WO2021092140A1 - Méthodes de réduction du lactate chez des patients malades du foie au moyen de variants et de fusions de polypeptides fgf19/fgf21 - Google Patents

Méthodes de réduction du lactate chez des patients malades du foie au moyen de variants et de fusions de polypeptides fgf19/fgf21 Download PDF

Info

Publication number
WO2021092140A1
WO2021092140A1 PCT/US2020/059060 US2020059060W WO2021092140A1 WO 2021092140 A1 WO2021092140 A1 WO 2021092140A1 US 2020059060 W US2020059060 W US 2020059060W WO 2021092140 A1 WO2021092140 A1 WO 2021092140A1
Authority
WO
WIPO (PCT)
Prior art keywords
seq
amino acid
sequence
fgf19
terminal region
Prior art date
Application number
PCT/US2020/059060
Other languages
English (en)
Inventor
Lei Ling
Alexander Mark DEPAOLI
Original Assignee
Ngm Biopharmaceuticals, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Ngm Biopharmaceuticals, Inc. filed Critical Ngm Biopharmaceuticals, Inc.
Publication of WO2021092140A1 publication Critical patent/WO2021092140A1/fr

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/18—Growth factors; Growth regulators
    • A61K38/1825—Fibroblast growth factor [FGF]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/475—Growth factors; Growth regulators
    • C07K14/50—Fibroblast growth factor [FGF]
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
    • C12N15/62—DNA sequences coding for fusion proteins
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K2319/00—Fusion polypeptide
    • C07K2319/30—Non-immunoglobulin-derived peptide or protein having an immunoglobulin constant or Fc region, or a fragment thereof, attached thereto

