Antisense oligonucleotides targeting bmper for treatment of glomerular diseases

Abstract

Compositions and methods for decreasing a level of expression or activity of BMP binding endothelial regulator (BMPER) are provided. Also provided are antisense oligonucleotides (ASOs) that target BMPER.

Description

[0001] Docket No. 084284.00345

[0002] ANTISENSE OLIGONUCLEOTIDES TARGETING BMPER FOR TREATMENT OF GLOMERULAR DISEASES CROSS-REFERENCE TO RELATED APPLICATIONS

[0003] This application is entitled to priority pursuant to 35 U. S. C. §119(e) to U. S. Provisional Application No. 63 / 719,469, filed on November 12, 2024. The content of the application is incorporated herein by reference in its entirety.

[0004] STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT

[0005] This invention was made with government support under Grant No. W81XWH2010836 awarded by the Department of Defense and under Grant No. R01DK097253 awarded by the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK). The government has certain rights in the invention.

[0006] REFERENCE TO AN ELECTRONIC SEQUENCE LISTING

[0007] The contents of the electronic sequence listing (084284.00345 SeqList.xml; Size: 39,552 bytes; and Date of Creation: October 28, 2025) is herein incorporated by reference in its entirety.

[0008] FIELD OF THE INVENTION

[0009] This disclosure relates generally to antisense oligonucleotides and methods of use thereof for treating glomerular diseases.

[0010] BACKGROUND OF THE INVENTION

[0011] Chronic kidney disease (CKD) is a progressive disorder affecting more than one in seven US adults, or 37 million people. Unfortunately, there is only a limited number of drugs currently approved for the treatment of CKD, including renin-angiotensin system (RAS) antagonists, which provide the only class of compounds available to reduce proteinuria and a slow loss of renal function. However, despite the widespread clinical use of RAS blockade, the prevalence of advanced stages of CKD continues to increase. There are also new therapies emerging, such as sodium-glucose cotransporter 2 (SGLT2) inhibitors and glucagon-like peptide 1 agonist, which can slow the progression. However, the absolute risk of renal and cardiovascular morbidity and mortality, as well as the need for renal replacement therapy, remains high. Notably, these are non- 1

[0012] 178599064.1 Docket No. 084284.00345

[0013] specific therapies. Hence, new therapies averting progression of CKD constitute an extraordinary unmet need in clinical practice as well as have a significant clinical and financial impact.

[0014] SUMMARY OF THE INVENTION

[0015] This disclosure relates to compositions for decreasing a level of expression or activity of BMP binding endothelial regulator (BMPER) and methods of using such compositions to treat a glomerular disease in a subject in need thereof.

[0016] In some embodiments, provided is an antisense oligonucleotide that specifically hybridizes to a nucleic acid encoding BMPER, wherein the oligonucleotide decreases a level of expression or activity of BMPER. In some embodiments, the oligonucleotide is unmodified. In some embodiments, the oligonucleotide is modified. In some embodiments, the modification comprises at least one modified internucleotide linkage. In some embodiments, the modified internucleotide linkage is a phosphorothioate linkage. In some embodiments, the modification further comprises at least a locked nucleic acid (LNA), a lipid, a peptide, an aptamer, an antibody or antigen-binding fragment thereof, or a sugar.

[0017] In some embodiments, the oligonucleotide comprises a polynucleotide sequence of any one of SEQ ID NOs: 1-11 or comprises a polynucleotide sequence having at least 75% sequence identity to any one of SEQ ID NOs: 1-11. In some embodiments, the oligonucleotide consists of 16 linked nucleotides. In some embodiments, the nucleic acid encodes human BMPER.

[0018] In some embodiments, provided is a pharmaceutical composition comprising the antisense oligonucleotide as described herein and optionally a pharmaceutically acceptable carrier or diluent. In some embodiments, the composition is a liposome composition.

[0019] In some embodiments, provided is a kit comprising the antisense oligonucleotide as described herein.

[0020] In some embodiments, provided is a method of treating a glomerular disease in a subject in need thereof, comprising administering to the subject one or more doses of the antisense oligonucleotide or the pharmaceutical composition as described herein. In some embodiments, the subject is a human. In some embodiments, the glomerular disease is chronic kidney disease (CKD). In some embodiments, the antisense oligonucleotide or composition as described herein is administered to the subject parenterally. In some embodiments, the antisense oligonucleotide or 2

[0021] 178599064.1 Docket No. 084284.00345

[0022] composition as described herein is administered to the subject intravenously. In some embodiments, the subject has primary or secondary focal segmental glomerulosclerosis (FSGS). In some embodiments, the one or more doses of the antisense oligonucleotide as described herein comprises about 0.0001 μg to about 100 g per kg of body weight of the subject.

[0023] BRIEF DESCRIPTION OF THE DRAWINGS FIGS. 1A-1E illustrate that BMP binding endothelial regulator (BMPER) is pathogenic and a therapeutic target for CKD in vivo. FIG. 1A is an immunohistochemistry (IHC) image of BMPER expression in control human glomeruli. FIG. IB is an IHC image of BMPER expression in diabetic human patients with CKD. FIG. 1C is an IHC image of BMPER expression in the vasculature of diabetic human patients with CKD. FIG. ID is a graph of the plasma levels of BMPER protein in 20 healthy controls and in 20 patients with early CKD. FIG. IE is a graph of human urine BMPER relative to urine creatine in controls, diabetics with non-progressive and fast progressive diabetic kidney disease (DKD). (*P<0.05).

[0024] FIG. 2 is a graph of albumin: creatinine (ACR) of PodTgfbrI + Dox mice with Scrbl (scramble control) with CKD or BMPER shRNA (n=3 mice per group). *P<0.05, **P< 0.01.

[0025] FIG. 3 is a graph of BMPER mRNA expression in immortalized mouse glomerular endothelial cells (mGECs) that were reverse transfected with BMPER ASO candidates (30 nM) overnight or with BMPER siRNA and treated with 10 pg / ml puromycin amino-nucleoside (PAN). Non-target ASO-NC5 is a negative control. PAN is a podocytotoxin. BMPER mRNA expression was determined via RT-PCR.

[0026] FIGS.4A-4C show the results of studies aimed to block BMPER release by stressed GECs using antisense oligos (ASOs) in a db / db type 2 diabetes (T2D) experimental mouse model for 16 weeks. FIG. 4A is a schematic of the experimental plan of ASONC5 (control), ASO70, and ASO 1982 doses that were subcutaneously injected (20 mg / kg) on 3 consecutive days every 4 weeks. FIG. 4B is a graph that confirms no change in fasting blood glucose (FBG) in db / db diabetic mice in either treatment. FIG. 4C is a graph of mouse body weights (BW) in db / m and db / db diabetic mice with ASOs. ** P<0.01, *** P<0.0001, **** P<0.0001.

[0027] FIGS. 5A and 5B illustrate albumimcreatine in db / db mice treated with ASO-NC5, ASO70, or ASO1982. FIG. 5A is a graph of albumin: creatinine (ACR) in db / db mice. A 3

[0028] 178599064.1 Docket No. 084284.00345

[0029] significant decrease in albuminuria was observed in BMPER ASO70 and ASO1982 treated mice (**p<0.01). FIG. 5B shows images of PAS staining in db / db mice treated as indicated. PAS staining of db / db Ctrl ASONC5 shows glomerular sclerotic lesions, tubular injury with loss of brush boarders compared to db / m mice. In contrast, BMPER ASO70 and ASO1982 treated db / db mice had reduced glomerular lesions, no tubular injury or presence of proteinaceous casts, which aligns with the reduced proteinuria and is associated with BMPER expression inhibition.

[0030] FIGS. 6A-6C demonstrate the results of a podocyte TGFbRl mouse model + Dox, a podocyte injury mouse model of focal segmental glomerulosclerosis (FSGS) treated with ASO70 or ASO1982. FIG. 6A shows the experimental plan of the podocyte TGFbRl model + Dox, a podocyte injury model of FSGS treated with ASONC5 (control), ASO70, and ASO1982. Doses were subcutaneously injected at 20 mg / kg on 3 consecutive days or with saline vehicle. FIG. 6B is a graph of ACR of the mice treated as indicated. BMPER ASO70 treated mice exhibited a significant decrease in albuminuria (*P<0.05). FIG. 6C shows images of PAS staining of the mouse model treated as indicated. PAS staining of mice shows glomerular sclerotic lesions, tubular injury with loss of brush boarders in Dox treated mice compared to vehicle control mice. In contrast, BMPER ASO70 and ASO1982 treated mice exhibited reduced glomerular lesions, no tubular injury or presence of proteinaceous casts, which was associated with reduced BMPER expression.

[0031] FIGS. 7A-7D demonstrate the results of an adriamycin (ADR; lOmg / kg) induced FSGS in Babl / c mice treated with ASO70. FIG. 7A is a schematic of the experimental plan of an ADR induced FSGS model and treatment with ASONC5 (control), and ASO70 (injected subcutaneously at 20 mg / kg on 3 consecutive days). FIG. 7B is a graph of ACR of the indicated mice. BMPER ASO-70 treated mice exhibited a significant decrease in albuminuria (**P<0.01). FIG.7C are IHC images of BMPER staining in ADR treated Balb / c mice with ASONC5 or ASO70. FIG.7D shows images of PAS staining in Balb / c mice treated with ASO70 or ADR. PAS staining shows glomerular sclerotic lesions, tubular injury with loss of brush boarders in ADR treated mice compared to ASO70 treated mice, which had reduced glomerular lesions, no tubular injury or presence of proteinaceous casts.

[0032] 4

[0033] 178599064.1 Docket No. 084284.00345

[0034] DETAILED DESCRIPTION OF THE INVENTION

[0035] This disclosure is based at least in part on an unexpected discovery that downregulating BMP binding endothelial regulator (BMPER; CV2; CV-2; or CRIM3) gene expression improves glomerular filtration barrier function in CKD. Also provided are antisense oligonucleotides (ASOs) that target BMPER and methods of use of such ASOs for treating glomerular diseases.

