Drugs and pharmaceutical compositions

Abstract

Provided herein are compounds, compositions, uses, and methods for reducing obesity in a subject, or for preventing or treating obesity. In certain examples, provided are methods for reducing obesity in a subject and / or for preventing or treating obesity in a subject in need thereof, which involve the administration of GDP-bound Rab1a (Rab1a GDP ), Rab1a GDP The method may include the step of treatment with one or more expressible nucleic acids encoding the

Description

[Technical Field]

[0001] The present invention relates generally to the treatment of obesity. More specifically, the present invention relates to compositions and methods for enhancing microautophagy to treat obesity and related diseases or disorders. [Background technology]

[0002] In eukaryotes, there are three known cellular processes by which cytosolic contents and / or intracellular organelles can be delivered to or captured by lysosomes for degradation: macroautophagy (commonly known as autophagy), microautophagy, and chaperone-mediated autophagy. Unlike macroautophagy or chaperone-mediated autophagy, microautophagy is a type of autophagic process mediated by direct lysosomal (in mammals) or vacuolar (in plants and fungi) engulfment of cytoplasmic targets or cargo (e.g., proteins, lipids, glycogen, or pathogens) into the vacuole.

[0003] Microautophagy can contribute to cytosolic protein degradation via multivesicular bodies (MVBs) in late endosomes. In addition, microautophagy can also support direct glycogen delivery to lysosomes and its degradation. In this regard, it is contemplated that dysfunctional or insufficient microautophagy may be associated with the development of various metabolic and / or neurological diseases, for example.

[0004] Clearly, the accumulation of specific proteins, lipids, and / or glycogen can be associated with a variety of important diseases, disorders, and conditions, and methods for targeting such proteins, lipids, and / or glycogen are desirable.

[0005] Obesity is a leading preventable cause of death worldwide, with rates increasing among both adults and children. Indeed, the prevalence of obesity has increased significantly over the past few decades, reaching epidemic proportions. Globally, more than one-third of adults are overweight (BMI 25-29.9 kg / m²) or obese (BMI ≥ 30 kg / m²) (Non-Patent Document 1). Obesity is a complex disease involving excessive body fat, which increases the risk of other diseases and health problems, such as heart disease, diabetes, hypertension, and certain cancers (Non-Patent Document 2). Because the efficacy of current treatments remains poor and focuses primarily on reducing caloric intake, obesity is widely considered one of the most serious public health problems of the 21st century. Furthermore, sibutramine, one of the most effective drugs for weight loss, was withdrawn from the U.S. market due to cardiovascular safety concerns. Therefore, clinical management of obesity remains a major challenge in the absence of effective medications. Therefore, anti-obesity drugs that target the molecular and cellular mechanisms involved in obesity are highly desirable.

[0006] Alternative, additional, and / or improved anti-obesity agents, compositions, and / or methods for treating obesity and related diseases or disorders are desirable. Summary of the Invention

[0007] As described in detail herein, GDP-bound forms of Rab1a, e.g., Rab1a S25N , Rab1a N124I , Rab1a D41N , Rab1a D47N or another dominant-negative (DN) GDP-bound form of Rab1a, or such Rab1a GDP It has now been identified that treatment with a microautophagy-enhancing agent comprising one or more expressible nucleic acids encoding: can be used to reduce obesity in obese subjects.

[0008] In certain embodiments, provided herein is a method for reducing obesity, or preventing or treating obesity in a subject in need thereof, said method comprising: GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP treating a subject with one or more expressible nucleic acids encoding This results in Rab1a GDP increasing cellular levels of and resulting in a reduction in obesity in a subject.

[0009] In certain embodiments, obesity is associated with at least one of the following conditions: inflammatory response, increased abdominal obesity, high blood pressure, hyperglycemia, high serum triglycerides, low serum high-density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0010] In another embodiment, provided herein is a method for reducing weight in an overweight subject, said method comprising: GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP or a combination thereof, Thereby, Rab1a in overweight subjects GDP increasing cellular levels of and resulting in weight loss in a subject.

[0011] In another embodiment of any of the above methods, Rab1a GDP Rab1a S25N , Rab1a N124I , Rab1a D41N , Rab1a D47N , or may be or include another dominant-negative (DN) GDP-bound form of Rab1a.

[0012] In yet another embodiment of any of the above methods, Rab1a GDP may comprise the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41N or MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0013] In yet another embodiment of any of the above methods, Rab1a GDP can consist of the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41N or MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0014] In another embodiment of any of the above methods, Rab1a GDP may comprise or consist of the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0015] In yet another embodiment of any of the above methods, Rab1a GDP may be in the form of a fusion protein, wherein Rab1a GDP is fused or otherwise linked, directly or indirectly, optionally via a linker, to a signal or targeting peptide, a fluorescent peptide, or other marker or tracer, or to another peptide or non-peptide moiety for targeted delivery, to facilitate cellular uptake, to increase stability or in vivo half-life, or to enhance Rab1a GDP improve another therapeutic, diagnostic, or in vivo property of

[0016] In yet another embodiment of any of the above methods, the fusion protein may comprise the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0017] In another embodiment of any of the above methods, Rab1a GDP may be in the form of a fusion protein and may comprise the following amino acid sequence: MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAHIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSMYTFVTSHLPPGFNEYDFVPESFDR DKTIALIMNSSGSTGLPKGVALPHRTACVRFSHARDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYKIQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYGLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLD TGKTLGVNQRGELCVRGPMIMSGYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILLQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVFVDEVPKGLTGKLDARKIREILIKAKKGGKSKLMSSMNPEYDYLFKLLL IGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNVKQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 21), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0018] In yet another embodiment of any of the above methods, the one or more expressible nucleic acids are Rab1a, Rab2a, Rab3a, Rab4a, Rab5a, Rab6a, Rab7a, Rab8a, Rab9a, Rab10a, Rab11a, Rab12a, Rab13a, Rab14a, Rab15a, Rab16a, Rab17a, Rab18a, Rab19a, Rab GDP It may encode any one or more of:

[0019] In yet another embodiment of any of the above methods, the one or more expressible nucleic acids may be DNA-based or RNA-based.

[0020] In another embodiment of any of the above methods, the one or more expressible nucleic acids express Rab1a in one or more cells of the subject. GDP or wherein one or more expressible nucleic acids are integrated into the genome of one or more cells and express Rab1a in one or more cells of a subject. GDP can be expressed.

[0021] In yet another embodiment of any of the above methods, the one or more expressible nucleic acids is Rab1a GDP The gene may include one or more expression vectors, plasmids, or mRNAs encoding and capable of expressing the gene in one or more cells of the subject.

[0022] In yet another embodiment of any of the above methods, the one or more expressible nucleic acids may comprise the following nucleic acid sequence: ATGTCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGACTCAGGGGTTGGAAAGAATTGCCTTCTTCTTAGGTTTGCAGATGATACATATACAGAAAGCTACATCAGCACAATTGGTGTGGATTTCAAAATAAGAA CTATAGAGTTAGACGGGAAAACAATCAAGCTTCAAATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACCTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAGGAGTCCTTCAATAATGTTAAA CAGTGGCTGCAGGAAATAGATCGTTATGCCAGTGAAAATGTCAACAAATTGTTGGTAGGGAACAAATGTGATCTGACCACAAAGAAAGTAGTAGACTACACAACAGCGAAGGAATTTGCTGATTCCCTTGGAATTCCGTTTTTGGAAACCAGTG CTAAGAATGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAGCGAATGGGTCCCGGAGCAACAGCTGGTGGTGCTGAGAAGTCCAATGTTAAAATTCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGTTGCTGCTAA (Human Rab1a S25N ORF codon sequence, SEQ ID NO: 5), ATGGGGGACTACAAGGACGACGATGACAAGGGGGGTAGCGGTGGATCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGATTCTGGGGTTGGAAAGTCCTGCCTTCTCCTTAGGTTTGCAGATGATACGTATACGGAAAGCTACATC AGCACAATTGGTGTGGATTTCAAGATACGAACTATAGAGTTAGATGGGAAAACAATCAAGCTACAGATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACTTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAG GAGTCCTTCAATAACGTTAAACAGTGGCTGCAGGAGATAGATCGCTACGCCAGTGAAAATGTCAACAAGTTGTTGGTAGGGATCAAATGTGACCTGACCACAAAGAAAGTAGTAGACTACACAACAGCAAAGGAATTTGCAGATTCCCTTGGAATTCCATTTTTG GAAACCAGTGCTAAGAACGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAAGCGAATGGGTCCTGGAGCTACAGCTGGTGGTGCCGAGAAGTCCAATGTTAAAATCCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGCTGCTGCTAA (Mouse Rab1a N124I ORF codon sequence, SEQ ID NO: 8), or ATGTCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGACTCAGGGGTTGGAAAGTCTTGCCTTCTTCTTAGGTTTGCAGATGATACATATACAGAAAGCTACATCAGCACAATTGGTGTGAACTTCAAAATAAGAA CTATAGAGTTAGACGGGAAAACAATCAAGCTTCAAATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACCTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAGGAGTCCTTCAATAATGTTAAA CAGTGGCTGCAGGAAATAGATCGTTATGCCAGTGAAAATGTCAACAAATTGTTGGTAGGGAACAAATGTGATCTGACCACAAAGAAAGTAGTAGACTACACAACAGCGAAGGAATTTGCTGATTCCCTTGGAATTCCGTTTTTGGAAACCAGTG CTAAGAATGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAGCGAATGGGTCCCGGAGCAACAGCTGGTGGTGCTGAGAAGTCCAATGTTAAAATTCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGTTGCTGCTAA (Human Rab1a D47N ORF codon sequence, SEQ ID NO: 17), or a Rab1a having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which binds preferentially to GDP. GDP a nucleic acid sequence encoding Alternatively, a nucleic acid sequence equivalent to any of the above sequences due to codon redundancy.

[0023] In another embodiment of any of the above methods, the one or more expressible nucleic acids may comprise the following nucleic acid sequence: (SEQ ID NO: 19, DNA sequence of MG-008 ORF with 5' luciferase tag), or (SEQ ID NO: 20, mRNA sequence of MG-008 ORF with 5' luciferase tag), or a Rab1a having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which binds preferentially to GDP. GDP a nucleic acid sequence encoding Alternatively, a nucleic acid sequence equivalent to any of the above sequences due to codon redundancy.

