Biomarkers for Parkinson's Disease

Abstract

The present disclosure relates to the treatment of Parkinson's disease with granulocyte-macrophage colony-stimulating factor, as well as methods of diagnosis, prognosis, and patient selection.

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Patent Application No. 63 / 240,233, filed September 2, 2021, and U.S. Provisional Patent Application No. 63 / 303,102, filed January 26, 2022, the entire contents of each of which are incorporated by reference herein for all purposes.

[0002] The present disclosure relates, in part, to the treatment and / or amelioration of Parkinson's disease, as well as methods of diagnosis, prognosis and patient selection.

[0003] Government support This invention was made with Government support under Grant No. R01NS034239 awarded by the National Institutes of Health. The Government has certain rights in the invention.

[0004] Description of electronically submitted text files The contents of the text file submitted electronically herewith are incorporated herein by reference in their entirety: Copy of Sequence Listing in Computer Readable Format (Filename: Sequence_Listing_PNR-011PC / 127114-5011.xml; Recorded Date: August 26, 2022; File Size: 5,000 bytes). [Background technology]

[0005] Neurodegenerative diseases are increasingly recognized as the leading cause of death and disability (including disability-adjusted life-years (DALYs; the sum of Years of life lost [YLLs] and Years lived with disability [YLDs])) worldwide. Globally, in 2016, neurological disorders were the leading contributor to DALYs (~276 million DALYs) and the second leading cause of death (~9 million). See Global Burden of Diseases, Injuries, and Risk Factors Study (GBD), Lancet Neurol 2019; 18: 459-80.

[0006] Neurodegenerative disorders can be broadly classified according to their clinical manifestations, with extrapyramidal and pyramidal motor disorders, and cognitive or behavioral disorders being the most common. Few patients have pure syndromes, and most have complex clinical features. Although neurodegenerative diseases are usually defined by the accumulation of specific proteins and anatomical vulnerabilities, they share many fundamental processes associated with progressive neuronal dysfunction and death, including proteotoxicity and associated abnormalities in the ubiquitin-proteasome and autophagosome / lysosomal systems, oxidative stress, programmed cell death, and neuroinflammation. See Dugger BN and Dickson DW. Cold Spring Harb Perspect Biol. 2017. 9(7): a028035.

[0007] Parkinson's disease (PD) is a progressive neurodegenerative disorder characterized by progressive loss of substantia nigra dopaminergic neurons. The predominant presence of α-synuclein (a-syn) aggregates in intracellular inclusions, both within and outside the central nervous system (CNS), is a disease driver that characterizes a systemic disease with multiorgan involvement and consequent immune response. The link between innate (monocytes and microglia) and adaptive immunity (T cells), inflammation, and disease is well established. The main drivers are α-syn misfolding and aggregation, impaired protein clearance, mitochondrial dysfunction, and inflammation. All of these affect dopaminergic neuronal function with secondarily effects on noradrenergic, glutamatergic, serotonergic, and adenosine neuronal vitality. The disease is accentuated by the release of α-syn aggregates into the systemic circulation, which disrupts immune tolerance. In particular, activated monocytes enter the meninges and choroid plexus and exert early detrimental effects on neuroimmune homeostasis. Based on these findings, efforts to maintain immune homeostasis are attractive targets for PD-modifying therapies. One avenue to restore immune homeostasis within and outside the CNS focuses on immune alterations. In particular, a balanced shift from monocyte-macrophages, effector T cells (Teff) to regulatory T cells (Treg), and the interplay between these two, is presumed to have clinical benefits not only in PD but also in Alzheimer's disease (AD), traumatic brain injury, stroke, and amyotrophic lateral sclerosis (ALS). Enhancing Treg numbers and function leads to restoration of innate microglial homeostasis and control of neuroinflammation and neuroprotection.See Schabitz WR, et al. J Cereb Blood Flow Metab. 2008. 28, 29-43; Brochard V, et al. J Clin Invest. 2009. 119, 182-192; Jain S. Parkinsonism Relat Disord. 2011. 17, 77-83; Waschbisch, A, et al. J Immunol. 2016. 196, 1558-1567; Houser MC and Tansey MG. NPJ Parkinsons Dis. 2017. 3(3); Rocha EM, et al. Neurobiol Dis. 2018. 109, 249-257; Harms AS, et al. Exp Neurol. 2018. 300, 179-187; Nissen SK et al. Mov Disord. 2019. 34, 1711 -1721 ; Machhi J, et al. Mol Neurodegener. 2020. 15(32); J. Jankovic J and Tan EK. J Neurol Neurosurg Psychiatry. 2020. 91, 795-808.

[0008] The growing recognition that inflammation may play an important role in neurodegenerative diseases of the CNS is highlighted by the observation of both adaptive vs. innate immune responses at various stages of neurodegenerative diseases. These distinct immune responses may not only drive the disease process but also serve as therapeutic targets. Ongoing research into the specific inflammatory mechanisms that play a role in disease pathogenesis and progression is revealing lessons about inflammatory neurodegeneration that will help understand the emergence of these diseases and therapies to treat them. A growing number of immunotherapeutic strategies that have been successful in MS are now being applied to other neurodegenerative diseases. Some approaches suppress the immune mechanisms of the CNS, while others harness the immune system to remove harmful products and cells. See Mamun AA and Liu F. Neurol Neurother. 2017. 2(1); Chitnas T and Weiner HL. J Clin Invest. 2017. 127(10): 3577-3587.

[0009] It is important to note that not all immune responses in the CNS are harmful, and in many cases they are actually beneficial for repair and regeneration. For example, microglia clear debris after myelin damage, and if this is prevented, delayed regeneration occurs. Immune activation is also important for limiting neurotropic viral infections and clearing necrotic cells after ischemia. Thus, microglia may exert a dual role in neurodegeneration, both as instigators of damage and as protectors of brain homeostasis. In addition to microglia, T cells may also help in the recovery of neurodegenerative diseases, although the exact mechanism of this beneficial role of T cells is unclear. Detailed studies of neuroimmune interactions at both cellular and molecular levels have revealed complex interactions and shown that immune cells can secrete both neurotoxic and neuroprotective molecules. Thus, control of immune responses in neurological diseases is of therapeutic value. See Neumann H et al. Brain. 2009. 132: 288-95; Schwartz M et al. Neuroscience. 2009. 158: 1133-42; Amor S et al. Immunology. 2010. 129(2): 154-69.

[0010] The clinical and pathobiological heterogeneity of Parkinson's disease presents a major challenge in the development of relevant biomarkers to monitor disease progression and disease-modifying therapies. Furthermore, because of the variability in treatment response based on different clinical and molecular phenotypes, a shift towards a personalized or precision medicine approach, including the development and validation of biomarkers, has been thought to improve the management of many neurodegenerative diseases such as PD.

[0011] Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a blood growth factor that regulates the production, migration, proliferation, differentiation, and function of hematopoietic cells. It was first identified in vitro for its ability to induce the proliferation and differentiation of bone marrow progenitor cells into granulocytes and macrophages. In response to inflammatory stimuli, GM-CSF is released by various cell types, including T lymphocytes, macrophages, fibroblasts, and endothelial cells. In turn, GM-CSF activates and enhances the production and survival of neutrophils, eosinophils, and macrophages. Native GM-CSF is usually produced near the site of action where it regulates the proliferation, differentiation, and survival of hematopoietic progenitor cells in vitro, but is present in the circulating blood at picomolar concentrations (10 -10 ~10 -12 M). Several studies have shown that GM-CSF has a broad range of functions across various tissues in its action on myeloid cells, and GM-CSF deletion / depletion approaches have demonstrated its potential as an important therapeutic target in several inflammatory and autoimmune diseases. A Metcalf D. Immunol Cell Biology. 1987, 65:35-43; Gasson JC. Blood. 1991 , 77:1131 -1145; Shannon MF et al. Crit Rev Immunol. 1997, 17:301 -323; Alexander WS. Int Rev Immunol. 1998, 16:651 -682; Barreda DR et al. Dev Comp Immunol. 2004, 28:509-554; Lee KMC et al. Immunotargets Then 2020. 9:225-240.

