in mice treated with mAb 5.50.3 in mice treated with mAb levels in mice in and in mice treated with mAb 5.50.3 in mice treated with mAb not serum levels in mice in and in mice treated with mAb 5.50.3 in mice treated with mAb not In mice and in mice treated with mAb 5.50.3, the levels to not these results that monoclonal antibodies that ANGPTL3-mediated inhibition of LPL activity plasma and that hyperlipidemia. The SE1 of ANGPTL3 and ANGPTL4 are the that the SE1 in vivo. To the of 14D12 and 5.50.3 in we treated Angptl3-/- mice mice with mAb 14D12 5.50.3 and serum TG levels and fasting treatment of Angptl3-/- mice with mAb 14D12 lowered fasting serum TG levels, treatment of Angptl3-/- mice with mAb 5.50.3 on serum TG levels treatment of mice with mAb 5.50.3 lowered serum TG levels, treatment of mice with mAb 14D12 on serum TG levels These results that mAb 14D12 is with ANGPTL4 and mAb 5.50.3 is with ANGPTL3 in vivo. of ANGPTL3 on TG in mAb and mAb that to the of ANGPTL3 in C-terminal to and of the SE1 domain ability to with ANGPTL3, the serum TG of these was more that for mAb 5.50.3 not ANGPTL3 and ANGPTL4 regulate lipid metabolism by inhibiting the activity of LPL, which in plasma lipid levels A. Chao Y.B. Mosior M. Ford A. Gonzalez-DeWhitt P.A. Hale J.E. Li D. Qiu Y. Fraser C.C. Yang D.D. Heuer J.G. Jaskunas S.R. Eacho P. Endocrinology. 2005; 146: 4943-4950Crossref PubMed Scopus (354) Google Scholar, S. Y. A. Nat. Genet. 2007; PubMed Scopus Google Scholar, T. Ono M. Shimamura M. Yoshida K. Ando Y. Koishi R. K. T. H. T. Furukawa H. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, K. Shimizugawa T. Ono M. Furukawa H. J. Lipid Res. 2002; 43: Full Text Full Text PDF PubMed Scopus Google Scholar). we showed that mAb 14D12 neutralized the inhibition of LPL by ANGPTL4 and that mice treated with this mAb recapitulated the phenotype of mice (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). These in vitro and in that mAb 14D12 a region in ANGPTL4 that is important for its as an of LPL activity. the ANGPTL4 recognized by mAb 14D12, we to ANGPTL4 inhibits LPL activity and to similar domains angiopoietin angiopoietin-like family members that inhibit We showed that the epitope recognized by mAb 14D12 is amino acids Gln29–His53, which we designate as This region is the signal and the of coiled-coil domains and The of the SE1 region in the of ANGPTL4 is not only by the activity of mAb 14D12 by the of a which is the of the SE1 The ANGPTL4 E40K variant is with serum TG levels in S. Y. A. Nat. Genet. 2007; PubMed Scopus Google and the LPL of the human E40K variant is in vitro Yu Y. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). We showed that mice with the E40K variant have serum TG levels in consistent with the human the SE1 region of ANGPTL4 likely a domain in inhibition of all angiopoietin and angiopoietin-like family members ANGPTL4, we found that only ANGPTL3 contains an SE1-like To whether the SE1 region of ANGPTL3 in LPL we produced mAb 5.50.3, which with this SE1 This mAb neutralized the LPL-inhibitory activity of ANGPTL3 in vitro and inhibited the binding of ANGPTL3 with LPL, as mAb 14D12 inhibited the binding of ANGPTL4 to mAb 5.50.3 is an of ANGPTL3 in vivo. Treatment of C57BL/6J mice with mAb 5.50.3 was of serum to levels found in Angptl3-/- mice. Moreover, mAb 5.50.3 treatment LPL activity in U. J. N. B. P. G. D. R. and K. these in vitro and in results that the SE1 present in ANGPTL4 and ANGPTL3, a functional domain in inhibition of LPL activity. Recently, and S. J. J. Google on in ANGPTL3 and ANGPTL4 of the SE1 domain that to the LPL-inhibitory activity of these We that to the of ANGPTL3 ANGPTL4 in that are C-terminal to the SE1 domain and that some of these of these to serum TG levels to SE1 In of these to inhibit the binding of LPL to in to the SE1 these that the SE1 domain in ANGPTL3 and ANGPTL4 plays a and role in inhibiting LPL activity. results not the of N-terminal amino acids in SE1 in inhibition of TG levels of mice significantly lowered by mAb 5.50.3, and serum TG levels of Angptl3-/- mice significantly lowered by mAb mAb in mice the by that These indicate that mAb 5.50.3 ANGPTL3 and mAb 14D12 ANGPTL4. The the activity of serum ANGPTL3 in the and activity of ANGPTL4 in the consistent with observations in A. Chao Y.B. Mosior M. Ford A. Gonzalez-DeWhitt P.A. Hale J.E. Li D. Qiu Y. Fraser C.C. Yang D.D. Heuer J.G. Jaskunas S.R. Eacho P. Endocrinology. 2005; 146: 4943-4950Crossref PubMed Scopus (354) Google Scholar, U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). these that a significantly in serum TG be with inhibition of with inhibition of we that and mice a reduced with of the and (9Desai U. Lee E.C. Chung K. Gao C. Gay J. Key B. Hansen G. Machajewski D. Platt K.A. Sands A.T. Schneider M. Van Sligtenhorst I. Suwanichkul A. Vogel P. Wilganowski N. Wingert J. Zambrowicz B.P. Landes G. Powell D.R. Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 11766-11771Crossref PubMed Scopus (152) Google Scholar). mice with mAb 14D12 recapitulated the phenotype found in mice. the for this phenotype is not clearly the of with these in mice that ANGPTL4 may be for LPL activity to of acids from In the present we not reduced in Angptl3-/- mice in mice treated with mAb 5.50.3, the mAb not These results indicate that ANGPTL3, ANGPTL4, is not in lipid metabolism in mouse have shown that of ANGPTL3 with plasma lipid and of and in S. J. J. Google Scholar, S. A. R. J. A. A. G. N. D. S. H. P. P. S. D. Li Y. P.A. J. R. S. Y. G. R. M. J. A. M. D. Nat. Genet. PubMed Scopus Google Scholar, S. C. A. C. B. T. D. G. P. B. C. M. Nat. Genet. PubMed Scopus Google Scholar, S. T. K. K. S. H. M. Y. J. Res. 2007; PubMed Scopus Google Scholar). Moreover, have shown that TG levels with an of of M. P. A. J. 2007; PubMed Scopus Google Scholar, S. J.E. N. S. J. 2007; PubMed Scopus Google Scholar). In the of fasting TG levels with was for the of The mechanisms for the TG levels and however, it likely that TG levels, a in of to an of lipid Chem. PubMed Scopus Google Scholar). These observations that may have in the treatment of In this study, we that mAb 5.50.3 is of the lipid of and of hyperlipidemia. These results that by of LPL, a for treatment of human with