[0036] BMPER is an endothelial cell stress signal in CKD. BMPER is a secreted glycoprotein protein that inhibits bone morphogenetic protein (BMP) function. BMPER contains 5 cysteine-rich domains, followed by a von Willebrand D (vWD) domain, and a trypsin inhibitor (TI) domain. In particular, BMPER plays a key role in BMP-mediated endothelial events, glucose homeostasis, and chronic inflammation in the vasculature. BMPER null mice are embryonic lethal, reflecting its importance. Additionally, cleavage of BMPER may dictate its pro or anti-BMP activity (Moser, M. et al. J Mol Cell Cardiol. 2007. 43(3):243-53).

[0037] Exemplary sequences of BMPER include, but are not limited to:

[0038] SEQ ID NO: 12 (Mus musculus BMP -binding endothelial regulator (Bmper), mRNA; NM_028472.2)

[0039] GCCGGGGCTGGGCTAGACCGCCTGGCTGGGCGTTCGCGTGCGCCGCTGTCGCTTCTGAGCCGGC ATAGCTCCGGCGCCAGGATCGTAGTCCCTCGTGCGACCCTGCGGATTCCGCGCAGAGCCTAGTA CGCCCCTCGGTCCCGCACGGAGCAAGCGCGCGGGCCGGACACCGCGCTCGGGGGCCGGGCCGGA GCTGCGCGTGCGGGAGCCGGAGCTCAGCCGGGCTCCAGAGCTGCGAGCGCACTCGGCAGTCGCC GCCGCCGCTCCCCAGATCCCGCCGCTCAGCCACCCCTCCTCCCGGCCCAGCTCTGGCGCGGAGG GGCTGACGGCTAACCCGCTGAGAGCAACTCCCTTAGAAAAAAGCCGCCGCATTGGCCGAGGCTG GCCGCAGGACCCCTCCTCCCGGGGCGCCCCCACTCCTCCCCCGCCGCTCCGGGCGCCCGCCTCC CTCAGCCGACTGTCTCCCGGCACGCCGGCCCACAGCCCTTTTGACTGCGAGCAGCGGCTGCCGA GCAGAGGCGGCGACGCGGGACCTGCAGTCGCCCGGGATTCCCTCCAGGTGACGATGCTCTGGTT CTTCAGCGTCAGGGCTCTGGCTGAGCGTCCTTGCCGACGCTCGCCTGGGATTACCTGCTGCGTC TTGCTGCTGCTCAATTGCTCCGGGGTCCCCATGTCTCTGGCTTCCTCCTTCCTGACAGGGTCTG TTGCAAAATGTGAAAATGAAGGGGAGGTCCTCCAAATTCCATTTATCACAGACAACCCTTGCAT AATGTGTGTGTGTTTGAACAAAGAGGTGACTTGTAAGAGGGAGAAGTGCCCTGTGCTGTCACGG GACTGTGCGCTGGCCATCAAGCAGAGAGGCGCCTGCTGTGAACGATGCAAAGGTTGTACCCATG

[0040] 5

[0041] 178599064.1 Docket No. 084284.00345

[0042] AAGGAAGAACCTACAACAGTTCCTTCAAGTGGCAAACCCCGGCTGAGCCATGTGTCCTTCGGCA GTGCCAGGAGGGTGTGGTGACAGAGTCTGAGGTGCGCTGTGTTGTTCATTGTAAAAATCCCGCA GAGCATCAAGGAGCCTGCTGCCCTACCTGCCCAGGCTGTGTGTTTGAAGGTGTGCAATACCGGG AAGGAGAAGAATTTCAGCCAGAAGGAAACAAATGTATTACATGCTCCTGCGTTGGAGGCAGGAC ACAGTGCGTGAGAGAAGTCTGTCCCATTCTCTCTTGTCCCCAGCACCTTAGTCACACACCCTCA GGACAGTGCTGCCCCAAATGTTTGGGTCAGAGAAAAGTATTCGACCTTCCCTTTGGGAGCTGCC T G T T T C G GAG TGATGTTTAT GAG AAT GGAG CCTCATTTGTCTAT GAT AAC T G GAG AG TAT GC AC TTGCAAGGACTCAACAATGGTTTGTAAGAAGAAGTGCTCCCACCCTGGTGTCTGTAACAGTGAT GAGGATGCCTGTTGTGAAGACTGTCTCTTACGGGTGCCCCCTGAAGACATCAAAGTGTGCAAGT TCGGTAGCAAGATTTTCAGGGATGGAGAGATGTGGTCCTCAGTCAACTGCAGCATCTGTGCTTG TGTGAAGGGGAAGACAGAGTGTCGAAAGAAGCAGTGTGTTCCTGTCAGCAGCTGCCCACAGGGT AAAATTCTCAACAGGAAAGGATGCTGTCCTATCTGCACTGAAAAACCTGGTGTGTGCACAGTAT TCGGAGATCCCCACTACAACACTTTTGATGGGCGGACATTTAACTTTCAGGGGACGTGTCAGTA TGTTTTGAC AAG CTGCTCCTCCCCAGCCTCACCCTTCCAGGTGCTGGTGA G TG TGCA CGGAGGACCCGTTCCTTCTCCTGGACCAAGTCTGTGGAGCTGATGCTGGGCGAAAGCACAGTCA GCCTGCAGCAGCACCTCACTGTGCGCTGGAATGGCTCTCGCATTGCCCTTCCCTGCCACACACC CCACTTCCACATCGACCTGGACGGCTACCTTTTGAAAGTGACTACCAGAGCAGGTTTGGAGATA TCGTGGGATGGAGACAGTTTTGTGGAGGTCATGGCTGCCCCTCATCTCAAGGGCAAGCTCTGTG GTCTTTGTGGCAACTACAATGGTCATAAACGTGATGACTTGATTGGTGGAGATGGGAACTTCAA GTTTGACGTGGATGACTTTGCTGAATCGTGGAGGGTGGAATCCAACGAATTCTGCAACCGACCA CAGAGAAAACCAGTACCTGAGCTCTGTCAAGGCACAGTGAAGGTGAAGCTTCGAGCCCATCGAG AATGCCAGAAACTCAAGTCCTGGGAGTTCCAGACCTGCCACTCTACAGTGGACTATACCACTTT CT CCGGTCCTGTGT ACAG C TGTGTGAATGTCC GTCCAT AA CTGTT CTGTG GTCA TTTCTGGCGTATACCCGTGCCTGTCAGAGAGAGGGAATCAAAGTGCACTGGGAACCTCAGCAGA GCTGTGCAGCCACCCAGTGCAAACATGGTGCTGTGTATGATACCTGTGGCCCAGGATGTGTGAA GACATGTGATAACTGGAATGAGATCGGCCCATGCAATAAGCCCTGCATTGCAGGTTGCCACTGC CCAGCAAATCTGGTCCTTCATAAGGGAAGGTGCATCAAGCCAGTCCTCTGTCCTCAGCGATGAC CTTTGTTCTAGTCCATAGACTTTGAAATCCAGTGTCTTTGACACTGACATGCAAGAGCCAGTGA AGGACTGTTGTATTTGTGTGCCTGATTCTGCAAATACACAGAGTATATATGTGTACATATATAT AGAT AT AT AGAT AT AT T C GAAAC AT T T C AT C AT T T AT AT AAAC TATAGGTGGATTATTATATGT AT AT T T T T T GC T AT AAGACAT G TAT TAT T T C TAGAAT CC T AAT AT C T AAGC CAT T GGAT GCAT C

[0043] 6

[0044] 178599064.1 Docket No. 084284.00345

[0045] ATATACATAGGT GC T T T TAAT T TAATAAGAT GGCAT GCAGAC T CAT CAGGT C T C T T TAAGAGT G CGTTCATGTGTCATTCTGGGGACATTTCCTAAGAGCCCTCATGCCTACCTGCATTAATTAAAGA TTTGAGTCAGGACCTTTGAGTGAGATTTTCTTGGAGAAGGGTACTTTATCTGGGGGTATTTTGT ATTTCCAAAGACTGGAATGTATGCCAGAGAAAGATGGCATGAGATTGGGGATGAGCCCTCGTGA GTGCAAACCAAGGAGAGTCTGTCAGATTTCACCGGGTCAGAGCTGGATATTGTCCCCCAAACTG GATAGTTGCCACGGAATATGTATTTTTCACTAGCATATTGCCAAATAATTTCTTTTCATTTTTT GCATATGTCATGAGTCTCCTCATTACTTATGTGTGCAATGGATATGACCATTTGCATGTCCTGC AC T GAAG T CAAAGGAT G T C T GGAT AACAT T T TAT T AAACAC GAAAAGC TAT T T GGAGAGAACAT TGCACAACATATAAAGAATCTCAGAGCAATTTAGGATTGGCAAATCCATTGAGTGGCTAGGGGC CATCTACGTTTTGTCACTATCTCATGGTGTATAAAATGAAAGCCTTCGGCTATTCTGTCTCATT ACTGTCCAGTGTTGTTTGCTGGGTACTCATATGTTATGGGTATTGGGCAGGAGGGAAGGGACTG CTGACCAGTTGTTTTAGCTTCATTCCAAAAGAGAATTGTTTCTGTATTTGTTCTAATTTACTGA T G T T AC T AAC AG T T AT T G G T T C T T G T T T T C T G T T G GAT AAAT AAAAAT G T CAAAAAG C AAAAAC AAAAAAC C C C AC C C C C AAC T C AC T GAT G T T GAAAAG T AAAAT G T T C T G CAT T G T AG T AAAT AAA AGGCT TAAGAT T TAAGCAAACAGT T T TCAA

[0046] The underlined sequence is the coding sequence (CDS) (SEQ ID NO: 13).