[0012] Recombinant human granulocyte macrophage colony stimulating factor (rhuGM-CSF) is FDA approved for the treatment of neutropenia, hematopoietic dysfunction, and malignancies such as leukemia, in combination with chemotherapy. In clinical practice, GM-CSF used to treat post-chemotherapy neutropenia and aplastic anemia significantly reduces the risk of infection associated with bone marrow transplantation. Its utility in the treatment of myeloid leukemia and as a vaccine adjuvant is also well established. See Dorr RT. Clin Therapeutics. 1993. 15(1): 19-29; Armitage JO. Blood 1998, 92:4491-4508; Kovacic JC et al. J Mol Cell Cardiol. 2007, 42:19-33; Jacobs PP et al. Microbial Cell Factories 2010, 9:93.

[0013] Identification of specific biomarkers can be used for the diagnosis, prognosis, or theranosis of neurodegenerative diseases such as Parkinson's disease. There remains a need for new and more effective biomarkers and combination therapies for Parkinson's disease. Summary of the Invention

[0014] Thus, in some embodiments, the present disclosure relates to a method for treating Parkinson's disease comprising administering to a patient in need thereof an effective amount of a composition comprising GM-CSF, wherein the patient is characterized by alterations in expression and / or activity of various biomarkers.

[0015] In some embodiments, the disclosure relates to a method of treating one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, down syndrome cognition, and encephalitis comprising administering an effective amount of a composition comprising GM-CSF to a patient in need thereof, wherein the patient is characterized by altered expression and / or activity of various biomarkers.

[0016] In some embodiments, the present disclosure relates to a method for treating one or more diseases or disorders characterized by an imbalance between Teff and Treg, comprising administering to a patient in need thereof an effective amount of a composition comprising GM-CSF, wherein the patient is characterized by altered expression and / or activity of various biomarkers.

[0017] In some embodiments, there is provided a method of selecting a patient for treatment with an agent for Parkinson's disease and / or a method of assessing a therapeutic response to an agent for Parkinson's disease, comprising determining the presence, absence or amount of one or more biomarkers in a biological sample from said patient, wherein said patient is suitable for said treatment if an alteration in expression and / or activity of said biomarkers is demonstrated compared to a pre-treatment and / or disease-free state, and wherein said agent comprises an effective amount of granulocyte-macrophage colony-stimulating factor (GM-CSF).

[0018] In some embodiments, there is provided a method of selecting a patient for treatment with an agent for one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, cognition of Down's syndrome, and encephalitis, and / or a method of evaluating a therapeutic response of an agent for one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, cognition of Down's syndrome, and encephalitis, comprising determining the presence, absence, or amount of one or more biomarkers in a biological sample from a patient, wherein the patient is suitable for said treatment if a change in expression and / or activity of said biomarkers is demonstrated compared to pre-treatment and / or disease-free state, and wherein said agent comprises an effective amount of granulocyte-macrophage colony-stimulating factor (GM-CSF).

[0019] In some embodiments, methods are provided for selecting a patient for treatment with an agent for one or more diseases or disorders characterized by an imbalance between Teff and Treg, and / or for assessing a therapeutic response to an agent for one or more diseases or disorders characterized by an imbalance between Teff and Treg, comprising determining the presence, absence, or amount of one or more biomarkers in a biological sample from the patient, wherein the patient is suitable for said treatment if an alteration in expression and / or activity of said biomarkers is demonstrated compared to pre-treatment and / or a disease-free state, and wherein said agent comprises an effective amount of granulocyte-macrophage colony-stimulating factor (GM-CSF).

[0020] In some embodiments, the biomarkers are HMOX1, TLR2, TLR8, RELA (NK-κB p65), IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, ATP 5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, optionally said biomarkers are selected from HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2.

[0021] In some embodiments, one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA are downregulated during or after treatment with GM-CSF. In some embodiments, one or more of HMOX1, TLR2, TLR8, RELA, and LRRK2 are downregulated during or after treatment with GM-CSF.

[0022] In some embodiments, one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA are downregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF. In some embodiments, one or more of HMOX1, TLR2, TLR8, RELA, and LRRK2 are downregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF.

[0023] In some embodiments, one or more of ATG3, ATG7, and GABARAPL2 are upregulated during or after treatment with GM-CSF. In some embodiments, one or more of ATG7 and GABARAPL2 are upregulated during or after treatment with GM-CSF.

[0024] In some embodiments, ATG3, ATG7, and GABARAPL2 are all upregulated during or after treatment with GM-CSF. In some embodiments, both ATG7 and GABARAPL2 are upregulated during or after treatment with GM-CSF.

[0025] In some embodiments, ATG3, ATG7, and GABARAPL2 are upregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF. In some embodiments, ATG7 and GABARAPL2 are upregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF.

[0026] In some embodiments, (i) one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA are downregulated during or after treatment with GM-CSF, and optionally one or more of HMOX1, TLR2, TLR8, RELA, and LRRK2 are downregulated during or after treatment with GM-CSF, and (ii) one or more of ATG3, ATG7, and GABARAPL2 are upregulated during or after treatment with GM-CSF, and optionally one or more of ATG7 and GABARAPL2 are upregulated during or after treatment with GM-CSF.

[0027] In some embodiments, the biomarkers are associated with one or more pathways selected from the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, the sirtuin signaling pathway, and the oxidative phosphorylation pathway.

[0028] In some embodiments, the biomarkers associated with the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway are downregulated or inhibited during or after treatment with GM-CSF.

[0029] In some embodiments, the biomarkers associated with the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway are downregulated or inhibited at about 1 month, about 2 months, about 3 months, about 4 months, about 5 months, and / or about 6 months after treatment with GM-CSF.

[0030] In some aspects, methods are provided for treating Parkinson's disease in a patient, comprising identifying a patient having symptoms of Parkinson's disease, determining the presence, absence, or amount of one or more biomarkers in a biological sample from the patient, and administering an effective amount of a GM-CSF agent to a patient exhibiting altered expression and / or activity of one or more biomarkers compared to a pre-treatment and / or disease-free state.

[0031] In some embodiments, the biomarkers are HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, LRRK2, GABALAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, The biomarkers are selected from ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, and optionally, the biomarkers are selected from HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2.

[0032] In some embodiments, HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and one or more of ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA are downregulated during or after treatment with GM-CSF. In some embodiments, HMOX1, TLR2, TLR8, RELA, and LRRK2 are downregulated during or after treatment with GM-CSF.

[0033] In some embodiments, one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA are downregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF. In some embodiments, one or more of HMOX1, TLR2, TLR8, RELA, and LRRK2 are downregulated during or after treatment with GM-CSF and are downregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF.

[0034] In some embodiments, one or more of ATG3, ATG7, and GABARAPL2 are upregulated during or after treatment with GM-CSF. In some embodiments, one or more of ATG7 and GABARAPL2 are upregulated during or after treatment with GM-CSF. In some embodiments, all of ATG3, ATG7, and GABARAPL2 are upregulated during or after treatment with GM-CSF. In some embodiments, both ATG7 and GABARAPL2 are upregulated during or after treatment with GM-CSF. In some embodiments, one or more of ATG3, ATG7, and GABARAPL2 are upregulated about 1 month, about 2 months, about 3 months, about 4 months, about 5 months, and / or about 6 months after treatment with GM-CSF. In some embodiments, one or more of ATG7 and GABARAPL2 are upregulated at about 1 month, about 2 months, about 3 months, about 4 months, about 5 months, and / or about 6 months after treatment with GM-CSF.

[0035] In some embodiments, (i) one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA are downregulated during or after treatment with GM-CSF, and optionally one or more of HMOX1, TLR2, TLR8, RELA, and LRRK2 are downregulated during or after treatment with GM-CSF, and (ii) one or more of ATG3, ATG7, and GABARAPL2 are upregulated during or after treatment with GM-CSF, and optionally one or more of ATG7 and GABARAPL2 are upregulated during or after treatment with GM-CSF.

[0036] In some embodiments, the biomarkers are associated with one or more pathways selected from the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, the sirtuin signaling pathway, and the oxidative phosphorylation pathway.

[0037] In some embodiments, the biomarkers associated with the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway are downregulated or inhibited during or after treatment with GM-CSF.

[0038] In some embodiments, the biomarkers associated with the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway are downregulated or inhibited at 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF.