[0024] In another embodiment, a GDP-bound form of Rab1a (Rab1a) is administered to a subject in need thereof for reducing obesity in the subject or for preventing or treating obesity. GDP ), Rab1a GDP Provided herein are uses of one or more expressible nucleic acids encoding the

[0025] In another embodiment, provided herein is the use of the GDP-bound form of Rab1a (Rab1aGDP), one or more expressible nucleic acids encoding Rab1aGDP, or a combination thereof, to reduce, prevent, or treat at least one of the following conditions in a subject in need thereof: inflammatory response, increased abdominal obesity, high blood pressure, hyperglycemia, high serum triglycerides, low serum high-density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0026] In another embodiment, a GDP-bound form of Rab1a (Rab1a GDP ), one or more expressible nucleic acids encoding Rab1aGDP, or combinations thereof are provided herein.

[0027] In another embodiment, use is made of a GDP-bound form of Rab1a (Rab1a) in the manufacture of a medicament for reducing, or preventing or treating obesity in a subject in need thereof. GDP ), Rab1a GDP Provided herein are uses of one or more expressible nucleic acids encoding the

[0028] In another embodiment, provided herein is the use of a GDP-bound form of Rab1a (Rab1aGDP), one or more expressible nucleic acids encoding Rab1aGDP, or a combination thereof, in the manufacture of a medicament for reducing, preventing, or treating at least one of the following conditions in a subject in need thereof: inflammatory response, increased abdominal obesity, hypertension, hyperglycemia, high serum triglycerides, low serum high-density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0029] In another embodiment, the use of a GDP-bound form of Rab1a (Rab1a) in the manufacture of a medicament for reducing weight in an overweight subject. GDP ), Rab1a GDP Provided herein are uses of one or more expressible nucleic acids encoding the

[0030] In another embodiment of any of the above uses, Rab1a GDP Rab1a S25N , Rab1a N124I , Rab1a D41N , Rab1a D47N , or may be or include another dominant-negative (DN) GDP-bound form of Rab1a.

[0031] In yet another embodiment of any of the above uses, Rab1a GDP may comprise the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41N or MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0032] In yet another embodiment of any of the above uses, Rab1a GDP can consist of the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41N or MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0033] In another embodiment of any of the above uses, Rab1a GDP may comprise or consist of the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0034] In yet another embodiment of any of the above uses, Rab1a GDP may be in the form of a fusion protein, wherein Rab1a GDP is fused or otherwise linked, directly or indirectly, optionally via a linker, to a signal or targeting peptide, a fluorescent peptide, or other marker or tracer, or to another peptide or non-peptide moiety for targeted delivery, to facilitate cellular uptake, to increase stability or in vivo half-life, or to enhance Rab1a GDP improve another therapeutic, diagnostic, or in vivo property of

[0035] In yet another embodiment of any of the above uses, the fusion protein may comprise the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0036] In another embodiment of any of the above uses, Rab1a GDP may be in the form of a fusion protein and may comprise the following amino acid sequence: MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAHIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSMYTFVTSHLPPGFNEYDFVPESFDR DKTIALIMNSSGSTGLPKGVALPHRTACVRFSHARDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYKIQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYGLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLD TGKTLGVNQRGELCVRGPMIMSGYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILLQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVFVDEVPKGLTGKLDARKIREILIKAKKGGKSKLMSSMNPEYDYLFKLLL IGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNVKQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 21), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0037] In yet another embodiment of any of the above uses, the one or more expressible nucleic acids are one or more Rab1a as defined herein. GDP may be coded.

[0038] In yet another embodiment of any of the above uses, the one or more expressible nucleic acids may be DNA-based or RNA-based.

[0039] In another embodiment of any of the above uses, the one or more expressible nucleic acids express Rab1a in one or more cells of the subject. GDP or wherein the one or more expressible nucleic acids are integrated into the genome of one or more cells of the subject and express Rab1a in one or more cells of the subject. GDP can be expressed.

[0040] In another embodiment of any of the above uses, the one or more expressible nucleic acids is Rab1a GDP The gene may include one or more expression vectors, plasmids, or mRNAs encoding and capable of expressing the gene in one or more cells of the subject.

[0041] In another embodiment of any of the above uses, the one or more expressible nucleic acids may comprise the following nucleic acid sequence: ATGTCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGACTCAGGGGTTGGAAAGAATTGCCTTCTTCTTAGGTTTGCAGATGATACATATACAGAAAGCTACATCAGCACAATTGGTGTGGATTTCAAAATAAGAA CTATAGAGTTAGACGGGAAAACAATCAAGCTTCAAATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACCTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAGGAGTCCTTCAATAATGTTAAA CAGTGGCTGCAGGAAATAGATCGTTATGCCAGTGAAAATGTCAACAAATTGTTGGTAGGGAACAAATGTGATCTGACCACAAAGAAAGTAGTAGACTACACAACAGCGAAGGAATTTGCTGATTCCCTTGGAATTCCGTTTTTGGAAACCAGTG CTAAGAATGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAGCGAATGGGTCCCGGAGCAACAGCTGGTGGTGCTGAGAAGTCCAATGTTAAAATTCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGTTGCTGCTAA (Human Rab1a S25N ORF codon sequence, SEQ ID NO: 5), ATGGGGGACTACAAGGACGACGATGACAAGGGGGGTAGCGGTGGATCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGATTCTGGGGTTGGAAAGTCCTGCCTTCTCCTTAGGTTTGCAGATGATACGTATACGGAAAGCTACATC AGCACAATTGGTGTGGATTTCAAGATACGAACTATAGAGTTAGATGGGAAAACAATCAAGCTACAGATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACTTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAG GAGTCCTTCAATAACGTTAAACAGTGGCTGCAGGAGATAGATCGCTACGCCAGTGAAAATGTCAACAAGTTGTTGGTAGGGATCAAATGTGACCTGACCACAAAGAAAGTAGTAGACTACACAACAGCAAAGGAATTTGCAGATTCCCTTGGAATTCCATTTTTG GAAACCAGTGCTAAGAACGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAAGCGAATGGGTCCTGGAGCTACAGCTGGTGGTGCCGAGAAGTCCAATGTTAAAATCCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGCTGCTGCTAA (Mouse Rab1a N124I ORF codon sequence, SEQ ID NO: 8), or ATGTCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGACTCAGGGGTTGGAAAGTCTTGCCTTCTTCTTAGGTTTGCAGATGATACATATACAGAAAGCTACATCAGCACAATTGGTGTGAACTTCAAAATAAGAA CTATAGAGTTAGACGGGAAAACAATCAAGCTTCAAATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACCTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAGGAGTCCTTCAATAATGTTAAA CAGTGGCTGCAGGAAATAGATCGTTATGCCAGTGAAAATGTCAACAAATTGTTGGTAGGGAACAAATGTGATCTGACCACAAAGAAAGTAGTAGACTACACAACAGCGAAGGAATTTGCTGATTCCCTTGGAATTCCGTTTTTGGAAACCAGTG CTAAGAATGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAGCGAATGGGTCCCGGAGCAACAGCTGGTGGTGCTGAGAAGTCCAATGTTAAAATTCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGTTGCTGCTAA (Human Rab1a D47N ORF codon sequence, SEQ ID NO: 17), or a Rab1a having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which binds preferentially to GDP. GDP a nucleic acid sequence encoding Alternatively, a nucleic acid sequence equivalent to any of the above sequences due to codon redundancy.

[0042] In yet another embodiment of any of the above uses, the one or more expressible nucleic acids may comprise the following nucleic acid sequence: (SEQ ID NO: 19, DNA sequence of MG-008 ORF with 5' luciferase tag), or (SEQ ID NO: 20, mRNA sequence of MG-008 ORF with 5' luciferase tag), or a Rab1a having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which binds preferentially to GDP. GDP a nucleic acid sequence encoding Alternatively, a nucleic acid sequence equivalent to any of the above sequences due to codon redundancy.

[0043] In another embodiment, provided herein is a polypeptide comprising the following amino acid sequence for use in reducing, preventing, or treating obesity in a subject in need thereof, or for use in reducing weight in an overweight subject: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41Nor MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0044] In certain embodiments, the above polypeptides are for use in reducing obesity or preventing or treating obesity in a subject in need thereof, wherein the obesity is associated with at least one of the following conditions: inflammatory response, increased abdominal obesity, high blood pressure, hyperglycemia, high serum triglycerides, low serum high-density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0045] In another embodiment, provided herein is a pharmaceutical composition comprising: GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP or a combination thereof, and Another anti-obesity drug.

[0046] In yet another embodiment, provided herein is a kit comprising any one or more of the following: GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP one or more expressible nucleic acids encoding anti-obesity drugs, instructions for carrying out any of the methods described herein; or Any combination of them.

[0047] These and other characteristics suggest that obese mice express the Rab1a mRNA GDP This will be further understood in connection with the following drawings, which are illustrated by: [Brief explanation of the drawings]

