REFERENCE TO SEQUENCE LISTING

This application includes a Sequence Listing submitted electronically as a text file named 18923802001SEQ, created on Mar. 7, 2020, with a size of 191 kilobytes. The Sequence Listing is incorporated herein by reference.

FIELD

The present disclosure relates generally to the treatment of subjects having increased lipid levels with Sterol Regulatory Element Binding Transcription Factor 1 (SREBF1) inhibitors, methods of identifying subjects having an increased risk of developing increased lipid levels, methods of detecting SREBF1 variant nucleic acid molecules and variant polypeptides, and SREBF1 variant nucleic acid molecules and SREBF1 variant polypeptides.

BACKGROUND

Lipid metabolism disorders are a well-known complication of obesity. Lipid metabolism disorders are often characterised by hyperinsulinaemia, elevated apolipoprotein B levels, high triglycerides concentration, high low-density lipoproteins (LDL) cholesterol concentration, and low high-density lipoproteins (HDL) cholesterol concentration.

Sterol Regulatory Element Binding Proteins (SREBPs) are transcriptional factors that control lipogenesis and lipid uptake. There are two SREBP genes in mammals, SREBP-1 and SREBP-2. The SREBP-1 gene transcribes two isoforms SREBP-1a and SREBP-1c encoded from different promoters, which regulate genes that control fatty acid synthesis. SREBP-2 regulates genes involved in cholesterol synthesis. The roles of SREBP-1 and SREBP-2, however, significantly overlap in the regulation of lipid metabolism. In normal tissues, SREBPs levels and activity are tightly controlled by endogenous sterol levels via negative feedback regulation. SREBPs are located in the endoplasmic reticulum (ER) membrane in association with SREBPs cleavage-activating protein (SCAP) in which they are retained by Insulin-induced gene (Insig) when cellular sterol levels are sufficient. Once sterol levels decrease, SCAP protein dissociates with Insig protein and escorts SREBPs to the Golgi, in which they are sequentially cleaved by site-1 and site-2 proteases (S1P and S2P) thereby releasing the N-terminus, which then enters into the nucleus to transcribe lipogenesis genes and low-density lipoprotein receptor (LDLR).

SUMMARY

The present disclosure also provides methods of treating a subject having increased total cholesterol, the method comprising administering an SREBF1 inhibitor to the subject.

The present disclosure also provides methods of treating a subject having increased low density lipoprotein (LDL), the method comprising administering an SREBF1 inhibitor to the subject.

The present disclosure also provides methods of treating a subject with a therapeutic agent that treats or inhibits an increased lipid level, wherein the subject is suffering from an increased lipid level, the method comprising the steps of determining whether the subject has an SREBF1 variant nucleic acid molecule encoding a human SREBF1 polypeptide by: obtaining or having obtained a biological sample from the subject; and performing or having performed a genotyping assay on the biological sample to determine if the subject has a genotype comprising the SREBF1 variant nucleic acid molecule; and when the subject is SREBF1 reference, then administering or continuing to administer to the subject the therapeutic agent that treats or inhibits the increased lipid level in a standard dosage amount, and administering to the subject an SREBF1 inhibitor; and when the subject is heterozygous for the SREBF1 variant, then administering or continuing to administer to the subject the therapeutic agent that treats or inhibits the increased lipid level in an amount that is the same as or lower than a standard dosage amount, and administering to the subject an SREBF1 inhibitor; wherein the presence of a genotype having the SREBF1 variant nucleic acid molecule encoding the human SREBF1 polypeptide indicates the subject has a reduced risk of developing the increased lipid level; wherein the increased lipid level is increased serum lipid level, increased total cholesterol, or increased LDL; and wherein the SREBF1 variant nucleic acid molecule is: i) a genomic nucleic acid molecule having a nucleotide sequence comprising a thymine at a position corresponding to position 17,922 according to SEQ ID NO:2, or the complement thereof, ii) an mRNA molecule having a nucleotide sequence comprising a uracil at a position corresponding to position 1,185 according to SEQ ID NO:6, or the complement thereof, iii) an mRNA molecule having a nucleotide sequence comprising a uracil at a position corresponding to position 1,260 according to SEQ ID NO:7, or the complement thereof, iv) an mRNA molecule having a nucleotide sequence comprising a uracil at a position corresponding to position 1,056 according to SEQ ID NO:8, or the complement thereof, v) a cDNA molecule produced from an mRNA molecule in the sample, wherein the cDNA molecule has a nucleotide sequence comprising a thymine at a position corresponding to position 1,185 according to SEQ ID NO:12, or the complement thereof, vi) a cDNA molecule produced from an mRNA molecule in the sample, wherein the cDNA molecule has a nucleotide sequence comprising a thymine at a position corresponding to position 1,260 according to SEQ ID NO:13, or the complement thereof, and/or vii) a cDNA molecule produced from an mRNA molecule in the sample, wherein the cDNA molecule has a nucleotide sequence comprising a thymine at a position corresponding to position 1,056 according to SEQ ID NO:14, or the complement thereof.