[0039] In some aspects, the method includes: identifying a patient who is undergoing or has undergone treatment with a neurological agent for a neurological condition, and who is showing failure, intolerance, resistance, or refractoriness to treatment with the neurological agent; determining the presence, absence or amount of one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA, ATG3, ATG7, and / or GABARAPL2; administering an effective amount of a GM-CSF agent to a patient exhibiting increased or higher expression and / or activity of one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and / or SDHA compared to a pre-treatment and / or disease-free state; and / or exhibiting decreased or lower expression and / or activity of ATG3, ATG7, and / or GABARAPL2 compared to a pre-treatment and / or disease-free state; The present invention provides a method for treating Parkinson's disease in a patient, comprising:

[0040] In some embodiments, identifying a patient undergoing or having undergone treatment with a neurological agent for a neurological condition, the patient exhibiting failure, intolerance, resistance, or refractoriness to treatment with the neurological agent; determining the presence, absence or amount of one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA, ATG3, ATG7, and / or GABARAPL2; administering an effective amount of a GM-CSF agent to a patient that exhibits decreased or lower expression and / or activity of one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and / or SDHA compared to a pre-treatment and / or disease-free state; and / or that exhibits increased or higher expression and / or activity of ATG3, ATG7, and / or GABARAPL2 compared to a pre-treatment and / or disease-free state; The present invention provides a method for treating Parkinson's disease in a patient, comprising:

[0041] In some embodiments, identifying a patient undergoing or having undergone treatment with a neurological agent for a neurological condition, the patient exhibiting failure, intolerance, resistance, or refractoriness to treatment with the neurological agent; determining an increase or decrease in expression and / or activity of biomarkers from one or more pathways including the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, the sirtuin signaling pathway, and the oxidative phosphorylation pathway; administering an effective amount of a GM-CSF agent to a patient exhibiting increased or elevated expression and / or activity of a neuroinflammatory signaling pathway, an IL-8 signaling pathway, a pathway for the generation of nitric oxide and reactive oxygen species, an integrin-linked kinase (ILK) signaling pathway, and an oxidative phosphorylation pathway, compared to a pre-treatment and / or disease-free state; The present invention provides a method for treating Parkinson's disease in a patient, comprising:

[0042] In some embodiments, (a) identifying a patient who is undergoing or has undergone treatment with a neurological agent for a neurological condition, and who is exhibiting failure, intolerance, resistance, or refractoriness to treatment with the neurological agent; (b) determining a decrease in expression and / or activity of a biomarker from one or more pathways including the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway, and / or an increase in expression and / or activity of a biomarker from the sirtuin signaling pathway; (c) administering an effective amount of a GM-CSF agent to a patient exhibiting decreased or lower expression and / or activity of the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway, compared to a pre-treatment and / or disease-free state; The present invention provides a method for treating Parkinson's disease in a patient, comprising:

[0043] In some embodiments, identifying a patient undergoing or having undergone treatment with a neurological agent for a neurological condition, the patient exhibiting failure, intolerance, resistance, or refractoriness to treatment with the neurological agent; determining a decrease in expression and / or activity of a biomarker from one or more pathways including the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway, and / or an increase in expression and / or activity of a biomarker from the sirtuin signaling pathway; administering an effective amount of a GM-CSF agent to a patient exhibiting decreased or lower expression and / or activity of a neuroinflammatory signaling pathway, an IL-8 signaling pathway, a pathway for generation of nitric oxide and reactive oxygen species, an integrin-linked kinase (ILK) signaling pathway, and an oxidative phosphorylation pathway, compared to a pre-treatment and / or disease-free state; The present invention provides a method for treating Parkinson's disease in a patient, comprising:

[0044] In some embodiments, a method for monitoring regression, progression, elimination, or recurrence of symptoms of Parkinson's disease in a patient following treatment with a GM-CSF agent comprises: determining a baseline expression and / or activity level of one or more biomarkers in a biological sample from the patient at a first time point; determining the expression and / or activity level of one or more biomarkers in the biological sample at the second and subsequent time points; determining whether expression and / or activity levels of one or more biomarkers change between said first and second time points; Methods are provided wherein the one or more biomarkers are selected from HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA, ATG3, ATG7, and GABARAPL2.

[0045] In some embodiments, the methods herein further comprise administering an effective amount of a drug or therapeutic agent to treat Parkinson's disease.

[0046] In some embodiments, the patient is characterized by having one or more of the following: oxidative stress, loss of neurite integrity, apoptosis, neuronal loss, and / or inflammatory responses, cognitive impairment, cognitive decline, behavioral and personality changes, tremors, bradykinesia, rigidity, impaired posture and balance, loss of automatic movements, decreased coordination, changes in speech, photophobia, difficulty controlling the eye muscles, slowed saccadic eye movements, dysphagia, blepharospasm, fainting or lightheadedness due to orthostatic hypotension, dizziness, bladder control problems, well-formed hallucinations and delusions, changes in memory, concentration, and judgment, memory loss, depression, irritability, anxiety, rapid eye movement (REM) sleep disorder, seizures, paresthesia, numbness or tingling, convulsions, difficulty chewing or swallowing, muscle spasms and weakness in the limbs, and / or prickling or tingling in the feet or hands.

[0047] In some embodiments, the presence, absence, or amount of the one or more biomarkers is determined by protein and / or nucleic acid detection. In some embodiments, the presence, absence, or amount of the one or more biomarkers is determined by one or more of ELISA, Luminex multiplex assay, immunohistochemical staining, Western blotting, in-cell Western, immunofluorescence staining, or fluorescence activated cell sorting (FACS). In some embodiments, the presence, absence, or amount of the one or more biomarkers is determined by one or more of polymerase chain reaction (PCR) amplification reaction, reverse transcriptase PCR analysis, quantitative real-time PCR, droplet digital PCR (ddPCR), single strand conformation polymorphism analysis (SSCP), mismatch cleavage detection, heteroduplex analysis, deoxyribonucleic acid (DNA) sequencing, ribonucleic acid (RNA) sequencing, Northern blot analysis, in situ hybridization, array analysis, and restriction fragment length polymorphism analysis. In some embodiments, the presence, absence, or amount of the one or more biomarkers is determined by single-cell RNA sequencing and / or next-generation sequencing (NGS). In some embodiments, the presence, absence, or amount of the one or more biomarkers is determined by RNA sequencing, e.g., single-cell RNA sequencing.

[0048] In some embodiments, the biological sample is or comprises blood, a skin or tissue sample, plasma, serum, pus, urine, sweat, tears, mucus, sputum, saliva, cerebrospinal fluid (CSF), and / or other bodily fluids. In some embodiments, the biological sample is or comprises monocytes. In some embodiments, the biological sample is or comprises a monocyte population.

[0049] In some embodiments, the method prevents, treats, and / or ameliorates the progression and / or onset of Parkinson's disease in a patient. In some embodiments, the method induces a disease-modifying response, and / or slows cognitive decline, either temporarily or persistently, and / or results in amelioration of symptoms of a neurodegenerative disease, and / or slows the onset and / or progression of a neurodegenerative disease or disorder, and / or reverses or prevents chronic inflammation in the central nervous system (CNS), and / or reduces or ameliorates dysfunction of endogenous or extrinsic CNS immune cells, and / or reduces or ameliorates activation of CNS astrocytes and mononuclear phagocytes (e.g., perivascular macrophages and / or microglial cells). and / or reducing, alleviating or reversing astrogliopathy, and / or regulating, maintaining or supporting glutamine-glutamate balance in the CNS, and / or reducing, alleviating or reversing chronic microglial cell activation, and / or reducing or reversing axonal injury, and / or reducing or preventing excessive production and / or signaling of one or more inflammatory cytokines and / or proteins, and / or reducing or preventing the formation of protein plaques, and / or reducing or preventing tauopathy.

[0050] In some embodiments, the GM-CSF has an amino acid sequence of one of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, and SEQ ID NO:4, or a variant having at least about 90%, or at least about 93%, or at least about 95%, or at least about 97%, or at least about 98% identity thereto. In some embodiments, the GM-CSF is one of molgramostim, sargramostim, and regramostim. In some embodiments, the GM-CSF is sargramostim. In some embodiments, the GM-CSF is administered by an intravenous route.

[0051] In some embodiments, the method further comprises administering one or more additional therapeutic and / or neurological agents selected from dopamine precursors such as levodopa, carbidopa (LODOSYN); dopamine antagonists such as selegiline (ZELAPAR); MAOB inhibitors such as selegiline (ZELAPAR); catechol-O-methyltransferase (COMT) inhibitors such as entacapone (COMTAN); anticholinergics such as benztropine (COGENTIN); amantadine; adenosine receptor antagonists (A2A receptor antagonists) such as istradefylline (NOURIANZ); and / or pimavanserin (NUPLAZID).