[0047] SEQ ID NO: 14 (Homo sapiens BMP binding endothelial regulator (BMPER), transcript variant 1, mRNA; NM_001365308.1)

[0048] GACTGTGAGCTGCGGCAGCTGAGCAGAGGCGGCGGCGCGGGACCTGCAGTCGCCAGGGATTCCC T C C AG G T G C GATGCTCTGGTTCTCCGGCGTCGGGGCTCTGGCTGAGCGTTACTGCCGCCGCTC GCCTGGGATTACGTGCTGCGTCTTGCTGCTACTCAATTGCTCGGGGGTCCCCATGTCTCTGGCT TCCTCCTTCTTGACAGGTTCTGTTGCAAAATGTGAAAATGAAGGTGAAGTCCTCCAGATTCCAT T T AT C AC AGAC AAC C C T T G C AT AAT G T G T G T C T G C T T GAAC AAGGAAG T GAC AT G T AAG GAGA GAAGTGCCCCGTGCTGTCCCGAGACTGTGCCCTGGCCATCAAGCAGAGGGGAGCCTGTTGTGAA CAGTGCAAAGGTTGCACCTATGAAGGAAATACCTATAACAGCTCCTTCAAATGGCAGAGCCCGG CTGAGCCTTGTGTTCTACGCCAGTGCCAGGAGGGCGTTGTCACAGAGTCTGGGGTGCGCTGTGT TGTTCATTGTAAAAACCCTTTGGAGCATCTGGGAATGTGCTGCCCCACATGTCCAGGCTGTGTG TTTGAGGGTGTGCAGTATCAAGAAGGGGAGGAATTTCAGCCAGAAGGAAGCAAATGTACCAAGT GTTCCTGCACTGGAGGCAGGACACAATGTGTGAGAGAAGTCTGTCCCATTCTCTCCTGTCCCCA GCACCTTAGTCACATACCCCCAGGACAGTGCTGCCCCAAATGTTTGGGTCAGAGGAAAGTGTTT

[0049] 7

[0050] 178599064.1 Docket No. 084284.00345

[0051] GACCTCCCTTTTGGGAGCTGCCTCTTTCGAAGTGATGTTTATGACAATGGATCCTCATTTCTGT ACGATAACTGCACAGCTTGTACCTGCAGGGACTCTACTGTGGTTTGCAAGAGGAAGTGCTCCCA CCCTGGTGGCTGTGACCAAGGCCAGGAGGGCTGTTGTGAAGAGTGCCTCCTACGAGTGCCCCCA GAAGACATCAAAGTATGCAAATTTGGCAACAAGATTTTCCAGGATGGAGAGATGTGGTCCTCTA TCAATTGTACCATCTGTGCTTGTGTGAAAGGCAGGACGGAGTGTCGCAATAAGCAGTGCATTCC CATCAGTAGCTGCCCACAGGGCAAAATTCTCAACAGAAAAGGATGCTGTCCTATTTGCACTGAA AAGCCCGGCGTTTGCACGGTGTTTGGAGATCCCCACTACAACACTTTTGACGGTCGGACATTTA ACTTTCAGGGGACGTGTCAGTACGTTTTGACAAAAGACTGCTCCTCCCCTGCCTCGCCCTTCCA GGTGCTGGTGAAGAACGACGCCCGCCGGACACGCTCCTTCTCGTGGACCAAGTCGGTGGAGCTG GTGCTGGGCGAGAGCAGGGTCAGCCTGCAGCAGCACCTCACCGTGCGCTGGAACGGCTCGCGCA TCGCGCTCCCCTGCCGCGCGCCACACTTCCACATCGACCTGGATGGCTACCTCTTGAAAGTGAC CACCAAAGCAGGTTTGGAAATATCTTGGGATGGAGACAGTTTTGTAGAAGTCATGGCTGCGCCG CATCTCAAGGGCAAGCTCTGTGGTCTTTGTGGCAACTACAATGGACATAAACGTGATGACTTAA TTGGTGGAGATGGAAACTTCAAGTTTGATGTGGATGACTTTGCTGAATCTTGGAGGGTGGAGTC CAATGAGTTCTGCAACAGACCTCAGAGAAAGCCAGTGCCTGAACTGTGTCAAGGGACAGTCAAG GTAAAGCTCCGGGCCCATCGAGAATGCCAAAAGCTCAAATCCTGGGAGTTTCAGACCTGCCACT CGACTGTGGACTACGCCACTTTCTACCGGTCCTGTGTGACAGACATGTGTGAATGTCCAGTCCA TAAAAACTGTTATTGCGAGTCATTTTTGGCATATACCCGGGCCTGCCAGAGAGAGGGCATCAAA GTCCACTGGGAGCCTCAGCAGAATTGTGCAGCCACCCAGTGTAAGCATGGTGCTGTGTACGATA CCTGTGGTCCGGGATGTATCAAGACGTGTGACAACTGGAATGAAATTGGTCCATGCAACAAGCC GTGCGTTGCTGGGTGCCACTGTCCAGCAAACTTGGTCCTTCACAAGGGAAGGTGCATCAAGCCA GTCCTTTGTCCCCAGCGGTGACCTTTGTTTCGATCCTTAAGACTCTGAAATCTGGTGACTTTGA CACTGAAGCGGAAGAGCCAATGAAGGACTGCAGTATTTGTGTGCCCGATTCTGTAAACACACAC AC AC AGAG TATATATGTG TAT AT AT AT AT AGAT AT AT T C AAAAAC AT TGCATCATTTATAT GAA C TATAGGGGGAT TAT TATAT GTATAT T T T T T GCTATAAGACAT GTAT T GT T T C TAGGAT CCTAA CCTGTAAGCCATTGAACATGTTGTATAAATACACCAGGTGTTTTTAATTTAATAAGGTGGCATG C AGAT AC AT T G GAT AG T G T T AAC AT C AC AT ACAT TTGTCATTTT T AAG GAAG T T T T C T AAGAG C CCTCAATTGCCTGCCTGTATTAATTTTAGTTTTGAGTCAGGATTTGTAATGGAGTGGGAAATGT GTTTCTCTGGAGAAGGGCACATTTATCTAGGGGCATTTCAGGTTTCCAAAGAAAGGAATGTATG CCTGGGAAAGACAGCAGGAGATTGGTGACCGGCCCTAATGGTGCATGAAAAGCGAGTGATAGGC TGTTAGAATGTATTAGGTCATGGGCTAACATTATTTCCAAAATTGATTGGCTGGTTGCCAAGAA

[0052] 8

[0053] 178599064.1 Docket No. 084284.00345

[0054] ATATATATTTGTCACTAGGGCATAACCGAAGAATCAAGTGATTTCTTGCCATCCTTGCGTAGTC CTCAAGTCTTCTAATCATCTATGTGTGTATGATGGATGTGACCATTTATATGTCATGTGTTGAA AT T AAAG GGTGTCTGGATGCCATGTTAATTT TAT T GAGC AT GAGAAAG TAT T T GAAGAAAC T G T T G C AC AAC AT AT GAT GAAAT TTTGATTTCTCT GAAT C T C AGAG T AAT T T AAGAT GG G C AAAT C C AAT GAG T T GAT G TAAC C C AT C T AC AT T T T G T G G C T AT T T C AT G T ATAAAAT GAAAG G C T T T AG T T AT T T C T AGAG GAG T C C C AT C C AAC AC TATTTGGC GAC T G T T AAT AT GTTATAATTAT T AGAC A GTGGAAAAGCCTTTATTGACTAATTGATTTAACCTCAGTCCAAAGAACAGTCTCTATACTTGCC CTCATTTACTGATAGGCTGCTACTAGTTACCATTTGATTGTCTTGACTTTCGTCTTCAGTTGAA T T G G T GAAAAC AT C AAC AAAAAC AAAAC C C AC T G T G T C T T AAAAT AAAAT GTTCTGCATTGTAG T CAATAAAG GG C T T AAGAT G TAAAAC AAT G CAT T T T C T C T C T C AAGAT G T C C T G T G T TAT AC T T TAG AAT TAGTTGTAT C AAC T T C AGAT T C AT T AAAT T T TAAC AG C T G T T T T AG T T AT AAT T T T T G T AT C T TAG AAC AG T AT AT AT AAAGAC TAT TAT T AAT AGAC AAAAG C AAAC C C AC G T T T T T GAC AAGCCAGCATATTGCCAATGCTCGTCAAAGACATTTATGTTTCTTTTACATTCATGACAGCTAA AGTATTAACTTGGAATTTTTTTTTTTTTTTTTGACTGGAAAATCCCTAGTTGTTTGAATTAACT GAATAAAGTGCAGGGCGTAATAGTGACTGGCTGGGGGCACTGGGCCTTTGTGTTTTTTTTCATT CCTGTGTTTGGATACTAATCACCCTCGTATCTGCAGCCTGAGGTAGTCAGGTCATTGTTTCAAT GCTGCCCTTGGAAAGAAGACTTTGACTTAGGTGCTTTGGGATCTAAGTGTGGTTTGTGACTATC CCCATTATTTAAGCATTTGGGGGTCTCACTTTTGCTTATCTGTAACACAAAGAGGCCATGTAGT ATAGTGAGCAGGGTTGAGAATCGGGAAAACTAGACCCCTGCCACTAACAGGCTAGGAGACTTTG GGCAGTAATGGGAGGCACTGAGGGTTTGCAAATACTTTCCTACTTGTAATCTCATTGATTCCCT GCTCTGCCCTGACAGGAGGGGTAGGGAAAATCAGTTGACGTTGCCTGGTGGTTCCTACCTTCTG TAAAACGAACAGATTCCTTAAGATTAGCTTTAAATCCCTTTTCAGTTCACTGTTGGTTCTCATC T T T GAAAGAGGGCAGCATAAGGGAAATGAT GGAGAAGGGAGAAT T GT GGAT GGT TAT T TATAGC TCATGATTCATCCAAGGACTGAGTTATTCTCCCCATGAGGCTAACCGGCCTCTTTCCCAACACT TAATCATGGCAGCCAGGATGGGTGAATGATGAAAGCTCTTCTGCATGAGCCGGTTTCATCATCT TGGGTGGGTTGATTACAAGTTTGTGTGGCATTCCTTTAGCTGGGGATTAGCTTAATCTCTGTAC AT T T T T C T CAT T T C TAAT T GAAGAAGAAAGGGAGGGCAGAGAGGACAGAAAAT TAG T T GGGT T T ACTAATTCAAAGAGCCATCTCTGATGGGAGAGGGTGAGGAAATCAGCTGAATGGTTGGCCTGCA TATTCTTTTTTAGGCCACAGGCTACAGCTGAGCAATGGCTTAGTCTAATGATTGGGCACAATGA GGATTTACCAAGTGCTTCAATGAGAGCAGTCCTGTTTCATTCACATGGGATGTTTTCAATTGGT T C T T C AAC T AAG C T C T T G C AGAG T GAGAT GAC T T GAAAT AAAAAT G T T C T AT GAAT AT T GAAT T