[0048] [Figure 1] FIG. 1 shows that obese mice treated with MG-008 lost weight 12 days after injection, as confirmed as described in Example 1. [Figure 2A] FIG. 2A shows certain amino acid or nucleic acid sequences described herein. [Figure 2B] FIG. 2B shows certain amino acid or nucleic acid sequences described herein. [Figure 2C] FIG. 2C shows certain amino acid or nucleic acid sequences described herein. [Figure 2D] FIG. 2D shows certain amino acid or nucleic acid sequences described herein. [Figure 2E] FIG. 2E shows certain amino acid or nucleic acid sequences described herein. [Figure 2F] FIG. 2F shows certain amino acid or nucleic acid sequences described herein. [Figure 2G] FIG. 2G shows certain amino acid or nucleic acid sequences described herein. [Figure 2H] FIG. 2H shows certain amino acid or nucleic acid sequences described herein. [Figure 2I] FIG. 2I shows certain amino acid or nucleic acid sequences described herein. [Figure 2J] FIG. 2J shows certain amino acid or nucleic acid sequences described herein. [Figure 2K] FIG. 2K shows certain amino acid or nucleic acid sequences described herein. [Figure 2L] FIG. 2L shows certain amino acid or nucleic acid sequences described herein. [Figure 2M] FIG. 2M shows certain amino acid or nucleic acid sequences described herein. [Figure 2N] FIG. 2N shows certain amino acid or nucleic acid sequences described herein. [Figure 2O] FIG. 2O depicts certain amino acid or nucleic acid sequences described herein. [Figure 2P] FIG. 2P shows certain amino acid or nucleic acid sequences described herein. [Figure 2Q] Figure 2Q shows certain amino acid or nucleic acid sequences described herein. [Figure 2R] FIG. 2R shows certain amino acid or nucleic acid sequences described herein. [Figure 2S] FIG. 2S shows certain amino acid or nucleic acid sequences described herein. [Figure 2T] FIG. 2T shows certain amino acid or nucleic acid sequences described herein. [Figure 2U] Figure 2U shows certain amino acid or nucleic acid sequences described herein. [Figure 3] FIG. 3 shows the timeline of mice treated with F11 and the corresponding time points at which data were collected. [Figure 4]Figure 4 shows baseline fat and lean mass in diet-induced obese (DIO) mice before F11 treatment as assessed by NMR analysis. Figure 4A shows the correlation between total fat and body weight. Figure 4B shows the correlation between total lean mass and body weight. Figure 4C shows the correlation between total fat and total lean mass. Figure 4D shows the correlation between percentage of total fat and body weight. Figure 4E shows the correlation between percentage of total lean mass and body weight. Figure 4F shows the correlation between percentage of total fat and percentage of total lean mass. [Figure 5] Figure 5 shows fat and lean mass in diet-induced obese (DIO) mice after 3 days of F11 treatment confirmed by NMR analysis. Figure 5A shows the correlation between total fat and body weight. Figure 5B shows the correlation between total lean mass and body weight. Figure 5C shows the correlation between total fat and total lean mass. Figure 5D shows the correlation between the percentage of total fat and body weight. Figure 5E shows the correlation between the percentage of total lean mass and body weight. Figure 5F shows the correlation between the percentage of total fat and the percentage of total lean mass. [Figure 6] Figure 6 shows fat and lean mass in diet-induced obese (DIO) mice after 17 days of F11 treatment confirmed by NMR analysis. Figure 6A shows the correlation between total fat and body weight. Figure 6B shows the correlation between total lean mass and body weight. Figure 6C shows the correlation between total fat and total lean mass. Figure 6D shows the correlation between the percentage of total fat and body weight. Figure 6E shows the correlation between the percentage of total lean mass and body weight. Figure 6F shows the correlation between the percentage of total fat and the percentage of total lean mass. [Figure 7]Figure 7 shows the changes in body weight, fat weight, and lean weight values ​​in diet-induced obese (DIO) mice after 3 days of F11 treatment, as assessed by NMR analysis. Figure 7A shows the body weight changes in DIO mice after 3 days of F11 treatment. Figure 7B shows the total fat changes in DIO mice after 3 days of F11 treatment. Figure 7C shows the total lean weight changes in DIO mice after 3 days of F11 treatment. The changed values ​​were calculated by the formula: changed value (grams) = value (grams) on day 3 after IV administration - value (grams) at baseline (before IV administration). [Figure 8] Figure 8 shows the changes in body weight, fat weight, and lean weight values ​​in diet-induced obese (DIO) mice after 17 days of F11 treatment, as assessed by NMR analysis. Figure 8A shows the body weight changes in DIO mice after 17 days of F11 treatment. Figure 8B shows the total fat changes in DIO mice after 17 days of F11 treatment. Figure 8C shows the total lean weight changes in DIO mice after 17 days of F11 treatment. The changed values ​​were calculated by the formula: changed value (grams) = value at 17 days after IV (grams) - baseline (pre-IV) value (grams). [Figure 9] Figure 9 shows a multiple logistic regression analysis of F11 efficacy in diet-induced obese (DIO) mice after 3 days of F11 treatment. Figure 9A shows a multiple logistic regression analysis of F11 efficacy in DIO mice after 3 days of F11 treatment for body weight change. Figure 9B shows a multiple logistic regression analysis of F11 efficacy in DIO mice after 3 days of F11 treatment for fat weight change. Figure 9C shows a multiple logistic regression analysis of F11 efficacy in DIO mice after 3 days of F11 treatment for lean weight change. [Figure 10]Figure 10 shows a multiple logistic regression analysis of F11 efficacy in diet-induced obese (DIO) mice after 17 days of F11 treatment. Figure 10A shows a multiple logistic regression analysis of F11 efficacy in DIO mice after 17 days of F11 treatment for body weight change. Figure 10B shows a multiple logistic regression analysis of F11 efficacy in DIO mice after 17 days of F11 treatment for fat weight change. Figure 10C shows a multiple logistic regression analysis of F11 efficacy in DIO mice after 17 days of F11 treatment for lean body weight change. [Figure 11] FIG. 11 shows the body weight (A) and percentage body weight (B) changes in diet-induced obese (DOI) mice before and after F11 treatment. DETAILED DESCRIPTION OF THE INVENTION

[0049] Described herein are compounds, compositions, uses, and methods for reducing obesity in a subject and / or for preventing or treating obesity in a subject in need thereof. It will be understood that the embodiments and examples are provided for illustrative purposes intended for those skilled in the art and are not intended to be limiting in any way.

[0050] Obesity is a complex disease because its causes are often multifactorial. Indeed, several factors, including diet, physical activity, automation, urbanization, genetic predisposition, drugs, psychiatric disorders, economic policies, endocrine abnormalities, and exposure to endocrine-disrupting chemicals, can contribute to an individual's obesity. One of the hallmarks of obesity is the accumulation of dysfunctional adipose tissue when energy intake exceeds energy expenditure. This induces an inflammatory response and metabolic stress by increasing levels of fatty acids, triglycerides, and LDL cholesterol, leading to a cluster of interrelated complications, including insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, nonalcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0051] Autophagy is the primary intracellular degradation system, and its degradative capacity is derived from lysosomes. It is a homeostatic and evolutionarily conserved mechanism of self-digestion, allowing cells to degrade and recycle long-lived proteins and excess or damaged organelles, adapting to adverse microenvironmental conditions, including limited nutrient supplies (Allyson et al., “Cleaning House: Selective Autophagy of Organelles,” Developmental Cell, (US), 2017, Vol. 41, pp. 10-22). Several sensors interacting with the autophagy machinery have evolved to detect fluctuations in important metabolic parameters. Indeed, because lysosomal disposal of intracellular macromolecules always leads to their degradation into essential metabolic intermediates, including amino acids, glucose, nucleotides, and free fatty acids (FAs), autophagy stands out as a key regulator of responses to energy stress, both at tissue-specific and systemic levels (Rabinowitz JD, White E, “Autophagy and Metabolism,” Science, (US), 2010, Vol. 330, pp. 1344-1348; Galluzzi L, Pietrocola F, Levine B, Kroemer G, “Metabolic Control of Autophagy,” Cell, (US), 2014, Vol. 159, pp. 1263-1276). Thus, autophagy fulfills tissue-specific metabolic tasks within key organs involved in maintaining the organism's energy balance, including adipose tissue, liver, and exocrine pancreas. Thus, insufficient autophagy is associated with metabolic syndrome, which is characterized by at least three of the following five medical conditions: abdominal obesity, hypertension, hyperglycemia, high serum triglycerides, and low serum high-density lipoprotein (HDL).

[0052] Autophagy is a highly regulated process by which misfolded proteins and organelles reach the lysosome for their degradation (Kim & Lee, 2014). There are three different types of autophagy: macroautophagy (commonly known as autophagy), chaperone-mediated autophagy, and microautophagy. In chaperone-mediated autophagy and microautophagy, lysosomal degradation of substrates occurs directly within the lysosome (Parzych & Klionsky, 2014).

[0053] Global and tissue-specific deletion and / or mutation of specific autophagy genes or autophagy-regulating molecules results in altered lipid metabolism, hepatic steatosis, and obese or diabetic phenotypes in animal models (Kim & Lee, 2014; Lee et al., 2016). More specifically, genetic ablation or loss-of-function of several autophagy-related genes, including Atg7 (Lim et al., 2014), Atg4b (Fernandez et al., 2017), Becn2 (He et al., 2013), and Tfeb (Settembre et al., 2013), predisposes these animals to metabolic disorders, both on a normal and high-fat diet, at the systemic level or in a tissue-restricted manner. Furthermore, obesity is associated with increased plasma levels of autophagy inhibitors, including DBI / ACBP, in both humans and mice (Bravo-San Pedro et al., 2019; Joseph et al., 2020). These findings support a role for autophagy in the development of obesity (obesogenesis) and obesity-related complications (Zhang et al., 2018).

[0054] Autophagy is regulated by several signaling molecules, particularly the mechanistic target of rapamycin (mTOR) kinase and the autophagy-related protein (ATG) family. Dietary intake transiently increases plasma levels of branched-chain amino acids (BCAAs), including leucine, which subsequently activate mTOR signaling and thereby inhibit autophagy (Nicklin et al., 2009; Broer & Broer, 2017). Activation of mTOR as a result of increased growth factor and insulin signaling and / or increased BCAA consumption is common in human obesity and experimental models and is thought to be the primary driver of nutrition-induced suppression of autophagy (Shimobayashi & Hall, 2016; Meijer et al., 2015). In contrast, autophagy is capable of maintaining normal amino acid levels during short-term fasting. Similarly, high levels of plasma glucose activate insulin-like growth factor 1 (IGF1) signaling, leading to AKT1-dependent activation of mTOR via inhibition of TSC2-RHEB signaling. Interestingly, both glucose and amino acids share similar means of activating mTORC1 in a RAG-dependent manner. Furthermore, autophagy also controls glucose and energy metabolism through regulation of gluconeogenesis (Kim & Lee, 2014).

[0055] High lipid levels are common in obesity and can suppress autophagy by blocking autophagolysosome fusion, lysosomal acidification, and hydrolase activity (Wang, 2016). Indeed, high lipid levels inhibit autophagosome-lysosome fusion (Koga et al., 2010). Autophagy can regulate lipid homeostasis through the selective degradation of lipid droplets by lipophagy and the utilization of lipids as an energy source (Nguyen & Olzmann, 2017). Conversely, evidence points to a role for adipose autophagy in the regulation of adipose tissue development, adipogenesis, and lipid metabolism (Tao et al., 2016). Furthermore, adipose lysosomal dysfunction has been shown to contribute to autophagosome accumulation and early adipose pathology in obesity (Mizunoe et al., 2017). Furthermore, autophagy dysfunction can facilitate the transition from obesity to diabetes, highlighting the therapeutic potential of autophagy regulators for the prevention and treatment of diabetes and obesity ( Lim et al., 2014 ).