[0052] In some aspects, methods are provided for treating Parkinson's disease comprising: selecting a patient having Parkinson's disease and having altered expression and / or activity of one or more biomarkers compared to a disease-free state; and administering to the patient an effective amount of a composition comprising GM-CSF, wherein the one or more biomarkers are selected from HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA, ATG3, ATG7, and GABARAPL2.

[0053] In some embodiments, altered expression and / or activity of said one or more biomarkers indicates discontinuation of administration of GM-CSF, hi some embodiments, the level of any of said biomarkers is assayed in a biological sample from said patient.

[0054] In some embodiments, the biological sample comprises blood, a tissue sample, plasma, serum, pus, urine, sweat, tears, mucus, sputum, saliva, cerebrospinal fluid (CSF), and / or other bodily fluids.

[0055] In some embodiments, the methods prevent, treat, and / or alleviate the progression and / or development of Parkinson's disease.

[0056] In some embodiments, the method ameliorates a symptom of Parkinson's disease in the patient, hi some embodiments, the method reduces sequelae of Parkinson's disease in the patient compared to before treatment.

[0057] In some embodiments, the GM-CSF has an amino acid sequence of one of SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, and SEQ ID NO:4, or a variant having at least about 90%, or at least about 93%, or at least about 95%, or at least about 97%, or at least about 98% identity thereto. In some embodiments, the GM-CSF is one of molgramostim, sargramostim, and regramostim. In some embodiments, the GM-CSF is sargramostim. In some embodiments, the GM-CSF is administered by an intravenous route.

[0058] In some embodiments, the method further comprises administering one or more additional therapeutic and / or neurological agents selected from dopamine precursors such as levodopa, carbidopa (LODOSYN); dopamine antagonists such as selegiline (ZELAPAR); MAOB inhibitors such as selegiline (ZELAPAR); catechol-O-methyltransferase (COMT) inhibitors such as entacapone (COMTAN); anticholinergics such as benztropine (COGENTIN); amantadine; adenosine receptor antagonists (A2A receptor antagonists) such as istradefylline (NOURIANZ); and / or pimavanserin (NUPLAZID).

[0059] In some aspects, a companion diagnostic, complementary diagnostic, or co-diagnostic test kit is provided that includes an array of nucleic acids or proteins suitable for detection of one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA, ATG3, ATG7, and GABARAPL2, and instructions for use.

[0060] In some embodiments, companion, complementary, or co-diagnostic test kits are provided that contain reagents and instructions for use in one or more of the methods described herein. [Brief description of the drawings]

[0061] [Figure 1A] Pathway enrichment of differentially expressed proteins in monocytes of PD patients after 2 months of sargramostim treatment. Functional enrichment of Gene Ontology (GO) terms with five categories (immune response, biological process, cellular component, KEGG, and Reactome) was performed using Cytoscape with the plugin ClueGO.

[0062] [Figure 1B] 1 shows canonical pathway enrichment analysis performed using IPA (Qiagen) showing differentially expressed proteins in monocytes of PD patients after 2 months of sargramostim treatment. Black arrows point to the canonical pathway status shown in FIG. 1B. Positive Z-score (activated), negative Z-score (inhibited), and light gray (no activity pattern).

[0063] [Figure 2A]Pathway enrichment of differentially expressed proteins / genes in monocytes from PD patients after 6 months of sargramostim treatment. Functional enrichment of Gene Ontology (GO) terms with five categories (immune response, biological process, cellular component, KEGG, and Reactome) was performed using Cytoscape with the plugin ClueGO.

[0064] [Figure 2B] 2B shows canonical pathway enrichment analysis performed using IPA (Qiagen) showing differentially expressed proteins in monocytes of PD patients after 6 months of sargramostim treatment. Black arrows point to the canonical pathway status shown in FIG. 2B. Positive Z-score (activated), negative Z-score (inhibited), and light gray (no activity pattern). [Diagram 3] Graphs showing gene and protein expression of potential biomarkers in monocytes after 2 and 6 months of sargramostim treatment. ddPCR assays were performed to measure gene expression of LRRK2, HMOX1, TLR2, TLR8, RELA, ATG7, and GABARAPL2 after 2 months (A) and 6 months (B) of sargramostim treatment compared to baseline. Gene expression was normalized to HPRT1 and ddPCR assays were performed four times (n=4 technical replicates). Western blot analysis was performed to measure protein expression of β-actin, LRRK2, HMOX1, TLR2, TLR8, RELA, ATG7, and GABARAPL2 after 2 months (C) and 6 months (D) of sargramostim treatment compared to baseline. Protein expression was normalized to β-actin and densitometric quantification is shown. Western blot analysis was performed in triplicate (n=3 technical replicates). Data represent mean ± SD. The horizontal line in each image represents baseline expression: values ​​above the line represent upregulation and values ​​below the line represent downregulation.

[0065] [Figure 4A]1 shows integration of scRNA-seq and proteomics data showing overlapping genes between scRNA-seq and proteomics datasets for patients 2003, 2004, and 2005 after 6 months of sargramostim treatment compared to baseline.

[0066] [Figure 4B] 1 shows a graph depicting the correlation of overlapping genes in both the scRNA-seq and proteomics datasets for patients 2003, 2004, and 2005 after 6 months of sargramostim treatment compared to baseline. Correlation was determined using the Pearson product moment correlation coefficient (r).

[0067] [Diagram 5] Figure 5A shows a graph depicting the correlation between gene expression of LRRK2, HMOX1, TLR2, TLR8, RELA, ATG7, and GABARAPL2 and changes in MDS-UPDRSIII scores. r = Pearson product moment correlation coefficient.

[0068] Figure 5B shows a graph depicting the correlation between gene expression of LRRK2, HMOX1, TLR2, TLR8, RELA, ATG7, and GABARAPL2 and raw MDS-UPDRSIII scores. r = Pearson product moment correlation coefficient.

[0069] FIG. 5C shows multiple linear regression analysis of the effect of gene expression of LRRK2, HMOX1, TLR2, TLR8, and ATG7 on change in MDS-UPDRSIII score.

[0070] Figure 5D shows multiple linear regression analysis of the effect of gene expression of LRRK2, HMOX1, TLR2, TLR8, and ATG7 on raw MDS-UPDRSIII scores. r = regression coefficient.

[0071] [Figure 6]Figure 6A shows a graph depicting the correlation between protein expression of LRRK2, RELA, and ATG7 and changes in MDS-UPDRSIII scores. r = Pearson product moment correlation coefficient.

[0072] Figure 6B shows a graph depicting the correlation between protein expression of LRRK2, RELA, and ATG7 and raw MDS-UPDRSIII scores. r = Pearson product moment correlation coefficient.

[0073] Figure 6C shows multiple linear regression analysis of the effect of LRRK2, HMOX1, RELA, and GABARAPL2 protein expression on change in MDS-UPDRSIII score. r = regression coefficient.

[0074] Figure 6D shows multiple linear regression analysis of the effect of TLR2, TLR8, and ATG7 protein expression on raw MDS-UPDRSIII scores. r = regression coefficient. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0075] This disclosure relates, in part, to the use of GM-CSF as an effective treatment for Parkinson's disease, selected using specific biomarkers as predictive clinical markers of disease sequelae and responsiveness to current treatments (e.g., presence or absence of treatment with a Parkinson's disease medication).

[0076] In some aspects, the present disclosure relates to improved methods of selecting patients in need of treatment for one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, encephalitis, and / or one or more conditions characterized by an imbalance between Teff and Tregs, and / or improved methods of assessing a therapeutic response to an agent for one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, encephalitis, and / or one or more conditions characterized by an imbalance between Teff and Tregs. In one aspect, the disclosure relates to improved treatment of one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, encephalitis, and / or one or more conditions characterized by an imbalance between Teff and Treg in a patient based on selected predictive biomarkers.For example, in some embodiments, HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, LRRK2, GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG Assessment of the presence, absence, level or activity of one or more biomarkers, such as HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and / or GABARAPL2, optionally one or more biomarkers, such as HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2, is informative or predictive of a patient's condition and indicates administration of GM-CSF to a patient having one or more of, but not limited to, Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, encephalitis, and / or one or more conditions characterized by an imbalance between Teff and Treg. In other embodiments, evaluation of biomarkers from one or more pathways, including the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, the sirtuin signaling pathway, and the oxidative phosphorylation pathway, informs or predicts a patient's condition and directs administration of GM-CSF to patients with one or more of, but not limited to, Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, encephalitis, and / or one or more conditions characterized by an imbalance between Teff and Treg.