[0055] 9

[0056] 178599064.1 Docket No. 084284.00345

[0057] T GATGAGAAACAAGCACCC T CAAT GAAGAACAAGCATAAT TAT GAT T C TAAT T GCCCAT T GT T G GTACTCAGTAAAACTAAGTTCTTTCTTCTCTCCCTCTGTAAATAGGAATATTTGAGGGTCCTGA CCTAAAATTTTTTAGCTGGGAAGAAAAGATATCTCTTTGGCATCGACAAGAGCAATTTTTGTTA AAACTGTGGTACACTAAGAATCACTAACTCTTCAGGTTGAAGGCCTCACTCATCTTAACTCGTT GAG TGCTTCCT GAG CAT C AGAC C G T C C T G T AAGAG GGCATCTTT C AT AAGAAC GAGAAAGAC C CAGGAGGTTGACTAGCTTCTCCCAGCTTTCACTGAAACACGGATAAACCATTTCAACTGGAAAA CTCTTCCTTTTTAT GAGAT AT T T C AGAC AC ACAAAAAGG C AT AGAGAAAAAT G T T G C AAAC AT C CAT GAC TAG AT T AC C C T AT T AAGAAAC T AAC C T G G T AAG T AG T C AAT AAAT T T T T C T T GAAT C AAT AAAT AAAAC C TAAT AAAT GCTGTCTCATGGTTGCA

[0058] The underlined sequence is the coding sequence (CDS) (SEQ ID NO: 15).

[0059] Antisense Oligonucleotides

[0060] In one embodiment, provided are antisense oligonucleotides that decrease a level of expression or activity of BMPER. Antisense oligonucleotides are short single-stranded oligonucleotides that modify gene expression of target genes. Antisense oligonucleotides are designed to hybridize with a specific RNA sequence for degradation by RNase H as a method to inhibit expression of a target protein, or alternatively to sterically block translation of its complementary RNA sequence. In one embodiment, the antisense oligonucleotides comprise 15 to 25 nucleotides such as 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25 nucleotides. In one embodiment, the antisense oligonucleotides described herein consist of 16 nucleotides. Table 1 depicts exemplary sequences of antisense oligonucleotides that target BMPER.

[0061] Table 1. Example antisense oligonucleotide sequences that target BMPER.

[0062] SEQ Name Species Sequence Modifications

[0063] ID NO:

[0064] 1 ASO- Human TATTGCGACACTCCGT +T*+A*+T*T*G*C*G*A*C*A*C*T 1055 *C*+c*+G*+T

[0065] 2 ASO- Human CTTCGGTTATGCCCTA +C*+T*T*C*G*G*T*T*A*T*G*C 2763 *C*+c*+T*+A

[0066] 3 ASO- Human TAGTGTTGGATGGGAC +T*+G*T*G*T*T*G*G*A*T*G 3081 *G*+G*+A*+C

[0067] 4 ASO- Human TATCGTACACAGCACC +T*+A*+T*C*G*T*A*C*A*C*A*G

[0068]

[0069] 1964 *C*+A*+C*+C

[0070] 10

[0071] 178599064.1 Docket No. 084284.00345

[0072] 5 ASO- Human GCCATGACTTCTACAA +Q*+C*+C*A*T*G*A*C*T*T*C*T 1513 *A*+C*+A*+A

[0073] 6 ASO- Human AACCAGAGCATCGTCA +A*+A*+C*C*A*G*A*G*C*A*T*C 70 *G*+T*+C*+A

[0074] 7 ASO- Mouse TTATTGCATGGGCCGA _|_ y * _|_ y * _|_y^ * y * y * Q * Q * A * T * G * G * G 2515 *c*+c*+G*+A

[0075] 8 ASO- Mouse TGTTGTAGTGGGGATC _|_y *+G * +T * T * G* T * A * G* T * G* G* G 1665 *G*+A*+T*+C

[0076] 9 ASO- Mouse AACCAGAGCATCGTCA +A*+A*+C*C*A*G*A*G*C*A*T*C 557 *G*+T*+C*+A

[0077] 10 ASO- Mouse GATGCACCTTCCCTTA +G*+A*+T*G*C*A*C*C*T*T*C*C 2580 *C*+T*+T*+A

[0078] 11 ASO- Mouse TCTCCATCCCACGATA+T*+C*+T*C*C*A*T*C*C*C*A*C

[0079]

[0080] 1982 *G*+A*+T*+A

[0081] * = Phosphor othioate bonds

[0082] += affinity plus locked nucleic acid base

[0083] The following antisense oligonucleotides have target regions within SEQ ID NO: 12: ASO-1665, ASO-1982, ASO-2580, and ASO-70. Table 2 depicts the target sequences of these antisense oligonucleotides.

[0084] Table 2. Target sequences of antisense oligonucleotides within SEQ ID NO: 12 Mus musculus BMP -binding endothelial regulator (BMPER), mRNA; NM_028472.2).

[0085] ASO Target Sequence SEQ ID NO:

[0086] ASO-1665 GATCCCCACTACAACA 16

[0087] ASO-1982 TATCGTGGGATGGAGA 17

[0088] ASO-2580 TAAGGGAAGGTGCATC 18

[0089] ASO-70 TGACGATGCTCTGGTT 19

[0090]

[0091] The following antisense oligonucleotides have target regions within SEQ ID NO: 14: ASO- 1055, ASO-2763, ASO-3081, ASO-1964, ASO-1513, and ASO-70. Table 3 depicts the target sequences of these antisense oligonucleotides.

[0092] 11

[0093] 178599064.1 Docket No. 084284.00345

[0094] Table 3. Target sequences of antisense oligonucleotides within SEQ ID NO: 14 ((Homo sapiens BMP binding endothelial regulator (BMPER), transcript variant 1, mRNA; NM_001365308.1).

[0095] ASO Target Sequence SEQ ID NO:

[0096] ASO-1055 ACGGAGTGTCGCAATA 20

[0097] ASO-2763 TAGGGCATAACCGAAG 21

[0098] ASO-3081 GTCCCATCCAACACTA 22

[0099] ASO-1964 GGTGCTGTGTACGATA 23

[0100] ASO-1513 TTGTAGAAGTCATGGC 24

[0101] ASO-70 TGACGATGCTCTGGTT 19

[0102]

[0103] ASO-70 has shared target regions between human and murine BMPER.

[0104] In some embodiments, provided are antisense oligonucleotides that have at least 70% (e.g., 70%, 75%, 80%, 85%, 90%, 95%, 99%, or more, or any intermediate values therebetween) identity to those described in Table 1. The sequence variation can be such that the antisense oligonucleotides retain their ability to target BMPER and decrease a level of BMPER activity or expression.

[0105] As used herein, the term “oligonucleotide” refers to an oligomer or polymer of ribonucleic acid (RNA) or deoxyribonucleic acid (DNA) or mimetics thereof. This term includes oligonucleotides composed of naturally occurring nucleobases, sugars and covalent internucleoside (backbone) linkages as well as oligonucleotides having non-naturally-occurring portions, which function similarly. Such modified or substituted oligonucleotides are often preferred over native forms because of desirable properties such as, for example, enhanced cellular uptake, enhanced affinity for nucleic acid target and increased stability in the presence of nucleases.

[0106] As used herein, the phrase “internucleotidic linkage” refers generally to a linkage linking nucleoside units of an oligonucleotide or a nucleic acid. In some embodiments, an internucleotidic linkage is a phosphodiester linkage, as extensively found in naturally occurring DNA and RNA 12

[0107] 178599064.1 Docket No. 084284.00345

[0108] molecules (natural phosphate linkage (-OP(=O)(OH)O-), which as appreciated by those skilled in the art may exist as a salt form). In some embodiments, an internucleotidic linkage is a modified internucleotidic linkage (not a natural phosphate linkage). In some embodiments, an internucleotidic linkage is a “modified internucleotidic linkage” wherein at least one oxygen atom or -OH of a phosphodiester linkage is replaced by a different organic or inorganic moiety. In some embodiments, such an organic or inorganic moiety is selected from =S, =Se, =NR’,-SR’,-SeR’,-N(R’)2, B(R’)3,-S-,- Se-, and-N(R’)-, wherein each R’ is independently as defined and described in the present disclosure. In some embodiments, an internucleotidic linkage is a phosphotriester linkage, phosphorothioate linkage (or phosphorothioate diester linkage, -OP(=O)(SH)O-, which as appreciated by those skilled in the art may exist as a salt form), or phosphorothioate triester linkage. In some embodiments, a modified internucleotidic linkage is a phosphorothioate linkage. In some embodiments, an internucleotidic linkage is one of, e.g., PNA (peptide nucleic acid) or PMO (phosphorodiamidate Morpholino oligomer) linkage. In some embodiments, a modified internucleotidic linkage is a non-negatively charged internucleotidic linkage. In some embodiments, a modified internucleotidic linkage is a neutral internucleotidic linkage (e.g., nOOl in certain provided oligonucleotides). It is understood by a person of ordinary skill in the art that an internucleotidic linkage may exist as an anion or cation at a given pH due to the existence of acid or base moieties in the linkage. In some embodiments, a modified internucleotidic linkage is a modified internucleotidic linkage designated as s, si, s2, s3, s4, s5, s6, s7, s8, s9, slO, si 1, s!2, si 3, sl4, sl5, sl6, sl7 or si 8 as described in WO2017 / 210647, which is herein incorporated by reference in its entirety.