[0056] The specialized functions of autophagy in metabolic regulation include, among other processes, adipocyte differentiation, hepatic fat accumulation, maintenance of pancreatic beta-cell health, central nervous system (CNS)-mediated regulation of food intake, and inflammatory responses (Klionsky et al., “Autophagy in major human diseases,” The EMBO Journal, (UK), 2021, Vol. 40: pp. e108863-e108863). Furthermore, regulation of autophagy in the CNS has been shown to contribute to weight control. More specifically, inhibition of autophagy in proopiomelanocortin (POMC)-producing neuronal subtypes has been shown to promote the development of obesity by stimulating increased feeding (Quan et al., 2012).

[0057] Reactive oxygen species (ROS) have been shown to play an important role in obesity (Lavallard et al., 2012). Excessive ROS production promotes the release of cytochrome c from mitochondria into the cytoplasm, contributing to the initiation and development of insulin resistance and diabetes in obesity (Houstis et al., 2006; Petersen et al., 2004). Unfortunately, ROS are an unavoidable by-product of the bioenergetic process, which is most produced in mitochondria during the ATP synthesis process known as oxidative phosphorylation.

[0058] In this regard, the present inventors recognized that in addition to suppressing caloric intake, catabolic processes mediated by autophagy lysosomes, particularly the process of microautophagy, could be used to develop effective therapeutic strategies for obesity.

[0059] Stimulating microautophagy (which in certain embodiments may involve engulfment of target membrane fragments) by promoting lysosome movement to target membranes may be particularly desirable for the treatment of various diseases and / or disorders and / or conditions. Specific lysosomal location within cells may be associated with different types of lysosomal activity. In addition, lysosomal location may correlate with mTOR activity and regulate autophagic flux. During macroautophagy, mTORC1 is inactive, and lysosomes may accumulate in the perinuclear region of cells. This may result in macroautophagy by stimulating fusion of the encased target membrane with lysosomes. In contrast, in the process of microautophagy, engulfment of target membrane fragments by lysosomes may occur through lysosomal movement and direct interaction with the target membrane, without the need for autophagosome formation. Lysosomes can move in a bidirectional manner on microtubules. Such lysosomal motility is governed by different sets of motor proteins recruited by different mechanisms.

[0060] In certain embodiments, microautophagy can involve the direct engulfment of cytoplasmic cargo at the limiting membrane by autophagic tubes. This can mediate both invagination into the lysosomal lumen and vesicle scission (see Li, W.-W., Li, J. & Bao, J.-K., "Microautophagy: lesser-known self-eating," Cellular and Molecular Life Sciences, (Switzerland), 2011, Vol. 69, pp. 1125-1136). Direct lysosomal degradation of target substrates, such as DNA, can occur (referred to as piecemeal autophagy; see Fujiwara, Y. et al., "Direct uptake and degradation of DNA by lysosomes," PubMed-NCBI, Autophagy, (USA), 2014, Vol. 9, pp. 1167-1171).In addition, lysosomes may be capable of moving to different organelles and / or membrane substrates (e.g., plasma membrane, mitochondria) through the recruitment of motor proteins and SNARE proteins, and may interact directly with them (Andrews, NW, "Lysosomes and the plasma membrane", Journal of Cell Biology, (USA), 2002, Vol. 158, pp. 389-394; Hofmann, I. & Munro, S., "An N-terminally acetylated Arf-like GTPase is localized to lysosomes and affects their motility", Journal of Cell Science, (UK), 2006, Vol. 119, pp. 1494-1503; Fraldi, A. et al., "Lysosomal fusion and SNARE function are impaired by cholesterol accumulation in lysosomal storage disorders", The EMBO Journal, (UK), 2010, Vol. 29, pp. 3607-3620; and Pankiv, S. et al., "FYCO1 is a Rab7 effector that binds to LC3 and PI3P to mediate microtubule plus end-directed vesicle transport," Journal of Cell Biology, (USA), 2010, Vol. 188, pp. 253-269). The movement of lysosomes to the cell periphery and their positioning within the cell may be related to signal transduction (see R. & Bonifacino, JS, "Lysosome Positioning Influences mTORC2 and AKT Signaling," Molecular Cell, (USA), 2019, Vol. 75, pp. 26-38.e3).In this regard, it is contemplated that mTORC1, mTORC2, and AKT activation may be important for lysosome peripheral distribution (see Pous, C. & Codogno, P., “Lysosome positioning coordinates mTORC1 activity and autophagy,” Nature Cell Biology, (UK), 2011, Vol. 13, pp. 342-344; and Cabukusta, B. & Neefjes, J., “Mechanisms of lysosomal positioning and movement,” Traffic, (US), 2018, Vol. 19, pp. 761-769).

[0061] In certain embodiments, microautophagy and / or peumaric degradation of target substrates and / or membranes can involve movement of lysosomes to the periphery in the cytosol and to the target membrane (e.g., direct interaction with the target membrane) (see Pu, J., Guardia, C.M., Keren-Kaplan, T. & Bonifacino, J.S., "Mechanisms and functions of lysosome positioning," Journal of Cell Science, (UK), 2016, Vol. 129, pp. 4329-4339; and Katherine R Parzych, D.J.K., "An Overview of Autophagy: Morphology, Mechanism, and Regulation," Antioxidants & Redox Signaling, (US), 2014, Vol. 20, pp. 460-473). In certain embodiments, the movement of lysosomes (from the perinuclear region of the cell) to the cell periphery and their interaction with target membranes / substrates (e.g., glycogen, lipids, proteins) at the cell periphery may be associated with the activation of mTORC1 / mTORC2 (see Rabanal-Ruiz, Y. & Korolchuk, VI, "mTORC1 and Nutrient Homeostasis: The Central Role of the Lysosome", International Journal of Molecular Sciences, (Russia), 2018, Vol. 19, p. 818; Jia, R. & Bonifacino, JS, "Lysosome Positioning Influences mTORC2 and AKT Signaling", Molecular Cell, (USA), 2019, Vol. 75, pp. 26-38.e3).

[0062] Without wishing to be bound by theory, it is contemplated that Rab1a DN (dominant negative form of Rab1a) may stimulate the peripheral distribution of lysosomes (from the perinuclear region) by activating mTORC1 / mTORC2 proteins within the cell, without the need for external (or extracellular) signals for their activation (to support the peripheral distribution of lysosomes).

[0063] Rab1a in reducing obesity GDP or Rab1a GTP Use of: As described in detail herein, GDP-bound forms of Rab1a, e.g., Rab1a S25N , Rab1a N124I (mouse Rab1 sequence), Rab1a D41N , Rab1a D47N or another dominant-negative (DN) GDP-bound form of Rab1a, or such Rab1a GDP It has now been identified that treatment with a microautophagy-enhancing agent comprising one or more expressible nucleic acids encoding can be used to reduce obesity in animal models.

[0064] As will be appreciated, in certain embodiments, the methods described herein may be in vitro methods, in vivo methods, or both.

[0065] In certain embodiments, provided herein is a method for reducing obesity in a subject in need thereof, said method comprising: GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP treating a subject with one or more expressible nucleic acids encoding This results in Rab1a GDP increasing cellular levels of and resulting in a reduction in obesity in a subject.

[0066] In certain embodiments of the above methods, the obesity is associated with at least one of the following conditions: increased inflammatory response, increased abdominal obesity, high blood pressure, hyperglycemia, high serum triglycerides, low serum high-density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0067] In another embodiment, provided herein is a method for reducing weight in an overweight subject, said method comprising: GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP or a combination thereof, Thereby, Rab1a in overweight subjects GDP Increases cellular levels of α-glucan, resulting in weight loss.

[0068] In another embodiment of any of the above methods, Rab1a GDP Rab1a S25N , Rab1a N124I , Rab1a D41N , Rab1a D47N , or may be or include another dominant-negative (DN) GDP-bound form of Rab1a.

[0069] In yet another embodiment of any of the above methods, Rab1a GDP may comprise the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41N or MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0070] In yet another embodiment of any of the above methods, Rab1a GDP can consist of the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41N or MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0071] In another embodiment of any of the above methods, Rab1a GDP may comprise or consist of the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0072] In yet another embodiment of any of the above methods, Rab1a GDP may be in the form of a fusion protein, wherein Rab1a GDP is fused or otherwise linked, directly or indirectly, optionally via a linker, to a signal or targeting peptide, a fluorescent peptide, or other marker or tracer, or to another peptide or non-peptide moiety for targeted delivery, to facilitate cellular uptake, to increase stability or in vivo half-life, or to enhance Rab1a GDP or improve another therapeutic, diagnostic, or in vivo property of the fusion protein. In certain embodiments, which are not intended to be limiting in any way, the fusion protein may include any protein or tag for identification or delivery to a specific organ, such as GFP, YFP, mCherry, a luciferase-specific antibody, or an aptamer.

[0073] In yet another embodiment of any of the above methods, the fusion protein may comprise the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0074] In another embodiment of any of the above methods, Rab1a GDP may be in the form of a fusion protein and may comprise the following amino acid sequence: MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAHIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSMYTFVTSHLPPGFNEYDFVPESFDR DKTIALIMNSSGSTGLPKGVALPHRTACVRFSHARDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYKIQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYGLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLD TGKTLGVNQRGELCVRGPMIMSGYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILLQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVFVDEVPKGLTGKLDARKIREILIKAKKGGKSKLMSSMNPEYDYLFKLLL IGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNVKQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 21), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0075] In yet another embodiment of any of the above methods, the one or more expressible nucleic acids are Rab1a, Rab2a, Rab3a, Rab4a, Rab5a, Rab6a, Rab7a, Rab8a, Rab9a, Rab10a, Rab11a, Rab12a, Rab13a, Rab14a, Rab15a, Rab16a, Rab17a, Rab18a, Rab19a, Rab GDP It may encode any one or more of:

[0076] In yet another embodiment of any of the above methods, the one or more expressible nucleic acids may be DNA-based or RNA-based.

[0077] In another embodiment of any of the above methods, the one or more expressible nucleic acids express Rab1a in one or more cells of the subject. GDP or wherein one or more expressible nucleic acids are integrated into the genome of one or more cells and express Rab1a in one or more cells of a subject. GDP can be expressed.

[0078] In yet another embodiment of any of the above methods, the one or more expressible nucleic acids is Rab1a GDP The gene may include one or more expression vectors, plasmids, or mRNAs that encode and are capable of expressing in one or more cells of the subject.