Definitions

  • FGF19 fibroblast growth factor 19
  • FGF21 fibroblast growth factor 21
  • Lactate homeostasis is maintained at various levels. Generally, lactate levels are kept within a normal range due to clearance by the liver and kidney. Lactate can also be cleared by gut bacteria. It is important to regulate lactate levels because an elevation of lactate, also known as hyperlactemia, can exacerbate existing disease among other damaging effects. In fact, hyperlactemia is associated with many diseases and is used clinically to evaluate severity of disease. Such diseases include liver diseases. Examples of liver diseases that are associated with hyperlactemia include cirrhosis, non-alcoholic steatohepatitis (NASH), primary biliary cirrhosis (PBC), and acute liver failure, among others. Therefore, there is a need in the art for a method of reducing lactate levels, e.g., to provide a protective effect in advanced liver disease. The present disclosure satisfies this and other needs.
  • NASH non-alcoholic steatohepatitis
  • PBC primary biliary cirrhosis
  • the present disclosure is based, in part, on variants of fibroblast growth factor 19 (FGF19) peptide sequences, fusions of fibroblast growth factor 19 (FGF19) and/or fibroblast growth factor 21 (FGF21) peptide sequences and variants of fusions (chimeras) of fibroblast growth factor 19 (FGF19) and/or fibroblast growth factor 21 (FGF21) peptide sequences having one or more activities, such as modulating the gut microbiome.
  • variants and fusions (chimeras) of FGF19 and/or FGF21 peptide sequences also include sequences that do not substantially or significantly increase or induce hepatocellular carcinoma (HCC) formation or HCC tumorigenesis.
  • HCC hepatocellular carcinoma
  • the gut microbiome plays an important role in liver disease pathogenicity.
  • Dysbiosis is a common feature of liver disease.
  • the imbalance between harmful and beneficial bacteria can cause severe consequences for a patient with liver disease.
  • the loss of beneficial bacteria can exacerbate certain liver diseases like cirrhosis.
  • Veillonella which ferments lactate to propionate and acetate, is a bacterium in the gastrointestinal tract that plays a role in lactate homeostasis. Lactate accumulation in liver disease may lead to acidosis, neurotoxicity and cardiac arrhythmia. It is known that the variants and fusions (chimeras) of FGF19 and/or FGF21 peptide sequences provided herein significantly inhibit bile acid synthesis.
  • provided herein are methods for enriching Veillonella in a subject in need thereof that include administering an effective amount of a peptide sequence disclosed herein. In some embodiments, provided herein are methods for reducing lactate in a subject in need thereof that include administering an effective amount of a peptide sequence disclosed herein. In some embodiments, provided herein are methods for treating or preventing disease associated with elevated lactate in a subject in need thereof that include administering an effective amount of a peptide sequence disclosed herein. In some embodiments, the subject has liver disease. In further embodiments, the liver disease is cirrhosis. In further embodiments, the liver disease is NASH.
  • a peptide sequence has amino-terminal amino acids 1-16 of SEQ ID NO: 100 (FGF21) fused to carboxy -terminal amino acids 21-194 of SEQ ID NO:99 (FGF19), or the peptide sequence has amino-terminal amino acids 1-147 of SEQ ID NO:99 (FGF19) fused to carboxy-terminal amino acids 147-181 of SEQ ID NO: 100 (FGF21) (M41), or the peptide sequence has amino-terminal amino acids 1-20 of SEQ ID NO: 99 (FGF19) fused to carboxy -terminal amino acids 17-181 of SEQ ID NO: 100 (FGF21) (M44), or the peptide sequence has amino-terminal amino acids 1-146 of SEQ ID NO: 100 (FGF21) fused to carboxy -terminal amino acids 148-194 of SEQ ID NO: 99 (FGF19) (M45), or the peptide sequence has amino-terminal amino acids 1-20 of SEQ ID NO
  • a peptide sequence has at least one amino acid substitution to amino acid residues 125-129 of SEQ ID NO:99 (FGF19), EIRPD; at least one amino acid substitution to amino acid residues 126-128 of SEQ ID NO:99 (FGF19), IRP; or at least one amino acid substitution to amino acid residues 127-128 of SEQ ID NO:99 (FGF19),
  • RP or at least one amino acid substitution to amino acid residues 1-124 of SEQ ID NO:99 (FGF19) and/or to amino acid residues 130-194 of SEQ ID NO:99 (FGF19). More specifically, for example, a peptide sequence with a substitution to one of amino acid residues 127-128 of SEQ ID NO:99 (FGF19), RP, wherein at least one amino acid substitution is R127L or P128E. Said substitutions within a corresponding FGF19 sequence (e.g., EIRPD, IRP or RP) of a peptide variant provided herein is also contemplated.
  • FGF19 sequence e.g., EIRPD, IRP or RP
  • the peptide comprises both a R127L and P128E substitution to amino acid residues 127-128 of SEQ ID NO:99 (FGF19), RP, or the corresponding FGF19 sequence thereof in a variant peptide provided herein.
  • the amino acid sequence of the peptide comprises at least one amino acid substitution in the Loop-8 region of FGF19, or the corresponding FGF19 sequence thereof in a variant peptide provided herein.
  • the amino acid sequence of the peptide comprises one amino acid substitution to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises two amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In other embodiments, the amino acid sequence of the peptide comprises three amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid sequence of the peptide comprises four amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises five amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid sequence of the peptide comprises one amino acid substitution to the IRP (amino acids 3-5 of SEQ ID NO:190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises two amino acid substitutions to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises three amino acid substitutions to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid sequence of the peptide comprises one amino acid substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises two amino acid substitutions to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is an Arg (R) to Leu (L) substitution.
  • the substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is a Pro (P) to Glu (E) substitution.
  • the substitutions to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is an Arg (R) to Leu (L) substitution and a Pro (P) to Glu (E) substitution.
  • the foregoing substitution(s) in the Loop-8 region of FGF19 is in the corresponding FGF19 sequence thereof in a variant peptide provided herein. That is, said substitutions within a corresponding FGF19 sequence (e.g., EIRPD, IRP or RP) of a peptide variant provided herein is also contemplated.
  • a corresponding FGF19 sequence e.g., EIRPD, IRP or RP
  • the peptide sequence comprises or consists of any sequence set forth in Table 1.
  • the peptide sequence comprises or consists of any sequence set forth in the Sequence Listing herein.
  • a peptide or chimeric sequence of any suitable length can be practiced using a peptide or chimeric sequence of any suitable length.
  • the N-terminal or C-terminal region of the peptide or chimeric sequence is from about 20 to about 200 amino acid residues in length.
  • a peptide or chimeric sequence has 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more amino acid deletions from the amino terminus, the carboxy-terminus or internally.
  • a peptide or chimeric sequence has an N-terminal region, or a C-terminal region that comprises or consists of an amino acid sequence of about 5 to 10, 10 to 20, 20 to 30, 30 to 40, 40 to 50, 60 to 70, 70 to 80, 80 to 90, 90 to 100 or more amino acids.
  • a peptide or chimeric sequence has a FGF19 sequence portion, or a FGF21 sequence portion that comprises or consists of an amino acid sequence of about 5 to 10, 10 to 20, 20 to 30, 30 to 40, 40 to 50, 50 to 60, 60 to 70, 70 to 80, 80 to 90, 90 to 100 or more amino acids ofFGF19 or FGF21.
  • a peptide sequence or a chimeric peptide sequence has a WGDPI (SEQ ID NO: 170) sequence motif corresponding to the WGDPI (SEQ ID NO: 170) sequence of amino acids 16-20 of SEQ ID NO:99 (FGF19); has a substituted, mutated or absent WGDPI (SEQ ID NO: 170) sequence motif corresponding to FGF19 WGDPI (SEQ ID NO: 170) sequence of amino acids 16-20 of FGF19; has a WGDPI (SEQ ID NO: 170) sequence with one or more amino acids substituted, mutated or absent.
  • the peptide sequence is distinct from a FGF19 variant sequence having any of GQV, GDI, WGPI (SEQ ID NO: 171), WGDPV (SEQ ID NO: 172), WGDI (SEQ ID NO: 173), GDPI (SEQ ID NO: 174), GPI, WGQPI (SEQ ID NO: 175), WGAPI (SEQ ID NO: 176), AGDPI (SEQ ID NO: 177), WADPI (SEQ ID NO: 178), WGDAI (SEQ ID NO: 179), WGDPA (SEQ ID NO: 180), WDPI (SEQ ID NO: 181), WGDI (SEQ ID NO: 182), WGDP (SEQ ID NO: 183) or FGDPI (SEQ ID NO: 184) substituted for the FGF19 WGDPI (SEQ ID NO: 170) sequence at amino acids 16-20.
  • a peptide sequence or a chimeric peptide sequence has N- terminal region comprises amino acid residues VHYG (SEQ ID NO: 101), wherein the N- terminal region comprises amino acid residues DASPHVHYG (SEQ ID NO: 102), or the N- terminal region comprises amino acid residues DSSPLVHYG (SEQ ID NO: 103). More particularly, in one aspect the G corresponds to the last position of the N-terminal region.
  • the N-terminal region comprises amino acid residues DSSPLLQ (SEQ ID NO: 104), where the Q residue is the last amino acid position of the N- terminal region, or comprises amino acid residues DSSPLLQFGGQV (SEQ ID NO: 105), where the V residue corresponds to the last position of the N-terminal region.
  • an N-terminal region comprises or consists of (or further comprises or consists of): RHPIP (SEQ ID NO: 106), where R is the first amino acid position of the N-terminal region; or HPIP (SEQ ID NO: 107), where H is the first amino acid position of the N-terminal region; or RPLAF (SEQ ID NO: 108), where R is the first amino acid position of the N-terminal region; or PLAF (SEQ ID NO: 109), where P is the first amino acid position of the N- terminal region; or R, where R is the first amino acid position of the N-terminal region.
  • RHPIP SEQ ID NO: 106
  • HPIP SEQ ID NO: 107
  • H the first amino acid position of the N-terminal region
  • RPLAF SEQ ID NO: 108
  • PLAF SEQ ID NO: 109
  • a peptide or chimeric sequence has: amino acid residues HPIP (SEQ ID NO: 107), which are the first 4 amino acid residues of the N-terminal region.
  • a peptide or chimeric sequence has: an R residue at the first position of the N-terminal region, or the first position of the N-terminal region is an M residue, or the first and second positions of the N-terminal region is an MR sequence, or the first and second positions of the N-terminal region is an RM sequence, or the first and second positions of the N-terminal region is an RD sequence, or the first and second positions of the N-terminal region is an DS sequence, or the first and second positions of the N-terminal region is an MD sequence, or the first and second positions of the N-terminal region is an MS sequence, or the first through third positions of the N-terminal region is an MDS sequence, or the first through third positions of the N-terminal region is an RDS sequence, or the first through
  • a peptide or chimeric sequence has at the N- terminal region first amino acid position an “M” residue, a “R” residue, a “S” residue, a “H” residue, a “P” residue, a “L” residue or a “D” residue.
  • a peptide or chimeric sequence peptide sequence does not have a “M” residue or an “R” residue at the first amino acid position of the N-terminal region.
  • a peptide or chimeric sequence has an N-terminal region with any one of the following sequences: MDSSPL (SEQ ID NO: 110), MSDSSPL (SEQ ID NO: 111), SDSSPL (SEQ ID NO: 112), MSSPL (SEQ ID NO: 113) or SSPL (SEQ ID NO: 114).
  • a peptide sequence or chimeric peptide sequence has a residue at the last position of the C-terminal region that corresponds to about residue 194 of SEQ ID NO:99 (FGF19).
  • a peptide sequence or a chimeric peptide sequence an addition of amino acid residues 30-194 of SEQ ID NO: 99 (FGF19) at the C- terminus, resulting in a chimeric polypeptide having a residue at the last position of the C- terminal region that corresponds to about residue 194 of SEQ ID NO: 99 (FGF19).
  • a chimeric peptide sequence or peptide sequence comprises all or a portion of a FGF19 sequence (e.g ., SEQ ID NO: 99), positioned at the C-terminus of the peptide, or where the amino terminal “R” residue is deleted from the peptide.
  • a FGF19 sequence e.g ., SEQ ID NO: 99
  • a chimeric peptide sequence or peptide sequence comprises or consists of any of M1-M98 variant peptide sequences, or a subsequence or fragment of any of the M1-M98 variant peptide sequences.
  • Methods and uses provided herein can also be practiced using a peptide or chimeric sequence, as set forth herein.
  • a sequence that comprises or consists of any peptide sequence set forth herein as Ml to M98
  • a peptide sequence comprises or consists of any one of the following sequences:
  • RPL AF SD S SPL VH Y GW GDPIRLRHL YT S GPHGL S S CFLRIRADGVVDC ARGQ S AH SLLEI KAVALRTVAIKGVHSVRYLCMGADGKMQGLLQYSEEDCAFEEEIRPDGYNVYRSEKH RLP V SL S S AKQRQL YKNRGFLPL SHFLPMLPM VPEEPEDLRGHLE SDMF S SPLETD SMDP F GLVTGLEAVRSPSFEK (M2) (SEQ ID NO:2 or 140);
  • the R terminal residue (R residue at the N-terminus) is deleted.
  • the peptide comprises or consists of:
  • the N-terminal R residue is deleted.
  • the peptide comprises or consists of: RPL AF SD S SPL VH Y GW GDPIRLRHL YT S GPHGL S S CFLRIRADGVVDC ARGQ S AH SLLEI KAVALRTVAIKGVHSVRYLCMGADGKMQGLLQYSEEDCAFEEEILEDGYNVYRSEKHR LP V SL S S AKQRQL YKNRGFLPL SHFLPMLPM VPEEPEDLRGHLE SDMF S SPLETD SMDPF GLVTGLEAVRSPSFEK (M201) (SEQ ID NO: 198); or a subsequence or fragment thereof.
  • the N-terminal R residue is deleted.
  • the peptide comprises or consists of:
  • RPL AF SD ASPHVHY GW GDPIRLRHL YT S GPHGL S SCFLRIRADGVVDC ARGQ S AHSLLEI KAVALRTVAIKGVHSVRYLCMGADGKMQGLLQYSEEDCAFEEEILEDGYNVYRSEKHR LP V SL S S AKQRQL YKNRGFLPL SHFLPMLPM VPEEPEDLRGHLE SDMF S SPLETD SMDPF GLVTGLEAVRSPSFEK (M202) (SEQ ID NO: 199); or a subsequence or fragment thereof.
  • the N-terminal R residue is deleted.
  • the peptide comprises or consists of:
  • the peptide comprises or consists of: RHPIPDSSPLLQFGDQVRLRHLYTSGPHGLSSCFLRIRADGVVDCARGQSAHSLLEIKAV ALRTVAIKGVHSVRYLCMGADGKMQGLLQYSEEDCAFEEEILEDGYNVYRSEKHRLPV SLS S AKQRQL YKNRGFLPL SHFLPMLPMVPEEPEDLRGHLE SDMF S SPLETD SMDPF GL V TGLEAVRSPSFEK (M204) (SEQ ID NO:201); or a subsequence or fragment thereof.
  • the N-terminal R residue is deleted.
  • the peptide comprises or consists of:
  • the N-terminal R residue is deleted.
  • the peptide comprises or consists of: RHPIPDSSPLLQFGGQVRLRHLYTSGPHGLSSCFLRIRADGVVDCARGQSAHSLLEIKAV ALRTVAIKGVHSVRYLCMGADGKMQGLLQYSEEDCAFEEEILEDGYNVYRSEKHRLPV SL S S AKQRQL YKNRGFLPL SHFLPMLPMVPEEPEDLRGHLE SDMF S SPLETD SMDPF GL V TGLEAVRSPSFEK (M206) (SEQ ID NO:203); or a subsequence or fragment thereof.
  • the N-terminal R residue is deleted.
  • the peptide comprises or consists of:
  • the peptide is a variant peptide designated M139. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO: 193. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO: 193. In some embodiments, the peptide is a variant peptide designated M140. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO: 194. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO: 194. In some embodiments, the peptide is a variant peptide designated M141. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO: 195.
  • the peptide consists of an amino acid sequence set forth in SEQ ID NO: 195. In some embodiments, the peptide is a variant peptide designated Ml 60. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO: 196. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO: 196. In some embodiments, the peptide is a variant peptide designated M200. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO: 197. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO: 197. In some embodiments, the peptide is a variant peptide designated M201.
  • the peptide comprises an amino acid sequence set forth in SEQ ID NO: 198. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO: 198. In other embodiments, the peptide is a variant peptide designated M202. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO: 199. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO: 199. In certain embodiments, the peptide is a variant peptide designated M203. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID N0:200.
  • the peptide consists of an amino acid sequence set forth in SEQ ID NO:200. In some embodiments, the peptide is a variant peptide designated M204. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO:201. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO:201. In another embodiment, the peptide is a variant peptide designated M205. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO:202. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO:202. In other embodiments, the peptide is a variant peptide designated M206.
  • the peptide comprises an amino acid sequence set forth in SEQ ID NO:203. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO:203. In yet other embodiments, the peptide is a variant peptide designated M207. In some embodiments, the peptide comprises an amino acid sequence set forth in SEQ ID NO:204. In other embodiments, the peptide consists of an amino acid sequence set forth in SEQ ID NO:204.
  • the N-terminus of the peptide sequence comprises or consists of any of:
  • HPIPD S SPLLQF GGQ VRLRHL YT S G (M5-R) (amino acids 1-25 of SEQ ID NO: 160);
  • RPL AF SD S SPLLQF GGQ VRLRHL YT S G (amino acids 1-27 of SEQ ID NO:7);
  • HPIPD S SPLLQF GW GDPIRLRHL YT SG (M9-R) (amino acids 2-28 of SEQ ID NO:9);
  • HPIPD S SPHVH Y GW GDPIRLRHL YT S G (M10-R) (amino acids 2-28 of SEQ ID NO: 10); RPLAFSDAGPLLQWGDPIRLRHLYTSG (Ml 1) (amino acids 1-27 of SEQ ID NO: 11);
  • RPL AF SD AGPLLQF GW GDPIRLRHL YT S G (M12) (amino acids 1-29 of SEQ ID NO: 12); RPLAF SDAGPLLQF GGQ VRLRHL YTSG (M13) (amino acids 1-27 of SEQ ID NO: 13); HPIPD S SPHVH Y GGQ VRLRHL YT S G (M14-R) (amino acids 2-26 of SEQ ID NO: 14); RPLAF SDAGPHVHYGGQ VRLRHL YTSG (Ml 5) (amino acids 1-27 of SEQ ID NO: 15); RPLAF SDAGPHVHWGDPIRLRHL YTSG (M16) (amino acids 1-27 of SEQ ID NO: 16); RPLAF SDAGPHVGWGDPIRLRHL YTSG (M17) (amino acids 1-27 of SEQ ID NO: 17); RPLAF SDAGPHYGWGDPIRLRHL YTSG (M18) (amino acids
  • VH Y GW GDPIRLRHL YT S G (M75-R) (amino acids 2-19 of SEQ ID NO:75);
  • RLRHLYTSG (M77-R) (amino acids 2-10 of SEQ ID NO:77);
  • RHPIPD S SPLLQF GW GDPIRLRHL YT SG (M9) (amino acids 1-28 of SEQ ID NO:9)
  • RHPIPD S SPLLQ W GDPIRLRHL YT S G (M8) (amino acids 1-26 of SEQ ID NO:8);
  • RPLAF SD AGPLLQF GW GDPIRLRHL YT S G (M12) (amino acids 1-29 of SEQ ID NO: 12); RHPIPD S SPHVH Y GW GDPIRLRHL YT S G (M10) (amino acids 1-28 of SEQ ID NO: 10); RPLAF SDAGPLLQFGGQ VRLRHL YTSG (M13) (amino acids 1-27 of SEQ ID NO: 13); RHPIPDSSPHVHYGGQ VRLRHL YTSG (M14) (amino acids 1-26 of SEQ ID NO: 14); RPL AF SD AGPHVHY GGDIRLRHL YT SG (M43) amino acids 1-27 of SEQ ID NO:43); or RD S SPLLQF GGQ VRLRHL YT SG (M6) (amino acids 1-22 of SEQ ID NO:6); or any of the foregoing peptide sequences where the amino terminal R residue is deleted. [0029] In certain embodiments,
  • HPIPD S SPLLQF GGQ VRLRHL YT S G (M5-R) (amino acids 1-25 of SEQ ID NO: 160); DSSPLLQFGGQ VRLRHL YTSG (M6-R) (amino acids 2-22 of SEQ ID NO:6);
  • RPL AF SD S SPLLQF GGQ VRLRHL YT S G (M7) (amino acids 1-27 of SEQ ID NO:7); HPIPD S SPLLQ W GDPIRLRHL YT S G (M8-R) (amino acids 2-26 of SEQ ID NO:8);
  • HPIPD S SPLLQF GW GDPIRLRHL YT SG (M9-R) (amino acids 2-28 of SEQ ID NO:9); HPIPD S SPHVH Y GW GDPIRLRHL YT S G (M10-R) (amino acids 2-28 of SEQ ID NO: 10); RPL AFSDAGPLLQWGDPIRLRHL YTSG (Ml 1) (amino acids 1-27 of SEQ ID NO: 11); RPL AF SD AGPLLQF GW GDPIRLRHL YT S G (M12) (amino acids 1-29 of SEQ ID NO: 12); RPLAFSDAGPLLQFGGQ VRLRHL YTSG (M13) (amino acids 1-27 of SEQ ID NO: 13); HPIPDSSPHVHYGGQ VRLRHL YTSG (M14-R) (amino acids 2-26 of SEQ ID NO: 14); RPLAFSDAGPHVHYGGQ VRLRHL YTSG (Ml 5)
  • RHPIPD S SPLLQF GPQ VRLRHL YT S G (M33) (amino acids 1-26 of SEQ ID NO:33);
  • RHPIPD S SPLLQF GGEVRLRHL YT SG (amino acids 1-26 of SEQ ID NO:35);
  • RHPIPD S SPLLQF GGQ ARLRHL YT S G (M37) (amino acids 1-26 of SEQ ID NO:37);
  • RHPIPD S SPLLQF GW GQP VRLRHL YT S G (M40) (amino acids 1-28 of SEQ ID NO:40);
  • RLRHLYTSG (M77-R) (amino acids 2-10 of SEQ ID NO:77);
  • RHPIPD S SPLLQF GW GDPIRLRHL YT SG (M9) (amino acids 1-28 of SEQ ID NO:9);
  • RPL AF SD AGPLLQF GW GDPIRLRHL YT S G (M12) (amino acids 1-29 of SEQ ID NO: 12); RHPIPD S SPHVH Y GW GDPIRLRHL YT S G (M10) (amino acids 1-28 of SEQ ID NO: 10); RPLAF SD AGPLLQF GGQ VRLRHL YTSG (M13) (amino acids 1-27 of SEQ ID NO: 13); RHPIPD S SPHVHY GGQ VRLRHL YT SG (M14) (amino acids 1-26 of SEQ ID NO: 14);
  • RPLAF SD AGPHVHY GGDIRLRHL YT SG (M43) amino acids 1-27 of SEQ ID NO:43); or RDSSPLLQFGGQ VRLRHL YTSG (M6) (amino acids 1-22 of SEQ ID NO:6).
  • the peptide comprise one of the foregoing sequences. In another embodiment, the peptide consists of one of the foregoing sequences.
  • the peptide comprises a C-terminal region comprising a portion of SEQ ID NO:99 (FGF19), the C-terminal region having a first amino acid position and a last amino acid position, wherein the C-terminal region comprises amino acid residues 16-29 of SEQ ID NO:99 (FGF19), WGDPIRLRHLYTSG (SEQ ID NO: 169), wherein the W residue corresponds to the first amino acid position of the C-terminal region.
  • a peptide sequence comprises or consists of: HPIPDS SPLLQF GGQVRLRHLYTSGPHGLS SCFLRIRADGVVDC ARGQ SAHSLLEIKAVA LRTVAIKGVHS VRYLCMGADGKMQGLLQ Y SEEDC AFEEEIRPDGYNVYRSEKHRLP V S L S S AKQRQL YKNRGFLPL SHFLPMLPMVPEEPEDLRGHLE SDMF S SPLETD SMDPF GL VT GLEAVRSP SFEK (SEQ ID NO: 160);
  • a peptide sequence comprises or consists of:
  • a peptide sequence includes the addition of amino acid residues 30-194 of SEQ ID NO: 99 (FGF19) at the C-terminus, resulting in a chimeric polypeptide.
  • a peptide sequence has at least one amino acid substitution to amino acid residues 125-129 of SEQ ID NO:99 (FGF19), EIRPD. In other embodiments, the peptide sequence has at least one amino acid substitution to amino acid residues 126-128 of SEQ ID NO:99 (FGF19), IRP. In other embodiments, the peptide sequence has at least one amino acid substitution to amino acid residues 127-128 of SEQ ID NO:99 (FGF19), RP. In other embodiments, the peptide sequence has at least one amino acid substitution to amino acid residues 1-124 of SEQ ID NO:99 (FGF19) and/or to amino acid residues 130-194 of SEQ ID NO:99 (FGF19).
  • a peptide sequence comprises substitution to one of amino acid residues 127-128 of SEQ ID NO:99 (FGF19), RP, wherein at least one amino acid substitution is R127L or P128E.
  • Said substitutions within a corresponding FGF19 sequence e.g., EIRPD, IRP or RP
  • the peptide comprises both a R127L and P128E substitution to amino acid residues 127-128 of SEQ ID NO:99 (FGF19), RP, or the corresponding FGF19 sequence thereof in a variant peptide provided herein.
  • the amino acid sequence of the peptide comprises at least one amino acid substitution in the Loop-8 region of FGF19, or the corresponding FGF19 sequence thereof in a variant peptide provided herein.
  • the amino acid sequence of the peptide comprises one amino acid substitution to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises two amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises three amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid sequence of the peptide comprises four amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises five amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises one amino acid substitution to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises two amino acid substitutions to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In other embodiments, the amino acid sequence of the peptide comprises three amino acid substitutions to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises one amino acid substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises two amino acid substitutions to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is an Arg (R) to Leu (L) substitution.
  • the substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is a Pro (P) to Glu (E) substitution.
  • the substitutions to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is an Arg (R) to Leu (L) substitution and a Pro (P) to Glu (E) substitution.
  • the foregoing substitution(s) in the Loop-8 region of FGF19 is in the corresponding FGF19 sequence thereof in a variant peptide provided herein. That is, said substitutions within a corresponding FGF19 sequence (e.g., EIRPD, IRP or RP) of a peptide variant provided herein is also contemplated.
  • Peptide or chimeric sequences provided herein can be of any suitable length.
  • the N-terminal or C-terminal region of the peptide or chimeric sequence is from about 20 to about 200 amino acid residues in length.
  • a chimeric peptide sequence or peptide sequence has at least one amino acid deletion.
  • a peptide or chimeric sequence has 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more amino acid deletions from the amino terminus, the carboxy- terminus or internally.
  • the amino acid substitution, or deletion is at any of amino acid positions 8-20 of FGF19 (AGPHVHYGWGDPI) (SEQ ID NO: 187).
  • a peptide or chimeric sequence has an N-terminal region, or a C- terminal region that comprises or consists of an amino acid sequence of about 5 to 10, 10 to 20, 20 to 30, 30 to 40, 40 to 50, 60 to 70, 70 to 80, 80 to 90, 90 to 100 or more amino acids.
  • a peptide or chimeric sequence has a FGF19 sequence portion, or a FGF21 sequence portion that comprises or consists of an amino acid sequence of about 5 to 10, 10 to 20, 20 to 30, 30 to 40, 40 to 50, 50 to 60, 60 to 70, 70 to 80, 80 to 90, 90 to 100 or more amino acids ofFGF19 orFGF21.
  • a peptide or chimeric sequence has an amino acid substitution, an addition, insertion or is a subsequence that has at least one amino acid deleted.
  • Such amino acid substitutions, additions, insertions and deletions of a peptide sequence can be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more amino acid residues (10-20, 20-30, 30-40, 40-50, etc.), for example, at the N- or C-terminus, or internal.
  • the amino acid substitution, or deletion is at any of amino acid positions 8-20 of FGF19 (AGPHVHYGWGDPI) (SEQ ID NO: 187).
  • a peptide or chimeric sequence includes all or a portion of a FGF19 sequence set forth as:
  • a peptide or chimeric sequence has a function or activity greater or less than a comparison sequence.
  • chimeric peptide sequences and peptide sequences have particular functions or activities.
  • a chimeric peptide sequence or peptide sequence maintains or increases a fibroblast growth factor receptor 4 (FGFR4) mediated activity.
  • FGFR4 fibroblast growth factor receptor 4
  • a chimeric peptide sequence or peptide sequence binds to FGFR4 or activates FGFR4, or does not detectably bind to FGFR4 or activate FGFR4, or binds to FGFR4 with an affinity less than, comparable to or greater than FGF19 binding affinity for FGFR4, or activates FGFR4 to an extent or amount less than, comparable to or greater than FGF19 activates FGFR4.
  • a chimeric peptide sequence or peptide sequence provided herein activates FGFR4 to an extent or amount less than the extent or amount that FGF19 activates FGFR4.
  • a chimeric peptide sequence or peptide sequence provided herein activates FGFR4 to an extent or amount comparable to the extent or amount that FGF19 activates FGFR4. In some embodiments, a chimeric peptide sequence or peptide sequence provided herein activates FGFR4 to an extent or amount greater than the extent or amount that FGF19 activates FGFR4. [0037] In one embodiment, a chimeric peptide sequence or peptide sequence provided herein maintains a FGFR4 mediated activity. In one embodiment, a chimeric peptide sequence or peptide sequence provided herein increases a FGFR4 mediated activity.
  • a chimeric peptide sequence or peptide sequence provided herein binds to FGFR4 with an affinity less than FGF19 binding affinity for FGFR4. In some embodiments, a chimeric peptide sequence or peptide sequence provided herein binds to FGFR4 with an affinity comparable to FGF19 binding affinity for FGFR4. In some embodiments, a chimeric peptide sequence or peptide sequence provided herein binds to FGFR4 with an affinity greater than FGF19 binding affinity for FGFR4. In some embodiments, a chimeric peptide sequence or peptide sequence provided herein does not detectably bind to FGFR4.
  • a chimeric peptide sequence or peptide sequence has reduced HCC formation compared to FGF19, or a FGF19 variant sequence having any of GQV, GDI, WGPI (SEQ ID NO: 171), WGDPV (SEQ ID NO: 172), WGDI (SEQ ID NO: 173), GDPI (SEQ ID NO: 174), GPI, WGQPI (SEQ ID NO: 175), WGAPI (SEQ ID NO: 176), AGDPI (SEQ ID NO: 177), WADPI (SEQ ID NO: 178), WGDAI (SEQ ID NO: 179), WGDPA (SEQ ID NO: 180), WDPI (SEQ ID NO: 181), WGDI (SEQ ID NO: 182), WGDP (SEQ ID NO: 183) or FGDPI (SEQ ID NO: 184) substituted for the WGDPI (SEQ ID NO: 170) sequence at amino acids 16-20 of FGF19; or has greater glucose
  • a peptide or chimeric sequence has a function or activity greater or less than a comparison sequence.
  • the comparison sequence is FGF19.
  • the comparison sequence is FGF19 variant sequence having any of GQV, GDI, WGPI (SEQ ID NO: 171), WGDPV (SEQ ID NO: 172), WGDI (SEQ ID NO: 173), GDPI (SEQ ID NO: 174), GPI, WGQPI (SEQ ID NO: 175), WGAPI (SEQ ID NO: 176), AGDPI (SEQ ID NO: 177), WADPI (SEQ ID NO: 178), WGDAI (SEQ ID NO: 179), WGDPA (SEQ ID NO: 180), WDPI (SEQ ID NO: 181), WGDI (SEQ ID NO: 182), WGDP (SEQ ID NO: 183) or FGDPI (SEQ ID NO: 184) substituted for the WGDPI (SEQ ID NO: 170
  • a peptide or chimeric peptide sequence provided herein has greater glucose lowering activity compared to a comparison sequence. In another embodiment, a peptide or chimeric peptide sequence provided herein has less lipid increasing activity compared to a comparison sequence. In another embodiment, a peptide or chimeric peptide sequence provided herein has lower or reduced lipid (e.g triglyceride, cholesterol, non- HDL) increasing activity compared to a comparison sequence. In other embodiments, a peptide or chimeric peptide sequence provided herein has more HDL increasing activity as compared to a comparison sequence.
  • lipid e.g triglyceride, cholesterol, non- HDL
  • a peptide or chimeric peptide sequence provided herein has less lean mass reducing activity compared to a comparison sequence or FGF21.
  • a peptide or chimeric sequence provided herein has greater Veillonella enrichment activity or lactate reducing activity compare to a comparison sequence, FGF19, or FGF21.
  • a peptide or chimeric sequence includes one or more L-amino acids, D-amino acids, non-naturally occurring amino acids, or amino acid mimetic, derivative or analogue.
  • a peptide or chimeric sequence has an N-terminal region, or a C-terminal region, or a FGF19 sequence portion, or a FGF21 sequence portion, joined by a linker or spacer.
  • the chimeric peptide sequence or peptide sequence is comprised in a pharmaceutical composition.
  • an additional therapeutic agent is administered.
  • the additional therapeutic agent is an inflammatory agent.
  • FIG. 1 shows that patients with non-alcoholic steatohepatitis (NASH) treated with the FGF19 analogue M70 had stable gut microbial composition by alpha diversity over 12 weeks.
  • FIGS. 2A and 2B show that patients with non-alcoholic steatohepatitis (NASH) treated with the FGF19 analogue M70 had stable gut microbial composition by beta diversity over 12 weeks with unweighted UniFrac (FIG. 2A) and weighted UniFrac (FIG. 2B). Day 1 shown in light gray, week 12 shown in dark gray.
  • FIGS. 3A and 3B show that patients with non-alcoholic steatohepatitis (NASH) treated with the FGF19 analogue M70 had no change in the phyla of the gut microbiome over 12 weeks with Kruskal-Wallis test (FIG. 3A) and Mann-Whitney U-test (FIG. 3B; day 1 (left), week 12 (right)), both with FDR corrections using Benjamini-Hoxhberg.