[0109] As used herein, the terms “linker,” “linking moiety” and the like refer to any chemical moiety, which connects one chemical moiety to another. As appreciated by those skilled in the art, a linker can be bivalent or trivalent or more, depending on the number of chemical moieties the linker connects. In some embodiments, a linker is a moiety which connects one oligonucleotide to another oligonucleotide in a multimer. In some embodiments, a linker is a moiety optionally positioned between the terminal nucleoside and another nucleoside, nucleotide, or nucleic acid. In some embodiments, in an oligonucleotide a linker connects a chemical moiety (e.g., a targeting moiety, a lipid moiety, a carbohydrate moiety, etc.) with an oligonucleotide chain (e.g., through its 5’-end, 3’-end, nucleobase, sugar, internucleotidic linkage, etc.)

[0110] 13

[0111] 178599064.1 Docket No. 084284.00345

[0112] As used herein, the terms “modified nucleobase”, “modified base” and the like refer to a chemical moiety, which is chemically distinct from a nucleobase, but which is capable of performing at least one function of a nucleobase. In some embodiments, a modified nucleobase is a nucleobase, which comprises a modification. In some embodiments, a modified nucleobase is capable of at least one function of a nucleobase, e.g., forming a moiety in a polymer capable of base-pairing to a nucleic acid comprising an at least complementary sequence of bases. In some embodiments, a modified nucleobase is substituted A, T, C, G, or U, or a substituted tautomer of A, T, C, G, or U. In some embodiments, a modified nucleobase in the context of oligonucleotides refer to a nucleobase that is not A, T, C, G or U.

[0113] As used herein, the term “modified nucleoside” refers to a moiety derived from or chemically similar to a natural nucleoside, but which comprises a chemical modification, which differentiates it from a natural nucleoside. Non-limiting examples of modified nucleosides include those which comprise a modification at the base and / or the sugar. Non-limiting examples of modified nucleosides include those with a 2’ modification at a sugar. Non-limiting examples of modified nucleosides also include abasic nucleosides (which lack a nucleobase). In some embodiments, a modified nucleoside is capable of at least one function of a nucleoside, e.g., forming a moiety in a polymer capable of base-pairing to a nucleic acid comprising at least a complementary sequence of bases.

[0114] As used herein, the term “modified nucleotide” includes any chemical moiety which differs structurally from a natural nucleotide, but is capable of performing at least one function of a natural nucleotide. In some embodiments, a modified nucleotide comprises a modification at a sugar, base and / or internucleotidic linkage. In some embodiments, a modified nucleotide comprises a modified sugar, modified nucleobase and / or modified internucleotidic linkage. In some embodiments, a modified nucleotide is capable of at least one function of a nucleotide, e.g., forming a subunit in a polymer capable of base-pairing to a nucleic acid comprising at least a complementary sequence of bases.

[0115] As used herein, the term “modified sugar” refers to a moiety that can replace a sugar. A modified sugar mimics the spatial arrangement, electronic properties, or some other physicochemical property of a sugar. In some embodiments, a modified sugar is substituted ribose or deoxyribose. In some embodiments, a modified sugar comprises a 2’-modification. Examples 14

[0116] 178599064.1 Docket No. 084284.00345

[0117] of useful 2 ’-modi fi cations are widely utilized in the art. In some embodiments, a 2 ’-modification is 2’-OR, wherein R is optionally substituted Ci-io aliphatic. In some embodiments, a 2’-modification is 2’-OMe. In some embodiments, a 2’-modification is 2’-MOE. In some embodiments, a modified sugar is a bicyclic sugar (e.g., a sugar used in LNA, BNA, etc.). In some embodiments, in the context of oligonucleotides, a modified sugar is a sugar that is not ribose or deoxyribose as typically found in natural RNA or DNA.

[0118] The term “antisense”, as used herein, refers to a characteristic of an oligonucleotide or other nucleic acid having a base sequence complementary or substantially complementary to a target nucleic acid to which it is capable of hybridizing. In some embodiments, a target nucleic acid is a target gene mRNA. In some embodiments, hybridization is required for or results in at least one activity, e.g., a decrease in the level, expression or activity of the target nucleic acid or a gene product thereof. The term “antisense oligonucleotide”, as used herein, refers to an oligonucleotide complementary to a target nucleic acid. In some embodiments, an antisense oligonucleotide is capable of directing a decrease in the level, expression or activity of a target nucleic acid or a product thereof. In some embodiments, an antisense oligonucleotide is capable of directing a decrease in the level, expression or activity of the target nucleic acid or a product thereof, via a mechanism that involves RNase H, steric hindrance and / or RNA interference.

[0119] In one embodiment, the antisense oligonucleotides described herein comprise modification of their nucleic acid backbone, the ribose sugar moiety, and / or the nucleobase itself. In one embodiment, the antisense nucleotides described herein are chemically modified. Chemically modified nucleotides can be used to increase the binding affinity or stability of an antisense oligonucleotide. Chemical modifications include, but are not limited to phosphorothioate (PS) bond modifications. PS linkages involve replacement of one of the non-bridging oxygen atoms of the inter-nucleotide phosphate group with sulfur. Examples of nucleobase modifications include, but are not limited to, pyrimidine methylation. Examples of ribose sugar modifications include, but are not limited to, 2'-O-methyl (2'-0Me), 2'-O-methoxyethyl (2'-M0E), 2'-Fluoro (2'-F), locked nucleic acid (LNA) (See W02005002507, which is incorporated herein by reference in its entirety). LNA modification results in conformational constraint due to the connecting bridge and that of its methylated analog.

[0120] 15

[0121] 178599064.1 Docket No. 084284.00345

[0122] As used herein, “unmodified” or “natural” nucleobases include the purine bases guanine (G) and adenine (A), and the pyrimidine bases thymine (T), cytosine (C) and uracil (U). Modified nucleobases also referred herein as heterocyclic base moieties include other synthetic and natural nucleobases.

[0123] Antisense oligonucleotides are classified into three generations based on their modifications. First-generation antisense oligonucleotides comprise modification of the phosphate backbone linking the nucleotides. Specifically, one of the non-bridging oxygen atoms in the phosphodiester bond is replaced by a sulfur, methyl or amine group, generating phosphorothioates (PS), methyl-phosphonates, and phosphoramidates, respectively. Second-generation antisense oligonucleotides comprise alkyl modifications at the 2' position of the ribose. Lastly, third-generation antisense oligonucleotides comprise modifications that are not only nuclease resistant, but also improve binding affinity.

[0124] In some embodiments, provided are third-generation antisense oligonucleotides designed as gapmers with locked nucleic acid-modified nucleotides in the flanks, which increase the affinity of the antisense oligonucleotides to BMPER. These chemical modifications that increase stability and affinity to the target RNA, are phosphorothioated backbones and bridged nucleic acids (e.g., locked nucleic acid and constrained ethyl modifications). In contrast to earlier generations, third-generation antisense oligonucleotides do not require transfection reagents or conjugations for efficient target knockdown in vitro and in vivo without the use of complex formulations.

[0125] As used herein, the term “gapmer” refers to an oligonucleotide characterized in that it comprises a core flanked by a 5’ and a 3’ wing. In some embodiments, in a gapmer, at least one internucleotidic phosphorus linkage of the oligonucleotide is a natural phosphate linkage. In some embodiments, more than one internucleotidic phosphorus linkage of the oligonucleotide strand is a natural phosphate linkage. In some embodiments, a gapmer is a sugar modification gapmer, wherein each wing sugar independently comprises a sugar modification, and no core sugar comprises a sugar modification found in a wing sugar. In some embodiments, each core sugar comprises no modification and is 2’- unsubstituted (as in natural DNA). In some embodiments, each wing sugar is independently a 2’-modified sugar. In some embodiments, at least one wing sugar is a bicyclic sugar. In some embodiments, sugar units in each wing have the same sugar modification (e.g., 2’-OMe (a 2’-OMe wing), 2’-M0E (a 2’-M0E wing), etc.). In some 16

[0126] 178599064.1 Docket No. 084284.00345

[0127] embodiments, each wing sugar has the same modification. Core and wing can have various lengths. In some embodiments, a wing is 2, 3, 4, 5, 6, 7, 8, 9, 10 or more nucleosides (in many embodiments, 3, 4, 5, or 6 or more) in length, and a core is 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more nucleosides (in many embodiments, 8, 9, 10, 11, 12, or more) in length. In some embodiments, an oligonucleotide comprises or consists of a wing-core-wing structure of 2-9-6, 3- 9-3, 3-9-4, 3-9-5, 4-7-4, 4- 9-4, 4-9-5, 4-10-5, 4-11-4, 4-11-5, 5-7-5, 5-8-6, 5-9-3, 5-9-5, 5-10-4, 5-10-5, 6-7-6, 6-8-5, or 6-9-2. In some embodiments, an oligonucleotide is a gapmer.

[0128] Moreover, antisense oligonucleotides can be conjugated to various molecules (such as antibodies, peptides, fatty acids, sugars, etc.) to improve their delivery to specific tissues. Examples of such conjugates include, but are not limited to cell -penetrating peptides, sugars (e.g., GalNac), cholesterol moieties, and aptamers. Additionally, antisense oligonucleotides can be conjugated to fluorescent dyes as known in the art for tracking purposes.

[0129] The antisense oligonucleotides as described herein can be prepared using standard methods as known in the art. The antisense oligonucleotides are synthesized in vitro.