[0077] In some embodiments, the biomarkers are associated with one or more proteins and / or biomarkers and signaling and / or regulatory pathways. In some embodiments, the biomarkers of the present disclosure are used in combination with one or more other of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, encephalitis, and / or one or more conditions characterized by an imbalance between Teff and Treg to aid in monitoring disease progression and response to treatment.

[0078] Thus, in some aspects, the present disclosure provides methods for treating one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, encephalitis, and / or one or more conditions characterized by an imbalance between Teff and Treg by assessing selected clinical biomarkers.

[0079] composition In some embodiments, the present disclosure relates to compositions for treating Parkinson's disease, such as pharmaceutical compositions comprising GM-CSF and / or an additional therapeutic agent.

[0080] In some embodiments, the additional neurological agent and / or additional therapeutic agent is selected from dopamine precursors such as levodopa, carbidopa (LODOSYN); dopamine antagonists such as selegiline (ZELAPAR); MAOB inhibitors such as selegiline (ZELAPAR); catechol-O-methyltransferase (COMT) inhibitors such as entacapone (COMTAN); anticholinergics such as benztropine (COGENTIN); amantadine; adenosine receptor antagonists (A2A receptor antagonists) such as istradefylline (NOURIANZ); and / or pimavanserin (NUPLAZID).

[0081] In some embodiments, the neurological agent and / or additional therapeutic agent for treating Parkinson's disease is an antibody or antibody format selected from one or more of a monoclonal antibody, a polyclonal antibody, an antibody fragment, Fab, Fab', Fab'-SH, F(ab')2, Fv, single chain Fv, diabody, linear antibody, bispecific antibody, multispecific antibody, chimeric antibody, humanized antibody, human antibody, and a fusion protein comprising an antigen-binding portion of an antibody.

[0082] Composition of GM-CSF In some embodiments, GM-CSF includes any pharma- ceutically safe and effective GM-CSF, or any derivative thereof that has the biological activity of GM-CSF. In some embodiments, GM-CSF is rhuGM-CSF, such as sargramostim (LEUKINE). Sargramostim is a recombinant human GM-CSF biosynthesized from yeast, and has a single 127 amino acid glycoprotein that differs from endogenous human GM-CSF in that it has a leucine instead of a proline at position 23. Other natural and synthetic GM-CSFs, and derivatives thereof that have the biological activity of natural human GM-CSF, may be similarly useful in some embodiments.

[0083] In some embodiments, GM-CSF is produced or producible in bacteria, yeast, plants, insect cells, and mammalian cells. In some embodiments, GM-CSF is produced or producible in Escherichia coli cells. In some embodiments, GM-CSF is produced or producible in yeast cells. In some embodiments, GM-CSF is produced or producible in Chinese Hamster Ovary cells (CHO). In some embodiments, GM-CSF is not produced in E. coli cells. In some embodiments, GM-CSF is produced in cells that allow for glycosylation, such as yeast cells or CHO cells.

[0084] In some embodiments, the GM-CSF has the amino acid sequence of SEQ ID NO: 1, or a variant having at least about 90%, or at least about 93%, or at least about 95%, or at least about 97%, or at least about 98% identity thereto. In some embodiments, the GM-CSF has the amino acid sequence of SEQ ID NO: 2, SEQ ID NO: 3, and SEQ ID NO: 4, or a variant having at least 90%, or at least about 93%, or at least about 95%, or at least about 97%, or at least about 98% identity thereto. In some embodiments, the GM-CSF is one of sargramostim, molgramostim, and regramostim. In some embodiments, the GM-CSF is sargramostim.

[0085] Without wishing to be bound by theory, the core of hGM-CSF consists of four helices packed at angles. The crystal structure and mutagenesis analysis of rhGM-CSF (Rozwarski DA et al., Proteins 26:304-13,1996) showed that in addition to the nonpolar side chains of the protein core, 10 buried hydrogen-bonding residues are involved in intramolecular hydrogen bonding to the main chain and are more conserved than residues hydrogen-bonding to other side-chain atoms. Twenty-four solvation sites were observed at equivalent positions in the two molecules in the asymmetric unit, among which the strongest solvation sites were located in the clefts between the secondary structure elements. Two surface clusters of hydrophobic side chains are located near the predicted receptor-binding region. Mutagenesis of residues on the helix A / helix C face confirmed the importance of specific Glu, Gly, and Gln residues. Thus, these residues should not be replaced in functional substitution mutants of hGM-CSF for use in this disclosure, and these helices should be retained in functional fragment or deletion mutants of hGM-CSF for use in this disclosure. Additionally, in some embodiments, one of skill in the art can refer to UniProtKB entry P04141 for structural information to learn the identity of the variants.

[0086] The N-terminal helix of hGM-CSF governs high affinity binding to its receptor (Shanafelt AB et al., EMBO J 10:4105-12, 1991). Transduction of the biological effects of GM-CSF requires interaction with at least two cell surface receptor components, one of which is shared with the cytokine IL-5. In the above studies, we identified receptor binding determinants in GM-CSF by localizing unique receptor binding domains in a series of human-mouse hybrid GM-CSF cytokines. The interaction of GM-CSF with the shared subunit of its high affinity receptor complex was governed by a small portion of the peptide chain. The presence of several key residues in the N-terminal α-helix was sufficient to confer specificity to the interaction.

[0087] In some embodiments, the amino acid mutations are amino acid substitutions, which can include conservative and / or non-conservative substitutions.

[0088] "Conservative substitutions" can be made, for example, on the basis of similarity in polarity, charge, size, solubility, hydrophobicity, hydrophilicity, and / or the amphipathic nature of the amino acid residues involved. The 20 naturally occurring amino acids can be classified into six standard amino acid groups: (1) hydrophobic: Met, Ala, Val, Leu, Ile; (2) neutral hydrophilic: Cys, Ser, Thr; Asn, Gln; (3) acidic: Asp, Glu; (4) basic: His, Lys, Arg; (5) residues that affect chain orientation: Gly, Pro; and (6) aromatic: Trp, Tyr, Phe.

[0089] As used herein, "conservative substitution" is defined as the replacement of an amino acid with another amino acid listed in the same group of the six standard amino acid groups above. For example, the replacement of Asp with Glu retains one negative charge in the polypeptide so modified. Furthermore, glycine and proline can be substituted for each other based on their ability to disrupt α-helices.

[0090] As used herein, a "non-conservative substitution" is defined as the replacement of an amino acid with another amino acid listed in a different group of the six standard amino acid groups (1) to (6) above.

[0091] In some embodiments, substitutions may also include non-classical amino acids (e.g., selenocysteine, pyrrolysine, N-formylmethionine, β-alanine, GABA, and δ-aminolevulinic acid, 4-aminobenzoic acid (PABA), D-forms of common amino acids, 2,4-diaminobutyric acid, α-aminoisobutyric acid, 4-aminobutyric acid, Abu, 2-aminobutyric acid, γ-Abu, ε-Ahx, 6-aminohexanoic acid, Aib, 2-aminoisobutyric acid, 3-aminopropionic acid, ornithine, norleucine, norvaline, hydroxyproline, sarcosme, citrulline, homocitrulline, cysteic acid, t-butylglycine, t-butylalanine, phenylglycine, cyclohexylalanine, β-alanine, fluoroamino acids, engineered amino acids such as β-methylamino acids, C α-methylamino acids, N α-methylamino acids, and amino acid analogs in general).

[0092] Amino acid sequence modifications can be accomplished using any technique known in the art, such as site-directed mutagenesis or PCR-based mutagenesis, as described, for example, in Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Press, Plainview, NY, 1989, and Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, New York, NY, 1989. Without intending to be bound by theory, the degree of glycosylation of biosynthetic GM-CSF appears to affect half-life, distribution, and clearance (Lieschke and Burgess, N. Engl. J. Med. 327:28-35, 1992; Dorr, RT, Clin. Ther. 15:19-29, 1993; Horgaard et al., Eur. J. Hematol. 50:32-36, 1993). In some embodiments, the GM-CSF molecule is glycosylated.