The present disclosure also provides methods of identifying a human subject having an increased risk for developing an increased lipid level, wherein the method comprises determining or having determined the presence or absence of an SREBF1 variant nucleic acid molecule encoding a human SREBF1 polypeptide in a biological sample obtained from the subject; wherein: when the human subject is SREBF1 reference, then the human subject has an increased risk for developing the increased lipid level; and when the human subject is heterozygous for the SREBF1 variant nucleic acid molecule or homozygous for the variant nucleic acid molecule, then the human subject has a decreased risk for developing the increased lipid level; wherein the SREBF1 variant nucleic acid molecule is: i) a genomic nucleic acid molecule having a nucleotide sequence comprising a thymine at a position corresponding to position 17,922 according to SEQ ID NO:2, or the complement thereof, ii) an mRNA molecule having a nucleotide sequence comprising a uracil at a position corresponding to position 1,185 according to SEQ ID NO:6, or the complement thereof, iii) an mRNA molecule having a nucleotide sequence comprising a uracil at a position corresponding to position 1,260 according to SEQ ID NO:7, or the complement thereof, iv) an mRNA molecule having a nucleotide sequence comprising a uracil at a position corresponding to position 1,056 according to SEQ ID NO:8, or the complement thereof, v) a cDNA molecule produced from an mRNA molecule in the sample, wherein the cDNA molecule has a nucleotide sequence comprising a thymine at a position corresponding to position 1,185 according to SEQ ID NO:12, or the complement thereof, vi) a cDNA molecule produced from an mRNA molecule in the sample, wherein the cDNA molecule has a nucleotide sequence comprising a thymine at a position corresponding to position 1,260 according to SEQ ID NO:13, or the complement thereof, and/or vii) a cDNA molecule produced from an mRNA molecule in the sample, wherein the cDNA molecule has a nucleotide sequence comprising a thymine at a position corresponding to position 1,056 according to SEQ ID NO:14, or the complement thereof.

The present disclosure also provides methods of detecting a human SREBF1 variant nucleic acid molecule in a human subject comprising assaying a sample obtained from the human subject to determine whether a nucleic acid molecule in the sample comprises a nucleotide sequence comprising: i) a thymine at a position corresponding to position 17,922 according to SEQ ID NO:2, or the complement thereof; ii) a uracil at a position corresponding to position 1,185 according to SEQ ID NO:6, or the complement thereof; iii) a uracil at a position corresponding to position 1,260 according to SEQ ID NO:7, or the complement thereof; iv) a uracil at a position corresponding to position 1,056 according to SEQ ID NO:8, or the complement thereof; v) a thymine at a position corresponding to position 1,185 according to SEQ ID NO:12, or the complement thereof; vi) a thymine at a position corresponding to position 1,260 according to SEQ ID NO:13, or the complement thereof; and/or vii) a thymine at a position corresponding to position 1,056 according to SEQ ID NO:14, or the complement thereof.