[0130] As used herein, the terms “decrease,” “reduce,” “downregulate,” and “inhibit” all generally refer to a decrease by a statistically significant amount. However, for avoidance of doubt, the term “reduced,” “decrease,” “reduce,” or “inhibit” means a decrease by at least 5% (e.g., 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%) as compared to a reference level, for example, a decrease by at least about 10%, a decrease by at least about 20%, or at least about 30%, or at least about 40%, or at least about 50%, or at least about 60%, or at least about 70%, or at least about 80%, or at least about 90% or up to and including a 100% decrease (e.g, absent level as compared to a reference sample), or any decrease of 10-100% as compared to a reference level. In some embodiments, these terms refer to a decrease of 10-20%, 10-30%, 10-40%, 10-50%, 10-60%, 10-70%, 10-80%, 10-90%, 10-100%, 10-110%, 10-120%, 10-130%, 10-140%, 10-150%, 10-160%, 10-170%, 10-180%, 10-190%, 10-200%, 10-210%, 10-220%, 10-230%, 10-240%, 10-250%, 10-260%, 10-270%, 10-280%, 10-290%, or 10-300%, as compared to a reference level. In some embodiments, these terms refer to a decrease of 10-300%, 20-300%, 30-300%, 40-300%, 50-300%, 60-300%, 70-300%, 80-300%, 90-300%, 100-300%, 110-300%, 120-300%, 130-300%, 140-300%, 150-300%, 160-300%, 170-300%, 180-300%, 190-300%, 200-300%, 210-300%, 220-300%, 230-300%, 240-300%, 250-300%, 260- 17

[0131] 178599064.1 Docket No. 084284.00345

[0132] 300%, 270-300%, 280-300%, or 290-300%, as compared to a reference level. In some embodiments, these terms refer to a decrease of 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, 70-80%, 80-90%, 90-100%, 100-110%, 110-120%, 120-130%, 130-140%, 140-150%, 150-160%, 160-170%, 170-180%, 180-190%, 190-200%, 200-210%, 210-220%, 220-230%, 230-240%, 240-250%, 250-260%, 260-270%, 270-280%, 280-290%, or 290-300%, as compared to a reference level.

[0133] In some embodiments, the expression level may include a mRNA or protein level.

[0134] As used herein, the “amount” or “level” of a biomarker is a detectable level or amount in a sample. These can be measured by methods known to one skilled in the art. These terms encompass a quantitative amount or level (e.g., weight or moles), a semi-quantitative amount or level, a relative amount or level (e.g, weight % or mole % within class), a concentration, and the like. Thus, these terms encompass absolute or relative amounts or levels or concentrations of a biomarker in a sample. The expression level or amount of biomarker assessed can be used to determine the response to treatment.

[0135] In some embodiments, the mRNA level is determined by at least one technique selected from reverse transcription polymerase chain reaction (RT-PCR), competitive RT-PCR, real-time RT-PCR, ribonucleic acid sequencing (RNA-seq), immunohistochemistry (IHC), immunofluorescence, RNaseprotection assay (RPA), northern blotting, and DNA chip.

[0136] In some embodiments, the protein level is determined by at least one technique selected from western blot, enzyme-linked immunosorbent assay (ELISA), radioimmunoassay (RIA), radioimmunodiffusion, Ouchterlony immunodiffusion, rocket immunoelectrophoresis, immunohistochemical staining, immunoprecipitation assay, complement fixation assay, fluorescence-activated cell sorter (FACS), and protein chip.

[0137] Pharmaceutical Comnositions and Kits

[0138] In some embodiments, provided are pharmaceutical compositions comprising an antisense oligonucleotide disclosed herein and a pharmaceutically acceptable carrier. As used herein, the term “pharmaceutical composition” refers to a combination of substances suitable for administering to an individual. For example, a pharmaceutical composition may comprise an antisense oligonucleotide and a sterile aqueous solution.

[0139] 18

[0140] 178599064.1 Docket No. 084284.00345

[0141] In some embodiments, the pharmaceutical composition is a nanoparticle. In some embodiments, the pharmaceutical composition is a liposome composition. In some embodiments, the pharmaceutical composition includes, but is not limited to lipid-based formulations for antisense oligonucleotide delivery such as endogenously derived exosomes, spherical nucleic acids (SNAs) and self-assembling DNA nanostructures.

[0142] In some embodiments, provided are pharmaceutical compositions that further comprise emulsions and / or surfactants. Emulsions comprise two immiscible liquid phases intimately mixed and dispersed with each other. Emulsions can have little or no thermodynamic stability. There are different types of emulsions such as oil in water (O / W) emulsion and emulsion water in oil (W / O). Emulsion oil in water (O / W) comprises an oil phase dispersed in an aqueous one (direct emulsion). Emulsion water in oil (W / O) comprises an aqueous phase dispersed in the oil phase. In some embodiments, the emulsions are microemulsions. Microemulsions form spontaneously and are thermodynamically stable. Emulsions and their uses are further described in U. S. Pat. No.

[0143] 6,287,860, which is incorporated herein in its entirety.

[0144] Surfactants form micelles in a solution and adsorb to the interface between a solution and a different phase (gases / solids). Generally, surfactants are amphiphilic and contain a hydrophobic portion and a hydrophilic portion. Surfactants are classified into different classes based on the nature of the hydrophilic group: anionic, nonionic, cationic, and amphoteric. Examples of surfactants include, but are not limited to Brij 96, polyoxyethylene oleyl ethers, polyglycerol fatty acid esters, tetraglycerol monolaurate (ML310), tetraglycerol monooleate (MO310), hexaglycerol monooleate (PO310), hexaglycerol pentaoleate (PO500), decaglycerol monocaprate (MCA750), decaglycerol monooleate (MO750), decaglycerol sequioleate (SO750), and decaglycerol decaoleate (DAO750). Surfactants and their uses are further described in U. S. Pat. No. 6,287,860, which is incorporated herein in its entirety.

[0145] A “pharmaceutically acceptable carrier,” after administered to or upon a subject, does not cause undesirable physiological effects. The carrier in the composition must be “acceptable” also in the sense that it is compatible with the active ingredient and is capable of stabilizing it. One or more solubilizing agents can be utilized as pharmaceutical carriers for the delivery of the abovedescribed composition. Examples of a pharmaceutically acceptable carrier include, but are not limited to, biocompatible vehicles, adjuvants, additives, and diluents to achieve a composition 19

[0146] 178599064.1 Docket No. 084284.00345

[0147] usable as a dosage form. Examples of other carriers include colloidal silicon oxide, magnesium stearate, cellulose, and sodium lauryl sulfate.

[0148] The pharmaceutical compositions described herein can be administered in various ways including, but not limited to subcutaneous administration, topical administration (e.g., transdermal patches, ointments, lotions, creams, gels, drops, etc.), parenteral administration, and intravenous administration. The method of administration of the pharmaceutical compositions described herein depends on whether systemic or local treatment is desired and upon the area to be treated.

[0149] The pharmaceutical compositions described herein can be prepared according to conventional techniques well known in the pharmaceutical industry. For example, the active ingredients are brought into association with the pharmaceutical carrier(s) or excipient(s).

[0150] The present disclosure provides a kit with packaging material and an antisense oligonucleotide as described herein. The kit can further comprise instructions for use. Specifically, the kit can include a label or packaging insert that describes the components.

[0151] A kit refers to a physical structure that contains one or more components of the kit. Packaging material can maintain the components in a sterile manner and can be made of material commonly used for such purposes (e.g., paper, glass, plastic, foil, ampules, vials, tubes, etc.).

[0152] A label or insert can include identifying information of one or more components therein, dose amounts, and clinical pharmacology of the active ingredients(s) including mechanism of action, pharmacokinetics, and pharmacodynamics. A label or insert can include information identifying manufacture, lot numbers, manufacture location and date, and expiration dates. A label or insert can include information on a glomerular disease for which a kit component may be used. A label or insert can include instructions for a clinician or subject for using one or more of the kit components in a method, use or treatment protocol, or therapeutic regimen. Instructions can include dosage amounts, frequency of duration, and instructions for practicing any of the methods, uses, treatment protocols, or therapeutic regimens described herein.

[0153] A label or insert can include information on potential adverse side effects, complications, or reactions, such as a warning to a subject or clinician regarding situations where it would not be appropriate to use a particular composition.

[0154] 20

[0155] 178599064.1 Docket No. 084284.00345

[0156] In some embodiments, the label or insert includes information on the suitable subjects that can be treated with the composition.

[0157] Methods of Treatment

[0158] Provided are methods of treating a glomerular disease in a subject in need thereof using the antisense oligonucleotides or pharmaceutical compositions described herein. In some embodiments, the glomerular disease is chronic kidney disease (CKD). In some embodiments, the antisense oligonucleotides or pharmaceutical compositions block renal microvasculature derived BMPER and therefore prevent its pathologic interaction with neighboring podocytes preventing their depletion, improving glomerular filtration barrier function in CKD. In some embodiments, the subject has primary or secondary focal segmental glomerulosclerosis (FSGS). Primary (idiopathic) FSGS occurs on its own without a known cause. Secondary FSGS is caused by another disease or a drug (e.g., HIV infection, diabetes, lupus, sickle cell disease, etc.). In some embodiments, the subject has diabetes. In some embodiments, the subject is a human.

[0159] As used herein, the terms “treat,” “treatment,” “treating,” and “amelioration” refer to therapeutic treatments, wherein the object is to reverse, alleviate, ameliorate, inhibit, slow down or stop the progression or severity of a condition associated with a disease or disorder. The term “treating” includes reducing or alleviating at least one adverse effect or symptom of a condition, disease or disorder associated with a disorder. Treatment is generally “effective” if one or more symptoms or clinical markers are reduced. Alternatively, treatment is “effective” if the progression of a disease is reduced. That is, “treatment” includes not just the improvement of symptoms or markers, but also a slowing of progress or worsening of symptoms compared to what would be expected in the absence of treatment. Beneficial or desired clinical results include, but are not limited to, alleviation of one or more symptom(s), diminishment of extent of disease, stabilized (i.e., not worsening) state of disease, delay or slowing of disease progression, amelioration or palliation of the disease state, remission (whether partial or total), and / or decreased mortality, whether detectable or undetectable. The term “treatment” of a disease also includes providing relief from the symptoms or side-effects of the disease (including palliative treatment).

[0160] The antisense oligonucleotides and pharmaceutical compositions described herein that target BMPER can be delivered to a subject via various modes of delivery. For example, the

[0161] 21

[0162] 178599064.1 Docket No. 084284.00345

[0163] antisense oligonucleotides and pharmaceutical compositions can be administered to a subject subcutaneously, intravenously, parenterally, intraperitoneally or systemically. In particular embodiments, the antisense oligonucleotides and pharmaceutical compositions described herein are administered to the subject intravenously or subcutaneously.