[0093] Biomarkers In some embodiments, the methods relate to the utility of novel predictive biomarkers for determining the use of GM-CSF in the treatment of Parkinson's disease.

[0094] In some aspects, the methods relate to the utility of novel predictive biomarkers for determining the use of GM-CSF in the treatment of one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, and encephalitis.

[0095] In some aspects, the methods relate to the utility of novel predictive biomarkers to determine the use of GM-CSF in the treatment of one or more diseases or disorders characterized by an imbalance between Teff and Treg.

[0096] In one aspect, the disclosure relates to improved methods of selecting patients in need of treatment for Parkinson's disease. In another aspect, the disclosure relates to improved treatment of Parkinson's disease in patients based on selected predictive biomarkers. In other aspects, evaluating a patient who has failed or is intolerant or refractory to treatment for Parkinson's disease comprises measuring a biomarker in a biological sample of the patient.

[0097] In some aspects, there are provided methods of selecting a patient for treatment with an agent for one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, cognition of Down's syndrome, and encephalitis, and / or methods of assessing a therapeutic response to an agent for one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, cognition of Down's syndrome, and encephalitis, comprising determining the presence, absence, or amount of one or more biomarkers in a biological sample from the patient, wherein the patient is suitable for the treatment if a change in expression and / or activity of the biomarkers is demonstrated compared to a pre-treatment and / or disease-free state, and wherein the agent comprises an effective amount of granulocyte-macrophage colony-stimulating factor (GM-CSF).

[0098] In some aspects, methods are provided for selecting a patient for treatment with an agent for one or more diseases or disorders characterized by an imbalance between Teff and Treg, and / or for assessing a therapeutic response to an agent for one or more diseases or disorders characterized by an imbalance between Teff and Treg, comprising determining the presence, absence, or amount of one or more biomarkers in a biological sample from said patient, wherein said patient is suitable for said treatment if an alteration in expression and / or activity of said biomarkers is exhibited compared to pre-treatment and / or a disease-free state, and wherein said agent comprises an effective amount of granulocyte-macrophage colony-stimulating factor (GM-CSF).

[0099] In some embodiments, the evaluation of a patient with Parkinson's disease includes measuring various patient parameters. In some embodiments, a patient's biological sample can be analyzed, for example, using immunohistochemical or immunofluorescence techniques, which can be used to detect immune infiltrates, such as CD4 + Th cells (T helper cells), IL-17 producing CD4 + Th cells (Th17 cells), CD8 + T-cell (cytotoxic T-cell) and systemic or circulating intermediate monocyte immune subsets can be assessed. In some embodiments, multicolor flow cytometry can be used to measure multiple surface and intracellular markers, allowing characterization of cell phenotype and activation state. In some embodiments, whole blood can be used to assess changes in cell numbers due to treatment, or changes in cytokine levels, such as IL-1, IL-4, IL-6, IL-10, IL-12, IL-18, IL-33, IFN-γ, IP-10, M-CSF, TGF-β, VEGF, and TNFα levels. In some embodiments, deep sequencing technology can be used to quantify changes in individual cell clonotypes.

[0100] In some embodiments, the evaluation of a patient with Parkinson's disease involves determining the presence, absence, or amount of various biomarkers in a biological sample from the patient.

[0101] In some embodiments, the present disclosure relates to a method of treating Parkinson's disease in a patient, comprising administering to the patient an effective amount of one or more of the following: HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, LRRK2, GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK 2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and / or GABARAPL2, e.g., about 1, or about 2, or about 3, or about 4, or about 5, or about 10, or about 15, or about 20, or about 25, or about 30, or about 35, or about 40, or about 45. In some embodiments, the disclosure relates to a method of treating Parkinson's disease in a patient, wherein one or more, e.g., about 1, or about 2, or about 3, or about 4, or about 5, of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2 are used as biomarkers to predict or determine the need for treatment with GM-CSF.

[0102] In some embodiments, the present disclosure relates to methods of treating Parkinson's disease, in which biomarkers from one or more pathways, including the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, the sirtuin signaling pathway, and / or the oxidative phosphorylation pathway, are used to predict or determine the need for treatment with GM-CSF.

[0103] In some embodiments, the present disclosure relates to a method of selecting a patient with Parkinson's disease and / or assessing a therapeutic response to an agent for Parkinson's disease, comprising the steps of: HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, LRRK2, GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA One or more of ATG3, ATG7, and / or GABARAPL2, for example, about 1, or about 2, or about 3, or about 4, or about 5, or about 10, or about 15, or about 20, or about 25, or about 30, or about 35, or about 40, or about 45, optionally one or more of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2, for example, about 1, or about 2, or about 3, or about 4, or about 5, are used as biomarkers to predict or determine the need for treatment with GM-CSF.

[0104] In some embodiments, the present disclosure relates to methods of selecting patients with Parkinson's disease and / or evaluating therapeutic response to drugs for Parkinson's disease by assaying biomarkers from one or more pathways, including the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, the sirtuin signaling pathway, and / or the oxidative phosphorylation pathway, which are used to predict or determine the need for treatment with GM-CSF.

[0105] In some embodiments, the biomarkers of the present disclosure are used in combination with the biomarkers described in U.S. Patent Application Publication No. 2014 / 0349877, the entire contents of which are incorporated herein by reference, and U.S. Patent Application Publication No. 2019 / 0117735, the entire contents of which are incorporated herein by reference.

[0106] In some embodiments, the presence, absence, or amount of one or more predictive clinical biomarkers is determined by detection of protein and / or nucleic acid in a patient biological sample.

[0107] In some embodiments, the presence, absence, or amount of one or more predictive clinical biomarkers is determined in a patient biological sample by ELISA, immunohistochemical staining, Western blotting, in-cell Western, immunofluorescence staining, or fluorescence activated cell sorting (FACS), etc.

[0108] In some embodiments, the presence, absence, or amount of one or more predictive clinical biomarkers is determined by one or more of polymerase chain reaction (PCR) amplification reaction, reverse transcriptase PCR analysis, quantitative real-time PCR, single-stranded conformation polymorphism analysis (SSCP), mismatch cleavage detection, heteroduplex analysis, deoxyribonucleic acid (DNA) sequencing, ribonucleic acid (RNA) sequencing, Northern blot analysis, in situ hybridization, array analysis, and restriction fragment length polymorphism analysis.

[0109] In some embodiments, the presence, absence, or amount of one or more biomarkers is determined by next generation sequencing (NGS) methods. In some embodiments, the presence, absence, or amount of one or more biomarkers is determined by deep sequencing methods.

[0110] In some embodiments, the presence, absence, or amount of one or more biomarkers is determined by including ribonucleic acid (RNA) sequencing.

[0111] In some embodiments, the method of determining the presence, absence, or amount of one or more predictive clinical biomarkers is a method of characterizing a patient or selecting a patient for treatment that includes GM-CSF.

[0112] In some embodiments, the method of determining the level of one or more predictive clinical biomarkers comprises assaying the biomarkers in a biological sample from the patient.

[0113] In some embodiments, the methods, e.g., methods of determining the presence, absence, level or activity of one or more predictive clinical biomarkers for patient selection purposes, use a biological sample selected from blood, a skin sample or tissue sample, plasma, serum, pus, urine, sweat, tears, mucus, sputum, saliva, cerebrospinal fluid (CSF), and / or other bodily fluids.

[0114] In some embodiments, the method of patient selection is performed using a patient biological sample, said sample being selected from blood, a skin or tissue sample, a tissue biopsy specimen, a formalin-fixed or paraffin-embedded tissue specimen, a cytological sample, cultured cells, plasma, serum, pus, urine, sweat, tears, mucus, sputum, saliva, cerebrospinal fluid (CSF), and / or other bodily fluids.

[0115] In some embodiments, the method directs treatment decisions for a patient. For example, in some embodiments, the method includes monitoring expression and / or activity of one or more predictive clinical biomarkers during the course of treatment. In some embodiments, the method may detect changes in expression and / or activity of one or more predictive clinical biomarkers that correlate with disease state or treatment outcome in a Parkinson's disease patient. In such embodiments, without limitation, this directs treatment of the patient with a GM-CSF agent.