  • FIG. 3A depicts M70 concentration left to right per phyla as 0.3 mg, 1 mg, 3 mg, 6 mg and placebo.
  • FIG. 3C and 3D show that patients with NASH treated with the FGF19 analogue M70 had no change in the genera of the gut microbiome over 12 weeks with Kruskal-Wallis test (FIG. 3C) and Mann-Whitney U-test (FIG. 3D; day 1 (left), week 12 (right)), both with FDR corrections using Benjamini-Hoxhberg.
  • FIG. 3C depicts M70 concentration left to right per genera as 0.3 mg, 1 mg, 3 mg, 6 mg and placebo.
  • FIG. 4A shows that patients with non-alcoholic steatohepatitis (NASH) treated with the FGF19 analogue M70 had enriched populations of lactate-consuming Veillonella bacteria.
  • FIG. 4B shows that patients with NASH treated with the FGF19 analogue M70 had fecal samples that were positive for Veillonella at baseline and week 12.
  • FIG. 4C shows that patients with NASH treated with the FGF19 analogue M70, did not have any changes in other microbes that are of oral origin, associated with Veillonella , ferment other substrates, or with ethanol producing properties. Day 1 shown in black (left), week 12 shown in gray (right).
  • FIG. 5A shows that patients with non-alcoholic steatohepatitis (NASH) treated with FGF19 analogue M70 had enriched populations of lactate-consuming Veillonella , which was inversely correlated with toxic, hydrophobic bile acids compared to baseline. Day 1 shown in light gray (left), week 12 shown in dark gray (right).
  • FIG. 5B shows that patients with NASH treated with the FGF19 analogue M70 had enriched populations of lactate-consuming Veillonella and its operational units were inverselty correlated with concentrations of GCA, GCDCA,
  • FIG. 6A shows that patients with non-alcoholic steatohepatitis (NASH) treated with FGF19 analogue M70 had no change in alpha diversity between low liver fat (Ql) and high liver fat (Q4).
  • FIG. 6B shows that patients with NASH treated with FGF19 analogue M70 showed no separation between Ql and Q4 at baseline. Ql shown in dark gray and Q4 shown in light gray.
  • FIG. 6C and FIG. 6D show that patients with NASH treated with FGF19 analogue M70 showed no significant difference at the phylum and genus level between Ql and Q4 populations. Ql shown in dark gray (left) and Q4 shown in light gray (right).
  • FIG. 7A shows that patients with non-alcoholic steatohepatitis (NASH) treated with FGF19 analogue M70 had a gut microbiota composition that was not significantly different between mild fibrosis (FI) and advanced fibrosis (F3) subjects by alpha diversity.
  • FIG. 7B shows that patients with NASH treated with FGF19 analogue M70 had beta diversity that was not significantly different between FI (stage 1) and F3 (stage 3) subjects. Stage 1 shown in dark gray and stage 3 shown in light gray.
  • FIG. 7C and FIG. 7D show that patients with NASH treated with FGF19 analogue M70 had relative abundance of the top bacterial phylotypes at the phylum and genus leels in the FI and F3 groups. Stage 1 shown in dark gray (left) and stage 3 shown in light gray (right).
  • FIG. 8A shows that patients with non-alcoholic steatohepatitis (NASH) that had 70% or greater reduction in liver fat content with FGF19 analogue M70 treatment, had a reduction in alanine aminotransferase (ALT) and aspartate aminotransferase (AST).
  • FIG. 8B shows that there is no correlation between the reduction in liver enzymes and Veillonella bacteria. DESCRIPTION
  • treat refers to a course of action (such as administering a polypeptide or a pharmaceutical composition comprising a polypeptide) initiated after a disease, disorder or condition, or a symptom thereof, has been diagnosed, observed, and the like so as to eliminate, reduce, suppress, mitigate, or ameliorate, either temporarily or permanently, at least one of the underlying causes of a disease, disorder, or condition afflicting a subject, or at least one of the symptoms associated with a disease, disorder, condition afflicting a subject.
  • treatment includes inhibiting (/. ., arresting the development or further development of the disease, disorder or condition or clinical symptoms association therewith) an active disease.
  • the term “in need of treatment” as used herein refers to a judgment made by a physician or other medical professional that a subject requires or will benefit from treatment.
  • the terms “prevent”, “preventing”, “prevention” and the like refer to a course of action (such as administering a polypeptide or a pharmaceutical composition comprising a polypeptide) initiated in a manner ( e.g ., prior to the onset of a disease, disorder, condition or symptom thereof) so as to prevent, suppress, inhibit or reduce, either temporarily or permanently, a subject’s risk of developing a disease, disorder, condition or the like (as determined by, for example, the absence of clinical symptoms) or delaying the onset thereof, generally in the context of a subject predisposed to having a particular disease, disorder or condition.
  • the terms also refer to slowing the progression of the disease, disorder or condition or inhibiting progression thereof to a harmful or otherwise undesired state.
  • in need of prevention refers to a judgment made by a physician or other medical professional that a subject requires or will benefit from preventative care.
  • administer refers to the act of injecting or otherwise physically delivering a substance as it exists outside the body into a patient, such as by mucosal, intradermal, intravenous, intramuscular delivery and/or any other method of physical delivery described herein or known in the art.
  • administration of the substance typically occurs after the onset of disease, disorder or condition or symptoms thereof.
  • administration of the substance typically occurs before the onset of the disease, disorder or condition or symptoms thereof.
  • the term “therapeutically effective amount” refers to the administration of an agent to a subject, either alone or as a part of a pharmaceutical composition and either in a single dose or as part of a series of doses, in an amount that is capable of having any detectable, positive effect on any symptom, aspect, or characteristics of a disease, disorder or condition when administered to a patient.
  • the therapeutically effective amount can be ascertained by measuring relevant physiological effects.
  • metabolic syndrome refers to an associated cluster of traits that includes, but is not limited to, hyperinsulinemia, abnormal glucose tolerance, obesity, redistribution of fat to the abdominal or upper body compartment, hypertension, dysfibrinolysis, and dyslipidemia characterized by high triglycerides, low high density lipoprotein (HDL)-cholesterol, and high small dense low density lipoprotein (LDL) particles.
  • Subjects having metabolic syndrome are at risk for development of type 2 diabetes and/or other disorders (e.g ., atherosclerosis).
  • polypeptide refers to a polymeric form of amino acids of any length, which can include genetically coded and non- genetically coded amino acids, chemically or biochemically modified or derivatized amino acids, and polypeptides having modified polypeptide backbones.
  • the terms include fusion proteins, including, but not limited to, fusion proteins with a heterologous amino acid sequence, fusion proteins with heterologous and homologous leader sequences, with or without N-terminus methionine residues; immunologically tagged proteins; and the like. It will be appreciated that throughout this disclosure reference is made to amino acids according to the single letter or three letter codes.
  • amino acids forming all or a part of a peptide may be from among the known 21 naturally occurring amino acids, which are referred to by both their single letter abbreviations and their common three-letter abbreviation.
  • conventional amino acid residues have their conventional meaning. Thus, “Leu” is leucine, “He” is isoleucine, “Me” is norleucine, and so on.
  • alanine Ala arginine Arg (R) asparagine Asn (N) aspartic acid Asp (D) cysteine Cys (C) glutamic acid Glu (E) glutamine Gin (Q) glycine Gly (G) histidine His (H) isoleucine lie (I) leucine Leu (L) lysine Lys (K) methionine Met (M) phenylalanine Phe (F) proline Pro (P) serine Ser (S) threonine Thr (T) tryptophan Trp (W) tyrosine Tyr (Y) valine Val (V)
  • variant encompasses naturally-occurring variants (e.g ., homologs and allelic variants) and non-naturally-occurring variants (e.g., muteins).
  • Naturally- occurring variants include homologs, i.e., nucleic acids and polypeptides that differ in nucleotide or amino acid sequence, respectively, from one species to another.
  • Naturally-occurring variants include allelic variants, i.e., nucleic acids and polypeptides that differ in nucleotide or amino acid sequence, respectively, from one individual to another within a species.
  • Non-naturally-occurring variants include nucleic acids and polypeptides that comprise a change in nucleotide or amino acid sequence, respectively, where the change in sequence is artificially introduced, e.g, the change is generated in the laboratory or other facility by human intervention (“hand of man”).
  • the term “native”, in reference to FGF19 refers to biologically active, naturally- occurring FGF19, including biologically active, naturally-occurring FGF19 variants. The term includes the 194 amino acid human FGF19 mature sequence.
  • label when use in the context of a polypeptide or nucleic acid (or antibody, as appropriate) of the present disclosure are meant to refer broadly to any means useful in, for example, polypeptide purification, identification, isolation and synthesis. Labels are generally covalently bound to the polypeptide of interest and can be introduced in any manner known in the art, including attachment to a mature polypeptide (generally at the N- or C-terminus), incorporation during solid-phase peptide synthesis, or through recombinant means. Examples include, but are not limited to, fluorescence, biotinylation, and radioactive isotopes.
  • Polypeptide and nucleic acid molecules can be labeled by both in vitro and in vivo methods. Labeling reagents and kits can be obtained from a number of commercial sources (e.g, Thermo Fischer Scientific, Rockford, IL; and Molecular Probes/Life Technologies; Grand Island, NY).
  • muteins refers broadly to mutated recombinant proteins, i.e., a polypeptide comprising an artificially introduced change in amino acid sequence, e.g, a change in amino acid sequence generated in the laboratory or other facility by human intervention (“hand of man”). These proteins usually carry single or multiple amino acid substitutions and are frequently derived from cloned genes that have been subjected to site- directed or random mutagenesis, or from completely synthetic genes.
  • the terms “modified”, “modification” and the like refer to one or more changes that enhance a desired property of human FGF19, a naturally-occurring FGF19 variant, or a FGF19 mutein, wherein the change(s) does not alter the primary amino acid sequence of the FGF19.
  • desired properties include, for example, enhancing solubility, prolonging the circulation half-life, increasing the stability, reducing the clearance, altering the immunogenicity or allergenicity, improving aspects of manufacturability (e.g., cost and efficiency), and enabling the raising of particular antibodies (e.g, by introduction of unique epitopes) for use in detection assays.
  • Changes to human FGF19, a naturally-occurring FGF19 variant, or a FGF19 mutein that can be carried out include, but are not limited to, pegylation (covalent attachment of one or more molecules of polyethylene glycol (PEG), or derivatives thereof); glycosylation (e.g, N-glycosylation), polysialylation and hesylation; albumin fusion; albumin binding through, for example, a conjugated fatty acid chain (acylation); Fc-fusion; and fusion with a PEG mimetic.
  • DNA DNA
  • nucleic acid nucleic acid molecule
  • polynucleotide and the like are used interchangeably herein to refer to a polymeric form of nucleotides of any length, either deoxyribonucleotides or ribonucleotides, or analogs thereof.
  • Non-limiting examples of polynucleotides include linear and circular nucleic acids, messenger RNA (mRNA), complementary DNA (cDNA), recombinant polynucleotides, vectors, probes, primers and the like.
  • probe refers to a fragment of DNA or RNA corresponding to a gene or sequence of interest, wherein the fragment has been labeled radioactively (e.g, by incorporating 32 P or 35 S) or with some other detectable molecule, such as biotin, digoxygen or fluorescein. As stretches of DNA or RNA with complementary sequences will hybridize, a probe can be used, for example, to label viral plaques, bacterial colonies or bands on a gel that contain the gene of interest.
  • a probe can be cloned DNA or it can be a synthetic DNA strand; the latter can be used to obtain a cDNA or genomic clone from an isolated protein by, for example, microsequencing a portion of the protein, deducing the nucleic acid sequence encoding the protein, synthesizing an oligonucleotide carrying that sequence, radiolabeling the sequence and using it as a probe to screen a cDNA library or a genomic library.
  • heterologous refers to two components that are defined by structures derived from different sources.
  • a “heterologous” polypeptide can include operably linked amino acid sequences that are derived from different polypeptides.
  • a “heterologous” polynucleotide can include operably linked nucleic acid sequences that can be derived from different genes.
  • heterologous nucleic acids include expression constructs in which a nucleic acid comprising a coding sequence is operably linked to a regulatory element (e.g, a promoter) that is from a genetic origin different from that of the coding sequence (e.g ., to provide for expression in a host cell of interest, which can be of different genetic origin than the promoter, the coding sequence or both).
  • a regulatory element e.g, a promoter
  • heterologous can refer to the presence of a nucleic acid (or gene product, such as a polypeptide) that is of a different genetic origin than the host cell in which it is present.
  • operably linked refers to linkage between molecules to provide a desired function.
  • operably linked in the context of nucleic acids refers to a functional linkage between nucleic acid sequences.
  • a nucleic acid expression control sequence such as a promoter, signal sequence, or array of transcription factor binding sites
  • the expression control sequence affects transcription and/or translation of the second polynucleotide.
  • operably linked refers to a functional linkage between amino acid sequences (e.g., different domains) to provide for a described activity of the polypeptide.
  • N-terminus (or “amino terminus”) and “C-terminus” (or “carboxyl terminus”) refer to the extreme amino and carboxyl ends of the polypeptide, respectively, while the terms “N-terminal” and “C-terminal” refer to relative positions in the amino acid sequence of the polypeptide toward the N-terminus and the C-terminus, respectively, and can include the residues at the N-terminus and C-terminus, respectively.
  • Immediately N-terminal or “immediately C-terminal” refers to a position of a first amino acid residue relative to a second amino acid residue where the first and second amino acid residues are covalently bound to provide a contiguous amino acid sequence.
  • “Derived from”, in the context of an amino acid sequence or polynucleotide sequence is meant to indicate that the polypeptide or nucleic acid has a sequence that is based on that of a reference polypeptide or nucleic acid (e.g, a naturally occurring FGF19 polypeptide or a FGF 19-encoding nucleic acid), and is not meant to be limiting as to the source or method in which the protein or nucleic acid is made.
  • the term “derived from” includes homologues or variants of reference amino acid or DNA sequences.
  • isolated refers to a polypeptide of interest that, if naturally occurring, is in an environment different from that in which it can naturally occur. “Isolated” is meant to include polypeptides that are within samples that are substantially enriched for the polypeptide of interest and/or in which the polypeptide of interest is partially or substantially purified. Where the polypeptide is not naturally occurring, “isolated” indicates the polypeptide has been separated from an environment in which it was made by either synthetic or recombinant means.
  • “Enriched” means that a sample is non-naturally manipulated (e.g, by a scientist or a clinician) so that a polypeptide of interest is present in a) a greater concentration (e.g, at least 3- fold greater, at least 4-fold greater, at least 8-fold greater, at least 64-fold greater, or more) than the concentration of the polypeptide in the starting sample, such as a biological sample (e.g, a sample in which the polypeptide naturally occurs or in which it is present after administration), or b) a concentration greater than the environment in which the polypeptide was made (e.g., as in a bacterial cell).
  • a biological sample e.g, a sample in which the polypeptide naturally occurs or in which it is present after administration
  • a concentration greater than the environment in which the polypeptide was made e.g., as in a bacterial cell.
  • chimeric and peptide sequences in the modulation of the gut microbiome are uses of chimeric and peptide sequences in promoting enrichment of the gut microbiome.
  • uses of chimeric and peptide sequences in promoting enrichment of Veillonella are uses of chimeric and peptide sequences in reducing lactate.
  • uses of chimeric and peptide sequences in the treatment or prevention of liver disease are uses of chimeric and peptide sequences in the modulation of the gut microbiome.
  • uses of chimeric and peptide sequences in promoting enrichment of the gut microbiome are uses of chimeric and peptide sequences in promoting enrichment of Veillonella.
  • uses of chimeric and peptide sequences in reducing lactate are uses of chimeric and peptide sequences in the treatment or prevention of liver disease.
  • the liver disease is NASH. In certain embodiments, the liver disease is cirrhosis. In certain embodiments, the liver disease is PBC. In certain embodiments, the liver disease is acute liver failure.
  • the liver disease is uses of chimeric and peptide sequences in the treatment or prevention of a liver-related disease. In certain embodiments, the liver-related disease is hepatic fibrosis.
  • the liver-related disease is hepatic fibrosis.
  • the invention is based, in part, on the use of variants of FGF19 peptide sequences, fusions of FGF19 and/or FGF21 peptide sequences and variants of fusions (chimeras) of FGF19 and/or FGF21 peptide sequences having one or more activities associated with modulating the gut microbiome and lactate levels.
  • a chimeric peptide sequence comprises or consists of an N- terminal region having at least seven amino acid residues and the N-terminal region having a first amino acid position and a last amino acid position, where the N-terminal region has a DSSPL (SEQ ID NO: 121) or DASPH (SEQ ID NO: 122) sequence; and a C-terminal region having a portion of FGF19 and the C-terminal region having a first amino acid position and a last amino acid position, where the C-terminal region includes amino acid residues 16-29 of FGF19 (WGDPIRLRHLYTSG; SEQ ID NO: 169) and the W residue corresponds to the first amino acid position of the C-terminal region.
  • the variant is M70:
  • the variant is M69: RDS SPLVHYGWGDPIRLRHLYTSGPHGLS SCFLRIRADGVVDC ARGQ S AHSLLEIKAVA LRTVAIKGVHS VRYLCMGADGKMQGLLQ Y SEEDC AFEEEIRPDGYNVYRSEKHRLP V S L S S AKQRQL YKNRGFLPL SHFLPMLPMVPEEPEDLRGHLE SDMF S SPLETD S 16MDPF GL VTGLEAVRSPSFEK (SEQ ID NO:69).
  • the N-terminal region has a DSSPL (SEQ ID NO:121).
  • the N-terminal region has a DASPH (SEQ ID NO: 122) sequence.
  • the treatment peptide comprises: a) an N-terminal region comprising at least seven amino acid residues, the N-terminal region having a first amino acid position and a last amino acid position; and b) a C-terminal region comprising a portion of SEQ ID NO:99 [FGF19], the C-terminal region having a first amino acid position and a last amino acid position, wherein the C-terminal region comprises (i) a first C-terminal region sequence comprising WGDPIRLRHLYTSG (amino acids 16 to 29 of SEQ ID NO:99 [FGF19]), wherein the W residue corresponds to the first amino acid position of the C-terminal region; and (ii) a second C-terminal region sequence comprising PHGLSSCFLRIRADGVVDCARGQSAHSLLEIKAVALRTVAIKGVHSVRYLCMGADGKM QGLLQ Y SEEDC AFEEEIRPDGYNVYRSEKHRLP V SLS SLS SLS SLS SLS S
  • the treatment peptide comprises: a) an N-terminal region comprising at least seven amino acid residues, the N-terminal region having a first amino acid position and a last amino acid position, wherein the N-terminal region comprises DSSPL (SEQ ID NO: 121) or DASPH (SEQ ID NO: 122); and b) a C-terminal region comprising a portion of SEQ ID NO:99 [FGF19], the C-terminal region having a first amino acid position and a last amino acid position, wherein the C-terminal region comprises (i) a first C-terminal region sequence comprising WGDPIRLRHLYTSG (amino acids 16 to 29 of SEQ ID NO:99 [FGF19]), wherein the W residue corresponds to the first amino acid position of the C-terminal region; and (ii) a second C-terminal region sequence comprising
  • the peptide (i) binds to FGFR4 with an affinity equal to or greater than FGF19 binding affinity for FGFR4; (ii) activates FGFR4 to an extent or amount equal to or greater than FGF19 activates FGFR4; (iii) has at least one of reduced HCC formation; greater glucose lowering activity, less lipid increasing activity, less triglyceride activity, less cholesterol activity, less non-HDL activity or less HDL increasing activity; greater lactate reducing activity or greater Veillonella enrichment activity, as compared to FGF19, or as compared to a FGF19 variant sequence having any of GQV, GDI, WGPI (SEQ ID NO: 171), WGDPV(SEQ ID NO: 172), WGDI (SEQ ID NO: 173), GDPI (SEQ ID NO: 174), GPI, WGQPI (SEQ ID NO: 175), WGAPI (SEQ ID NO: 176), AGDPI (SEQ ID NO: 177
  • the second C-terminal region sequence comprises at least one amino acid substitution to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) sequence.
  • the at least one amino acid substitution is to the IRP sequence of the EIRPD (amino acids 2-6 of SEQ ID NO: 190) sequence.
  • the at least one amino acid substitution is to the RP sequence of the EIRPD sequence (amino acids 2-6 of SEQ ID NO: 190).
  • the at least one amino acid substitution is R to L substitution.
  • the at least one amino acid substitution is P to E substitution.
  • the at least one amino acid substitution is RP to LE substitution.
  • the second C-terminal region sequence comprises from 2 to 5 amino acid substitutions, deletions or insertions. In other embodiments, the peptide is less than about 250 amino acids in length.
  • the treatment peptide has an amino acid sequence comprising or consisting of
  • the treatment peptide has an amino acid sequence comprising SEQ ID NO:70. In other embodiments, the treatment peptide has an amino acid sequence consisting of SEQ ID NO:70. In some embodiments, the treatment peptide is fused with an immunoglobulin Fc region.
  • the treatment peptide has an amino acid sequence comprising or consisting of
  • the treatment peptide has an amino acid sequence comprising SEQ ID NO:69. In other embodiments, the treatment peptide has an amino acid sequence consisting of SEQ ID NO:69. In some embodiments, the treatment peptide is fused with an immunoglobulin Fc region.
  • the treatment peptide comprises: a) an N-terminal region comprising at least seven amino acid residues, the N-terminal region having a first amino acid position and a last amino acid position,; and b) a C-terminal region comprising a first amino acid position and a last amino acid position, wherein the C-terminal region comprises (i) a first C- terminal region sequence comprising WGDPIRQRHLYTSG (SEQ ID NO: 169 with a L7Q substitution), wherein the W residue corresponds to the first amino acid position of the C- terminal region; and (ii) a second C-terminal region sequence comprising PHGLSSCFLRIRADGVVDCARGQSAHSLLEIKAVALRTVAIKGVHSVRYLCMGADGKM QGLLQ Y SEEDC AFEEEIRPDGYNVYRSEKHRLP V SLS S AKQRQL YKNRGFLPL SHFLPML PMVPEEPEDLRGHLE SDMF S
  • the treatment peptide comprises: a) an N-terminal region comprising at least seven amino acid residues, the N-terminal region having a first amino acid position and a last amino acid position, wherein the N-terminal region comprises DSSPL (SEQ ID NO: 121), DASPH (SEQ ID NO: 122), or DAGPH (amino acids 7 to 11 of SEQ ID NO: 99 [FGF19]); and b) a C-terminal region comprising a first amino acid position and a last amino acid position, wherein the C-terminal region comprises (i) a first C-terminal region sequence comprising WGDPIRQRHLYTSG (SEQ ID NO: 169 with a L7Q substitution), wherein the W residue corresponds to the first amino acid position of the C-terminal region; and (ii) a second C-terminal region sequence comprising
  • the peptide (i) binds to FGFR4 with an affinity equal to or greater than FGF19 binding affinity for FGFR4; (ii) activates FGFR4 to an extent or amount equal to or greater than FGF19 activates FGFR4; (iii) has at least one of reduced hepatocellular carcinoma (HCC) formation; greater glucose lowering activity, less lipid increasing activity, less triglyceride activity, less cholesterol activity, less non-HDL activity or less HDL increasing activity; greater lactate reducing activity or greater Veillonella enrichment activity as compared to FGF19, or as compared to a FGF19 variant sequence having any of GQV, GDI, WGPI (SEQ ID NO: 171), WGDPV (SEQ ID NO: 172), WGDI (SEQ ID NO: 173), GDPI (SEQ ID NO: 174), GPI, WGQPI (SEQ ID NO: 175), WGAPI (SEQ ID NO: 176), AGDPI (HCC) formation
  • the second C-terminal region sequence comprises at least one amino acid substitution to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) sequence.
  • the at least one amino acid substitution is to the IRP sequence of the EIRPD (amino acids 2-6 of SEQ ID NO: 190) sequence.
  • the at least one amino acid substitution is to the RP sequence of the EIRPD sequence (amino acids 2-6 of SEQ ID NO: 190).
  • the at least one amino acid substitution is R to L substitution.
  • the at least one amino acid substitution is P to E substitution. In yet other embodiments, the at least one amino acid substitution is RP to LE substitution.
  • the second C-terminal region sequence comprises from 2 to 5 amino acid substitutions, deletions or insertions. In other embodiments, the peptide is less than about 250 amino acids in length.
  • a chimeric peptide sequence comprises or consists of an N- terminal region having a portion of FGF21 and the N-terminal region having a first amino acid position and a last amino acid position, where the N-terminal region has a GQV sequence and the V residue corresponds to the last amino acid position of the N-terminal region; and a C- terminal region having a portion of FGF19 and the C-terminal region having a first amino acid position and a last amino acid position where the C-terminal region includes amino acid residues 21-29 of FGF19 (RLRHLYTSG; SEQ ID NO: 185) and the R residue corresponds to the first position of the C-terminal region.
  • modifications to the Loop-8 region of FGF19 are disclosed herein that possess favorable metabolic parameters without exhibiting substantial tumorigenicity.
  • FGF19 residues 127-129 are defined as constituting the Loop-8 region, although in the literature the Loop-8 region is sometimes defined as including or consisting of other residues ( e.g ., residues 125-129).
  • Certain combinations of R127L and P128E substitutions to the FGF19 framework had an unexpectedly positive effect on HCC formation. Even more surprisingly, a combination of R127L and P128E substitutions and a substitution of Gin (Q) for Leu (L) in the FGF19 core region had an even more significant effect on preventing HCC formation.
  • variants of FGF19 Loop-8 region are included since they can reduce or eliminate substantial, measurable or detectable HCC formation. Furthermore, the effect of reducing HCC formation may be enhanced by modifications to amino acid residues outside of the Loop 8 region (e.g., substitutions of amino acid residues in the core region).
  • variants of FGF19 Loop-8 region are included since they can reduce or eliminate substantial, measurable or detectable HCC formation. Furthermore, the effect of reducing HCC formation may be enhanced by modifications to amino acid residues outside of the Loop-8 region (e.g ., substitutions of amino acid residues in the core region, such as the region corresponding to amino acids 21-29 of SEQ ID NO: 99).
  • the Loop-8 modified variant comprises a substitution in the FGF19 Loop-8 region corresponding to amino acids 127-129 of SEQ ID NO:99.
  • the Loop-8 modified variant comprises a substitution in the FGF19 Loop-8 region corresponding to (i) a R127L substitution, (ii) a P128E substitution, or (iii) a R127L substitution and a P128E substitution.
  • the amino acid sequence of the peptide comprises at least one amino acid substitution in the Loop-8 region of FGF19, or the corresponding FGF19 sequence thereof in a variant peptide provided herein.
  • the amino acid sequence of the peptide comprises one amino acid substitution to the EIRPD (amino acids 2-6 of SEQ ID NO:190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises two amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises three amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid sequence of the peptide comprises four amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises five amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO:190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises one amino acid substitution to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises two amino acid substitutions to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In other embodiments, the amino acid sequence of the peptide comprises three amino acid substitutions to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises one amino acid substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises two amino acid substitutions to the RP (amino acids 4-5 of SEQ ID NO:190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is an Arg (R) to Leu (L) substitution.
  • the substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is a Pro (P) to Glu (E) substitution.
  • the substitutions to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is an Arg (R) to Leu (L) substitution and a Pro (P) to Glu (E) substitution.
  • the foregoing substitution(s) in the Loop-8 region of FGF19 is in the corresponding FGF19 sequence thereof in a variant peptide provided herein. That is, said substitutions within a corresponding FGF19 sequence (e.g., EIRPD, IRP or RP) of a peptide variant provided herein is also contemplated.
  • the FGF19 variant comprises or further comprises a substitution in the core region corresponding to amino acids 21-29 of SEQ ID NO:99. In certain embodiments, the FGF19 variant comprises or further comprises a substitution in the core region corresponding to a L22Q substitution.
  • the Loop-8 modified variant is M70:
  • the Loop-8 modified M70 variant comprises a substitution in the FGF19 Loop-8 region (RPD; underlined) corresponding to (i) an R to L substitution, (ii) a P to E substitution, or (iii) an R to L substitution and a P to E substitution (SEQ. ID NO:204).
  • the Loop-8 modified M70 variant further comprises or further comprises a substitution in the FGF19 core region.
  • the Loop-8 modified M70 variant comprises a L18Q substitution ( i.e SEQ ID NO:70 with an L18Q substitution).
  • the Loop-8 modified variant is M69:
  • the Loop-8 modified M69 variant comprises a substitution in the FGF19 Loop-8 region (RPD; underlined) corresponding to (i) an R to L substitution, (ii) a P to E substitution, or (iii) an R to L substitution and a P to E substitution.
