COMPOUNDS AND METHODS TARGETING HUMAN AND MOUSE INSL5
The present invention relates to the field of medicine. More particularly, the
present invention relates to compounds, diagnostics, and methods which include an antibody or
5 fragment thereof directed against human or mouse insulin-like peptide-5 (INSL5). The
compounds and methods of the present invention are expected to be useful in the
field of oncology, reproductive, and metabolic diseases, e.g., diabetes and obesity,
including diagnostics relating thereto.
The gut hormone INSL5 belongs to the Relaxin/Insulin-like family of peptides.
10 Like the other INSL family members, INSL5 is composed ofB- and A-chains connected
by two disulfide bonds. INSL5 is expressed in various tissues; however, it is primarily cosecreted
with other hormones from specialized enteroendocrine cells (EECs), called L
cells, in the distal intestine, particularly in the colon and rectum. These cells regulate
metabolic and physiological processes such as intestinal motility, hormone secretion,
15 glucose homeostasis and appetite. INSL5 is the ligand for the receptor
RXFR4/GPCR142/GPR100 and its receptor-ligand interaction results in the inhibition of
intracellular cAMP levels. While INSL5 is considered anorexigenic (appetitestimulating)
hormone and its secretion is elevated by calorie restriction, the biological
consequences of the INSL5-RXFP4 activated signaling pathway remain largely elusive.
20 In order to study INSL5, reliable and sensitive assays for INSL5 are needed.
INSL5 has been studied in patients with metabolic dysfunction, polycystic ovarian
syndrome (PCOS), colorectal cancer (CRC), and infertility, and preclinical models of
these diseases. However, there remains a need to provide antibodies that bind human
and mouse INSL5 to study these effects further. In particular, there remains a need for
25 INSL5 antibodies with high affinities to both human and mouse INSL5. Currently
available assays have sensitivity, reproducibility, performance, and/or quantitative
issues. Thus, additional methods of testing human or mouse samples are needed.
In a specific embodiment of the present disclosure, antibodies are provided which
bind INSL5 comprising a heavy chain variable region (VH) and a light chain variable
30 region (VL), wherein the VH comprises heavy chain complementarity determining
3
wo 2022/031674 PCT/US2021/044304
2
regions (HCDR) HCDR1, HCDR2, and HCDR3, and the VL comprises light chain
complementarity determining regions (LCDR) LCDR1, LCDR2, and LCDR3, wherein
the HCDR1 comprises TASGFSLSSYDNIG (SEQ ID NO: 10), the HCDR2 comprises
TlSAGGYTY (SEQ ID NO: 11), the HCDR3 comprises
5 ARERWNYDRSGGAGAGYFDL (SEQ ID NO: 12), the LCDR1 comprises
QASQSITSSYLS (SEQ ID N0:14), the LCDR2 comprises YPAANLAS (SEQ ID
N0:15), and the LCDR3 comprises LYGYFSSSIDFA (SEQ ID N0:16).
According to some embodiments, antibodies of the present disclosure comprise a VH that
comprises SEQ ID N0:9 and a VL which comprises SEQ ID NO: 13. According to some
10 embodiments, the antibodies of the present disclosure comprise a heavy chain (HC)
comprising SEQ ID N0:5 and a light chain (LC) comprising SEQ ID N0:7. According to
some embodiments, antibodies of the present disclosure comprise a heavy chain (HC)
comprising amino acids 2-446 of SEQ ID N0:5, and a light chain (LC) comprising SEQ
ID N0:7. According to some embodiments, the antibodies of the present disclosure
15 comprise a HC consisting of SEQ ID N0:5 and a LC consisting of SEQ ID N0:7.
According to some embodiments, the present disclosure provides a nucleic acid
comprising a sequence encoding SEQ ID N0:5 or SEQ ID N0:7, or both. According to
some embodiments, one or more vectors comprise one or more nucleic acids encoding
SEQ ID N0:5 or SEQ ID N0:7, or both. According to some embodiments, the present
20 disclosure provides a composition comprising a first vector comprising a nucleic acid
sequence encoding SEQ ID N0:5 and a second vector comprising a nucleic acid sequence
encoding SEQ ID N0:7. According to other embodiments, the present disclosure
provides a composition comprising a vector comprising a nucleic acid encoding SEQ ID
N0:5 and SEQ ID N0:7. In another embodiment, the present disclosure provides a cell
25 comprising one or more vectors comprising one or more nucleic acids encoding SEQ ID
N0:5 or SEQ ID N0:7, or both. According to some embodiments, the present disclosure
provides a cell comprising a first vector comprising a nucleic acid sequence encoding
SEQ ID N0:5 and a second vector comprising a nucleic acid sequence encoding SEQ ID
N0:7. According to other embodiments, the present disclosure provides a cell
30 comprising a vector comprising a nucleic acid encoding SEQ ID N0:5 and a second
4
wo 2022/031674 PCT/US2021/044304
3
nucleic acid sequence encoding SEQ ID N0:7. In further embodiments, the cell is a
mammalian cell.