The present disclosure also provides isolated alteration-specific probes or alteration-specific primers comprising at least about 15 nucleotides, wherein the alteration-specific probe or alteration-specific primer comprises a nucleotide sequence which is complementary to a portion of a nucleotide sequence encoding a human SREBF1 polypeptide, wherein the portion comprises a position corresponding to: i) position 17,922 according to SEQ ID NO:2, or the complement thereof; ii) position 1,185 according to SEQ ID NO:6, or the complement thereof; iii) position 1,260 according to SEQ ID NO:7, or the complement thereof; iv) position 1,056 according to SEQ ID NO:8, or the complement thereof; v) position 1,185 according to SEQ ID NO:12, or the complement thereof; vi) position 1,260 according to SEQ ID NO:13, or the complement thereof; or vii) position 1,056 according to SEQ ID NO:14, or the complement thereof.

The present disclosure also provides isolated nucleic acid molecules comprising a nucleotide sequence encoding a human SREBF1 polypeptide, wherein the polypeptide comprises: i) a cysteine at a position corresponding to position 334 according to SEQ ID NO:18, or the complement thereof; ii) a cysteine at a position corresponding to position 364 according to SEQ ID NO:19, or the complement thereof; or iii) a cysteine at a position corresponding to position 310 according to SEQ ID NO:20, or the complement thereof.

The present disclosure also provides isolated nucleic acid molecules comprising a nucleotide sequence encoding a human SREBF1 polypeptide, wherein the nucleotide sequence comprises a thymine at a position corresponding to position 17,922 according to SEQ ID NO:2, or the complement thereof.

The present disclosure also provides isolated mRNA molecules comprising a nucleotide sequence encoding a human SREBF1 polypeptide, wherein the nucleotide sequence comprises: i) a uracil at a position corresponding to position 1,185 according to SEQ ID NO:6, or the complement thereof; ii) a uracil at a position corresponding to position 1,260 according to SEQ ID NO:7, or the complement thereof; or iii) a uracil at a position corresponding to position 1,056 according to SEQ ID NO:8, or the complement thereof.

The present disclosure also provides cDNA molecules comprising a nucleotide sequence encoding a human SREBF1 polypeptide, wherein the nucleotide sequence comprises: i) a thymine at a position corresponding to position 1,185 according to SEQ ID NO:12, or the complement thereof; ii) a thymine at a position corresponding to position 1,260 according to SEQ ID NO:13, or the complement thereof; or iii) a thymine at a position corresponding to position 1,056 according to SEQ ID NO:14, or the complement thereof.

The present disclosure also provides isolated SREBF1 polypeptides having: i) an amino acid sequence at least about 90% identical to SEQ ID NO:18, wherein the polypeptide comprises a cysteine at a position corresponding to position 334 according to SEQ ID NO:18; ii) an amino acid sequence at least about 90% identical to SEQ ID NO:19, wherein the polypeptide comprises a cysteine at a position corresponding to position 364 according to SEQ ID NO:19; or iii) an amino acid sequence at least about 90% identical to SEQ ID NO:20, wherein the polypeptide comprises a cysteine at a position corresponding to position 310 according to SEQ ID NO:20.

The present disclosure also provides molecular complexes comprising an alteration-specific primer or an alteration-specific probe hybridized to: i) a genomic nucleic acid molecule comprising a nucleotide sequence encoding a human SREBF1 polypeptide, wherein the alteration-specific primer or the alteration-specific probe is hybridized to a thymine at a position corresponding to position 17,922 according to SEQ ID NO:2, or the complement thereof; ii) an mRNA molecule comprising a nucleotide sequence encoding a human SREBF1 polypeptide, wherein the alteration-specific primer or the alteration-specific probe is hybridized to: a uracil at a position corresponding to position 1,185 according to SEQ ID NO:6, or the complement thereof; a uracil at a position corresponding to position 1,260 according to SEQ ID NO:7, or the complement thereof; or a uracil at a position corresponding to position 1,056 according to SEQ ID NO:8, or the complement thereof; or iii) a cDNA molecule comprising a nucleotide sequence encoding a human SREBF1 polypeptide, wherein the alteration-specific primer or the alteration-specific probe is hybridized to: a thymine at a position corresponding to position 1,185 according to SEQ ID NO:12, or the complement thereof; a thymine at a position corresponding to position 1,260 according to SEQ ID NO:13, or the complement thereof; or a thymine at a position corresponding to position 1,056 according to SEQ ID NO:14, or the complement thereof.