  • the Loop-8 modified M69 variant further comprises or further comprises a substitution in the FGF19 core region.
  • the Loop-8 modified M69 variant comprises a L17Q substitution ( i.e SEQ ID NO:69 with an L17Q substitution).
  • the amino acid sequence of the peptide comprises one amino acid substitution to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises two amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In other embodiments, the amino acid sequence of the peptide comprises three amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO:190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises four amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In some embodiments, the amino acid sequence of the peptide comprises five amino acid substitutions to the EIRPD (amino acids 2-6 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid sequence of the peptide comprises one amino acid substitution to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises two amino acid substitutions to the IRP (amino acids 3-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In other embodiments, the amino acid sequence of the peptide comprises three amino acid substitutions to the IRP (amino acids 3- 5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19. In certain embodiments, the amino acid sequence of the peptide comprises one amino acid substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid sequence of the peptide comprises two amino acid substitutions to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19.
  • the amino acid substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is an Arg (R) to Leu (L) substitution.
  • the substitution to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is a Pro (P) to Glu (E) substitution.
  • the substitutions to the RP (amino acids 4-5 of SEQ ID NO: 190) amino acid sequence in the Loop-8 region of FGF19 is an Arg (R) to Leu (L) substitution and a Pro (P) to Glu (E) substitution.
  • the foregoing substitution(s) in the Loop-8 region of FGF19 is in the corresponding FGF19 sequence thereof in a variant peptide provided herein. That is, said substitutions within a corresponding FGF19 sequence (e.g EIRPD, IRP or RP) of a peptide variant provided herein is also contemplated.
  • a peptide sequence includes or consists of a FGF19 variant having one or more amino acid substitutions, insertions or deletions compared to a reference or wild type FGF19.
  • a peptide sequence includes or consists of a FGF21 sequence variant having one or more amino acid substitutions, insertions or deletions compared to a reference or wild type FGF21.
  • a peptide sequence includes or consists of a portion of a FGF19 sequence fused to a portion of a FGF21 sequence.
  • a peptide sequence includes or consists of a portion of a FGF19 sequence fused to a portion of a FGF21 sequence, where the FGF19 and/or FGF21 sequence portion(s) have one or more amino acid substitutions, insertions or deletions compared to a reference or wild type FGF19 and/or FGF21.
  • Examples of such sequences are disclosed in PCT Pub. No. WO 2013/006486 and US Pub. No. 2013/0023474, as well as PCT Publ. No. WO 2014/085365, published June 5, 2014. Table 1 and the Sequence Listing also set forth representative sequences that may be used in the methods provided herein.
  • the treatment peptides provided herein include variants and fusions of FGF19 and/or FGF21 peptide sequences.
  • the treatment peptides include one or more variant or fusion FGF19 and/or FGF21 peptide.
  • the methods provided herein include contacting or administering to a subject one or more nucleic acid molecules encoding a variant or fusion FGF19 and/or FGF21 peptide sequence (for example, an expression control element in operable linkage with the nucleic acid encoding the peptide sequence, optionally including a vector), in an amount effective for treating a disease associated with elevated lactate.
  • a representative reference or wild type FGF19 sequence is set forth as:
  • a representative reference or wild type FGF21 sequence is set forth as:
  • FGF21 allelic variants include, e.g., M70, M71 and M72.
  • a peptide sequence comprises or consists of a FGF19 variant having one or more amino acid substitutions, insertions or deletions compared to a reference or wild type FGF19.
  • a peptide sequence comprises or consists of a FGF21 sequence variant having one or more amino acid substitutions, insertions or deletions compared to a reference or wild type FGF21.
  • a peptide sequence comprises or consists of a portion of a FGF19 sequence fused to a portion of a FGF21 sequence.
  • a peptide sequence comprises or consists of a portion of a FGF19 sequence fused to a portion of a FGF21 sequence, where the FGF19 and/or FGF21 sequence portion(s) have one or more amino acid substitutions, insertions or deletions compared to a reference or wild type FGF19 and/or FGF21.
  • Examples of such sequences are disclosed in PCT Pub. No. WO 2013/006486 and US Pub. No. 2013/0023474, as well as PCT Publ. No. WO 2014/085365, published June 5, 2014. Table 1 and the Sequence Listing also sets forth representative sequences that may be used in the methods provided herein.
  • a peptide or chimeric sequence provided herein has at the N-terminal region first amino acid position an “M” residue, an “R” residue, a “S” residue, a “H” residue, a “P” residue, a “L” residue or an “D” residue.
  • a peptide or chimeric sequence peptide sequence does not have a “M” residue or an “R” residue at the first amino acid position of the N-terminal region.
  • the number of amino acids or residues in a peptide sequence provided herein will total less than about 250 (e.g., amino acids or mimetics thereof).
  • the number of residues comprise from about 20 up to about 200 residues (e.g, amino acids or mimetics thereof).
  • the number of residues comprise from about 50 up to about 200 residues (e.g, amino acids or mimetics thereof).
  • the number of residues comprise from about 100 up to about 195 residues ( e.g ., amino acids or mimetics thereof) in length.
  • Amino acids or residues can be linked by amide or by non-natural and non-amide chemical bonds including, for example, those formed with glutaraldehyde, N- hydroxysuccinimide esters, bifunctional maleimides, orN, N’-dicyclohexylcarbodiimide (DCC).
  • Non-amide bonds include, for example, ketomethylene, aminomethylene, olefin, ether, thioether and the like (see, e.g., Spatola in Chemistry and Biochemistry of Amino Acids. Peptides and Proteins. Vol. 7, pp 267-357 (1983), “Peptide and Backbone Modifications,” Marcel Decker, NY).
  • a peptide provided herein includes a portion of a FGF19 sequence and a portion of a FGF21 sequence
  • the two portions need not be joined to each other by an amide bond, but can be joined by any other chemical moiety or conjugated together via a linker moiety.
  • Also provided herein are subsequences, variants and modified forms of the exemplified peptide sequences (including the FGF19 and FGF21 variants and subsequences listed in the Sequence Listing, or Table 1), so long as the foregoing retains at least a detectable or measureable activity or function.
  • certain exemplified variant peptides for example, those having all or a portion of FGF21 sequence at the amino-terminus, have an “R” residue positioned at the N-terminus, which can be omitted.
  • certain exemplified variant peptides include an “M” residue positioned at the N-terminus, which can be appended to or further substituted for an omitted residue, such as an “R” residue.
  • peptide sequences at the N-terminus include any of: RDSS (SEQ ID NO: 115), DSS, MDSS (SEQ ID NO: 116) or MRDSS (SEQ ID NO: 117).
  • peptide sequences include those with the following residues at the N-terminus: MDSSPL (SEQ ID NO: 119), MSDSSPL (SEQ ID NO: 120) (cleaved to SDSSPL (SEQ ID NO: 112)) and MSSPL (SEQ ID NO: 113) (cleaved to SSPL (SEQ ID NO: 114)).
  • modified peptide sequences, nucleic acids and other compositions may have greater or less activity or function, or have a distinct function or activity compared with a reference unmodified peptide sequence, nucleic acid, or other composition, or may have a property desirable in a protein formulated for therapy (e.g . serum half-life), to elicit antibody for use in a detection assay, and/or for protein purification.
  • a protein formulated for therapy e.g . serum half-life
  • a peptide sequence provided herein can be modified to increase serum half-life, to increase in vitro and/or in vivo stability of the protein, etc.
  • Particular examples of such subsequences, variants and modified forms of the peptide sequences exemplified herein include substitutions, deletions and/or insertions/additions of one or more amino acids, to or from the amino-terminus, the carboxy-terminus or internally.
  • substitutions, deletions and/or insertions/additions of one or more amino acids include substitutions, deletions and/or insertions/additions of one or more amino acids, to or from the amino-terminus, the carboxy-terminus or internally.
  • substitutions, deletions and/or insertions/additions of one or more amino acids to or from the amino-terminus, the carboxy-terminus or internally.
  • substitutions, deletions and/or insertions/additions of one or more amino acids to or from the amino-terminus, the carboxy-terminus or internally.
  • substitution of an amino acid residue for another amino acid residue within the peptide sequence is a deletion of one or more amino acid residues from
  • the number of residues substituted, deleted or inserted/added are one or more amino acids (e.g, 1-3, 3-5, 5-10, 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- 225, 225-250, or more) of a peptide sequence.
  • amino acids e.g, 1-3, 3-5, 5-10, 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- 225, 225-250, or more
  • a FGF19 or FGF21 sequence can have few or many amino acids substituted, deleted or inserted/added (e.g, 1-3, 3-5, 5-10, 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, 200-225, 225-250, or more).
  • amino acids substituted, deleted or inserted/added e.g, 1-3, 3-5, 5-10, 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, 200-225, 225-250, or more).
  • a FGF19 amino acid sequence can include or consist of an amino acid sequence of about 1-3, 3-5, 5-10, 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, 200-225, 225-250, or more amino acids from FGF21; or a FGF21 amino acid or sequence can include or consist of an amino acid sequence of about 1-3, 3-5, 5-10, 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, 200-225, 225-250, or more amino acids from FGF19.
  • substitutions include substituting a D residue for an L-residue. Accordingly, although residues are listed in the L-isomer configuration, D-amino acids at any particular or all positions of the peptide sequences provided herein are included, unless a D- isomer leads to a sequence that has no detectable or measurable function. [00105] Additional specific examples are non-conservative and conservative substitutions.
  • a “conservative substitution” is a replacement of one amino acid by a biologically, chemically or structurally similar residue. Biologically similar means that the substitution is compatible with a biological activity, e.g ., activity that ameliorate disease associated with elevated lactate and/or the manifestations thereof.
  • Structurally similar means that the amino acids have side chains with similar length, such as alanine, glycine and serine, or having similar size, or the structure of a first, second or additional peptide sequence is maintained.
  • Chemical similarity means that the residues have the same charge or are both hydrophilic and hydrophobic. Particular examples include the substitution of one hydrophobic residue, such as isoleucine, valine, leucine or methionine, for another, or the substitution of one polar residue for another, such as the substitution of arginine for lysine, glutamic for aspartic acids, or glutamine for asparagine, serine for threonine, etc. Routine assays can be used to determine whether a subsequence, variant or modified form has activity, e.g. , activity that ameliorate disease associated with elevated lactate and/or the manifestations thereof.
  • Particular examples of subsequences, variants and modified forms of the peptide sequences exemplified herein have 50%-60%, 60%-70%, 70%-75%, 75%-80%, 80%-85%, 85%- 90%, 90%-95%, or 96%, 97%, 98%, or 99% identity to a reference peptide sequence.
  • identity and “homology” and grammatical variations thereof mean that two or more referenced entities are the same. Thus, where two amino acid sequences are identical, they have the identical amino acid sequence. “Areas, regions or domains of identity” mean that a portion of two or more referenced entities are the same.
  • a BLASTP algorithm is typically used in combination with a scoring matrix, such as PAMIOO, PAM 250, BLOSUM 62 or BLOSUM 50.
  • FASTA e.g, FASTA2 and FASTA3
  • SSEARCH sequence comparison programs are also used to quantitate the extent of identity (Pearson et al, Proc. Natl. Acad. Sci. USA 85:2444 (1988); Pearson, Methods Mol Biol. 132:185 (2000); and Smith et al. , J. Mol. Biol. 147:195 (1981)).
  • Programs for quantitating protein structural similarity using Delaunay-based topological mapping have also been developed (Bostick et al. , Biochem Biophys Res Commun. 304:320 (2003)).
  • an “amino acid” or “residue” includes conventional alpha- amino acids as well as beta-amino acids; alpha, alpha disubstituted amino acids; and N- substituted amino acids, wherein at least one side chain is an amino acid side chain moiety as defined herein.
  • An “amino acid” further includes N-alkyl alpha-amino acids, wherein the N- terminus amino group has a Ci to G linear or branched alkyl substituent.
  • amino acid therefore includes stereoisomers and modifications of naturally occurring protein amino acids, non-protein amino acids, post-translationally modified amino acids (e.g, by glycosylation, phosphorylation, ester or amide cleavage, etc.), enzymatically modified or synthesized amino acids, derivatized amino acids, constructs or structures designed to mimic amino acids, amino acids with a side chain moiety modified, derivatized from naturally occurring moieties, or synthetic, or not naturally occurring, etc. Modified and unusual amino acids are included in the peptide sequences provided herein (see, for example, in Synthetic Peptides: A User’s Guide: Hruby et al, Biochem. J. 268:249 (1990); and Toniolo C., Int. J. Peptide Protein Res. 35:287 (1990)).
  • amino acid side chain moiety includes any side chain of any amino acid, as the term “amino acid” is defined herein. This therefore includes the side chain moiety in naturally occurring amino acids. It further includes side chain moieties in modified naturally occurring amino acids as set forth herein and known to one of skill in the art, such as side chain moieties in stereoisomers and modifications of naturally occurring protein amino acids, non-protein amino acids, post-translationally modified amino acids, enzymatically modified or synthesized amino acids, derivatized amino acids, constructs or structures designed to mimic amino acids, etc. For example, the side chain moiety of any amino acid disclosed herein or known to one of skill in the art is included within the definition.
  • a “derivative of an amino acid side chain moiety” is included within the definition of an amino acid side chain moiety.
  • a single amino acid including stereoisomers and modifications of naturally occurring protein amino acids, non-protein amino acids, post-translationally modified amino acids, enzymatically-synthesized amino acids, non-naturally occurring amino acids including derivatized amino acids, an alpha, alpha disubstituted amino acid derived from any of the foregoing (i.e., an alpha, alpha disubstituted amino acid, wherein at least one side chain is the same as that of the residue from which it is derived), a beta-amino acid derived from any of the foregoing (i.e., a beta-amino acid which, other than for the presence of a beta-carbon, is the same as the residue from which it is derived) etc., including all of the foregoing can be referred to herein as a “residue.”
  • Suitable substituents, in addition to the side chain moiety of the alpha- amino acid include Ci to Ce linear or branched alkyl.
  • Aib is an example of an alpha, alpha disubstituted amino acid. While alpha, alpha disubstituted amino acids can be referred to using conventional L- and D-isomeric references, it is to be understood that such references are for convenience, and that where the substituents at the alpha-position are different, such amino acid can interchangeably be referred to as an alpha, alpha disubstituted amino acid derived from the L- or D-isomer, as appropriate, of a residue with the designated amino acid side chain moiety.
  • (S)-2-Amino-2-methyl-hexanoic acid can be referred to as either an alpha, alpha disubstituted amino acid derived from L-Nle (norleucine) or as an alpha, alpha disubstituted amino acid derived from D-Ala.
  • Aib can be referred to as an alpha, alpha disubstituted amino acid derived from Ala. Whenever an alpha, alpha disubstituted amino acid is provided, it is to be understood as including all (R) and (S) configurations thereof.
  • N- substituted amino acid includes any amino acid wherein an amino acid side chain moiety is covalently bonded to the backbone amino group, optionally where there are no substituents other than H in the alpha-carbon position.
  • Sarcosine is an example of an N- substituted amino acid.
  • sarcosine can be referred to as an N-substituted amino acid derivative of Ala, in that the amino acid side chain moiety of sarcosine and Ala is the same, i.e., methyl.
  • covalent modifications of the peptide sequences including subsequences, variants and modified forms of the peptide sequences exemplified herein are provided.
  • An exemplary type of covalent modification includes reacting targeted amino acid residues with an organic derivatizing agent that is capable of reacting with selected side chains or the N- or C-terminal residues of the peptide.
  • Derivatization with bifunctional agents is useful, for instance, for cross-linking peptide to a water-insoluble support matrix or surface for use in the method for purifying anti-peptide antibodies, and vice-versa.
  • cross linking agents include, e.g ., l,l-bis(diazoacetyl)-2-phenylethane, glutaraldehyde, N-hydroxysuccinimide esters, for example, esters with 4-azidosalicylic acid, homobifunctional imidoesters, including disuccinimidyl esters such as 3,3’-dithiobis(succinimidylpropionate), bifunctional maleimides such as bis-N-maleimido-1, 8-octane and agents such as methyl-3-[(p- azi dopheny l)dithi o]propi oimi date .
  • Exemplified peptide sequences, and subsequences, variants and modified forms of the peptide sequences exemplified herein can also include alterations of the backbone for stability, derivatives, and peptidomimetics.
  • peptidomimetic includes a molecule that is a mimic of a residue (referred to as a “mimetic”), including but not limited to piperazine core molecules, keto-piperazine core molecules and diazepine core molecules.
  • an amino acid mimetic of a peptide sequence provided herein includes both a carboxyl group and amino group, and a group corresponding to an amino acid side chain, or in the case of a mimetic of Glycine, no side chain other than hydrogen.
  • these would include compounds that mimic the sterics, surface charge distribution, polarity, etc. of a naturally occurring amino acid, but need not be an amino acid, which would impart stability in the biological system.
  • Proline may be substituted by other lactams or lactones of suitable size and substitution;
  • Leucine may be substituted by an alkyl ketone, N-substituted amide, as well as variations in amino acid side chain length using alkyl, alkenyl or other substituents, others may be apparent to the skilled artisan.
  • the essential element of making such substitutions is to provide a molecule of roughly the same size and charge and configuration as the residue used to design the molecule.
  • binding when used in reference to a peptide sequence, means that the peptide sequence interacts at the molecular level. Specific and selective binding can be distinguished from non-specific binding using assays known in the art (e.g., competition binding, immunoprecipitation, ELISA, flow cytometry, Western blotting).
  • Peptides and peptidomimetics can be produced and isolated using methods known in the art. Peptides can be synthesized, in whole or in part, using chemical methods (see, e.g, Caruthers (1980). Nucleic Acids Res. Symp. Ser. 215; Horn (1980); and Banga, A.K., Therapeutic Peptides and Proteins. Formulation. Processing and Delivery Systems (1995) Technomic Publishing Co., Lancaster, PA). Peptide synthesis can be performed using various solid-phase techniques (see, e.g ., Roberge Science 269:202 (1995); Merrifield, Methods Enzymol. 289:3 (1997)) and automated synthesis may be achieved, e.g.
  • Peptides and peptide mimetics can also be synthesized using combinatorial methodologies. Synthetic residues and polypeptides incorporating mimetics can be synthesized using a variety of procedures and methodologies known in the art (see, e.g. , Organic Syntheses Collective Volumes, Gilman, etal. (Eds) John Wiley & Sons, Inc., NY). Modified peptides can be produced by chemical modification methods (see, for example, Belousov, Nucleic Acids Res. 25:3440 (1997); Frenkel, Free Radic. Biol. Med.
  • Peptide sequence variations, derivatives, substitutions and modifications can also be made using methods such as oligonucleotide-mediated (site-directed) mutagenesis, alanine scanning, and PCR-based mutagenesis.
  • Site-directed mutagenesis (Carter et al ., Nucl. Acids Res., 13:4331 (1986); Zoller etal, Nucl. Acids Res. 10:6487 (1987)), cassette mutagenesis (Wells etal. , Gene 34:315 (1985)), restriction selection mutagenesis (Wells et al. , Philos. Trans. R. Soc. London SerA 317:415 (1986)) and other techniques can be performed on cloned DNA to produce peptide sequences, variants, fusions and chimeras provided herein, and variations, derivatives, substitutions and modifications thereof.
  • a “synthesized” or “manufactured” peptide sequence is a peptide made by any method involving manipulation by the hand of man. Such methods include, but are not limited to, the aforementioned, such as chemical synthesis, recombinant DNA technology, biochemical or enzymatic fragmentation of larger molecules, and combinations of the foregoing.
  • Peptide sequences provided herein including subsequences, sequence variants and modified forms of the exemplified peptide sequences (e.g., sequences listed in the Sequence Listing or Table 1), can also be modified to form a chimeric molecule.
  • peptide sequences that include a heterologous domain can be added to the amino-terminus or at the carboxyl-terminus of the peptide sequence.
  • heterologous domains can also be positioned within the peptide sequence, and/or alternatively flanked by FGF19 and/or FGF21 derived amino acid sequences.
  • peptide also includes dimers or multimers (oligomers) of peptides.
  • dimers or multimers (oligomers) of the exemplified peptide sequences are provided herein, as well as subsequences, variants and modified forms of the exemplified peptide sequences, including sequences listed in the Sequence Listing or Table 1.
  • a peptide sequence provided herein comprises an amino acid sequence set forth in Table 1. In other embodiments, a peptide sequence provided herein consists of an amino acid sequence set forth in Table 1.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:l. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:2. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:3. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:4. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:5. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:6. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:7.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:8. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:9. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 10. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 11. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 12. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 13. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 14. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 15.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 16. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 17. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 18. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 19. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:20. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:21. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:22. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:23.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:24. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:25. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:26. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:27. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:28. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:29. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:30.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:31. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:32. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:33. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:34. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:35. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:36. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:37.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:38. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:39. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:40. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:41. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:42. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:43. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:44.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:45. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:46. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:47. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:48. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:49. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:50. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:51.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:52. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:53. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:54. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:55. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:56. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:57. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:58.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:59. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:60. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:61. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:62. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:63. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:64. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:65.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:66. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:67. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:68. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:69. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:70. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:71. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:72.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:73. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:74. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:75. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:76. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:77. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:78. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:79.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:80. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:81. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:82. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:83. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:84. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:85. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:86.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:87. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:88. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:89. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:90. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:91. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:92. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:93.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:94. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:95. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:96. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:97. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:98. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 138. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 139.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 140. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 141. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 142. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 143. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 144. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 145. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 146.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 147. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 148. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 149. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 150. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 151. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 152. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 153.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 154. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 155. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 156. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 157. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 158. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 159. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 160.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 161. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 162. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 163. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 164. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 165. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 166. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 167.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 168. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 192. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 193. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 194. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 195. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 196. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 197.