[0164] In some embodiments, the antisense oligonucleotides and pharmaceutical compositions described herein can be administered to a subject with an additional therapeutic agent. In some embodiments, the antisense oligonucleotides and the additional therapeutic agent are administered simultaneously. In some embodiments, the antisense oligonucleotides are administered before the additional therapeutic agent. In some embodiments, the additional therapeutic agent is administered before the antisense oligonucleotides.

[0165] In some embodiments, one or more doses of the antisense oligonucleotides described herein are administered to a subject ranging from about 0.0001 pg to about 100 g per kg of body weight of the subject such as 0.0001 pg per body weight, 1 pg per body weight, 50 pg per body weight, 100 pg per body weight, 200 pg per body weight, 300 pg per body weight, 400 pg per body weight, 500 pg per body weight, 600 pg per body weight, 700 pg per body weight, 800 pg per body weight, 900 pg per body weight, 1,000 pg per body weight, 10,000 pg per body weight, 50,000 pg per body weight, 100,000 pg per body weight, 200,000 pg per body weight, 300,000 pg per body weight, 400,000 pg per body weight, 500,000 pg per body weight, 600,000 pg per body weight, 700,00 pg per body weight, 800,000 pg per body weight, 900,000 pg per body weight, 1 g per body weight, 10 g per body weight, 25 g per body weight, 50 g per body weight, 75 g per body weight, 100 g per body weight, or any amount in-between.

[0166] In some embodiments, at least one maintenance dose of the antisense oligonucleotides described herein is administered. A maintenance dose is a dose of the antisense oligonucleotides administered to the subject following a decrease in the level of expression or activity of BMPER to maintain said decrease in expression or activity. The number of maintenance doses depends on the level of expression or activity of BMPER that a subject exhibits and can be determined by a physician. In some embodiments, a maintenance dose is administered one month, two months, three months, four months, five months, six months, seven months, eight months, nine months, ten months, eleven months, one year, two years, three years or any time in-between or longer following the initial dose of the antisense oligonucleotides. In some embodiments, the at least one 22

[0167] 178599064.1 Docket No. 084284.00345

[0168] maintenance dose of the antisense oligonucleotides described herein are administered to a subject ranging from about 0.0001 pg to about 100 g per kg of body weight of the subject such as 0.0001 pg per body weight, 1 pg per body weight, 50 pg per body weight, 100 pg per body weight, 200 pg per body weight, 300 pg per body weight, 400 pg per body weight, 500 pg per body weight, 600 pg per body weight, 700 pg per body weight, 800 pg per body weight, 900 pg per body weight, 1,000 pg per body weight, 10,000 pg per body weight, 50,000 pg per body weight, 100,000 pg per body weight, 200,000 pg per body weight, 300,000 pg per body weight, 400,000 pg per body weight, 500,000 pg per body weight, 600,000 pg per body weight, 700,00 pg per body weight, 800,000 pg per body weight, 900,000 pg per body weight, 1 g per body weight, 10 g per body weight, 25 g per body weight, 50 g per body weight, 75 g per body weight, 100 g per body weight, or any amount in-between.

[0169] As used herein, a “subject” or “individual” means a human or animal. Usually, the animal is a vertebrate such as a primate, rodent, domestic animal or game animal. Primates include chimpanzees, cynomolgus monkeys, spider monkeys, and macaques, e.g., Rhesus. Rodents include mice, rats, woodchucks, ferrets, rabbits and hamsters. Domestic and game animals include cows, horses, pigs, sheep, goats, deer, bison, buffalo, feline species, e.g., domestic cat, canine species, e.g., dog, fox, wolf, avian species, e.g., chicken, emu, ostrich, and fish, e.g., trout, catfish and salmon. In some embodiments, the subject is a mammal, e.g., a human or a non-human mammal. The mammal can be a human, non-human primate, mouse, rat, dog, cat, horse, or cow, but is not limited to these examples. Mammals other than humans can be advantageously used as subjects that represent animal models of disorders. The terms “individual,” “patient” and “subject” can be used interchangeably herein. A subject can be male or female. In one embodiment, the subject is a human. In another embodiment, the subject is an experimental, non-human animal or animal suitable as a disease model.

[0170] A “subject in need” of treatment for a particular condition or disorder can be a subject having that condition or disorder, diagnosed as having that condition or disorder, or at risk of developing that condition or disorder.

[0171] 23

[0172] 178599064.1 Docket No. 084284.00345

[0173] Definitions

[0174] To aid in understanding the detailed description of the compositions and methods according to the disclosure, some express definitions are provided to facilitate an unambiguous disclosure of the various aspects of the disclosure. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.

[0175] As used herein, the percent homology between two amino acid sequences is equivalent to the percent identity between the two sequences. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences ( / .<?., % homology = # of identical positions / total # of positions x 100), taking into account the number of gaps, and the length of each gap, which need to be introduced for optimal alignment of the two sequences. The comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm, as described in the non-limiting examples below.

[0176] The percent identity between two amino acid sequences can be determined using the algorithm of E. Meyers and W. Miller (Comput. Appl. Biosci., 4:11-17 (1988)) which has been incorporated into the ALIGN program (version 2.0), using a PAM120 weight residue table, a gap length penalty of 12 and a gap penalty of 4. In addition, the percent identity between two amino acid sequences can be determined using the Needleman and Wunsch (J. Mol. Biol. 48:444-453 (1970)) algorithm which has been incorporated into the GAP program in the GCG software package (available at gcg.com), using either a Blossum 62 matrix or a PAM250 matrix, and a gap weight of 16, 14, 12, 10, 8, 6, or 4 and a length weight of 1, 2, 3, 4, 5, or 6.

[0177] The terms “sample” or “biological sample,” as used herein, include any biological specimen obtained (isolated, removed) from a subject. Samples may include, without limitation, organ tissue, whole blood, plasma, serum, whole blood cells, red blood cells, white blood cells (e.g., peripheral blood mononuclear cells), saliva, urine, stool (feces), tears, sweat, sebum, nipple aspirate, ductal lavage, synovial fluid, cerebrospinal fluid, lymph, fine needle aspirate, amniotic fluid, any other bodily fluid, exudate or secretory fluid, cell lysates, cellular secretion products, inflammation fluid, semen, and vaginal secretions. In some embodiments, a sample may be readily obtainable by non-invasive or minimally invasive methods, such as blood collection (“liquid

[0178] 24

[0179] 178599064.1 Docket No. 084284.00345

[0180] biopsy”), urine collection, feces collection, tissue biopsy or fine-needle aspiration, allowing the provision / removal / isolation of the sample from a subject. The term “tissue,” as used herein, encompasses all types of cells of the body, including cells of organs but also including blood and other body fluids recited above. The tissue may be healthy or affected by pathological alterations.

[0181] The term “agent” is used herein to denote a chemical compound, a mixture of chemical compounds, a biological macromolecule (such as a nucleic acid, an antibody, a protein or portion thereof, e.g., a peptide), or an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues. The activity of such agents may render it suitable as a “therapeutic agent,” which is a biologically, physiologically, or pharmacologically active substance (or substances) that acts locally or systemically in a subject.

[0182] The terms “therapeutic agent,” “therapeutic capable agent,” or “treatment agent” are used interchangeably and refer to a molecule or compound that confers some beneficial effect upon administration to a subject. The beneficial effect includes enablement of diagnostic determinations; amelioration of a disease, symptom, disorder or pathological condition; reducing or preventing the onset of a disease, symptom, disorder or condition; and generally counteracting a disease, symptom, disorder or pathological condition.

[0183] “Combination” therapy, as used herein, unless otherwise clear from the context, is meant to encompass administration of two or more therapeutic agents in a coordinated fashion and includes, but is not limited to, concurrent dosing. Specifically, combination therapy encompasses both co-admini strati on (e.g., administration of a co-formulation or simultaneous administration of separate therapeutic compositions) and serial or sequential administration, provided that administration of one therapeutic agent is conditioned in some way on administration of another therapeutic agent. For example, one therapeutic agent may be administered only after a different therapeutic agent has been administered and allowed to act for a prescribed period of time. See, e.g., Kohrt et al. (2011) Blood 117:2423.

[0184] Doses are often expressed in relation to bodyweight. Thus, a dose which is expressed as [g, mg, or other unit] / kg (or g, mg etc.) usually refers to [g, mg, or other unit] “per kg (or g, mg etc.) body weight,” even if the term “bodyweight” is not explicitly mentioned.

[0185] 25

[0186] 178599064.1 Docket No. 084284.00345

[0187] As used herein and in the appended claims, the singular forms “a,” “and,” and “the” include plural references unless the context clearly dictates otherwise.

[0188] The term “about” refers to a range of values which would not be considered by a person of ordinary skill in the art as substantially different from the baseline values. For example, the term “about” may refer to a value that is within 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.05%, or 0.01% of the stated value, as well as values intervening such stated values.

[0189] As disclosed herein, a number of ranges of values are provided. It is understood that each intervening value, to the tenth of the unit of the lower limit, unless the context clearly dictates otherwise, between the upper and lower limits of that range is also specifically disclosed. Each smaller range between any stated value or intervening value in a stated range and any other stated or intervening value in that stated range is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included or excluded in the range, and each range where either, neither, or both limits are included in the smaller ranges is also encompassed within the invention, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the invention.

[0190] As used herein, the term “each,” when used in reference to a collection of items, is intended to identify an individual item in the collection, but does not necessarily refer to every item in the collection. Exceptions can occur if explicit disclosure or context clearly dictates otherwise.

[0191] As used herein, the terms “including,” “comprising,” “containing,” or “having” and variations thereof are meant to encompass the items listed thereafter and equivalents thereof as well as additional subject matter unless otherwise noted.

[0192] The use of any and all examples, or exemplary language (e.g., “such as”) provided herein, is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0193] 26

[0194] 178599064.1 Docket No. 084284.00345

[0195] As used herein, the phrases “in one embodiment,” “in various embodiments,” “in some embodiments,” and the like are used repeatedly. Such phrases do not necessarily refer to the same embodiment, but they may unless the context dictates otherwise.

[0196] As used herein, the terms “and / or” or “ / ” means any one of the items, any combination of the items, or all of the items with which this term is associated.