[0079] In yet another embodiment of any of the above methods, the one or more expressible nucleic acids may comprise the following nucleic acid sequence: ATGTCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGACTCAGGGGTTGGAAAGAATTGCCTTCTTCTTAGGTTTGCAGATGATACATATACAGAAAGCTACATCAGCACAATTGGTGTGGATTTCAAAATAAGAA CTATAGAGTTAGACGGGAAAACAATCAAGCTTCAAATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACCTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAGGAGTCCTTCAATAATGTTAAA CAGTGGCTGCAGGAAATAGATCGTTATGCCAGTGAAAATGTCAACAAATTGTTGGTAGGGAACAAATGTGATCTGACCACAAAGAAAGTAGTAGACTACACAACAGCGAAGGAATTTGCTGATTCCCTTGGAATTCCGTTTTTGGAAACCAGTG CTAAGAATGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAGCGAATGGGTCCCGGAGCAACAGCTGGTGGTGCTGAGAAGTCCAATGTTAAAATTCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGTTGCTGCTAA (Human Rab1a S25N ORF codon sequence, SEQ ID NO: 5), ATGGGGGACTACAAGGACGACGATGACAAGGGGGGTAGCGGTGGATCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGATTCTGGGGTTGGAAAGTCCTGCCTTCTCCTTAGGTTTGCAGATGATACGTATACGGAAAGCTACATC AGCACAATTGGTGTGGATTTCAAGATACGAACTATAGAGTTAGATGGGAAAACAATCAAGCTACAGATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACTTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAG GAGTCCTTCAATAACGTTAAACAGTGGCTGCAGGAGATAGATCGCTACGCCAGTGAAAATGTCAACAAGTTGTTGGTAGGGATCAAATGTGACCTGACCACAAAGAAAGTAGTAGACTACACAACAGCAAAGGAATTTGCAGATTCCCTTGGAATTCCATTTTTG GAAACCAGTGCTAAGAACGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAAGCGAATGGGTCCTGGAGCTACAGCTGGTGGTGCCGAGAAGTCCAATGTTAAAATCCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGCTGCTGCTAA (Mouse Rab1a N124I ORF codon sequence, SEQ ID NO: 8), or ATGTCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGACTCAGGGGTTGGAAAGTCTTGCCTTCTTCTTAGGTTTGCAGATGATACATATACAGAAAGCTACATCAGCACAATTGGTGTGAACTTCAAAATAAGAA CTATAGAGTTAGACGGGAAAACAATCAAGCTTCAAATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACCTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAGGAGTCCTTCAATAATGTTAAA CAGTGGCTGCAGGAAATAGATCGTTATGCCAGTGAAAATGTCAACAAATTGTTGGTAGGGAACAAATGTGATCTGACCACAAAGAAAGTAGTAGACTACACAACAGCGAAGGAATTTGCTGATTCCCTTGGAATTCCGTTTTTGGAAACCAGTG CTAAGAATGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAGCGAATGGGTCCCGGAGCAACAGCTGGTGGTGCTGAGAAGTCCAATGTTAAAATTCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGTTGCTGCTAA (Human Rab1a D47N ORF codon sequence, SEQ ID NO: 17), or a Rab1a having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which binds preferentially to GDP. GDP a nucleic acid sequence encoding Alternatively, a nucleic acid sequence equivalent to any of the above sequences due to codon redundancy.

[0080] In another embodiment of any of the above methods, the one or more expressible nucleic acids may comprise the following nucleic acid sequence: (SEQ ID NO: 19, DNA sequence of MG-008 ORF with 5' luciferase tag), or (SEQ ID NO: 20, mRNA sequence of MG-008 ORF with 5' luciferase tag), or a Rab1a having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which binds preferentially to GDP. GDP a nucleic acid sequence encoding Alternatively, a nucleic acid sequence equivalent to any of the above sequences due to codon redundancy.

[0081] In another embodiment, a GDP-bound form of Rab1a (Rab1a) is administered to a subject in need thereof for reducing obesity in the subject or for preventing or treating obesity. GDP ), Rab1a GDP Provided herein are uses of one or more expressible nucleic acids encoding the

[0082] In another embodiment, a method for the treatment of at least one of the following conditions in a subject in need thereof, comprising administering to a subject a GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP Provided herein are uses of one or more expressible nucleic acids encoding, or combinations thereof, for treating: inflammatory response, increased abdominal obesity, high blood pressure, hyperglycemia, high serum triglycerides, low serum high density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0083] In another embodiment, provided herein is the use of a GDP-bound form of Rab1a (Rab1aGDP), one or more expressible nucleic acids encoding Rab1aGDP, or a combination thereof, for reducing weight in an overweight subject.

[0084] In another embodiment, use is made of a GDP-bound form of Rab1a (Rab1a) in the manufacture of a medicament for reducing, or preventing or treating obesity in a subject in need thereof. GDP ), Rab1a GDP Provided herein are uses of one or more expressible nucleic acids encoding the

[0085] In another embodiment, a GDP-bound form of Rab1a (Rab1a) is used in the manufacture of a medicament for reducing, preventing, or treating at least one of the following conditions in a subject in need thereof: GDP ), Rab1a GDP Provided herein are uses of one or more expressible nucleic acids encoding, or combinations thereof, for treating: inflammatory response, increased abdominal obesity, high blood pressure, hyperglycemia, high serum triglycerides, low serum high density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0086] In another embodiment, the use of a GDP-bound form of Rab1a (Rab1a) in the manufacture of a medicament for reducing weight or preventing or treating excess weight in an overweight subject. GDP ), Rab1a GDP Provided herein are uses of one or more expressible nucleic acids encoding the

[0087] In another embodiment of any of the above uses, Rab1a GDP Rab1a S25N , Rab1a N124I , Rab1a D41N , Rab1a D47N , or may be or include another dominant-negative (DN) GDP-bound form of Rab1a.

[0088] In yet another embodiment of any of the above uses, Rab1a GDP may comprise the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41N or MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0089] In yet another embodiment of any of the above uses, Rab1a GDP can consist of the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41N or MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0090] In another embodiment of any of the above uses, Rab1a GDP may comprise or consist of the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0091] In yet another embodiment of any of the above uses, Rab1a GDPmay be in the form of a fusion protein, wherein Rab1a GDP is fused or otherwise linked, directly or indirectly, optionally via a linker, to a signal or targeting peptide, a fluorescent peptide, or other marker or tracer, or to another peptide or non-peptide moiety for targeted delivery, to facilitate cellular uptake, to increase stability or in vivo half-life, or to enhance Rab1a GDP improve another therapeutic, diagnostic, or in vivo property of

[0092] In yet another embodiment of any of the above uses, the fusion protein may comprise the following amino acid sequence: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0093] In another embodiment of any of the above uses, Rab1a GDP may be in the form of a fusion protein and may comprise the following amino acid sequence: MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAHIEVNITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPANDIYNERELLNSMNISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSMYTFVTSHLPPGFNEYDFVPESFDR DKTIALIMNSSGSTGLPKGVALPHRTACVRFSHARDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYKIQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYGLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLD TGKTLGVNQRGELCVRGPMIMSGYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILLQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVFVDEVPKGLTGKLDARKIREILIKAKKGGKSKLMSSMNPEYDYLFKLLL IGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNVKQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 21), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which preferentially binds GDP.

[0094] In yet another embodiment of any of the above uses, the one or more expressible nucleic acids are one or more Rab1a as defined herein. GDP may be coded.

[0095] In yet another embodiment of any of the above uses, the one or more expressible nucleic acids may be DNA-based or RNA-based.

[0096] In another embodiment of any of the above uses, the one or more expressible nucleic acids express Rab1a in the one or more cells. GDP or wherein one or more expressible nucleic acids are integrated into the genome of one or more cells and express Rab1a in one or more cells. GDP can be expressed.

[0097] In another embodiment of any of the above uses, the one or more expressible nucleic acids is Rab1a GDP The gene may comprise one or more expression vectors, plasmids, or mRNAs encoding and capable of expressing the gene in one or more cells.

[0098] In another embodiment of any of the above uses, the one or more expressible nucleic acids may comprise the following nucleic acid sequence: ATGTCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGACTCAGGGGTTGGAAAGAATTGCCTTCTTCTTAGGTTTGCAGATGATACATATACAGAAAGCTACATCAGCACAATTGGTGTGGATTTCAAAATAAGAA CTATAGAGTTAGACGGGAAAACAATCAAGCTTCAAATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACCTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAGGAGTCCTTCAATAATGTTAAA CAGTGGCTGCAGGAAATAGATCGTTATGCCAGTGAAAATGTCAACAAATTGTTGGTAGGGAACAAATGTGATCTGACCACAAAGAAAGTAGTAGACTACACAACAGCGAAGGAATTTGCTGATTCCCTTGGAATTCCGTTTTTGGAAACCAGTG CTAAGAATGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAGCGAATGGGTCCCGGAGCAACAGCTGGTGGTGCTGAGAAGTCCAATGTTAAAATTCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGTTGCTGCTAA (Human Rab1a S25N ORF codon sequence, SEQ ID NO: 5), ATGGGGGACTACAAGGACGACGATGACAAGGGGGGTAGCGGTGGATCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGATTCTGGGGTTGGAAAGTCCTGCCTTCTCCTTAGGTTTGCAGATGATACGTATACGGAAAGCTACATC AGCACAATTGGTGTGGATTTCAAGATACGAACTATAGAGTTAGATGGGAAAACAATCAAGCTACAGATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACTTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAG GAGTCCTTCAATAACGTTAAACAGTGGCTGCAGGAGATAGATCGCTACGCCAGTGAAAATGTCAACAAGTTGTTGGTAGGGATCAAATGTGACCTGACCACAAAGAAAGTAGTAGACTACACAACAGCAAAGGAATTTGCAGATTCCCTTGGAATTCCATTTTTG GAAACCAGTGCTAAGAACGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAAGCGAATGGGTCCTGGAGCTACAGCTGGTGGTGCCGAGAAGTCCAATGTTAAAATCCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGCTGCTGCTAA (Mouse Rab1a N124I ORF codon sequence, SEQ ID NO: 8), or ATGTCCAGCATGAATCCCGAATATGATTATTTATTCAAGTTACTTCTGATTGGCGACTCAGGGGTTGGAAAGTCTTGCCTTCTTCTTAGGTTTGCAGATGATACATATACAGAAAGCTACATCAGCACAATTGGTGTGAACTTCAAAATAAGAA CTATAGAGTTAGACGGGAAAACAATCAAGCTTCAAATATGGGACACAGCAGGCCAGGAAAGATTTCGAACAATCACCTCCAGTTATTACAGAGGAGCCCATGGCATCATAGTTGTGTATGATGTGACAGATCAGGAGTCCTTCAATAATGTTAAA CAGTGGCTGCAGGAAATAGATCGTTATGCCAGTGAAAATGTCAACAAATTGTTGGTAGGGAACAAATGTGATCTGACCACAAAGAAAGTAGTAGACTACACAACAGCGAAGGAATTTGCTGATTCCCTTGGAATTCCGTTTTTGGAAACCAGTG CTAAGAATGCAACGAATGTAGAACAGTCTTTCATGACGATGGCAGCTGAGATTAAAAAGCGAATGGGTCCCGGAGCAACAGCTGGTGGTGCTGAGAAGTCCAATGTTAAAATTCAGAGCACTCCAGTCAAGCAGTCAGGTGGAGGTTGCTGCTAA (Human Rab1a D47N ORF codon sequence, SEQ ID NO: 17), or a Rab1a having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which binds preferentially to GDP. GDP a nucleic acid sequence encoding Alternatively, a nucleic acid sequence equivalent to any of the above sequences due to codon redundancy.