  • the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 198. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO: 199. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:200. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:201. In other embodiments, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:202. In one embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:203. In another embodiment, the peptide sequence comprises an amino acid sequence set forth in SEQ ID NO:204. In certain embodiments of the various peptide sequences provided herein, the R residue at the N-terminus is deleted.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 1. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:2. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:3. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:4. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:5. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:6. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:7.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:8. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:9. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 10. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 11. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 12. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 13. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 14.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 15. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 16. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 17. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 18. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 19. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:20. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:21.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:22. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:23. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:24. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:25. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:26. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:27. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:28.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:29. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:30. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:31. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:32. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:33. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:34. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:35.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:36. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:37. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:38. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:39. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:40. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:41. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:42.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:43. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:44. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:45. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:46. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:47. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:48. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:49.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:50. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:51. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:52. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:53. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:54. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:55. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:56.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:57. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 58. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:59. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:60. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:61. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:62. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:63.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:64. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:65. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:66. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:67. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:68. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:69. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:70.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:71. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:72. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:73. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:74. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:75. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:76. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:77.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:78. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:79. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:80. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:81. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:82. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:83. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:84.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:85. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:86. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:87. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:88. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:89. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:90. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:91.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:92. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:93. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:94. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:95. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:96. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:97. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:98.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 138. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 139. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 140. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 141. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 142. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:143. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 144.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 145. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 146. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 147. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 148. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 149. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 150. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 151.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 152. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 153. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 154. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:155. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 156. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 157. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 158.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 159. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 160. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 161. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 162. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 163. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 164. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 165.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 166. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 167. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 168. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 192. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 193. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 194. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 195.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 196. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 197. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 198. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO: 199. In one embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:200. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:201. In other embodiments, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:202.
  • the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:203. In another embodiment, the peptide sequence consists of an amino acid sequence set forth in SEQ ID NO:204. In certain embodiments of the various peptide sequences provided herein, the R residue at the N-terminus is deleted.
  • nucleic acid molecules encoding peptide sequences provided herein, including subsequences, sequence variants and modified forms of the sequences listed in the Sequence Listing or Table 1, and in PCT Pub. No. WO 2013/006486 and US Pub. No. 2013/0023474, as well as PCT Publ. No. WO 2014/085365, and vectors that include nucleic acid encoding the peptides used in the methods described herein.
  • nucleic acids include those that encode the exemplified peptide sequences disclosed herein, as well as those encoding functional subsequences, sequence variants and modified forms of the exemplified peptide sequences, so long as the foregoing retain at least detectable or measureable activity or function useful in the modulation of lactate homeostasis, or in the treatment or prevention of disease associated with elevated lactate.
  • Nucleic acid which can also be referred to herein as a gene, polynucleotide, nucleotide sequence, primer, oligonucleotide or probe, refers to natural or modified purine- and pyrimidine-containing polymers of any length, either polyribonucleotides or polydeoxyribonucleotides or mixed polyribo-polydeoxyribo nucleotides and a-anomeric forms thereof.
  • the two or more purine- and pyrimidine-containing polymers are typically linked by a phosphoester bond or analog thereof.
  • the terms can be used interchangeably to refer to all forms of nucleic acid, including deoxyribonucleic acid (DNA) and ribonucleic acid (RNA).
  • the nucleic acids can be single strand, double, or triplex, linear or circular. Nucleic acids include genomic DNA and cDNA. RNA nucleic acid can be spliced or unspliced mRNA, rRNA, tRNA or antisense. Nucleic acids include naturally occurring, synthetic, as well as nucleotide analogs and derivatives.
  • SNP single nucleotide polymorphism
  • Single-nucleotide polymorphisms may fall within coding sequences of genes, non-coding regions of genes, or in the intergenic regions (regions between genes). SNPs within a coding sequence do not necessarily change the amino acid sequence of the protein that is produced, due to degeneracy of the genetic code.
  • the nucleic acid molecules provided herein include sequences degenerate with respect to nucleic acid molecules encoding the peptide sequences useful in the methods provided herein.
  • nucleic acid sequences encoding peptide sequences, including subsequences, variants and modified forms of the peptide sequences exemplified herein (e.g ., in the Sequence Listing or Table 1), are provided.
  • Nucleic acid can be produced using any of a variety of known standard cloning and chemical synthesis methods, and can be altered intentionally by site-directed mutagenesis or other recombinant techniques known to one skilled in the art. Purity of polynucleotides can be determined through, for example, sequencing, gel electrophoresis, and UV spectrometry.
  • Nucleic acids may be inserted into a nucleic acid construct in which expression of the nucleic acid is influenced or regulated by an “expression control element,” referred to herein as an “expression cassette.”
  • expression control element refers to one or more nucleic acid sequence elements that regulate or influence expression of a nucleic acid sequence to which it is operatively linked.
  • An expression control element can include, as appropriate, promoters, enhancers, transcription terminators, gene silencers, a start codon (e.g., ATG) in front of a protein-encoding gene, etc.
  • An expression control element operatively linked to a nucleic acid sequence controls transcription and, as appropriate, translation of the nucleic acid sequence.
  • the term “operatively linked” refers to a juxtaposition wherein the referenced components are in a relationship permitting them to function in their intended manner.
  • expression control elements are juxtaposed at the 5’ or the 3’ ends of the genes but can also be intronic.
  • Expression control elements include elements that activate transcription constitutively, that are inducible (i.e., require an external signal or stimuli for activation), or derepressible (i.e., require a signal to turn transcription off; when the signal is no longer present, transcription is activated or “derepressed”). Also included in the expression cassettes provided herein are control elements sufficient to render gene expression controllable for specific cell types or tissues (i.e., tissue-specific control elements). Typically, such elements are located up stream or downstream (i.e., 5’ or 3’) of the coding sequence. Promoters are generally positioned 5’ of the coding sequence. Promoters, produced by recombinant DNA or synthetic techniques, can be used to provide for transcription of the polynucleotides provided herein. A “promoter” typically means a minimal sequence element sufficient to direct transcription.
  • Nucleic acids may be inserted into a plasmid for transformation into a host cell and for subsequent expression and/or genetic manipulation.
  • a plasmid is a nucleic acid that can be stably propagated in a host cell; plasmids may optionally contain expression control elements in order to drive expression of the nucleic acid.
  • a vector is synonymous with a plasmid. Plasmids and vectors generally contain at least an origin of replication for propagation in a cell and a promoter.
  • Plasmids and vectors may also include an expression control element for expression in a host cell, and are therefore useful for expression and/or genetic manipulation of nucleic acids encoding peptide sequences, expressing peptide sequences in host cells and organisms, or producing peptide sequences, for example.
  • transgene means a polynucleotide that has been introduced into a cell or organism by artifice.
  • the transgene has been introduced by genetic manipulation or “transformation” of the cell.
  • a cell or progeny thereof into which the transgene has been introduced is referred to as a “transformed cell” or “transformant.”
  • the transgene is included in progeny of the transformant or becomes a part of the organism that develops from the cell.
  • Transgenes may be inserted into the chromosomal DNA or maintained as a self-replicating plasmid, YAC, minichromosome, or the like.
  • Bacterial system promoters include T7 and inducible promoters such as pL of bacteriophage l, plac, ptrp, ptac (ptrp-lac hybrid promoter) and tetracycline-responsive promoters.
  • Insect cell system promoters include constitutive or inducible promoters ( e.g ., ecdysone).
  • Mammalian cell constitutive promoters include SV40, RSV, bovine papilloma virus (BPV) and other virus promoters, or inducible promoters derived from the genome of mammalian cells (e.g., metallothionein IIA promoter; heat shock promoter) or from mammalian viruses (e.g, the adenovirus late promoter; the inducible mouse mammary tumor virus long terminal repeat).
  • a retroviral genome can be genetically modified for introducing and directing expression of a peptide sequence in appropriate host cells.
  • expression systems further include vectors designed for in vivo use.
  • adenoviral vectors U.S. Patent Nos. 5,700,470 and 5,731,172
  • adeno-associated vectors U.S. Patent No. 5,604,090
  • herpes simplex virus vectors U.S. Patent No. 5,501,979
  • retroviral vectors U.S. Patent Nos. 5,624,820, 5,693,508 and 5,674,703
  • BPV vectors U.S. Patent No. 5,719,054
  • CMV vectors U.S. Patent No. 5,561,063
  • parvovirus rotavirus, Norwalk virus and lentiviral vectors (see, e.g ., U.S. Patent No. 6,013,516).
  • Vectors include those that deliver genes to cells of the intestinal tract, including the stem cells (Croyle etal. , Gene Ther. 5:645 (1998); S.J. Henning, Adv. Drug Deliv. Rev. 17:341 (1997), U.S. Patent Nos. 5,821,235 and 6,110,456). Many of these vectors have been approved for human studies.
  • Yeast vectors include constitutive and inducible promoters (see, e.g. , Ausubel et al ., In: Current Protocols in Molecular Biology Vol. 2, Ch. 13, ed., Greene Publish. Assoc. & Wiley Interscience, 1988; Grant etal. Methods in Enzymology, 153:516 (1987), eds. Wu & Grossman; Bitter Methods in Enzymology, 152:673 (1987), eds. Berger & Kimmel, Acad. Press, N.Y.; and, Strathem etal. , The Molecular Biology of the Yeast Saccharomyces (1982) eds. Cold Spring Harbor Press, Vols.
  • a constitutive yeast promoter such as ADH or LEU2 or an inducible promoter such as GAL may be used (R. Rothstein In: DNA Cloning. A Practical Approach. Vol.11, Ch. 3, ed. D.M. Glover, IRL Press, Wash., D.C., 1986).
  • Vectors that facilitate integration of foreign nucleic acid sequences into a yeast chromosome, via homologous recombination for example, are known in the art.
  • Yeast artificial chromosomes YAC are typically used when the inserted polynucleotides are too large for more conventional vectors (e.g, greater than about 12 Kb).
  • Expression vectors also can contain a selectable marker conferring resistance to a selective pressure or identifiable marker (e.g, beta-galactosidase), thereby allowing cells having the vector to be selected for, grown and expanded.
  • a selectable marker can be on a second vector that is co-transfected into a host cell with a first vector containing a nucleic acid encoding a peptide sequence.
  • Selection systems include, but are not limited to, herpes simplex virus thymidine kinase gene (Wigler et al, Cell 11 :223 (1977)), hypoxanthine-guanine phosphoribosyltransferase gene (Szybalska etal, Proc. Natl.
  • adenine phosphoribosyltransferase genes that can be employed in tk-, hgprt or aprt cells, respectively.
  • antimetabolite resistance can be used as the basis of selection for dhfr, which confers resistance to methotrexate (O’Hare et al. , Proc. Natl. Acad. Sci. USA 78:1527 (1981)); the gpt gene, which confers resistance to mycophenolic acid (Mulligan et al, Proc. Natl. Acad. Sci.
  • neomycin gene which confers resistance to aminoglycoside G-418 (Colberre-Garapin etal. , J. Mol. Biol. 150:1(1981)); puromycin; and hygromycin gene, which confers resistance to hygromycin (Santerre etal. , Gene 30:147 (1984)).
  • Additional selectable genes include trpB, which allows cells to utilize indole in place of tryptophan; hisD, which allows cells to utilize histinol in place of histidine (Hartman etal. , Proc. Natl. Acad. Sci.
  • a transformed cell(s) in vitro , ex vivo and in vivo ) and host cells that produce a variant or fusion of FGF19 and/or FGF21 as set forth herein, where expression of the variant or fusion of FGF19 and/or FGF21 is conferred by a nucleic acid encoding the variant or fusion of FGF19 and/or FGF21.
  • a “transformed” or “host” cell is a cell into which a nucleic acid is introduced that can be propagated and/or transcribed for expression of an encoded peptide sequence. The term also includes any progeny or subclones of the host cell.
  • Transformed and host cells that express peptide sequences provided herein typically include a nucleic acid that encodes the peptide sequence.
  • a transformed or host cell is a prokaryotic cell.
  • a transformed or host cell is a eukaryotic cell.
  • the eukaryotic cell is a yeast or mammalian ( e.g ., human, primate, etc.) cell.
  • Transformed and host cells include but are not limited to microorganisms such as bacteria and yeast; and plant, insect and mammalian cells.
  • bacteria transformed with recombinant bacteriophage nucleic acid, plasmid nucleic acid or cosmid nucleic acid expression vectors yeast transformed with recombinant yeast expression vectors
  • plant cell systems infected with recombinant virus expression vectors e.g., cauliflower mosaic virus, CaMV; tobacco mosaic virus, TMV
  • transformed with recombinant plasmid expression vectors e.g., Ti plasmid
  • insect cell systems infected with recombinant virus expression vectors e.g, baculovirus
  • animal cell systems infected with recombinant virus expression vectors e.g., retroviruses, adenovirus, vaccinia virus, or transformed animal cell systems engineered for transient or stable propagation or expression.
  • a transformed cell can be in a subject.
  • a cell in a subject can be transformed with a nucleic acid that encodes a peptide sequence as set forth herein in vivo.
  • a cell can be transformed in vitro with a transgene or polynucleotide, and then transplanted into a tissue of subject in order to effect treatment.
  • a primary cell isolate or an established cell line can be transformed with a transgene or polynucleotide that encodes a variant of FGF19 and/or FGF21 or a fusion/chimeric sequence (or variant) thereof, such as a chimeric peptide sequence including all or a portion of FGF19, or including all or a portion of FGF21, and then optionally transplanted into a tissue of a subject.
  • a transgene or polynucleotide that encodes a variant of FGF19 and/or FGF21 or a fusion/chimeric sequence (or variant) thereof, such as a chimeric peptide sequence including all or a portion of FGF19, or including all or a portion of FGF21, and then optionally transplanted into a tissue of a subject.
  • Non-limiting target cells for expression of peptide sequences include pancreas cells (islet cells), muscle cells, mucosal cells and endocrine cells.
  • pancreas cells islet cells
  • muscle cells can provide inducible production (secretion) of a variant of FGF19 and/or FGF21, or a fusion/chimeric sequence (or variant) thereof, such as a chimeric peptide sequence including all or a portion of FGF19, or including all or a portion of FGF21.
  • Additional cells to transform include stem cells or other multipotent or pluripotent cells, for example, progenitor cells that differentiate into the various pancreas cells (islet cells), muscle cells, mucosal cells and endocrine cells. Targeting stem cells provides longer term expression of peptide sequences provided herein.
  • the term “cultured,” when used in reference to a cell, means that the cell is grown in vitro.
  • a particular example of such a cell is a cell isolated from a subject, and grown or adapted for growth in tissue culture.
  • Another example is a cell genetically manipulated in vitro , and transplanted back into the same or a different subject.
  • isolated when used in reference to a cell, means a cell that is separated from its naturally occurring in vivo environment. “Cultured” and “isolated” cells may be manipulated by the hand of man, such as genetically transformed. These terms include any progeny of the cells, including progeny cells that may not be identical to the parental cell due to mutations that occur during cell division. The terms do not include an entire human being. [00146] Nucleic acids encoding peptide sequences provided herein can be introduced for stable expression into cells of a whole organism. Such organisms, including non-human transgenic animals, are useful for studying the effect of peptide expression in a whole animal and therapeutic benefit.
  • a variant of FGF19 and/or FGF21 or a fusion/chimeric sequence (or variant) thereof such as a chimeric peptide sequence including all or a portion of FGF19, or including all or a portion of FGF21 as set forth herein, in mice.
  • mice strains that develop or are susceptible to developing a particular disease are also useful for introducing therapeutic proteins as described herein in order to study the effect of therapeutic protein expression in the disease- susceptible mouse.
  • Transgenic and genetic animal models that are susceptible to particular disease or physiological conditions such as streptozotocin (STZ)-induced diabetic (STZ) mice, are appropriate targets for expressing variants of FGF19 and/or FGF21, fusions/chimeric sequences (or variant) thereof, such as a chimeric peptide sequence including all or a portion of FGF19, or including all or a portion of FGF21, as set forth herein.
  • non-human transgenic animals that produce a variant of FGF19 and/or FGF21, or a fusion/chimeric sequence (or variant) thereof, such as a chimeric peptide sequence including all or a portion of FGF19, or including all or a portion of FGF21, the production of which is not naturally occurring in the animal which is conferred by a transgene present in somatic or germ cells of the animal.
  • transgenic animal refers to an animal whose somatic or germ line cells bear genetic information received, directly or indirectly, by deliberate genetic manipulation at the subcellular level, such as by microinjection or infection with recombinant virus.
  • the term “transgenic” further includes cells or tissues (i.e., “transgenic cell,” “transgenic tissue”) obtained from a transgenic animal genetically manipulated as described herein.
  • a “transgenic animal” does not encompass animals produced by classical crossbreeding or in vitro fertilization, but rather denotes animals in which one or more cells receive a nucleic acid molecule.
  • Transgenic animals provided herein can be either heterozygous or homozygous with respect to the transgene.
  • mice including mice, sheep, pigs and frogs
  • transgenic animals including mice, sheep, pigs and frogs
  • U.S. Patent Nos. 5,721,367, 5,695,977, 5,650,298, and 5,614,396 are well known in the art (see, e.g., U.S. Patent Nos. 5,721,367, 5,695,977, 5,650,298, and 5,614,396) and, as such, are additionally included.
  • Peptide sequences, nucleic acids encoding peptide sequences, vectors and transformed host cells expressing peptide sequences include isolated and purified forms.
  • isolated when used as a modifier of a composition provided herein, means that the composition is separated, substantially, completely, or at least in part, from one or more components in an environment.
  • compositions that exist in nature, when isolated are substantially free of one or more materials with which they normally associate with in nature, for example, one or more protein, nucleic acid, lipid, carbohydrate or cell membrane.
  • isolated does not exclude alternative physical forms of the composition, such as variants, modifications or derivatized forms, fusions and chimeras, multimers/oligomers, etc ., or forms expressed in host cells.
  • isolated also does not exclude forms ( e.g ., pharmaceutical compositions, combination compositions, etc.) in which there are combinations therein, any one of which is produced by the hand of man.
  • An “isolated” composition can also be “purified” when free of some, a substantial number of, or most or all of one or more other materials, such as a contaminant or an undesired substance or material.
  • the term “recombinant,” when used as a modifier of peptide sequences, nucleic acids encoding peptide sequences, etc ., means that the compositions have been manipulated (i.e., engineered) in a fashion that generally does not occur in nature (e.g., in vitro).
  • a particular example of a recombinant peptide would be where a peptide sequence provided herein is expressed by a cell transfected with a nucleic acid encoding the peptide sequence.
  • a particular example of a recombinant nucleic acid would be a nucleic acid (e.g, genomic or cDNA) encoding a peptide sequence cloned into a plasmid, with or without 5’, 3’ or intron regions that the gene is normally contiguous within the genome of the organism.
  • a recombinant peptide or nucleic acid is a hybrid or fusion sequence, such as a chimeric peptide sequence comprising a portion of FGF19 and a portion of FGF21.
  • Improvements of physical properties include, for example, modulating immunogenicity; methods of increasing solubility, bioavailability, serum half-life, and/or therapeutic half-life; and/or modulating biological activity. Certain modifications may also be useful to, for example, raise of antibodies for use in detection assays (e.g, epitope tags) and to provide for ease of protein purification. Such improvements must generally be imparted without adversely impacting the bioactivity of the treatment modality and/or increasing its immunogenicity.
  • Pegylation of is one particular modification contemplated herein, while other modifications include, but are not limited to, glycosylation (N- and O-linked); polysialylation; albumin fusion molecules comprising serum albumin (e.g ., human serum albumin (HSA), cyno serum albumin, or bovine serum albumin (BSA)); albumin binding through, for example a conjugated fatty acid chain (acylation); and Fc-fusion proteins.
  • serum albumin e.g ., human serum albumin (HSA), cyno serum albumin, or bovine serum albumin (BSA)
  • Fc-fusion proteins Fc-fusion proteins
  • Pegylation The clinical effectiveness of protein therapeutics is often limited by short plasma half-life and susceptibility to protease degradation.
  • Studies of various therapeutic proteins e.g., filgrastim
  • conjugating or linking the protein to any of a variety of nonproteinaceous polymers, e.g, polyethylene glycol (PEG), polypropylene glycol, or polyoxyalkylenes. This is frequently effected by a linking moiety covalently bound to both the protein and the nonproteinaceous polymer, e.g, a PEG.
  • PEG-conjugated biomolecules have been shown to possess clinically useful properties, including better physical and thermal stability, protection against susceptibility to enzymatic degradation, increased solubility, longer in vivo circulating half-life and decreased clearance, reduced immunogenicity and antigenicity, and reduced toxicity.
  • pegylation itself may enhance activity.
  • PEGs suitable for conjugation to a polypeptide sequence are generally soluble in water at room temperature, and have the general formula R(0-CH2-CH2)n0-R, where R is hydrogen or a protective group such as an alkyl or an alkanol group, and where n is an integer from 1 to 1000. When R is a protective group, it generally has from 1 to 8 carbons.
  • R is hydrogen or a protective group such as an alkyl or an alkanol group, and where n is an integer from 1 to 1000. When R is a protective group, it generally has from 1 to 8 carbons.
  • the PEG conjugated to the polypeptide sequence can be linear or branched. Branched PEG derivatives, “star-PEGs” and multi-armed PEGs are contemplated by the present disclosure.
  • a molecular weight of the PEG used in embodiments provided herein is not restricted to any particular range, and examples are set forth elsewhere herein; by way of example, certain embodiments have molecular weights between 5kDa and 20kDa, while other embodiments have molecular weights between 4kDa and lOkDa.
  • Such compositions can be produced by reaction conditions and purification methods know in the art. Cation exchange chromatography may be used to separate conjugates, and a fraction is then identified which contains the conjugate having, for example, the desired number of PEGs attached, purified free from unmodified protein sequences and from conjugates having other numbers of PEGs attached.
  • Pegylation most frequently occurs at the alpha amino group at the N-terminus of the polypeptide, the epsilon amino group on the side chain of lysine residues, and the imidazole group on the side chain of histidine residues. Since most recombinant polypeptides possess a single alpha and a number of epsilon amino and imidazole groups, numerous positional isomers can be generated depending on the linker chemistry.
  • PEG may be bound to a polypeptide provided herein via a terminal reactive group (a “spacer” or “linker”) which mediates a bond between the free amino or carboxyl groups of one or more of the polypeptide sequences and polyethylene glycol.
  • a terminal reactive group a “spacer” or “linker”
  • the PEG having the spacer which may be bound to the free amino group includes N-hydroxysuccinylimide polyethylene glycol which may be prepared by activating succinic acid ester of polyethylene glycol with N- hydroxysuccinylimide.
  • Another activated polyethylene glycol which may be bound to a free amino group is 2,4-bis(0-methoxypolyethyleneglycol)-6-chloro-s-triazine, which may be prepared by reacting polyethylene glycol monomethyl ether with cyanuric chloride.
  • the activated polyethylene glycol which is bound to the free carboxyl group includes polyoxyethylenediamine.
  • Conjugation of one or more of the polypeptide sequences provided herein to PEG having a spacer may be carried out by various conventional methods.
  • the conjugation reaction can be carried out in solution at a pH of from 5 to 10, at temperature from 4°C to room temperature, for 30 minutes to 20 hours, utilizing a molar ratio of reagent to protein of from 4:1 to 30: 1.