[0197] All methods described herein are performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. In regard to any of the methods provided, the steps of the method may occur simultaneously or sequentially. When the steps of the method occur sequentially, the steps may occur in any order, unless noted otherwise. In cases in which a method comprises a combination of steps, each and every combination or sub-combination of the steps is encompassed within the scope of the disclosure, unless otherwise noted herein.

[0198] All references, patents and applications cited herein are incorporated herein by reference in their entireties. Furthermore, where a definition or use of a term in a reference, which is incorporated by reference herein is inconsistent or contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.

[0199] To facilitate a better understanding of the present invention, the following examples of specific embodiments are given. The following examples should not be read to limit or define the entire scope of the invention.

[0200] EXAMPLES

[0201] Example 1: BMPER is pathogenic and a therapeutic target for CKD in vivo.

[0202] An unbiased state of the art proteomics study of the mouse glomerular endothelial cell secretome after exposure to physiologic stress identified a panel of proteins including BMPER, that are present specifically in supernatant fractions with the most pro-apoptotic activity derived from the stressed glomerular endothelial cells (GECs). Although BMPER is a secreted protein, an increase in BMPER mRNA and protein in glomeruli from diabetic mice with CKD and not in nondiabetic controls was detected, as well as in GECs treated with Ednl or high glucose. Increased BMPER expression in glomeruli endothelium and in larger vessels was confirmed in kidneys from a biopsy cohort of patients with CKD compared to normal biopsy nephrectomy controls (FIGS.

[0203] 27

[0204] 178599064.1 Docket No. 084284.00345

[0205] 1A-1C). Also, BMPER circulating levels were elevated in patients with early CKD (mean GFR 90ml / min; FIG. ID), that were shown to progress after 2 years (Nadkarni, G. N., et al. Association of Urinary Biomarkers of Inflammation, Injury, and Fibrosis with Renal Function Decline: The ACCORD Trial. Clin J Am Soc Nephrol 11, 1343-1352 (2016)). Levels of urine BMPER normalized for creatinine were significantly higher in cases with progressive CKD compared to controls, and diabetic non-progressors (FIG. IE). Thus, the data indicate that BMPER is upregulated in stressed / injured GECs in patients with progressive CKD. The effect of silencing BMPER in GECs in experimental CKD was then determined. For these studies, replicationincompetent 3rdgeneration lentivirus BMPER shRNA was used, and showed reduced proteinuria in mice with progressive CKD (PodTgfbrl+ Dox for 7 days; FIG.2 grey line). These data indicate that BMPER is pathogenic and a therapeutic target for CKD in vivo.

[0206] Example 2. Blocking BMPER with anti-sense oligonucleotides is a novel CKD therapy.

[0207] Antisense oligonucleotides (ASOs) have emerged as a powerful therapeutic for a number of diseases (Hong, D. et al. Science Translational Medicine 7, 314ral85-314ral85 (2015)). In this study, third-generation ASOs were designed as gapmers with locked nucleic acid-modified nucleotides in the flanks, which increase the affinity of the ASO to the target; BMPER expression and secretion by stressed GECs in the kidney in disease settings. These chemical modifications that increase stability and affinity to the target RNA are phosphorothioated backbones and bridged nucleic acids (e.g., locked nucleic acid and constrained ethyl modifications). In contrast to earlier generations, third-generation ASOs do not require transfection reagents or conjugations for efficient target knockdown in vitro and in vivo without the use of complex formulations. ASOs can be administered systemically, for example, by intravenous or subcutaneous injection.

[0208] ASOs have a single stranded DNA backbone, freely enter cells by endocytic pathways and form double stranded RNA-DNA hybrids that induce RNA cleavage by endogenous RNase H27, thereby degrading the target, i.e., BMPER. Once inside the cells, ASOs in vivo can exhibit long half-lives (1- 4 weeks) and prolonged biological activity (Jansson-Lofmark, R et al. Nucleic Acid Ther 28, 319-325 (2018)). Phosphorothioate bond modifications were included in the ASOs to reduce nuclease degradation and to increase their stability, and a ‘gapmer’ was designed to enable activation of RNase H through a recognizable sequence of 10 bases that are flanked by 3 locked nucleic acids or O-methoxy ethyl bases on each side (3-10-3). Four effective candidate ASOs were 28

[0209] 178599064.1 Docket No. 084284.00345

[0210] identified against human and mouse BMPER that show 75%-90% reduction of PAN (stressor) induced BMPER in mGECs (FIG.3).

[0211] Next a comprehensive animal study was performed to determine the effect of various ASOs on mouse models of CKD with proteinuria. Subcutaneous or intravenous delivery of ASOs results in high levels of biodistribution to liver and kidney (Hung, G. N. et al. Nucleic Acid Ther 23, 369-378 (2013)). It was hypothesized that inhibition of GEC derived BMPER with ASOs prevents podocyte depletion and CKD progression. Thus, the next study aimed to determine the effect of BMPER ablation with ASOs in experimental models of progressive CKD.

[0212] Studies were performed with a transgenic mouse model and genetic models of T2DM (db / db). For the diabetes study, mice were injected with 50 mg / kg ASOs, every 4 weeks as described in Shahzad, K. et al. CHOP-ASO Ameliorates Glomerular and Tubular Damage on Top of ACE Inhibition in Diabetic Kidney Disease. Journal of the American Society of Nephrology 32, 3066-3079 (2021). (FIG.4A). db / db mice had increased fasting blood glucose levels compared to db / m controls. ASOs did not impact fasting blood glucose levels at any time point tested compared to db / db NC5 scrambled control (FIG. 4B). db / db mice had increased body weight compared to db / m controls. ASO70 did not impact body weight. However, there was a significant reduction in body weight in ASO1982 treated mice after 4 weeks, and levels reduced over time (FIG. 4C). After 16 weeks of diabetes, both ASO70 and ASO1982 significantly reduced albuminuria and improved the histopathology in these mice compared to ASONC5 control (FIGS.

[0213] 5A and 5B).

[0214] In a glomerular disease model, the Nephrin-activated TGFbRl in podocytes FSGS disease model (Daehn, I. et al. J Clin Invest 124, 1608-1621 (2014); Ebefors, K. et al. KI 2019), mice were fed doxycycline food (Dox) to induce TGFbRl transgene activation in podocytes and were injected with vehicle or with ASONC5, ASO70, and ASO1982. This is a severe model of proteinuria. Mice were injected with 20 mg / kg of ASOs on 3 consecutive days, similar to the approach by Aghajan et al. using ASOs against APOL1 (Aghajan, M. et al. Jci Insight 4 (2019)). Both ASO70 and ASO1982 reduced BMPER expression and glomerular histopathology. ASO70 also significantly reduced albuminuria in this model. In another glomerular disease model of FSGS, adriamycin (ADR; lOmg / kg) induced nephropathy in Balb / c mice, mice were injected with vehicle or with ASONC5, ASO70 as described above. In this model of glomerular disease ASO70 reduced 29

[0215] 178599064.1 Docket No. 084284.00345

[0216] BMPER expression and glomerular histopathology, and significantly reduced albuminuria (FIGS.

[0217] 7A-7D). Therefore, in an experimental model of diabetic kidney disease and two models of FSGS with severe proteinuria ASOs targeting BMPER significantly ameliorated disease (FIGS. 6A-6C and 7A-7D).

[0218] While the foregoing written description of the invention enables one of ordinary skill to make and use what is considered presently to be the best mode thereof, those of ordinary skill will understand and appreciate the existence of variations, combinations, and equivalents of the specific embodiments, methods, and examples herein.

[0219] 30

[0220] 178599064.1

Claims

Docket No. 084284.00345CLAIMSWhat is claimed is:

1. An antisense oligonucleotide that specifically hybridizes to a nucleic acid encoding BMP binding endothelial regulator (BMPER), wherein the oligonucleotide decreases a level of expression or activity of BMPER.

2. The antisense oligonucleotide of claim 1, wherein the oligonucleotide is unmodified.

3. The antisense oligonucleotide of claim 1, wherein the oligonucleotide is modified.

4. The antisense oligonucleotide of claim 3, wherein the modification comprises at least one modified internucleotide linkage.

5. The antisense oligonucleotide of claim 4, wherein the modified internucleotide linkage is a phosphorothioate linkage.

6. The antisense oligonucleotide of claim 5, wherein the modification further comprises at least a locked nucleic acid (LNA), a lipid, a peptide, an aptamer, an antibody or antigen-binding fragment thereof, or a sugar.

7. The antisense oligonucleotide of claim 1, wherein the oligonucleotide comprises a polynucleotide sequence of any one of SEQ ID NOs: 1-11 or comprises a polynucleotide sequence having at least 75% sequence identity to any one of SEQ ID NOs: 1-11.

8. The antisense oligonucleotide of claim 1, wherein the oligonucleotide consists of 16 linked nucleotides.

9. The antisense oligonucleotide of claim 1, wherein the nucleic acid encodes human BMPER.31178599064.1Docket No. 084284.0034510. A pharmaceutical composition comprising the antisense oligonucleotide of any one of the preceding claims and optionally a pharmaceutically acceptable carrier or diluent.

11. The pharmaceutical composition of claim 10, wherein the composition is a liposome composition.

12. A kit comprising the antisense oligonucleotide of any one of claims 1-10.

13. A method of treating a glomerular disease in a subject in need thereof, comprising administering to the subject one or more doses of the antisense oligonucleotide of any one of claims 1-9 or the pharmaceutical composition of claim 10.

14. The method of claim 13, wherein the subject is a human.

15. The method of claim 13, wherein the glomerular disease is chronic kidney disease (CKD).

16. The method of claim 13, wherein the antisense oligonucleotide is administered to the subject parenterally.

17. The method of claim 13, wherein the antisense oligonucleotide is administered to the subject intravenously.

18. The method of claim 13, wherein the subject has primary or secondary focal segmental glomerulosclerosis (FSGS).

19. The method of claim 13, wherein the one or more doses of the antisense oligonucleotide comprises about 0.0001 μg to about 100 g per kg of body weight of the subject.32178599064.1

Images