[0099] In yet another embodiment of any of the above uses, the one or more expressible nucleic acids may comprise the following nucleic acid sequence: (SEQ ID NO: 19, DNA sequence of MG-008 ORF with 5' luciferase tag), or (SEQ ID NO: 20, mRNA sequence of MG-008 ORF with 5' luciferase tag), or a Rab1a having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity thereto and which binds preferentially to GDP. GDP a nucleic acid sequence encoding Alternatively, a nucleic acid sequence equivalent to any of the above sequences due to codon redundancy.

[0100] In another embodiment, provided herein is a polypeptide comprising the following amino acid sequence for use in reducing, preventing, or treating obesity in a subject in need thereof, or for use in reducing weight in an overweight subject: MSSMNPEYDYLFKLLLIGDSGVGKNCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 6; human Rab1a S25N ), MGDYKDDDDKGGSGGSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVDFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTD QESFNNVKQWLQEIDRYASENVNKLLVGIKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 9; mouse Rab1 N124I ), Human Rab1a D41Nor MSSMNPEYDYLFKLLLIGDSGVGKSCLLLRFADDTYTESYISTIGVNFKIRTIELDGKTIKLQIWDTAGQERFRTITSSYYRGAHGIIVVYDVTDQESFNNV KQWLQEIDRYASENVNKLLVGNKCDLTTKKVVDYTTAKEFADSLGIPFLETSAKNATNVEQSFMTMAAEIKKRMGPGATAGGAEKSNVKIQSTPVKQSGGGCC (SEQ ID NO: 18; human Rab1a D47N ), Alternatively, a polypeptide having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences and which preferentially binds GDP.

[0101] In another embodiment of the polypeptide for the above uses, the obesity is accompanied by at least one of the following conditions: inflammatory response, increased abdominal obesity, high blood pressure, hyperglycemia, high serum triglycerides, low serum high density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0102] In another embodiment, provided herein is a pharmaceutical composition comprising: GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP or a combination thereof, and Another anti-obesity drug.

[0103] In yet another embodiment, provided herein is a kit comprising any one or more of the following: GDP-bound form of Rab1a (Rab1a GDP ), Rab1aGDP one or more expressible nucleic acids encoding anti-obesity drugs, instructions for carrying out any of the methods described herein; or Any combination of them.

[0104] As will be appreciated, in certain embodiments of any of the methods, uses, or polypeptides for use described herein, obesity may be accompanied by an increased inflammatory response, increased abdominal obesity, high blood pressure, hyperglycemia, high serum triglycerides, low serum high density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, non-alcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, and sleep apnea.

[0105] As will be understood, lysosome-mediated microautophagy can refer to a cellular process in which cellular lipid, protein, or glycogen substrates, or portions thereof, are engulfed and degraded by lysosomes. Specifically in this regard, lysosome-mediated microautophagy is an essential component of cellular bioenergetic (i.e., ATP generation) and biosynthetic processes (i.e., synthesizing new biomass by recycling existing "old" material).

[0106] It will also be understood that increasing lysosomal-mediated microautophagy can refer to an increase in the rate, extent, capacity, or effectiveness of the lysosomal-mediated microautophagy process in a cell compared to the baseline level of the cell, or compared to corresponding treated or untreated control cells, or compared to levels in reference diseased cells or reference cells having accumulation of protein, lipid, or glycogen substrates.

[0107] Furthermore, it will be understood that increasing lysosome-mediated microautophagy can include restoring or increasing cellular lysosomal motility or bidirectional lysosomal motility, and / or enhancing, increasing, activating, or otherwise restoring or rescuing cellular lysosomal motility activity. Such restoration or rescuing can result in increased autophagy (e.g., micro- and / or macroautophagy) and / or lysosomal degradation capacity. As detailed herein, there are several diseases, conditions, and cellular states in which cellular lysosomal motility can be impaired, reduced, blocked, or inhibited. Because lysosome-mediated microautophagy can involve lysosomal motility and kiss-and-run events, microautophagy-enhancing agents can restore or increase cellular lysosomal motility. Lysosome motility can play an important role in several cellular functions, including lysosome-mediated microautophagy, lysosome-mediated macroautophagy, lysosome regeneration, and lysosome maturation process.Restoring lysosome motility and / or lysosome bidirectional motility can refer to adjusting the lysosome motility / bidirectional motility level of cells to return to that of corresponding normal cells or healthy control cells with baseline level of lysosome motility / bidirectional motility.In certain embodiments, this adjustment can also adjust the degradation ability (i.e., phagy) level to return to that of normal or healthy cells.

[0108] It will be understood that a lysosome binding-dissociation event between a lysosome and a lipid, protein, or glycogen substrate can refer to an event in which the lysosome binds to a lipid, protein, or glycogen substrate (i.e., for example, a lipid droplet or protein aggregate), captures at least a portion of the lipid, protein, or glycogen substrate, and then dissociates from the lipid, protein, or glycogen substrate. A lysosome binding-dissociation (i.e., "on" and "off") event between a lysosome and a lipid, protein, or glycogen substrate can be considered a "kiss-and-run" type event. As part of the binding (or "kissing") event, at least a small piece of the substrate (i.e., lipid) can be "caught" or engulfed by the lysosome from the substrate (i.e., for example, a cytosolic lipid droplet or CLD). In the case of a lipid droplet substrate, this can be achieved by the formation of a fusion pore between the lysosome and the CLD. As part of the dissociation (or "run") event, dissociation of the lysosome from the substrate (i.e., CLD) can occur. An increase in lysosome binding-dissociation events may refer to an increase in the rate, extent, or effectiveness of lysosome binding-dissociation events in a cell compared to the baseline levels of corresponding treated or untreated control cells, e.g., identical cells under identical conditions but without a microautophagy-modulating agent, or treated with a compound or composition known not to affect the process.

[0109] It will be appreciated that in certain embodiments, microautophagy-enhancing agents may be used to correct a cellular microautophagy deficiency or a cellular condition in which microautophagy is reduced.

[0110] Those skilled in the art will understand, in conjunction with the teachings herein, that a microautophagy-enhancing agent can be any suitable agent that increases or promotes the rate, activity, extent, or effectiveness of lysosome-mediated microautophagy in a cell, or that increases lysosomal motility or bidirectional motility. In certain embodiments, a suitable microautophagy-enhancing agent is a GDP-bound form of Rab1a (Rab1a GDP ), Rab1a GDP The nucleic acid may be or may comprise one or more expressible nucleic acids encoding the

[0111] The Ras-related protein Rab-1A (i.e., Rab1a) is a protein encoded by the RAB1A gene in humans. It can control vesicle trafficking from the endoplasmic reticulum (ER) to the Golgi compartment and then to the cell surface, and may play an important role in IL-8 and growth hormone secretion. In addition, when it is in its GTP-bound form, it may play a role in the assembly of autophagosomes in macroautophagy and in the cellular defense response against pathogenic bacteria. It can also control the motility of endocytic compartments.

[0112] As described in detail herein, lysosome-mediated microautophagy of target protein, lipid, or glycogen substrates in cells involves the lysis of GDP-bound forms of Rab1a, e.g., Rab1a S25N , Rab1a N124I , Rab1a D41N , Rab1a D47N It has now been identified that microautophagy can be increased by treatment with microautophagy enhancers, including Rab1a, or another dominant-negative (DN) GDP-bound form of Rab1a.

[0113] It will be understood that the specific amino acid or nucleic acid sequence of a particular gene may vary between species. For example, a human Rab1a amino acid sequence may have a homolog in another species that has a sequence variation from the human sequence. In some embodiments, although homologous sequences may vary between species, the general effect (e.g., phenotypic effect) of the homologous sequence may be substantially similar to the effect of the wild-type sequence in a given cell or subject.

[0114] In a specific embodiment, the microautophagy enhancer is a GDP-bound form of Rab1a (Rab1a GDP ), e.g., Rab1a S25N , Rab1a N124I , Rab1a D41N , Rab1a D47N The Rab1a GDP-bound form may be or may comprise a Rab1a GDP-bound form, such as Rab1a WT or a functional equivalent thereof, or another dominant-negative (DN) GDP-bound form of Rab1a. The sequences of human and / or mouse Rab1aWT and their specific GDP-bound forms are shown in Figure 2. Suitable GDP-bound forms of Rab1a may include any suitable Rab1a variant that is "dominant-negative" or that preferentially binds GDP over GTP. Such Rab1a GDPVariants can be identified using techniques known in the art (e.g., Chan, C.-C. et al., "Systematic Discovery of Rab GTPases with Synaptic Functions in Drosophila", Current Biology, (UK), 2011, Vol. 21, pp. 1704-1715; Tabancay, AP et al., "Identification of dominant negative mutants of Rheb GTPase and their use to implicate the involvement of human Rheb in the activation of p70S6K", Journal of Biological Chemistry, (US), 2003, Vol. 278, pp. 39921-39930; and Dumas, JJ, Zhu, Z., Connolly, JL & Lambright, DG, "Structural basis of activation and GTP hydrolysis in Rab proteins", Structure, (US), 1999, Vol. 7, pp. 413-s2, each of which is incorporated herein by reference in its entirety).