  • Reaction conditions may be selected to direct the reaction towards producing predominantly a desired degree of substitution.
  • short reaction time tend to decrease the number of PEGs attached
  • high temperature, neutral to high pH e.g., pH>7)
  • longer reaction time tend to increase the number of PEGs attached.
  • the reaction is terminated by acidifying the reaction mixture and freezing at, e.g ., -20°C.
  • Pegylation of various molecules is discussed in, for example, U.S. Pat. Nos. 5,252,714; 5,643,575; 5,919,455; 5,932,462; and 5,985,263.
  • PEG mimetics also provided herein are uses of PEG mimetics.
  • Recombinant PEG mimetics have been developed that retain the attributes of PEG (e.g, enhanced serum half- life) while conferring several additional advantageous properties.
  • simple polypeptide chains comprising, for example, Ala, Glu, Gly, Pro, Ser and Thr
  • the peptide or protein drug of interest e.g, XTEN technology; Amunix; Mountain View, CA.
  • This obviates the need for an additional conjugation step during the manufacturing process.
  • established molecular biology techniques enable control of the side chain composition of the polypeptide chains, allowing optimization of immunogenicity and manufacturing properties.
  • Glvcosylation is meant to broadly refer to the enzymatic process by which glycans are attached to proteins, lipids or other organic molecules.
  • the use of the term “glycosylation” herein is generally intended to mean adding or deleting one or more carbohydrate moieties (either by removing the underlying glycosylation site or by deleting the glycosylation by chemical and/or enzymatic means), and/or adding one or more glycosylation sites that may or may not be present in the native sequence.
  • the phrase includes qualitative changes in the glycosylation of the native proteins involving a change in the nature and proportions of the various carbohydrate moieties present.
  • Glycosylation can dramatically affect the physical properties (e.g, solubility) of polypeptides and can also be important in protein stability, secretion, and subcellular localization. Glycosylated polypeptides may also exhibit enhanced stability or may improve one or more pharmacokinetic properties, such as half-life. In addition, solubility improvements can, for example, enable the generation of formulations more suitable for pharmaceutical administration than formulations comprising the non-glycosylated polypeptide.
  • solubility improvements can, for example, enable the generation of formulations more suitable for pharmaceutical administration than formulations comprising the non-glycosylated polypeptide.
  • Addition of glycosylation sites can be accomplished by altering the amino acid sequence.
  • the alteration to the polypeptide may be made, for example, by the addition of, or substitution by, one or more serine or threonine residues (for O-linked glycosylation sites) or asparagine residues (for N-linked glycosylation sites).
  • the structures of N-linked and O-linked oligosaccharides and the sugar residues found in each type may be different.
  • One type of sugar that is commonly found on both is N-acetylneuraminic acid (hereafter referred to as sialic acid).
  • sialic acid is usually the terminal residue of both N-linked and O-linked oligosaccharides and, by virtue of its negative charge, may confer acidic properties to the glycoprotein.
  • a particular embodiment comprises the generation and use of N-glycosylation variants.
  • polypeptide sequences provided herein may optionally be altered through changes at the nucleic acid level, particularly by mutating the nucleic acid encoding the polypeptide at preselected bases such that codons are generated that will translate into the desired amino acids.
  • Various cell lines can be used to produce proteins that are glycosylated.
  • DHFR Dihydrofolate reductase
  • CHO Chinese Hamster Ovary
  • These cells do not express the enzyme beta-galactoside alpha-2, 6-sialyltransferase and therefore do not add sialic acid in the alpha-2,6 linkage to N-linked oligosaccharides of glycoproteins produced in these cells.
  • Polysialylation in certain embodiments, also provided herein is the use of polysialylation, the conjugation of polypeptides to the naturally occurring, biodegradable a- (2 8) linked polysialic acid (“PSA”) in order to improve the polypeptides’ stability and in vivo pharmacokinetics.
  • PSA biodegradable a- (2 8) linked polysialic acid
  • Albumin Fusion Additional suitable components and molecules for conjugation include albumins such as human serum albumin (HSA), cyno serum albumin, and bovine serum albumin (BSA).
  • HSA human serum albumin
  • BSA bovine serum albumin
  • albumin is conjugated to a drug molecule (e.g ., a polypeptide described herein) at the carboxyl terminus, the amino terminus, both the carboxyl and amino termini, and internally (see, e.g., US Pat Nos. 5,876,969 and 7,056,701).
  • a drug molecule e.g ., a polypeptide described herein
  • albumin secretion pre-sequences and variants thereof, fragments and variants thereof, and HSA variants. Such forms generally possess one or more desired albumin activities.
  • fusion proteins are provided herein comprising a polypeptide drug molecule fused directly or indirectly to albumin, an albumin fragment, an albumin variant, etc., wherein the fusion protein has a higher plasma stability than the unfused drug molecule and/or the fusion protein retains the therapeutic activity of the unfused drug molecule.
  • the indirect fusion is effected by a linker, such as a peptide linker or modified version thereof.
  • fusion of albumin to one or more polypeptides provided herein can, for example, be achieved by genetic manipulation, such that the nucleic acid coding for HSA, or a fragment thereof, is joined to the nucleic acid coding for the one or more polypeptide sequences.
  • albumin binding through a conjugated fatty acid chain (acylation) and fusion proteins which comprise an albumin binding domain (ABD) polypeptide sequence and the sequence of one or more of the polypeptides described herein.
  • Fusion of albumin to a peptide sequence can, for example, be achieved by genetic manipulation, such that the DNA coding for HSA (human serum albumin), or a fragment thereof, is joined to the DNA coding for a peptide sequence. Thereafter, a suitable host can be transformed or transfected with the fused nucleotide sequence in the form of, for example, a suitable plasmid, so as to express a fusion polypeptide.
  • the expression may be effected in vitro from, for example, prokaryotic or eukaryotic cells, or in vivo from, for example, a transgenic organism.
  • the expression of the fusion protein is performed in mammalian cell lines, for example, CHO cell lines.
  • dAbs are the smallest functional binding units of human antibodies (IgGs) and have favorable stability and solubility characteristics.
  • the technology entails a dAb(s) conjugated to HSA (thereby forming a “AlbudAb”; see, e.g., EP1517921B, W02005/118642 and W02006/051288) and a molecule of interest (e.g., a peptide sequence provided herein).
  • AlbudAbs are often smaller and easier to manufacture in microbial expression systems, such as bacteria or yeast, than current technologies used for extending the serum half-life of peptides. As HSA has a half-life of about three weeks, the resulting conjugated molecule improves the half-life.
  • Use of the dAb technology may also enhance the efficacy of the molecule of interest.
  • Additional suitable components and molecules for conjugation include, for example, thyroglobulin; tetanus toxoid; Diphtheria toxoid; polyamino acids such as poly(D-lysine:D-glutamic acid); VP6 polypeptides of rotaviruses; influenza virus hemagglutinin, influenza virus nucleoprotein; Keyhole Limpet Hemocyanin (KLH); and hepatitis B virus core protein and surface antigen; or any combination of the foregoing.
  • thyroglobulin thyroglobulin
  • tetanus toxoid Diphtheria toxoid
  • polyamino acids such as poly(D-lysine:D-glutamic acid)
  • VP6 polypeptides of rotaviruses include influenza virus hemagglutinin, influenza virus nucleoprotein; Keyhole Limpet Hemocyanin (KLH); and hepatitis B virus core
  • conjugation of one or more additional components or molecules at the N- and/or C-terminus of a polypeptide sequence such as another polypeptide e.g ., a polypeptide having an amino acid sequence heterologous to the subject polypeptide), or a carrier molecule is also contemplated.
  • another polypeptide e.g ., a polypeptide having an amino acid sequence heterologous to the subject polypeptide
  • a carrier molecule e.g., an exemplary polypeptide sequence can be provided as a conjugate with another component or molecule.
  • a polypeptide may also be conjugated to large, slowly metabolized macromolecules such as proteins; polysaccharides, such as sepharose, agarose, cellulose, or cellulose beads; polymeric amino acids such as polyglutamic acid, or polylysine; amino acid copolymers; inactivated virus particles; inactivated bacterial toxins such as toxoid from diphtheria, tetanus, cholera, or leukotoxin molecules; inactivated bacteria; and dendritic cells.
  • Such conjugated forms if desired, can be used to produce antibodies against a polypeptide provided herein.
  • Fc-fusion Molecules In certain embodiments, the amino- or carboxyl- terminus of a polypeptide sequence provided herein is fused with an immunoglobulin Fc region to form a fusion conjugate (or fusion molecule). In a specific embodiment, the immunoglobulin Fc region is a human Fc region. Fusion conjugates have been shown to increase the systemic half-life of biopharmaceuticals, and thus the biopharmaceutical product may require less frequent administration. In certain embodiments, the half-life is increased as compared to the same polypeptide that is not fused to an immunoglobulin Fc region.
  • Fc binds to the neonatal Fc receptor (FcRn) in endothelial cells that line the blood vessels, and, upon binding, the Fc fusion molecule is protected from degradation and re-released into the circulation, keeping the molecule in circulation longer.
  • This Fc binding is believed to be the mechanism by which endogenous IgG retains its long plasma half-life.
  • More recent Fc- fusion technology links a single copy of a biopharmaceutical to the Fc region of an antibody to optimize the pharmacokinetic and pharmacodynamic properties of the biopharmaceutical as compared to traditional Fc-fusion conjugates.
  • Fc-fusion drugs consist of two copies of a biopharmaceutical linked to the Fc region of an antibody to improve pharmacokinetics, solubility, and production efficiency. More recent Fc-fusion technology links a single copy of a biopharmaceutical to the Fc region of an antibody to optimize the pharmacokinetic and pharmacodynamic properties of the biopharmaceutical as compared to traditional Fc-fusion conjugates.
  • provided herein is a fusion of M70 to a human antibody Fc fragment.
  • a fusion of M69 to a human antibody Fc fragment Such fusions can be useful in the treatment of bile acid related disorders and other metabolic disorders provided herein.
  • the Fc-fusion of M70 has a longer half-life.
  • the longer half-life of the Fc-fusion of M70 is as compared to M70 that is not an Fc-fusion.
  • the Fc-fusion of M69 has a longer half-life.
  • the longer half life of the Fc-fusion of M69 is as compared to M69 that is not an Fc-fusion. Such a long half-life makes these fusions suitable for once weekly, or less frequent dosing.
  • the Fc-fusion comprises a linker.
  • exemplary flexible linkers include glycine polymers (G )n , glycine-serine polymers, glycine-alanine polymers, alanine-serine polymers, and other flexible linkers.
  • the linker is (G)4S.
  • the linker is ((G)4S)n, where n is an integer of at least one.
  • the linker is ((G)4S)2.
  • Glycine and glycine-serine polymers are relatively unstructured, and therefore may serve as a neutral tether between components.
  • the glycine- serine polymer is (GS)n, where n is an integer of at least one. In some embodiments, the glycine- serine polymer is GSGGSn(SEQ ID NO: 129), where n is an integer of at least one. In some embodiments, the glycine-serine polymer is GGGSn (SEQ ID NO: 130), where n is an integer of at least one. In certain embodiments, the linker comprises an additional G residue at the N’ terminus of SEQ ID NO: 130. In one embodiment, the linker is GGSG (SEQ ID NO: 131). In one embodiment, the linker is GGSGG (SEQ ID NO: 132).
  • the linker is GSGSG (SEQ ID NO: 133). In one embodiment, the linker is GSGGG (SEQ ID NO: 134). In one embodiment, the linker is GGGSG (SEQ ID NO: 189). In one embodiment, the linker is GSSSG (SEQ ID NO: 135).
  • Additional suitable components and molecules for conjugation include those suitable for isolation or purification.
  • Particular non-limiting examples include binding molecules, such as biotin (biotin-avidin specific binding pair), an antibody, a receptor, a ligand, a lectin, or molecules that comprise a solid support, including, for example, plastic or polystyrene beads, plates or beads, magnetic beads, test strips, and membranes.
  • cation exchange chromatography may be used to separate conjugates by charge difference, which effectively separates conjugates into their various molecular weights.
  • the cation exchange column can be loaded and then washed with ⁇ 20 mM sodium acetate, pH ⁇ 4, and then eluted with a linear (0 M to 0.5 M) NaCl gradient buffered at a pH from 3 to 5.5, such as at pH ⁇ 4.5.
  • the content of the fractions obtained by cation exchange chromatography may be identified by molecular weight using conventional methods, for example, mass spectroscopy, SDS-PAGE, or other known methods for separating molecular entities by molecular weight.
  • a fraction is then identified which contains the conjugate having the desired number of PEGs attached, purified free from unmodified protein sequences and from conjugates having other numbers of PEGs attached.
  • a peptide sequence provided herein is linked to a chemical agent (e.g ., an immunotoxin or chemotherapeutic agent), including, but are not limited to, a cytotoxic agent, including taxol, cytochalasin B, gramicidin D, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicin, doxorubicin, daunorubicin, and analogs or homologs thereof.
  • a chemical agent e.g ., an immunotoxin or chemotherapeutic agent
  • a cytotoxic agent including taxol, cytochalasin B, gramicidin D, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicin, doxorubicin, daunorubicin, and analogs or homologs thereof.
  • Other chemical agents include, for example, antimetabolites (e.g., methotrexate, 6-mercaptopurine, 6- thioguanine, cytarabine, 5-fluorouracil decarbazine); alkylating agents (e.g, mechlorethamine, carmustine and lomustine, cyclothosphamide, busulfan, dibromomannitol, streptozotocin, mitomycin C, and cisplatin); antibiotics (e.g, bleomycin); and anti-mitotic agents (e.g, vincristine and vinblastine). Cytotoxins can be conjugated to a peptide provided herein using linker technology known in the art and described herein.
  • antimetabolites e.g., methotrexate, 6-mercaptopurine, 6- thioguanine, cytarabine, 5-fluorouracil decarbazine
  • alkylating agents e.g, mechlorethamine, car
  • suitable components and molecules for conjugation include those suitable for detection in an assay.
  • detectable labels such as a radioisotope (e.g ., 125 I; 35 S, 32 P; 33 P), an enzyme which generates a detectable product (e.g, luciferase, b-galactosidase, horse radish peroxidase and alkaline phosphatase), a fluorescent protein, a chromogenic protein, dye (e.g., fluorescein isothiocyanate); fluorescence emitting metals (e.g, 152 Eu); chemiluminescent compounds (e.g, luminol and acridinium salts); bioluminescent compounds (e.g, luciferin); and fluorescent proteins.
  • Indirect labels include labeled or detectable antibodies that bind to a peptide sequence, where the antibody may be detected.
  • a peptide sequence provided herein is conjugated to a radioactive isotope to generate a cytotoxic radiopharmaceutical (radioimmunoconjugates) useful as a diagnostic or therapeutic agent.
  • radioactive isotopes include, but are not limited to, iodine 131 , indium 111 , yttrium 90 and lutetium 177 .
  • Methods for preparing radioimmunoconjugates are known to the skilled artisan. Examples of radioimmunoconjugates that are commercially available include ibritumomab, tiuxetan, and tositumomab.
  • Linkers Linkers and their use have been described above. Any of the foregoing components and molecules used to modify the polypeptide sequences provided herein may optionally be conjugated via a linker. Suitable linkers include “flexible linkers” which are generally of sufficient length to permit some movement between the modified polypeptide sequences and the linked components and molecules. The linker molecules are generally about 6-50 atoms long. The linker molecules may also be, for example, aryl acetylene, ethylene glycol oligomers containing 2-10 monomer units, diamines, diacids, amino acids, or combinations thereof.
  • Suitable linkers can be readily selected and can be of any suitable length, such as 1 amino acid (e.g., Gly), 2, 3, 4, 5, 6, 7, 8, 9, 10, 10-20, 20-30, 30-50 or more than 50 amino acids.
  • Exemplary flexible linkers include glycine polymers (G)n, glycine-serine polymers (for example, (GS)n, GSGGSn (SEQ ID NO: 129) and GGGSn (SEQ ID NO: 130), where n is an integer of at least one), glycine-alanine polymers, alanine-serine polymers, and other flexible linkers. Glycine and glycine-serine polymers are relatively unstructured, and therefore may serve as a neutral tether between components.
  • Exemplary flexible linkers include, but are not limited to GGSG (SEQ ID NO: 131), GGSGG (SEQ ID NO: 132), GSGSG (SEQ ID NO: 133), GSGGG (SEQ ID NO: 134), GGGSG (SEQ ID NO: 189), and GSSSG (SEQ ID NO: 135).
  • the linker is (G)4S.
  • the linker is ((G)4S)n), where n is an integer of at least one.
  • the linker is ((G)4S)2).
  • the glycine-serine polymer is (GS)n, where n is an integer of at least one.
  • the glycine-serine polymer is GSGGSn (SEQ ID NO: 129), where n is an integer of at least one. In some embodiments, the glycine-serine polymer is GGGSn (SEQ ID NO: 130), where n is an integer of at least one. In certain embodiments, the linker comprises an additional G residue at the N’ terminus of SEQ ID NO: 130. In one embodiment, the linker is GGSG (SEQ ID NO:131). In one embodiment, the linker is GGSGG (SEQ ID NO:132). In one embodiment, the linker is GSGSG (SEQ ID NO: 133). In one embodiment, the linker is GSGGG (SEQ ID NO: 134). In one embodiment, the linker is GGGSG (SEQ ID NO: 189). In one embodiment, the linker is GSSSG (SEQ ID NO: 135).
  • the peptide sequences set forth herein can be used in methods for, for example, modulating the gut microbiome.
  • the peptide sequences set forth herein can be used for, for example, enriching Veillonella.
  • the peptide sequences set forth herein can be used for, for example, reducing lactate.
  • the peptide sequences set forth herein can be used for, for example, negatively regulating bile acids.
  • the peptide sequences set forth herein can be used for, for example, enriching Veillonella by negatively regulating bile acids.
  • the peptide sequences set forth herein can be used for, for example, ameliorating hepatic fibrosis.
  • the peptide sequences set forth herein can be used for, for example, ameliorating hepatic steatosis.
  • the peptide sequences set forth herein can be used for, for example, ameliorating inflammation.
  • the peptide sequences set forth herein can be used for, for example, treating or preventing liver disease.
  • the peptide sequences set forth herein can be used for, for example, treating or preventing disease associated with an increase in lactate.
  • the methods include administering an effective amount of a peptide sequence, such as a FGF19 or FGF21 variant, fusion or chimera disclosed herein (e.g ., in the Sequence Listing, Table 1, or Examples I and II), or a subsequence, a variant or modified form of a FGF19 or FGF21 variant, fusion or chimera disclosed herein (e.g., the Sequence Listing or Table 1), to a subject in need thereof for modulating bile acid homeostasis, treating or preventing liver disease, or treating or preventing disease associated with an increase in lactate.
  • the peptide sequences used in methods as provided herein are fused with an immunoglobulin Fc region.
  • provided herein are methods for enriching Veillonella in a subject in need thereof that include administering an effective amount of a peptide sequence disclosed herein. In some embodiments, provided herein are methods for reducing lactate in a subject in need thereof that include administering an effective amount of a peptide sequence disclosed herein. In some embodiments, provided herein are methods for treating or preventing disease associated with elevated lactate in a subject in need thereof that include administering an effective amount of a peptide sequence disclosed herein.
  • a “disease associated with elevated lactate” refers to a condition in which lactate levels increase, e.g ., systemically or in the blood.
  • An increase in the level of lactate is an indication that organ systems are not functioning appropriately and coincides with more severe disease.
  • lactate accumulates in various liver diseases. Specifically, lactate accumulates in cirrhosis patients, which is associated with a worsening of the disease. Lactate also accumulates in patients with NASH, PBC and acute liver failure.
  • Non-liver diseases associated with elevated lactate include diabetes, renal failure, infectious disease and heart disease.
  • Subjects refers to an animal. Typically, the animal is a mammal that would benefit from treatment with a peptide sequence provided herein. Particular examples include primates (e.g, humans), dogs, cats, horses, cows, pigs, and sheep. [00193] In some embodiments, the subject is a human.
  • Subjects that can be treated with methods described herein can have a disease associated with elevated lactate.
  • the subject has or is at risk of having a disease associated with elevated lactate.
  • the subject is a patient having a disease associated with elevated lactate.
  • the subject is a patient having elevated lactate.
  • the subject can be a heathy individual.
  • Subjects at risk of developing a disease associated with elevated lactate include, for example, those who may have a family history or genetic predisposition toward a disease associated with elevated lactate, as well those whose diet or habits may contribute to development of such disorders.
  • the subject that has or is at risk for a disease associated with elevated lactate has a liver disease.
  • the subject has cirrhosis.
  • the subject has NASH.
  • the subject has primary biliary cirrhosis.
  • the subject has acute liver failure.
  • the subject is overweight.
  • the subject is obese.
  • the subject has a Body Mass Index (BMI) that is at least 25 0
  • the subject has a BMI that is at least 30 0
  • the subject that has or is at risk for a disease associated with elevated lactate has an infection. In some embodiments, the subject that has or is at risk for a disease associated with elevated lactate has an autoimmune disorder. In some embodiments, the subject that has or is at risk for a disease associated with elevated lactate has a cancer.
  • Subjects that can be treated with methods described herein can have a liver disease.
  • Subjects that can be treated with methods described herein can have a disease associated with elevated lactate.
  • Subjects that can be treated with the methods described herein can have a bile acid-related disorder.
  • a bile acid-related disorder comprises a liver disease.
  • a liver disease comprises a bile acid-related disorder.
  • the term “bile acid-related disorder,” or the like, when used in reference to a condition of a subject means a disruption of bile acid homeostasis, which may manifest itself as, for example, an acute, transient or chronic abnormal level of a bile acid or one or more bile acids.
  • subjects that can be treated with methods described herein can have a bile acid-related disorder, such as cholestasis, including, for example diseases of intrahepatic cholestasis (e.g ., biliary cirrhosis (PBC), primary familial intrahepatic cholestasis (PFIC), primary sclerosing choangitis (PSC), pregnancy intrahepatic cholestasis (PIC), neonatal cholestasis, and drug induced cholestasis (e.g., estrogen)), and diseases of extrahepatic cholestasis (e.g, bile cut compression from tumor, bile duct blockade by gall stones); bile acid malabsorption and other disorders involving the distal small intestine, including ileal
  • the subject has cholestasis. In some embodiments, the subject has PBC. In some embodiments, the subject has PFIC. In some embodiments, the subject has PSC. In some embodiments, the subject has neonatal cholestasis. In some embodiments, the subject has PIC. In some embodiments, the subject has bile acid malabsorption. In some embodiments, the subject has NASH. In some embodiments, the subject has NAFLD.
  • Additional bile acid-related disorders include metabolic syndrome, a lipid or glucose disorder, cholesterol or triglyceride metabolism, diabetes (e.g ., type 2 diabetes), other hyperglycemic-related disorders, including kidney damage (e.g., tubule damage or nephropathy), liver degeneration, eye damage (e.g, diabetic retinopathy or cataracts), and diabetic foot disorders, and dyslipidemias and their sequelae such as, for example, atherosclerosis, coronary artery disease, cerebrovascular disorders and the like.
  • diabetes e.g ., type 2 diabetes
  • other hyperglycemic-related disorders including kidney damage (e.g., tubule damage or nephropathy), liver degeneration, eye damage (e.g, diabetic retinopathy or cataracts), and diabetic foot disorders, and dyslipidemias and their sequelae such as, for example, atherosclerosis, coronary artery disease, cerebrovascular disorders and the like.
  • Other conditions associated with metabolic syndrome can also include such as obesity and elevated body mass (including the co-morbid conditions thereof such as, but not limited to, nonalcoholic fatty liver di. suave (NAFLD), nonalcoholic steatohepatitis (NASH), and polycystic ovarian syndrome (PCOS)), and also include thromboses, hypercoagulable and prothrombotic states (arterial and venous), hypertension (including portal hypertension (defined as a hepatic venous pressure gradient (HVPG) greater than 5 mm Hg), cardiovascular disease, stroke and heart failure; Disorders or conditions in which inflammatory reactions are involved, including atherosclerosis, chronic inflammatory bowel diseases (e.g, Crohn’s disease and ulcerative colitis), asthma, lupus erythematosus, arthritis, or other inflammatory rheumatic disorders; Disorders of cell cycle or cell differentiation processes such as adipose cell tumors, lipomatous carcinomas including, for example, liposarcomas,
  • treatment methods include administering a peptide as set forth herein (e.g ., a variant or fusion of FGF19 and/or FGF21 as set forth in the Sequence Listing or Table 1) in an amount effective to achieve a desired outcome or result in a subject.
  • a treatment that results in a desired outcome or result includes decreasing, reducing or preventing the severity or frequency of one or more symptoms of the condition in the subject, e.g., an improvement in the subject’s condition or a “beneficial effect” or “therapeutic effect.”
  • treatment can decrease or reduce or prevent the severity or frequency of one or more symptoms of the disorder, stabilize or inhibit progression or worsening of the disorder, and in some instances, reverse the disorder, transiently (e.g, for 1-6, 6-12, or 12-24 hours), for medium term (e.g., 1-6, 6-12, 12-24 or 24-48 days) or long term (e.g., for 1-6, 6-12, 12-24, 24-48 weeks, or greater than 24-48 weeks).
  • transiently e.g, for 1-6, 6-12, or 12-24 hours
  • medium term e.g., 1-6, 6-12, 12-24 or 24-48 days
  • long term e.g., for 1-6, 6-12, 12-24, 24-48 weeks, or greater than 24-48 weeks.
  • an “effective amount” or a “sufficient amount” for use and/or for treating a subject refers to an amount that provides, in single or multiple doses, alone, or in combination with one or more other agents, treatments, protocols, or therapeutic regimens, a detectable response of any duration of time (transient, medium or long term), a desired outcome in or an objective or subjective benefit to a subject of any measurable or detectable degree or for any duration of time (e.g, for hours, days, months, years, in remission or cured).
  • Such amounts typically are effective to ameliorate a disorder, or one, multiple or all adverse symptoms, consequences or complications of the disorder, to a measurable extent, although reducing or inhibiting a progression or worsening of the disorder, is considered a satisfactory outcome.
  • the term “ameliorate” means an improvement in the subject’s disorder, a reduction in the severity of the disorder, or an inhibition of progression or worsening of the disorder (e.g, stabilizing the disorder).
  • an improvement can be a lowering or a reduction in one or more symptoms or effects of the disorder, including lactate.
  • a therapeutic benefit or improvement therefore need not be complete ablation of any one, most or all symptoms, complications, consequences or underlying causes associated with the disorder or disease.
  • a satisfactory endpoint is achieved when there is a transient, medium or long term, incremental improvement in a subject’s condition, or a partial reduction in the occurrence, frequency, severity, progression, or duration, or inhibition or reversal, of one or more associated adverse symptoms or complications or consequences or underlying causes, worsening or progression (e.g ., stabilizing one or more symptoms or complications of the condition, disorder or disease), of the disorder or disease, over a duration of time (hours, days, weeks, months, etc.).
  • the amount of the peptide and the additional agent sufficient to ameliorate a disorder will depend on the type, severity and extent, or duration of the disorder, the therapeutic effect or outcome desired, and can be readily ascertained by the skilled artisan. Appropriate amounts will also depend upon the individual subject (e.g., the bioavailability within the subject, gender, age, etc.). For example, a transient, or partial, reduction of lactate in a subject can reduce the dosage amount or frequency of the peptides described herein in order to treat a disease associated with elevated lactate even though complete freedom from treatment has not resulted. A therapeutically effective amount can be ascertained, for example, by measuring one or more relevant physiological effects.
  • Peptide sequences provided herein including subsequences, sequence variants and modified forms of the exemplified peptide sequences (e.g., sequences listed in the Sequence Listing or Table 1), may be formulated in a unit dose or unit dosage form.
  • a peptide sequence is in an amount effective to treat a subject in need of treatment.
  • Exemplary unit doses range from about 25-250, 250-500, 500-1000, 1000-2500, 2500-5000, 5000-25,000, or 25,000-50,000 ng; from about 25-250, 250-500, 500-1000, 1000-2500, 2500- 5000, 5000-25,000, or 25,000-50,000 pg; and from about 25-250, 250-500, 500-1000, or 1000- 2500 mg.
  • Peptide sequences provided herein including subsequences, sequence variants and modified forms of the exemplified peptide sequences can be administered to provide the intended effect as a single dose or multiple dosages, for example, in an effective or sufficient amount.
  • Exemplary doses range from about 25-250, 250-500, 500-1000, 1000-2500, 2500-5000, 5000-25,000, or 25,000-50,000 pg/kg; from about 50-500, 500-5000, 5000-25,000 or 25,000-50,000 ng/kg; and from about 25-250, 250-500, 500-1000, or 1000-2500 pg/kg.