WE CLAIM:
1. An antibody that binds INSL5 comprising a heavy chain variable region (VH) and a
light chain variable region (VL), wherein the VH comprises heavy chain complementarity
determining regions (HCDR) HCDR1, HCDR2, and HCDR3, and the VL comprises light
5 chain complementarity determining regions (LCDR) LCDR1, LCDR2, and LCDR3,
wherein
the HCDR1 comprises TASGFSLSS\'DMG (SEQ ID NO: 10),
the HCDR2 comprises TISAGGYTY (SEQ ID NO: 11),
the HCDR3 comprises ARERv1lN\TIRSGGAGAG'{FDL (SEQ ID NO: 12),
10 the LCDR1 comprises QASQSITSSYLS (SEQ ID NO: 14),
the LCDR2 comprises YJ> AANLAS (SEQ ID NO: 15), and
the LCDR3 comprises LYGYFSSSlDF A (SEQ ID NO: 16).
2. The antibody of claim 1, wherein the VH comprises SEQ ID N0:9 and the VL
15 comprises SEQ IDNO:l3.
3. The antibody of claim 1 or 2, wherein the antibody comprises a heavy chain (HC)
comprising SEQ ID N0:5 and a light chain (LC) comprising SEQ ID N0:7.
20 4. The antibody of claim 1 or 2, wherein the antibody comprises a heavy chain (HC)
comprising amino acids 2-446 of SEQ ID N0:5, and a light chain (LC) comprising SEQ
IDN0:7.
5. The antibody of claim 1 or 2, wherein the antibody comprises a HC consisting of SEQ
25 ID N0:5 and a LC consisting of SEQ ID N0:7.
6. The antibody of any one of claims 1-5 that binds to human INSL5.
7. The antibody of any one of claims 1-5 wherein the antibody binds to the A chain (SEQ
30 ID NO: 1) and the B chain (SEQ ID N0:2) of human INSL5.
29
5
wo 2022/031674 PCT/US2021/044304
28
8. The antibody of any one of claims 1-5 that binds to mouse INSL5.
9. The antibody of any one of claims 1-5 wherein the antibody binds to the A chain (SEQ
ID N0:3) and the B chain (SEQ ID N0:4) of mouse INSL5.
10. A nucleic acid comprising a sequence encoding SEQ ID N0:5 or SEQ ID N0:7.
11. A vector comprising the nucleic acid of claim 10.
10 12. A composition comprising a first vector comprising a nucleic acid sequence encoding
SEQ ID N0:5 and a second vector comprising a nucleic acid sequence encoding SEQ ID
N0:7.
13. A cell comprising:
15 (a) a first vector comprising a nucleic acid sequence encoding SEQ ID N0:5 and a second
vector comprising a nucleic acid sequence encoding SEQ ID N0:7; or
(b) a vector comprising a first nucleic acid sequence encoding SEQ ID NO: 5 and a
second nucleic acid sequence encoding SEQ ID NO: 7.
20 14. The cell of claim 13, wherein the cell is a mammalian cell.
25
15. A process of producing an antibody comprising culturing the cell of any one of claims
13-14 under conditions such that the antibody is expressed and recovering the expressed
antibody from the culture medium.
16. An antibody produced by culturing the cell of any one of claims 13-14 under
conditions such that the antibody is expressed and recovering the expressed antibody
from the culture medium.
30
wo 2022/031674 PCT/US2021/044304
29
17. An antibody that binds INSL5 comprising a VH and a VL, wherein the VH comprises
HCDR1, HCDR2, and HCDR3, and the VL comprises LCDR1, LCDR2, and LCDR3,
wherein
the HCDR1 comprises TVSGIDLTTYAMG (SEQ ID N0:22),
5 the HCDR2 comprises IIGGGGRTY (SEQ ID N0:23),
10
the HCDR3 comprises VRGGDFFDL (SEQ ID N0:24),
the LCDR1 comprises QASEDISKYLS (SEQ ID N0:26),
the LCDR2 comprises YYVS:NL.EF (SEQ ID N0:27), and
the LCDR3 comprises HQGYTGVNVENV (SEQ ID N0:28).
18. The antibody of claim 17, wherein the VH comprises SEQ ID N0:21 and the VL
comprises SEQ ID N0:25.