[0115] 2A-2U show specific sequences of the nucleic acids and amino acids / proteins described herein. In FIG. 2A-2U, SEQ ID NOs: 1-3 correspond to human Rab1a, respectively. WT The DNA gene sequence, ORF codon sequence, and amino acid sequence are provided. SEQ ID NOs: 4 to 6 are human Rab1a S25N The DNA gene sequence, ORF codon sequence, and amino acid sequence are provided. SEQ ID NOS: 7 to 9 are the sequences of mouse Rab1, N124I The DNA gene sequence, ORF codon sequence, and amino acid sequence are provided. SEQ ID NOs: 10 to 12 are human Rab1a Q70LThe DNA gene sequence, ORF codon sequence, and amino acid sequence are provided. SEQ ID NOs: 13 to 15 are human Rab1a Q63L The DNA gene sequence, ORF codon sequence, and amino acid sequence are provided. SEQ ID NOs: 16 to 18 are human Rab1a D47N The DNA gene sequence, ORF codon sequence, and amino acid sequence are provided. SEQ ID NOs: 19-21 provide the DNA sequence of the MG-008 ORF with a 5' luciferase tag, the mRNA sequence of the MG-008 ORF with a 5' luciferase tag, and the MG-008 protein sequence with an N-terminal luciferase tag, respectively (see Example 1 for further discussion of SEQ ID NOs: 19-21). Human Rab1a D41N The array of Internet<URL:https: / / www.addgene.org / 49581 / > (which is incorporated herein by reference in its entirety) and are commercially available. In certain embodiments, nucleic acids or amino acids comprising any of these sequences are provided herein. In certain embodiments, nucleic acids or amino acids that share at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity to any of these sequences (i.e., any of SEQ ID NOS: 1-21 or Rab1a). D41N Provided herein are nucleic acids or amino acids comprising a nucleic acid or amino acid sequence having a sequence similar to that of the amino acid sequence of the present invention (with respect to the sequence of the amino acid sequence of the present invention), or active fragments thereof.

[0116] Rab1a S25N , Rab1a D41N , Rab1a D47N , and Rab1a N124I Suitable functional equivalents of Rab1a, for example, have at least 80% (or >85%, or >90%, or >95%, or >99%) sequence identity with Rab1a. WT , or Rab1a S25N , or Rab1a N124I , or Rab1a D41N , or Rab1a D47Nand preferentially binds GDP over GTP, while interacting with Rab1a as described in detail herein. S25N , or Rab1a N124I , or Rab1a D41N , or Rab1a D47N In a further embodiment, the microautophagy enhancer may comprise a suitable Rab1a variant or mutant that also retains the relevant cellular / biochemical function of Rab1a. GDP one or more expressible nucleic acids encoding, for example, a GDP-bound form of Rab1a (Rab1a GDP ), e.g., Rab1a S25N , or Rab1a N124I , or Rab1a D41N , or Rab1a D47N It will be understood that the nucleic acid may be or comprise any suitable nucleic acid / expression vector (i.e., e.g., vector, cassette, mRNA, modified mRNA, plasmid), etc. that encodes / expresses the nucleic acid sequence of the present invention, or a functional equivalent thereof.

[0117] As will be understood, sequences are described herein primarily with reference to human and / or mouse homologs. It will be understood that functional equivalents and / or variants may be found in a variety of different species, for example, in different mammals. As used herein, Rab1a GDP (DN) or Rab1a GTP References to specific sequence modifications and / or variants that allow for (DA) forms are often provided for convenience with reference to human and / or mouse homologues / sequences, providing the location and modification / mutation information (e.g., S25N, D41N, D47N, N124I; Q70L, Q67L, Q63L). However, it will be understood that equivalent DN and / or DA forms can be implemented in homologous sequences from other species and / or other sequences related to the human and / or mouse sequences provided herein, although the location and / or nature of the modification / mutation may vary somewhat depending on the specific sequence of interest. For example, Rab1N124I is a modification / mutation of the mouse sequence. Another example is Rab1 Q67L is a modification / mutation in the mouse sequence; human Rab1a does not have a Q at position 67. Rather, the human sequence has a Q at position 63, so the modification / mutation with reference to the human sequence is Rab1a. Q63L is.

[0118] Those skilled in the art will appreciate that microautophagy reducers may bind to the GTP-bound form of Rab1a (Rab1a GTP ), Rab1a GTP one or more expressible nucleic acids encoding Rab1a wild type (Rab1a WT ), or Rab1a WT It will be understood that the nucleic acid sequence may be or may comprise any one or more of one or more expressible nucleic acids encoding Rab1a. GTP Rab1a Q70L , Rab1a Q67L (mouse sequence), Rab1a Q63L (human Rab1a sequence), or a functional equivalent thereof, or another dominantly active (DA) GTP-bound form of Rab1a. The GTP-bound form of Rab1a can include any Rab1a variant that is "dominantly active" or that preferentially binds GTP over GDP. Such Rab1a GTPVariants can be identified by techniques known in the art (e.g., Chan, C.-C. et al., "Systematic Discovery of Rab GTPases with Synaptic Functions in Drosophila", Current Biology, (UK), 2011, Vol. 21, pp. 1704-1715; Tabancay, AP et al., "Identification of dominant negative mutants of Rheb GTPase and their use to implicate the involvement of human Rheb in the activation of p70S6K", Journal of Biological Chemistry, (US), 2003, Vol. 278, pp. 39921-39930; and Dumas, JJ, Zhu, Z., Connolly, JL & Lambright, DG, "Structural basis of activation and GTP hydrolysis in Rab proteins", Structure, (US), 1999, Vol. 7, pp. 413-s2, each of which is incorporated herein by reference in its entirety), e.g., Rab1a Q70L , and / or Rab1a Q67L , and / or Rab1a Q63L can be identified using the same used to identify

[0119] In certain embodiments, Rab1a GDP Treatment with microautophagy enhancers such as Rab1a GDP Introducing proteins into cells, Rab1a GDP It will be understood that the expression of a gene may include expressing a gene that is a nucleotide sequence, a nucleotide sequence, or both.

[0120] It will be understood that expression of a specific protein in a cell can refer to the production of a polypeptide from a nucleic acid sequence encoding the polypeptide. Gene expression can encompass both transcription and translation processes, and therefore gene expression can refer to the production of a nucleic acid sequence such as mRNA (i.e., transcription), the production of a protein (i.e., translation), or both. Furthermore, it will be understood that overexpression of a specific gene in a cell can refer to increasing the expression of the specific gene in the cell compared to wild-type, baseline, or untreated levels. Introduction or overexpression of a mutant gene into a cell can be achieved using any of several methods known in the art. For example, a vector (whether viral, plasmid, or other) containing one or more copies of a specific gene, each driven by a suitable promoter sequence (e.g., a constitutive or inducible promoter), or mRNA or a chemically modified version thereof, can be introduced into a cell by transfection, electroporation, viral infection, or another suitable method known in the art. Suitable expression vector technologies for introducing or overexpressing specific genes into cells are known in the art (see, for example, "Molecular Cloning: A Laboratory Manual," Cold Spring Harbor Laboratory Press, 2012, 4th ed.). In conjunction with the teachings herein, those skilled in the art will be able to easily identify Rab1a vectors that can be prepared for introduction into cells to provide expression of a protein of interest (e.g., either transiently or long-term via integration into the genome). GDP One will know a variety of expressible nucleic acids encoding specific proteins such as:

[0121] It will be understood that compounds and / or compositions comprising or consisting of one or more of the nucleic acids and / or proteins described herein can be used.The composition can additionally comprise one or more pharmaceutically acceptable diluents, carriers, excipients or buffers.The composition can be used to administer one or more nucleic acids and / or proteins to cells in vitro or in vivo.

[0122] In the context of inserting a nucleic acid sequence into a cell, the introduction of a gene may refer to "transfection," "transformation," or "transduction," and may encompass the incorporation or introduction of a nucleic acid sequence into a eukaryotic cell, where the nucleic acid sequence may optionally be integrated into the genome of the cell or may be transiently expressed (e.g., transfected mRNA). A protein or enzyme may be introduced into a cell by delivering the protein or enzyme itself into the cell, or by expressing an mRNA encoding the protein or enzyme within the cell, leading to its translation.

[0123] As will be known to those skilled in the art, expressible nucleic acids for expressing specific genes may encode or include features described in Lewin B., "Genes VII", Oxford University Press, (UK), 2000, or Sambrook et al., "Molecular Cloning: A Laboratory Manual", Cold Spring Harbor Laboratory Press, (US), 2001, 3rd Edition. The nucleotide sequence encoding the polypeptide or protein may be incorporated into a suitable vector, such as a commercially available vector. Vectors may also be individually constructed or modified using standard molecular biology techniques, for example, as outlined in Sambrook et al. (Cold Spring Harbor Laboratory Press, 2001, 3rd Edition). Those skilled in the art will recognize that a vector may include a nucleotide sequence encoding desired elements that can be operably linked to a nucleotide sequence encoding a polypeptide or protein. Such nucleotide sequences encoding desired elements may include a transcription promoter, a transcription enhancer, a transcription terminator, a translation initiation factor, a translation terminator, a ribosome binding site, a 5' untranslated region, a 3' untranslated region, a cap structure, a polyA tail, and / or an origin of replication. Selection of an appropriate vector can depend on several factors, including, without limitation, the size of the nucleic acid to be incorporated into the vector, the type of transcriptional and translational control elements desired, the desired expression level, the desired copy number, whether chromosomal integration is desired, the type of selection process desired, or the host cell or host range intended to be transformed.

[0124] Those skilled in the art will understand that the biomolecules and / or compounds described herein can be provided as pharmaceutical compositions, together with a pharmaceutically acceptable diluent, carrier, or excipient, and / or together with one or more separate active agents or drugs, as part of a combination or pharmaceutical composition. In certain embodiments, the biomolecules, compounds, and / or pharmaceutical compositions can be administered simultaneously, sequentially, or in combination with other drugs or pharmaceutical compositions in a treatment regimen, either separately or as a combination or pharmaceutical composition.

[0125] The biomolecules, compounds, and / or compositions described herein may include one or more pharmaceutically acceptable excipients, diluents, and / or carriers. Pharmaceutically acceptable carriers, diluents, or excipients may include any suitable carriers, diluents, or excipients known to those skilled in the art. Examples of pharmaceutically acceptable excipients may include, but are not limited to, cellulose derivatives, sucrose, and starch. Those skilled in the art will recognize that pharmaceutically acceptable excipients may include suitable fillers, binders, lubricants, buffers, flow agents, and disentanglers known in the art (see, for example, "Remington: The Science and Practice of Pharmacy," Lippincott Williams & Wilkins, (USA), 2006). Examples of pharmaceutically acceptable carriers, diluents, and excipients can be found, for example, in "Remington's Pharmaceutical Sciences", Mack Publishing Co., (USA), 2000, 20th Edition, and the United States Pharmacopeia: National Formulary (USP 24 NF19), 1999.