  • Single or multiple doses can be administered, for example, multiple times per day, on consecutive days, alternating days, weekly or intermittently (e.g, twice per week, once every 1, 2, 3, 4, 5, 6, 7 or 8 weeks, or once every 2, 3, 4, 5 or 6 months).
  • the peptide sequences provided herein can be administered to provide the intended effect as short-term therapies (e.g. daily administration for multiple days).
  • the peptide sequences provided herein can be administered daily for a period of 7 to 336 days, including any number of days in between.
  • the peptide sequences provided herein can be administered daily for a period of 7 to 280 days, 7 to 224 days, 7 to 168 days, 7 to 112 days, or 7 to 56 days.
  • the peptide sequences provided herein can be administered daily for a period of 14 to 336 days, 14 to 280 days, 14 to 224 days, 14 to 168 days, 14 to 112 days, or 14 to 56 days. In some embodiments, the peptide sequences provided herein can be administered daily for a period of 21 to 336 days, 21 to 280 days, 21 to 224 days, 21 to 168 days, 21 to 112 days, or 21 to 56 days. In some embodiments, the peptide sequences provided herein can be administered daily for a period of 28 to 336 days, 28 to 280 days, 28 to 224 days, 28 to 168 days, 28 to 112 days, or 28 to 56 days. In some embodiments, the peptide sequences provided herein can be administered daily for 7 days, 14 days, 21 days, 28 days, 35 days, 42 days, 49 days, 56 days, 63 days, 70 days, 77 days, 84 days,
  • the peptide sequences provided herein can be administered daily for 21 days. In some embodiments, the peptide sequences provided herein can be administered daily for 42 days. In some embodiments, the peptide sequences provided herein can be administered daily for 63 days. In some embodiments, the peptide sequences provided herein can be administered daily for 84 days. In some embodiments, the peptide sequences provided herein can be administered daily for 105 days.
  • the peptide sequences provided herein can be administered daily for 126 days. In some embodiments, the peptide sequences provided herein can be administered daily for 147 days. In some embodiments, the peptide sequences provided herein can be administered daily for 168 days.
  • the peptide sequences provided herein can be administered to provide the intended effect as longer-term therapies (e.g. weekly administration for multiple weeks).
  • the peptide sequences provided herein can be administered weekly for a period of 8 to 156 weeks, including any number of weeks in between.
  • the peptide sequences provided herein can be administered for a period of 2 to 36 months, 2 to 30 months, 2 to 24 months, 2 to 18 months, 2 to 12 months, or 2 to 6 months.
  • the peptide sequences provided herein can be administered for a period of 3 to 36 months, 3 to 30 months, 3 to 24 months, 3 to 18 months, 3 to 12 months, or 3 to 6 months.
  • the peptide sequences provided herein can be administered for a period of 4 to 36 months, 4 to 30 months, 4 to 24 months, 4 to 18 months, 4 to 12 months, or 4 to 6 months. In some embodiments, the peptide sequences provided herein can be administered for a period of 5 to 36 months, 5 to 30 months, 5 to 24 months, 5 to 18 months, 5 to 12 months, or 5 to 6 months. In some embodiments, the peptide sequences provided herein can be administered weekly for 2 months, 3 months, 4 months, 5 months, 6 months, 7 months, 8 months, 9 months,
  • the peptide sequences provided herein can be administered weekly for 12 weeks. In some embodiments, the peptide sequences provided herein can be administered weekly for 24 weeks. In some embodiments, the peptide sequences provided herein can be administered weekly for 36 weeks. In some embodiments, the peptide sequences provided herein can be administered weekly for 48 weeks. In some embodiments, the peptide sequences provided herein can be administered weekly for 60 weeks.
  • the peptide sequences provided herein can be administered weekly for 72 weeks. In some embodiments, the peptide sequences provided herein can be administered weekly for 84 weeks. In some embodiments, the peptide sequences provided herein can be administered weekly for 96 weeks. In some embodiments, the peptide sequences provided herein can be administered weekly for 108 weeks. In some embodiments, the peptide sequences provided herein can be administered weekly for 120 weeks.
  • the peptide sequences provided herein can be administered daily for 7 to 280 days, 7 to 224 days, 7 to 168 days, 7 to 112 days, 7 to 56 days, 14 to 336 days, 14 to 280 days, 14 to 224 days, 14 to 168 days, 14 to 112 days, 14 to 56 days, 21 to 336 days, 21 to 280 days, 21 to 224 days, 21 to 168 days, 21 to 112 days, 21 to 56 days, 28 to 336 days, 28 to 280 days, 28 to 224 days, 28 to 168 days, 28 to 112 days, or 28 to 56 days followed by weekly administration for 2 to 36 months, 2 to 30 months, 2 to 24 months, 2 to 18 months, 2 to 12 months, 2 to 6 months, 3 to 36 months, 3 to 30 months, 3 to 24 months, 3 to 18 months, 3 to 12 months, 3 to 6 months, 4 to 36 months, 4 to 30 months, 4 to 24 months, 4 to 18 months, 4 to 12 months, 4 to 6 months, 5 to 36 months, 5 to
  • the peptide sequences provided herein can be administered daily for 21 to 168 days followed by weekly administration for 2 to 36 months, 2 to 30 months, 2 to 24 months, 2 to 18 months, 2 to 12 months, 2 to 6 months, 3 to 36 months, 3 to 30 months, 3 to 24 months, 3 to 18 months, 3 to 12 months, 3 to 6 months, 4 to 36 months, 4 to 30 months, 4 to 24 months, 4 to 18 months, 4 to 12 months, 4 to 6 months, 5 to 36 months, 5 to 30 months, 5 to 24 months, 5 to 18 months, 5 to 12 months, or 5 to 6 months.
  • the peptide sequences provided herein can be administered daily for 7 to 280 days, 7 to 224 days, 7 to 168 days, 7 to 112 days, 7 to 56 days, 14 to 336 days, 14 to 280 days, 14 to 224 days, 14 to 168 days, 14 to 112 days, 14 to 56 days, 21 to 336 days, 21 to 280 days, 21 to 224 days, 21 to 168 days, 21 to 112 days, 21 to 56 days, 28 to 336 days, 28 to 280 days, 28 to 224 days, 28 to 168 days, 28 to 112 days, or 28 to 56 days followed by weekly administration for 12 to 104 weeks.
  • the peptide sequences provided herein can be administered daily for 21 to 168 days followed by weekly administration for 12 to 104 weeks.
  • Peptide sequences provided herein including subsequences, variants and modified forms of the exemplified peptide sequences can be administered and methods may be practiced via systemic, regional or local administration, by any route.
  • a peptide sequence can be administered parenterally (e.g., subcutaneously, intravenously, intramuscularly, or intraperitoneally), orally (e.g, ingestion, buccal, or sublingual), inhalation, intradermally, intracavity, intracranially, transdermally (topical), transmucosally or rectally.
  • Peptide sequences provided herein including subsequences, variants and modified forms of the exemplified peptide sequences (e.g, sequences listed in the Sequence Listing or Table 1) and methods provided herein including pharmaceutical compositions can be administered via a (micro)encapsulated delivery system or packaged into an implant for administration.
  • parenteral (e.g ., subcutaneous) administration entails the use of Intarcia’s subcutaneous delivery system (Intarcia Therapeutics, Inc.; Hayward, CA).
  • the system comprises a miniature osmotic pump that delivers a consistent amount of a therapeutic agent over a desired period of time.
  • the system can be used with formulations that maintain the stability of proteinaceous therapeutic agents at human body temperature for extended periods of time.
  • DUROS®-type implantable osmotic pumps from, e.g., DURECT Corp.
  • the DUROS® system can be used for therapies requiring systemic or site-specific administration of a drug.
  • the DUROS® system is placed just under the skin, for example in the upper arm, in an outpatient procedure that is completed in just a few minutes using local anesthetic.
  • miniaturized catheter technology can be used.
  • the catheter can be attached to the DUROS® system to direct the flow of a drug to the target organ, tissue or synthetic medical structure, such as a graft.
  • Site-specific delivery enables a therapeutic concentration of a drug to be administered to the desired target without exposing the entire body to a similar concentration.
  • the precision, size and performance of the DUROS® system will allow for continuous site-specific delivery to a variety of precise locations within the body.
  • parenteral administration entails the use of an on-body delivery system (e.g, the NEULASTA® Delivery Kit by Amgen).
  • This on-body delivery system includes an on-body injector, which is a small, lightweight, injection system applied on the same day as a doctor visit (such as the day of chemotherapy). It is designed to deliver a dose of the therapeutic agent the next day, or in the near future of the doctor visit, so that the patient does not need to return to the doctor’s office to receive the injection.
  • Various methods of controlled release is also contemplated herein. Encapsulation of therapeutic molecules within polymer particles is a well-established method for achieving controlled release and can be used in methods provided herein. Also, by taking advantage of the adsorption of protein therapeutics to poly(lactic-co-glycolic acid) (PLGA) nanoparticles, controlled release can also be achieved without encapsulation. In particular, extended-release for protein therapeutics can be applied with and without encapsulation in PLGA nanoparticles embedded within a hydrogel. The release profile tunable by modifying nanoparticle concentration, nanoparticle size, or environmental pH. Pakulska et al., Science Advances 2(5): el600519 (2016).
  • variants of FGF19 peptide sequences fusions of FGF19 and/or FGF21 peptide sequences and variants of fusions (chimeras) of FGF19 and/or FGF21 peptide sequences having one or more activities associated with the modulation of lactate homeostasis in combination with other therapeutic agents and/or treatment modalities.
  • variants of FGF19 peptide sequences fusions of FGF19 and/or FGF21 peptide sequences and variants of fusions (chimeras) of FGF19 and/or FGF21 peptide sequences having one or more activities associated with the enrichment of Veillonella in combination with other therapeutic agents and/or treatment modalities.
  • variants of FGF19 peptide sequences fusions of FGF19 and/or FGF21 peptide sequences and variants of fusions (chimeras) of FGF19 and/or FGF21 peptide sequences having one or more activities associated with the treatment and/or prevention of a disease associated with elevated lactate in combination with other therapeutic agents and/or treatment modalities.
  • additional therapeutic agents that can be used in combination with peptide sequences described herein in the methods of treating or preventing a disease associated with elevated lactate include, but are not limited to a lactate lowering agent, or a probiotic.
  • the additional therapeutic agent is a lactate-lowering agent.
  • the lactate-lowering agent is a lactate-lowering drug.
  • the lactate-lowering drug can be alkalinizing agents.
  • the alkalinizing agent can be sodium bicarbonate.
  • the alkalinizing agent can be tromethamine.
  • the additional therapeutic agent is a probiotic.
  • the probiotic can be a probiotic that enriches Veillonella.
  • the probiotic can contain Veillonella.
  • treatment methods can include administering one or more additional therapeutic agents or therapeutic modalities useful in reducing lactate or in the treatment or prevention of disease associated with elevated lactate, such as those agents or therapeutic modalities described herein, in an amount effective to achieve a desired outcome or result in a subject.
  • Treatment methods can include administering one or more additional therapeutic agents of therapeutic modalities useful in enriching Veillonella.
  • a treatment that results in a desired outcome or result includes decreasing, reducing or preventing the severity or frequency of one or more symptoms of the condition in the subject, e.g ., an improvement in the subject’s condition or a “beneficial effect” or “therapeutic effect.” Therefore, treatment can decrease or reduce or prevent the severity or frequency of one or more symptoms of the disorder, stabilize or inhibit progression or worsening of the disorder, and in some instances, reverse the disorder, transiently (e.g., for 1-6, 6-12, or 12-24 hours), for medium term (e.g., 1-6, 6-12, 12-24 or 24-48 days) or long term (e.g, for 1-6, 6-12, 12-24, 24-48 weeks, or greater than 24-48 weeks).
  • transiently e.g., for 1-6, 6-12, or 12-24 hours
  • medium term e.g., 1-6, 6-12, 12-24 or 24-48 days
  • long term e.g, for 1-6, 6-12, 12-24, 24-48 weeks, or greater than 24-48 weeks.
  • treatment with a peptide provided herein in combination with another therapeutic agent can prevent the formation, cause regression of or stabilize atherosclerotic plaques in a subject.
  • treatment with a peptide provided herein in combination with another therapeutic agent can lower or reduce one or more symptoms or effects of the disease.
  • treatment with a peptide provided herein in combination with another therapeutic agent can reduce serum level of lactate.
  • methods and uses provided herein for treating a subject having, or at risk of developing, a disease associated with lactate elevation can be practiced prior to, substantially contemporaneously with, or following administration or application of another agent useful for the treatment or prevention of a disease associated with lactate elevation, and/or can be supplemented with other forms of therapy.
  • Methods and uses provided herein for treating a subject having, or at risk of developing, a disease associated with lactate elevation can be practiced prior to, substantially contemporaneously with, or following administration or application of another agent useful for the treatment or prevention of a disease associated with lactate elevation, and/or can be supplemented with other forms of therapy.
  • Supplementary therapies can be administered prior to, contemporaneously with or following methods and uses provided herein.
  • compositions which include a peptide sequence (or sequences) provided herein, including subsequences, variants and modified forms of the exemplified peptide sequences (e.g ., sequences listed in the Sequence Listing or Table 1), and one or more pharmaceutically acceptable or physiologically acceptable diluents, carriers or excipients; in combination with, or separate from, one or more additional agents for the modulation of lactate homeostasis, for enrichment of Veillonella , for the treatment or prevention of a disease associated with lactate elevation, or a composition comprising such one or more additional agents and one or more pharmaceutically acceptable or physiologically acceptable diluents, carriers or excipients.
  • a peptide sequence or sequences and an additional agent(s) are present in a therapeutically acceptable amount.
  • the pharmaceutical compositions can be used in accordance with the methods and uses provided herein.
  • the pharmaceutical compositions can be administered ex vivo or in vivo to a subject in order to practice treatment methods and uses provided herein.
  • Pharmaceutical compositions provided herein can be formulated to be compatible with the intended method or route of administration; exemplary routes of administration are set forth herein.
  • the pharmaceutical compositions may further comprise an additional therapeutically active agents or compounds disclosed or known to the skilled artisan which can be used in the treatment or prevention of a disease associated with lactate elevation as set forth herein.
  • the pharmaceutical compositions can further comprise an additional therapeutically active agents or compounds disclosed or known to the skilled artisan which can be used in the treatment or prevention of an a disease associated with lactate elevation as set forth herein.
  • the additional agent can be a lactate lowering agent or a probiotic.
  • the additional therapeutically active agents or compounds can be present in a separate pharmaceutical composition(s). Exemplary dosing parameters and regimens are described herein.
  • compositions typically comprise a therapeutically effective amount of at least one of the peptide sequences provided herein, including subsequences, variants and modified forms of the exemplified peptide sequences (e.g., sequences listed in the Sequence Listing or Table 1) and/or one or more additional agents described herein, and one or more pharmaceutically and physiologically acceptable formulation agents.
  • Suitable pharmaceutically acceptable or physiologically acceptable diluents, carriers or excipients include, but are not limited to, antioxidants (e.g, ascorbic acid and sodium bisulfate), preservatives (e.g, benzyl alcohol, methyl parabens, ethyl or n-propyl, p-hydroxybenzoate), emulsifying agents, suspending agents, dispersing agents, solvents, fillers, bulking agents, buffers, vehicles, diluents, and/or adjuvants.
  • a suitable vehicle may be physiological saline solution or citrate buffered saline, possibly supplemented with other materials common in pharmaceutical compositions for parenteral administration.
  • Neutral buffered saline or saline mixed with serum albumin are further exemplary vehicles.
  • Typical buffers include, but are not limited to pharmaceutically acceptable weak acids, weak bases, or mixtures thereof.
  • Buffer components also include water soluble materials such as phosphoric acid, tartaric acids, lactic acid, succinic acid, citric acid, acetic acid, ascorbic acid, aspartic acid, glutamic acid, and salts thereof.
  • a primary solvent in a vehicle may be either aqueous or non-aqueous in nature.
  • the vehicle may contain other pharmaceutically acceptable excipients for modifying or maintaining the pH, osmolarity, viscosity, sterility or stability of the pharmaceutical composition.
  • the pharmaceutically acceptable vehicle is an aqueous buffer.
  • a vehicle comprises, for example, sodium chloride and/or sodium citrate.
  • compositions provided herein may contain still other pharmaceutically-acceptable formulation agents for modifying or maintaining the rate of release of a peptide and/or an additional agent, as described herein.
  • formulation agents include those substances known to artisans skilled in preparing sustained-release formulations.
  • Remington s Pharmaceutical Sciences. 18th Ed. (1990, Mack Publishing Co., Easton, Pa. 18042) pages 1435-1712, The Merck Index. 12th Ed. (1996, Merck Publishing Group, Whitehouse, NJ); and Pharmaceutical Principles of Solid Dosage Forms (1993, Technonic Publishing Co., Inc., Lancaster, Pa.). Additional pharmaceutical compositions appropriate for administration are known in the art and are applicable in the methods and compositions provided herein.
  • a pharmaceutical composition may be stored in a sterile vial as a solution, suspension, gel, emulsion, solid, or dehydrated or lyophilized powder. Such compositions may be stored either in a ready to use form, a lyophilized form requiring reconstitution prior to use, a liquid form requiring dilution prior to use, or other acceptable form.
  • a pharmaceutical composition is provided in a single-use container (e.g ., a single-use vial, ampoule, syringe, or autoinjector (similar to, e.g., an EpiPen®)), whereas a multi-use container (e.g, a multi-use vial) is provided in other embodiments.
  • Any drug delivery apparatus may be used to deliver peptides and the other agents described herein, including implants (e.g, implantable pumps) and catheter systems, both of which are known to the skilled artisan.
  • Depot injections which are generally administered subcutaneously or intramuscularly, may also be utilized to release peptides and/or other agents described herein over a defined period of time. Depot injections are usually either solid- or oil-based and generally comprise at least one of the formulation components set forth herein. The skilled artisan is familiar with possible formulations and uses of depot injections.
  • a pharmaceutical composition can be formulated to be compatible with its intended route of administration.
  • pharmaceutical compositions include carriers, diluents, or excipients suitable for administration by routes including parenteral (e.g, subcutaneous (s.c.), intravenous, intramuscular, or intraperitoneal), intradermal, oral (e.g, ingestion), inhalation, intracavity, intracranial, and transdermal (topical).
  • compositions may be in the form of a sterile injectable aqueous or oleagenous suspension.
  • This suspension may be formulated using suitable dispersing or wetting agents and suspending agents disclosed herein or known to the skilled artisan.
  • the sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1,3 -butane did.
  • Acceptable diluents, solvents and dispersion media include water, Ringer’s solution, isotonic sodium chloride solution, Cremophor ELTM (BASF, Parsippany, NJ) or phosphate buffered saline (PBS), ethanol, polyol (e.g, glycerol, propylene glycol, and liquid polyethylene glycol), and suitable mixtures thereof.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • any bland fixed oil may be employed, including synthetic mono- or diglycerides.
  • fatty acids such as oleic acid find use in the preparation of injectables. Prolonged absorption of particular injectable formulations can be achieved by including an agent that delays absorption (e.g, aluminum monostearate or gelatin).
  • compositions may be in a form suitable for oral use, for example, as tablets, capsules, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups, solutions, microbeads or elixirs.
  • Pharmaceutical compositions intended for oral use may be prepared according to any method known to the art for the manufacture of pharmaceutical compositions. Such compositions may contain one or more agents such as sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide pharmaceutically elegant and palatable preparations.
  • Tablets containing a peptide provided herein may be in admixture with non-toxic pharmaceutically acceptable excipients suitable for the manufacture of tablets.
  • excipients include, for example, diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, for example, corn starch, or alginic acid; binding agents, for example starch, gelatin or acacia, and lubricating agents, for example magnesium stearate, stearic acid or talc.
  • Tablets, capsules and the like suitable for oral administration may be uncoated or they may be coated by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period.
  • a time delay material such as glyceryl monostearate or glyceryl distearate may be employed. They may also be coated by techniques known in the art to form osmotic therapeutic tablets for controlled release.
  • Additional agents include biodegradable or biocompatible particles or a polymeric substance such as polyesters, carbohydrates, polyamine acids, hydrogel, polyvinyl pyrrolidone, polyanhydrides, polyglycolic acid, polyanhydrides, polyglycolic acid, ethylene-vinylacetate, methylcellulose, carboxymethylcellulose, protamine sulfate, or lactide/glycolide copolymers, polylactide/glycolide copolymers, or ethyl enevinyl acetate copolymers in order to control delivery of an administered composition.
  • a polymeric substance such as polyesters, carbohydrates, polyamine acids, hydrogel, polyvinyl pyrrolidone, polyanhydrides, polyglycolic acid, polyanhydrides, polyglycolic acid, ethylene-vinylacetate, methylcellulose, carboxymethylcellulose, protamine sulfate, or lactide/glycolide copolymers, polylactide/glycolide
  • the oral agent can be entrapped in microcapsules prepared by coacervation techniques or by interfacial polymerization, by the use of hydroxymethylcellulose or gelatin-microcapsules or poly (methylmethacrolate) microcapsules, respectively, or in a colloid drug delivery system.
  • Colloidal dispersion systems include macromolecule complexes, nano-capsules, non encapsulated nanoparticles, microspheres, microbeads, and lipid-based systems ( e.g N-fatty acyl groups such as N-lauroyl, N-oleoyl, fatty amines such as dodecyl amine, oleoyl amine, etc., see US Patent No.
  • Formulations for oral use may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate, kaolin or microcrystalline cellulose, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example peanut oil, liquid paraffin, or olive oil.
  • an inert solid diluent for example, calcium carbonate, calcium phosphate, kaolin or microcrystalline cellulose
  • water or an oil medium for example peanut oil, liquid paraffin, or olive oil.
  • Aqueous suspensions contain the active materials in admixture with excipients suitable for the manufacture thereof.
  • excipients are suspending agents, for example sodium carboxymethylcellulose, methylcellulose, hydroxy-propylmethylcellulose, sodium alginate, polyvinyl-pyrrolidone, gum tragacanth and gum acacia; dispersing or wetting agents may be a naturally-occurring phosphatide, for example lecithin, or condensation products of an alkylene oxide with fatty acids, for example polyoxy-ethylene stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example heptadecaethyleneoxycetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids and a hexitol such as polyoxyethylene sorbitol monooleate, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, for example polyethylene sorbitan monooleate.
  • Oily suspensions may be formulated by suspending the active ingredient in a vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in a mineral oil such as liquid paraffin.
  • the oily suspensions may contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol. Sweetening agents such as those set forth above, and flavoring agents may be added to provide a palatable oral preparation.
  • Dispersible powders and granules suitable for preparation of an aqueous suspension by addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives.
  • a dispersing or wetting agent e.g., kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin, kaolin,
  • compositions provided herein may also be in the form of oil-in-water emulsions.
  • the oily phase may be a vegetable oil, for example olive oil or arachis oil, or a mineral oil, for example, liquid paraffin, or mixtures of these.
  • Suitable emulsifying agents may be naturally-occurring gums, for example, gum acacia or gum tragacanth; naturally-occurring phosphatides, for example, soy bean, lecithin, and esters or partial esters derived from fatty acids; hexitol anhydrides, for example, sorbitan monooleate; and condensation products of partial esters with ethylene oxide, for example, polyoxyethylene sorbitan monooleate.
  • compositions can also include carriers to protect the composition against rapid degradation or elimination from the body, such as a controlled release formulation, including implants, liposomes, hydrogels, prodrugs and microencapsulated delivery systems.
  • a time delay material such as glyceryl monostearate or glyceryl stearate alone, or in combination with a wax, may be employed.
  • Prolonged absorption of injectable pharmaceutical compositions can be achieved by including an agent that delays absorption, for example, aluminum monostearate or gelatin.
  • Prevention of the action of microorganisms can be achieved by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like.
  • peptides and/or one or more additional agents described herein in the form of suppositories for rectal administration can be prepared by mixing a peptide and/or one or more additional agents described herein with a suitable non irritating excipient which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the drug.
  • suitable non irritating excipient include, but are not limited to, cocoa butter and polyethylene glycols.
  • range expressly includes all possible subranges, all individual numerical values within that range, and all numerical values or numerical ranges including integers within such ranges and fractions of the values or the integers within ranges, unless the context clearly indicates otherwise.
  • This construction applies regardless of the breadth of the range and in all contexts throughout this patent document.
  • reference to a range of 90-100% includes 91-99%, 92-98%, 93-95%, 91-98%, 91-97%, 91-96%, 91-95%, 91- 94%, 91-93%, and so forth.
  • Reference to a range of 90-100% also includes 91%, 92%, 93%, 94%, 95%, 96%, 97%, etc., as well as 91.1%, 91.2%, 91.3%, 91.4%, 91.5%, etc., 92.1%, 92.2%, 92.3%, 92.4%, 92.5%, etc., and so forth.
  • reference to a range of 25-250, 250-500, 500- 1000, 1000-2500, 2500-5000, 5000-25,000, or 5000-50,000 includes any numerical value or range within or encompassing such values, e.g, 25, 26, 27, 28, 29...250, 251, 252, 253,
  • a series of ranges are disclosed throughout this document.
  • the use of a series of ranges includes combinations of the upper and lower ranges to provide another range. This construction applies regardless of the breadth of the range and in all contexts throughout this patent document.
  • reference to a series of ranges such as 5-10, 10-20, 20-30, 30-40, 40-50, 50-75, 75-100, 100-150, includes ranges such as 5-20, 5-30, 5-40, 5-50, 5-75, 5- 100, 5-150, and 10-30, 10-40, 10-50, 10-75, 10-100, 10-150, and 20-40, 20-50, 20-75, 20-100, 20-150, and so forth.
  • the invention is generally disclosed herein using affirmative language to describe the numerous embodiments.
  • the invention also specifically includes embodiments in which particular subject matter is excluded, in full or in part, such as substances or materials, method steps and conditions, protocols, procedures, assays or analysis.
  • the invention is generally not expressed herein in terms of what the invention does not include, aspects that are not expressly included in the invention are nevertheless disclosed herein.
  • M70 an analogue of FGF19, promotes Veillonella enrichment in the gut microbiome of patients with NASH. It was further identified that Veillonella was negatively correlated with bile acid levels, specifically the more hydrophobic and toxic bile acids.
  • Veillonella as a Bile Acid-sensitive Bacteria in the Gut Microbiome Analysis from M70 Trials in Patients with Non-alcoholic Steatohepatitis [00251] The in vivo role of M70 on minor genera Veillonella was explored in the context of NASH, using the methods described in Example I.
  • M70 promotes the enrichment of Veillonella , a gram negative bacterium that ferments lactate. Therefore, M70 may reduce lactate levels.
  • M70 may enrich Veillonella and reduce lactate through its negative regulation of bile acids. Because lactate is elevated in patients with advanced liver disease, which is associated with organ failure and mortality, M70 may be a useful treatment option for liver diseases or other related diseases that have elevated levels of lactate (see, e.g., Drolz et al., J. Hepatol. 69:258 (2019)).
  • liver steatosis Previous studies have shown that several phyla were significantly associated with liver steatosis (see, Hoyles et al., Nat. Med., 24(7):1070-80 (2016)), including Proteobacteria, Actinobacteria, and Verrucomicrobia that were positively correlated with liver steatosis, and Firmicutes and Euryarchaeota that were negatively correlated with liver steatosis.
  • To assess liver fat content patients underwent MRI-PDFF at baseline and at 12 weeks. Steatosis in hepatocytes was scored as 0, 1, 2 or 3 if there were less than 5%, 5-33%, 33-66%, or greater than 66% hepatocytes with fat, respectively.
  • Non-invasive detecting of liver fibrosis among patients with NASH is an important clinical need.
  • Previous studies comparing the bacterial taxonomic composition between patients with mild/moderate fibrosis and patients with advanced fibrosis have yielded variable and often contradictory findings.
  • To investigate the relationship between gut microbiota and liver fibrosis baseline data from the pooled cohort of NASH patients was used. Differences in the composition of microbiota taxa in patients with mild (FI) or advanced fibrosis (F3) was assessed. No differences were observed between FI and F3 subjects in alpha diversity (FIG. 7A).