[0116] In some embodiments, the biomarkers of the present disclosure are proteomic-based biomarkers. In some other embodiments, the biomarkers comprise one or more proteins. In another embodiment, the biomarkers comprise one or more signaling and / or regulatory pathways in a cell. In some embodiments, the expression and / or activity of the biomarkers may be altered (increased or decreased) by treatment. Non-limiting examples of protein biomarkers include HMOX1, TLR2, TLR8, RELA (also referred to as RELA), IKBGG, ATG3, ATG7, LRRK2, GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and / or GABARAPL2.

[0117] In some embodiments, HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA are downregulated during or after treatment. In some embodiments, HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and / or SDHA are downregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF.

[0118] In some embodiments, ATG3, ATG7, and GABARAPL2 are upregulated during or after treatment with GM-CSF. In some embodiments, ATG7 and GABARAPL2 are upregulated during or after treatment with GM-CSF. In some embodiments, ATG3, ATG7, and GABARAPL2 are upregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF. In some embodiments, ATG7 and GABARAPL2 are upregulated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF.

[0119] In some embodiments, the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway are downregulated or inhibited during or after treatment with GM-CSF. In some embodiments, the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway are downregulated or inhibited 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF. In some embodiments, the sirtuin signaling pathway is upregulated and / or activated during or after treatment with GM-CSF. In some embodiments, the sirtuin signaling pathway is upregulated and / or activated 1 month, 2 months, 3 months, 4 months, 5 months, and / or 6 months after treatment with GM-CSF.

[0120] In some embodiments, the GM-CSF agents described herein enhance treatment with a therapy for treating Parkinson's disease. In some embodiments, the GM-CSF agents described herein are used to modulate a patient's immune system, for example, by decreasing or increasing the expression and / or activity of one or more, for example, about 1, or about 2, or about 3, or about 4, or about 5, or about 10, or about 15, or about 20, or about 25, of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and / or GABARAPL2. In some embodiments, the GM-CSF agents described herein are used to modulate a patient's immune system by decreasing or increasing the expression and / or activity of various signaling pathways, including, for example, the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway.

[0121] Methods of Treatment and Monitoring of Disease In one aspect, the disclosure relates to a method of treating Parkinson's disease comprising administering to a patient in need thereof an effective amount of the composition GM-CSF.

[0122] In some aspects, the disclosure relates to a method of treating one or more of Parkinson's disease, Alzheimer's disease, traumatic brain injury, stroke, amyotrophic lateral sclerosis, traumatic brain injury, progressive supranuclear palsy, Down's syndrome cognition, and encephalitis comprising administering to a patient in need thereof an effective amount of a composition comprising GM-CSF, wherein the patient is characterized by altered expression and / or activity of various biomarkers.

[0123] In some aspects, the present disclosure relates to a method of treating one or more diseases or disorders characterized by an imbalance between Teff and Treg, comprising administering to a patient in need thereof an effective amount of a composition comprising GM-CSF, wherein the patient is characterized by altered expression and / or activity of various biomarkers.

[0124] In another aspect, the disclosure relates to a method of treating Parkinson's disease comprising identifying a patient having symptoms of Parkinson's disease, determining the presence, absence, or amount of any of the biomarkers disclosed herein, and administering an effective amount of a GM-CSF agent to a patient exhibiting altered expression and / or activity of said one or more biomarkers compared to a pre-treatment and / or disease-free state.

[0125] In another aspect, the disclosure relates to a method of monitoring regression, progression, elimination, or recurrence of symptoms of Parkinson's disease in a patient following treatment with a GM-CSF agent, the method comprising determining a baseline expression and / or activity level of one or more biomarkers in a biological sample from the patient, determining the expression and / or activity level of the one or more biomarkers in the biological sample from the patient during treatment with GM-CSF, and determining whether the expression and / or activity level of the one or more biomarkers changes after initiation of treatment with GM-CSF compared to the baseline expression and / or activity level.

[0126] In some aspects, the present disclosure provides a method for producing a method for treating a cancer cell comprising: (a) identifying a patient who is undergoing or has undergone treatment with a neurological agent for a neurological condition, and who is experiencing failure, intolerance, resistance, or refractoriness to treatment with the neurological agent; (b) determining the presence, absence, or amount of one or more, e.g., about 1, or about 2, or about 3, or about 4, or about 5, or about 10, or about 15, or about 20, of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA, ATG3, ATG7, and / or GABARAPL2; (c) administering an effective amount of a GM-CSF agent to a patient that (i) exhibits increased or exhibits higher expression and / or activity of one or more of, e.g., about 1, or about 2, or about 3, or about 4, or about 5, or about 10, or about 15, or about 20, of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and / or SDHA, compared to a pre-treatment and / or disease-free state; and / or (ii) exhibits decreased or exhibits lower expression and / or activity of ATG3, ATG7, and / or GABARAPL2, compared to a pre-treatment and / or disease-free state.

[0127] In some aspects, the present disclosure provides a method for producing a method for treating a cancer cell comprising: (a) identifying a patient who is undergoing or has undergone treatment with a neurological agent for a neurological condition, and who is experiencing failure, intolerance, resistance, or refractoriness to treatment with the neurological agent; (b) determining an increase or decrease in expression and / or activity of biomarkers from one or more pathways, including a neuroinflammatory signaling pathway, an IL-8 signaling pathway, a nitric oxide and reactive oxygen species generation pathway, an integrin-linked kinase (ILK) signaling pathway, a sirtuin signaling pathway, and an oxidative phosphorylation pathway; (c) administering an effective amount of a GM-CSF agent to a patient exhibiting increased or elevated expression and / or activity of a neuroinflammatory signaling pathway, an IL-8 signaling pathway, a nitric oxide and reactive oxygen species generation pathway, an integrin-linked kinase (ILK) signaling pathway, and an oxidative phosphorylation pathway, compared to a pre-treatment and / or disease-free state; The present invention relates to a method of treating Parkinson's disease in a patient, comprising:

[0128] In some embodiments, the present disclosure relates to a method for treating Parkinson's disease comprising administering to a patient in need thereof an effective amount of a composition comprising GM-CSF, alone or in combination with a neurological agent and / or additional agent, wherein the patient is characterized as a partial responder or non-responder to a neurological treatment.

[0129] In some embodiments, the treatment method results in a decrease in expression and / or activity of one or more, e.g., about 1, or about 2, or about 3, or about 4, or about 5, or about 10, or about 15, or about 20, of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and / or SDHA.

[0130] In some embodiments, the method of treatment results in increased expression and / or activity of ATG3, ATG7, and / or GABARAPL2. In some embodiments, the method of treatment results in increased expression and / or activity of ATG7 and / or GABARAPL2.

[0131] In some embodiments, the treatment methods result in a decrease in expression and / or activity of biomarkers from the neuroinflammatory signaling pathway, the IL-8 signaling pathway, the nitric oxide and reactive oxygen species generation pathway, the integrin-linked kinase (ILK) signaling pathway, and the oxidative phosphorylation pathway.

[0132] In some embodiments, a method of monitoring regression, progression, elimination, or recurrence of symptoms of Parkinson's disease in a patient following treatment with a GM-CSF agent comprises: (a) determining a baseline expression and / or activity level of a biomarker in a biological sample from the patient; (b) determining the expression and / or activity level of one or more biomarkers in a biological sample from said patient during treatment with a GM-CSF agent; and (c) determining whether the expression and / or activity levels of one or more biomarkers change after initiation of treatment with GN-CSF compared to baseline expression and / or activity levels.

[0133] In some embodiments, the method of treatment prevents, treats, and / or ameliorates the progression and / or onset of Parkinson's disease in a patient. In some embodiments, the method of treatment ameliorates symptoms of Parkinson's disease in a patient. In some embodiments, the method of treatment induces a disease-modifying response in a patient. In other embodiments, the method of treatment slows cognitive decline in a patient, either temporarily or persistently. In yet other embodiments, the method of treatment results in the amelioration of symptoms of a neurodegenerative disease. In yet other embodiments, the method of treatment slows the onset and / or progression of Parkinson's disease.

[0134] In some embodiments, the method of treatment reduces, alleviates, reverses, or prevents chronic inflammation in the central nervous system (CNS). In some embodiments, the method of treatment reduces or alleviates dysfunction of endogenous or exogenous CNS immune cells. In some embodiments, the method reduces or alleviates activation of CNS astrocytes and mononuclear phagocytes, such as perivascular macrophages and / or microglial cells.