[0126] It will also be understood that one or more conservative amino acid substitutions may be possible. As will be recognized, conservative amino acid substitutions may include replacing an amino acid with another amino acid having similar properties such that protein folding, activity, or other functionality is not significantly affected. Examples of aromatic amino acids that may be substituted include phenylalanine, tryptophan, and tyrosine. Examples of interchangeable hydrophobic amino acids that may be substituted include leucine, isoleucine, methionine, and valine. Examples of interchangeable polar amino acids that may be substituted include glutamine and asparagine. Examples of interchangeable basic amino acids that may be substituted include arginine, lysine, and histidine. Examples of interchangeable acidic amino acids that may be substituted include aspartic acid and glutamic acid. Finally, examples of interchangeable small amino acids that may be substituted include alanine, serine, threonine, cysteine, and glycine.

[0127] As described in detail herein, dominant-negative (DN) Rab1a (e.g., GDP-bound Rab1a, Rab1a GDP ) can be used to increase microautophagy, which can allow for the degradation of cellular targets of interest, such as disease-related proteins, glycogen, or lipids. As will be appreciated, any suitable Rab1a DN or Rab1a GDP may be used. In light of the teachings herein, a variety of Rab1a DN proteins will be known to those of skill in the art. In certain embodiments, generally any suitable dominant-negative form of Rab1a (i.e., Rab1a fixed / locked in its GDP form) may be used, for example, to promote lysosome movement to the cytosol and periphery of the cell (from the perinuclear region of the cell) and stimulate direct lysosomal interaction with target substrates.

[0128] In certain embodiments, it is contemplated that expression of Rab1a DN (GDP form) can stimulate activation of mTORC1 / mTORC2 and AKT and promote lysosome movement to the periphery and target substrates based on their effects on lysosomal positioning (see also Jia, R. & Bonifacino, JS, "Lysosome Positioning Influences mTORC2 and AKT Signaling", Molecular Cell, (US), 2019, Vol. 75, pp. 26-38.e3).

[0129] It is contemplated herein that different genetic modifications of Rab1a that lock this protein in its constant GDP-bound state can be used to promote direct lysosomal interaction and piecemeal engulfment of different target substrates (e.g., one or more protein, lipid, and / or glycogen targets). The constant GDP-bound form (DN) of Rab1a is not generally available under normal physiological conditions, in which the native protein Rab1a constantly shifts between its GTP and GDP forms. In certain embodiments, it is contemplated that genetic mutations and / or amino acid substitutions / modifications can be used to lock this GTPase in its GDP-bound form (or constant GTP form) and prevent it from going to its GTP state (or GDP state). In certain embodiments, it is contemplated that any suitable modification that can substantially preserve the integrity of the GDP form / state of the GTPase (which can exert effects on signal transduction and on lysosomal movement to the periphery / cytosol) can promote microautophagy and degradation of target substrates by engulfment of lysosomal piecemeal.

[0130] In certain embodiments, Rab1a GDPcan generally be administered to a particular cell type or subject in need thereof in any suitable manner, which can be selected to suit the particular cell type, subject, and / or indication. In certain embodiments, in connection with the teachings herein, Rab1a GDP Nucleic acid sequences encoding and capable of expressing the protein can be administered to a subject or introduced into a cell type by any suitable transfection or nucleic acid delivery approach that would be known to one of skill in the art. In certain embodiments, delivery can be based on DNA or RNA transfection (e.g., using common transfection reagents such as lipofectamine (Invitrogen), FuGENE (Rosche), using DNA adenovirus (gene therapy), or using modified RNA (i.e., stabilized RNA) delivery to the body (e.g., using viruses, or microvesicles, exosomes, or ectosomes). In certain embodiments, the protein can be administered or delivered to cells in need thereof, optionally assisted by any suitable technique or delivery vehicle for facilitating protein delivery to cells.

[0131] In one embodiment of the present invention, F11 can be administered to an obese or overweight subject to reduce weight in the subject. F11 is a lipid nanoparticle encapsulating Rab1, commercially available from Precision NanoSystems (#50 655 W Kent Ave N, Vancouver, BC V6P 6T7). F11 can be administered intravenously, orally, subcutaneously, intramuscularly, sublingually, or by any other route known to those skilled in the art. F11 can be administered at 0.001 to 1 mg / kg (mpk), although other doses may be beneficial depending on the subject. F11 treatment can be administered as a single dose or multiple doses. When F11 is administered repeatedly, individual treatments can be separated by hours, days, weeks, months, or years as may be necessary to establish, maintain, or re-establish weight loss. Administration of F11 can reduce a subject's weight and / or fat content and / or positively affect HbA1c, alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), creatinine (CRE), or other blood chemistry measurements. F11 treatment can also improve other physical health indicators, such as, but not limited to, high blood pressure, high blood sugar, high cholesterol levels, inflammation, high serum triglycerides, high resting heart rate, low serum high-density lipoprotein (HDL), insulin resistance, glucose intolerance, diabetes, hypertension, dyslipidemia, nonalcoholic fatty liver disease (NAFLD), heart failure, atrial fibrillation, musculoskeletal disorders, sleep apnea, and / or high cancer progression markers.

[0132] In certain embodiments of the present invention, Rab1 encapsulated in lipid nanoparticles can be in the form of a fusion protein, where Rab1 is fused or otherwise linked, directly or indirectly, optionally via a linker, to a signal or targeting peptide, a fluorescent peptide, or other marker or tracer, or another peptide or non-peptide moiety for targeted delivery, to facilitate cellular uptake, increase stability or in vivo half-life, or improve another therapeutic, diagnostic, or in vivo property of Rab1. [Example]

[0133] Example 1: Rab1a DN (GDP form) reduces body weight in obese mice A diet-induced obesity (DIO) mouse model was established in 6-week-old female C57BL / 6 mice (n = 10) by feeding a high-fat diet for 10 months. Mice were divided into two groups: one group (n = 5) lacking Rab1a DN In this example, Rab1a DN is also referred to as MG-008. It is encoded by mRNA and encapsulated in the ANM formulation. The control group (n = 5) was injected with PBS. Mice continued on the HFD for an additional 18 days, and food intake and body weight were recorded every 2–3 days. The weight loss of mice on day 12 after MG008 injection (single dose) is shown on the left in Figure 1 (red bar). The weight of control mice on day 12 is shown on the right (black bar). Weight loss in MG008-treated mice was evident throughout the entire 18 days of treatment, with the earliest significant weight loss occurring as early as day 4 after MG008 injection (data not shown).

[0134] Example 2: Encapsulated Rab1 lipid nanoparticles (F11) reduce body weight in obese mice Approximately 6-week-old male C57BL / 6 mice were fed a high-fat diet (HFD) until they reached a body weight of at least 40 g (40–50 g). Mice were weighed at the start of the HFD and their weights were monitored weekly. Appropriate weight gain was usually achieved by approximately 18–26 weeks of age. At this time, body composition was assessed using NMR and adiposity was assessed using Vevo LAZR-X. Mice were then treated intravenously with F11 at 0.00625, 0.0125, 0.025, 0.05, or 0.5 mg / kg (mpk). NMR body composition measurements were then performed at 1, 3, 5, and 7 weeks after F11 treatment. LAZR-X adiposity measurements were performed at 4 and 7 weeks after F11 treatment. Additionally, at 7 weeks after F11 treatment, blood (200 μl) was drawn submandibularly to assess HbA1c levels. Eight weeks after F11 administration, mice were sacrificed and the following measurements were performed: (1) body weight, liver weight, and epididymal fat weight were measured, (2) blood was collected for measurement of alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), creatinine (CRE), and cytokines, (3) liver and epididymal fat were photographed, (4) optical coherence tomography (OCT) of the liver was performed, and (5) mice were perfused and tissues were processed for immunohistochemistry and H&E staining.

[0135] Results from these experiments showed that body weight was positively correlated with total fat, but not lean mass, at baseline, 3 days after F11 treatment, and 7 days after F11 treatment (Figures 4-6, panels A and B). Furthermore, the percentage of total fat mass and the percentage of total lean mass were negatively correlated at the same three time points (Figures 4-6, panel F). Consistent results from baseline, 3 days after F11 iv administration, and 17 days after F11 iv administration indicate that F11 proportionally alters fat and lean mass in the same mice (Figures 4-6, panel F). Total body weight changes were generally confirmed by lean mass changes after 3 days of F11 treatment at all F11 doses (Figures 7A and 7C). F11 administered at 0.05-0.5 mpk reduced body weight by decreasing either fat or lean mass, or both, after 3 days of F11 treatment (Figure 7). The most dramatic effect on body weight was observed when F11 was administered at 0.5 mpk, leading to a substantial decline in both fat and lean mass (Figure 7). Lower doses of F11 (0.025-0.05 mpk) did not significantly alter fat mass, and even lower levels of F11 (≤0.0125 mpk) appeared to induce fat accumulation (Figure 7B).

[0136] Because mice were continuously fed a 60% HFD, after 17 days of F11 treatment, the total body weight changes with all F11 doses were largely confirmed by changes in fat content (Figures 8A and 8B). Consistent with the results of 3-day treatment with F11, only 0.5mpk F11 treatment was effective in reducing body weight, fat content, and lean mass content (Figure 8). F11 doses below 0.5mpk were ineffective in reducing fat content compared to baseline (before iv administration of F11) (Figure 8).

[0137] Figures 9 and 10 show the weight change (R ) after 3 days of F11 treatment using multiple logistic regression analysis (Figures 9A and 9B). 2 =0.9821) and fat change (R 2A significant standard curve for F11 efficacy against body weight change (R = 0.9639) after 17 days of F11 treatment was obtained. 2 =0.7278) and fat change (R 2 = 0.7023) compared to a standard curve for F11 efficacy. Multiple regression analysis suggests that 0.32mpk and 0.4mpk may be the best F11 doses for reducing fat and body weight, respectively, after 3 days of treatment.

[0138] One or more exemplary embodiments have been described by way of example, and it will be understood by those skilled in the art that numerous variations and modifications can be made without departing from the scope of the invention, as defined in the claims.

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[0140] All references cited herein and throughout the specification are hereby incorporated by reference in their entirety.

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