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Genetics & Genomics (AREA)
  • Organic Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Zoology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biomedical Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • Wood Science & Technology (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Animal Behavior & Ethology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • General Engineering & Computer Science (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Epidemiology (AREA)
  • Plant Pathology (AREA)
  • Microbiology (AREA)
  • Toxicology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Peptides Or Proteins (AREA)

Abstract

L'invention concerne un procédé d'enrichissement de Veillonella et de réduction du lactate dans le microbiome intestinal chez un sujet, comprenant l'administration au sujet d'une quantité thérapeutiquement efficace d'une séquence peptidique chimérique, ce qui permet d'enrichir Veillonella et de réduire le lactate dans le microbiome intestinal chez le sujet. La présente invention inclut le traitement ou la prévention d'une maladie associée à des niveaux de lactate universellement élevés avec des variants des protéines du facteur de croissance des fibroblastes 19 (FGF19) et des séquences peptidiques (et des peptidomimétiques) et des fusions des protéines FGF19 et/ou des protéines du facteur de croissance des fibroblastes 21 (FGF21) et des séquences peptidiques (et des peptidomimétiques), et des variants de fusions de protéines FGF19 et/ou de protéines FGF21 et des séquences peptidiques (et des peptidomimétiques).
PCT/US2020/059060 2019-11-06 2020-11-05 Méthodes de réduction du lactate chez des patients malades du foie au moyen de variants et de fusions de polypeptides fgf19/fgf21 WO2021092140A1 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US201962931683P 2019-11-06 2019-11-06
US62/931,683 2019-11-06
US202063000948P 2020-03-27 2020-03-27
US63/000,948 2020-03-27

Publications (1)

Publication Number Publication Date
WO2021092140A1 true WO2021092140A1 (fr) 2021-05-14

Family

ID=75848225

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2020/059060 WO2021092140A1 (fr) 2019-11-06 2020-11-05 Méthodes de réduction du lactate chez des patients malades du foie au moyen de variants et de fusions de polypeptides fgf19/fgf21

Country Status (1)

Country Link
WO (1) WO2021092140A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2023085931A1 (fr) 2021-11-11 2023-05-19 Koninklijke Nederlandse Akademie Van Wetenschappen Organoïdes hépatiques

Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20110104152A1 (en) * 2009-10-15 2011-05-05 Sonoda Junichiro Chimeric fibroblast growth factors with altered receptor specificity
WO2016073855A1 (fr) * 2014-11-07 2016-05-12 Ngm Biopharmaceuticals, Inc. Procédés de traitement de troubles liés à l'acide biliaire et prédiction de la sensibilité clinique au traitement de troubles liés aux acides biliaires
US20160200788A1 (en) * 2012-11-28 2016-07-14 Ngm Biopharmaceuticals, Inc. Compositions, uses and methods for treatment of metabolic disorders and diseases
WO2018195390A1 (fr) * 2017-04-21 2018-10-25 Ngm Biopharmaceuticals, Inc. Méthodes de traitement des troubles liés à la motilité gastro-intestinale à l'aide de variants et de fusions de polypeptides fgf19/fgf21
US20190307847A1 (en) * 2016-08-29 2019-10-10 Ngm Biopharmaceuticals, Inc. Methods for treatment of bile acid-related disorders
WO2019195514A1 (fr) * 2018-04-04 2019-10-10 Genentech, Inc. Méthodes de détection et de quantification de fgf21

Patent Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20110104152A1 (en) * 2009-10-15 2011-05-05 Sonoda Junichiro Chimeric fibroblast growth factors with altered receptor specificity
US20180079806A1 (en) * 2009-10-15 2018-03-22 Genentech, Inc. Chimeric fibroblast growth factors with altered receptor specificity
US20160200788A1 (en) * 2012-11-28 2016-07-14 Ngm Biopharmaceuticals, Inc. Compositions, uses and methods for treatment of metabolic disorders and diseases
WO2016073855A1 (fr) * 2014-11-07 2016-05-12 Ngm Biopharmaceuticals, Inc. Procédés de traitement de troubles liés à l'acide biliaire et prédiction de la sensibilité clinique au traitement de troubles liés aux acides biliaires
US20190307847A1 (en) * 2016-08-29 2019-10-10 Ngm Biopharmaceuticals, Inc. Methods for treatment of bile acid-related disorders
WO2018195390A1 (fr) * 2017-04-21 2018-10-25 Ngm Biopharmaceuticals, Inc. Méthodes de traitement des troubles liés à la motilité gastro-intestinale à l'aide de variants et de fusions de polypeptides fgf19/fgf21
WO2019195514A1 (fr) * 2018-04-04 2019-10-10 Genentech, Inc. Méthodes de détection et de quantification de fgf21

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
ANDERSEN ET AL.: "Etiology and Therapeutic Approach to Elevated Lactate Levels", MAYO CLIN PROC, vol. 88, 1 October 2013 (2013-10-01), pages 1127 - 1140, XP055288931, DOI: 10.1016/j.mayocp.2013.06.012 *
BEYOGLU ET AL.: "The metabolomic window into hepatobiliary disease", J. HEPATHOL, vol. 59, 1 October 2013 (2013-10-01), pages 842 - 858, XP009173791, DOI: 10.1016/j.jhep.2013.05.030 *
DROLZ ANDREAS, HORVATITS THOMAS, RUTTER KAROLINE, LANDAHL FELIX, ROEDL KEVIN, MEERSSEMAN PHILIPPE, WILMER ALEXANDER, KLUWE JOHANNE: "Lactate Improves Prediction of Short-Term Mortality in Critically III Patients With Cirrhosis: A Multinational Study", HEPATOLOGY, vol. 69, 2 August 2018 (2018-08-02), pages 258 - 269, XP055824184 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2023085931A1 (fr) 2021-11-11 2023-05-19 Koninklijke Nederlandse Akademie Van Wetenschappen Organoïdes hépatiques

Similar Documents

Publication Publication Date Title
US20240050530A1 (en) Methods of Using Compositions Comprising Variants and Fusions of FGF19 Polypeptides for Treatment of Metabolic Disorders and Diseases
US11066454B2 (en) Compositions comprising variants and fusions of FGF19 polypeptides
US20220047675A1 (en) Pharmaceutical compositions comprising peptide variants and methods of use thereof
US9963494B2 (en) Methods of using compositions comprising variants and fusions of FGF19 polypeptides for reducing glucose levels in a subject
US20200330555A1 (en) Methods of treating gastrointestinal motility-related disorders using variants and fusions of fgf19/fgf21 polypeptides
AU2020260110A1 (en) Combination therapy for modulating bile acid homeostasis and treatment of bile acid disorders and diseases
WO2021092140A1 (fr) Méthodes de réduction du lactate chez des patients malades du foie au moyen de variants et de fusions de polypeptides fgf19/fgf21

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 20885485

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 20885485

Country of ref document: EP

Kind code of ref document: A1