[0135] In some embodiments, the method of treatment reduces, alleviates, or reverses astrogliopathy, hi some embodiments, the method of treatment modulates the expression and / or activity of one or more cytokines and / or proteins.

[0136] In some embodiments, the method of treatment modulates, maintains, or supports glutamine-glutamate balance in the CNS. In some embodiments, the method of treatment reduces, alleviates, or reverses chronic microglial cell activation. In some embodiments, the method of treatment reduces or reverses axonal injury.

[0137] In some embodiments, the method of treatment reduces or prevents a pathological condition of the protein, hi some embodiments, the method of treatment reduces a tauopathy or prevents a tauopathy.

[0138] In some embodiments, the method of treatment results in a reduction in the sequelae of Parkinson's disease in the patient compared to before treatment.

[0139] In some embodiments, the patient suffers from a chronic progressive disorder of the nervous system.

[0140] In some embodiments, the patient is characterized by having oxidative stress, loss of neurite integrity, apoptosis, neuronal loss, and / or inflammatory responses, cognitive impairment, cognitive decline, behavioral and personality changes, tremors, bradykinesia, rigidity, impaired posture and balance, loss of automatic movements, decreased coordination, changes in speech, photophobia, difficulty controlling the eye muscles, slowed saccadic eye movements, difficulty swallowing, blepharospasm, fainting or lightheadedness due to orthostatic hypotension, dizziness, bladder control problems, well-formed hallucinations and delusions, changes in memory, concentration, and judgment, memory loss, depression, irritability, anxiety, rapid eye movement (REM) sleep disorder, seizures, paresthesia, numbness or tingling, convulsions, difficulty chewing or swallowing, muscle spasms and weakness in the limbs, and / or prickling or tingling in the feet and hands.

[0141] In some embodiments, the method further comprises administering one or more additional therapeutic agents selected from dopamine precursors such as levodopa, carbidopa (LODOSYN); dopamine antagonists such as selegiline (ZELAPAR); MAOB inhibitors such as selegiline (ZELAPAR); catechol-O-methyltransferase (COMT) inhibitors such as entacapone (COMTAN); anticholinergics such as benztropine (COGENTIN); amantadine; adenosine receptor antagonists (A2A receptor antagonists) such as istradefylline (NOURIANZ); and / or pimavanserin (NUPLAZID).

[0142] Pharmaceutically acceptable salts and excipients The compositions described herein may have a sufficiently basic functional group that can react with inorganic or organic acids to form pharma- ceutically acceptable salts, or a carboxyl group that can react with inorganic or organic bases to form pharma-ceutically acceptable salts. Pharmaceutically acceptable acid addition salts are formed from pharma-ceutically acceptable acids as is well known in the art. Such salts include, for example, the pharma-ceutically acceptable salts listed in Journal of Pharmaceutical Science, 66, 2-19 (1977), and The Handbook of Pharmaceutical Salts; Properties, Selection, and Use. PH Stahl and CG Wermuth (eds.), Verlag, Zurich (Switzerland) 2002, which are incorporated herein by reference in their entirety.

[0143] Pharmaceutically acceptable salts include, but are not limited to, sulfate, citrate, acetate, oxalate, chloride, bromide, iodide, nitrate, bisulfate, phosphate, hydrophosphate, isonicotinate, lactate, salicylate, acid salt, and the like. citrate), tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisate, fumarate, gluconate, glucaronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, camphorsulfonate, pamoate, phenylacetate, trifluoroacetate, acrylate, chlorobenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, methylbenzoate, o-acetoxybenzoate, naphthalene-2-benzoate, isobutyrate, phenylbutyrate, α-hydroxybutyrate, butyrate, Examples of suitable salts include phenyl-1,4-dicarboxylate, hexyne-1,4-dicarboxylate, caprate, caprylate, cinnamate, glycolate, heptanoate, hippurate, malate, hydroxymaleate, malonate, mandelate, mesylate, nicotinate, phthalate, terephthalate, propiolate, propionate, phenylpropionate, sebacate, suberate, p-bromobenzenesulfonate, chlorobenzenesulfonate, ethylsulfonate, 2-hydroxyethylsulfonate, methylsulfonate, naphthalene-1-sulfonate, naphthalene-2-sulfonate, naphthalene-1,5-sulfonate, xylenesulfonate, and tartrate.

[0144] The term "pharmaceutically acceptable salt" also refers to a salt of a composition of the present disclosure having an acidic functional group, such as a carboxylic acid functional group, with a base. Suitable bases include, but are not limited to, hydroxides of alkali metals such as sodium, potassium, lithium, etc.; hydroxides of alkaline earth metals such as calcium, magnesium, etc.; hydroxides of other metals such as aluminum and zinc; ammonia, and unsubstituted or hydroxy-substituted mono-, di-, or trialkylamines, organic amines such as dicyclohexylamine; tributylamine; pyridine; N-methyl, N-ethylamine; diethylamine; triethylamine; mono-, bis-, or tris-(2-OH-lower alkylamines), such as mono-, bis-, or tris-(2-hydroxyethyl)amine, 2-hydroxy-tert-butylamine, or tris-(hydroxymethyl)methylamine, N,N-di-lower alkyl-N-(hydroxyl-lower alkyl)-amines, such as N,N-dimethyl-N-(2-hydroxyethyl)amine or tri-(2-hydroxyethyl)amine; N-methyl-D-glucamine; and amino acids such as arginine, lysine, etc.

[0145] In some embodiments, the compositions described herein are in the form of a pharma- ceutically acceptable salt.

[0146] Pharmaceutical Compositions and Formulations In some embodiments, the disclosure relates to a composition, e.g., a pharmaceutical composition, comprising GM-CSF and / or an additional therapeutic agent, e.g., a therapeutic agent described herein, and a pharma- ceutically acceptable carrier or excipient.

[0147] In some embodiments, the additional therapeutic agent is a dopamine precursor, such as levodopa; a cholinesterase inhibitor, such as donepezil (ARICEPT), rivastigmine (EXELON), or galantamine (RAZADYNE); an atypical / second generation antipsychotic, including serotonin-dopamine antagonists (SDAs), multi-receptor acting antipsychotics (MARTAs), and D2 partial agonists (e.g., ABILIFY / aripiprazole); memantine, an NMDA receptor antagonist; riluzole (RILUTEK); an NSAID (nonsteroidal anti-inflammatory drug); a caffeine A2A receptor antagonist; and CERE-120 (adeno-associated virus hematologic malignancies). serum type 2-neurturin; deep brain stimulation; TNF-α antagonists including etanercept, adalimumab, and infliximab; IFN-γ inhibitors; TGF-β modulators; IL-33 inhibitors; IL-18 inhibitors; VEGF inhibitors; IL-1 inhibitors; inhibitors of pathological beta amyloid (Ab) plaques, e.g., pathological beta amyloid (Ab) specific monoclonal antibodies such as aducanumab (ADUHELM); NSAIDs such as metacetamol and aspirin; diabetes drugs such as linagliptin; inhibitors of tau activation such as liraglutide; miRNAs targeting Ab plaque formation and tau protein phosphorylation; ginkgo biloba (ginkgo alpha-secretase enhancers, such as Salvia biloba, Salvia miltiorrhiza; beta-secretase inhibitors, such as Coptis Rhizome (Huanglian) and Yuanzhi; and pharmaceutically acceptable salts, acids, or derivatives of any of the above.

[0148] Any of the pharmaceutical compositions described herein can be administered to a patient as a component of a composition that includes a pharma- ceutically acceptable carrier or vehicle. Such compositions can optionally include a suitable amount of a pharma- ceutically acceptable excipient to provide the form for proper administration.

[0149] In some embodiments, pharmaceutical excipients can be liquids, such as water and oils, such as those of petroleum, animal, vegetable, or synthetic origin, for example, peanut oil, soybean oil, mineral oil, sesame oil, and the like. Pharmaceutical excipients can be, for example, saline, gum acacia, gelatin, starch paste, talc, keratin, colloidal silica, urea, and the like. Additionally, auxiliary, stabilizer, thickening, lubricating, and coloring agents can be used. In some embodiments, the pharma- ceutically acceptable excipients are sterile when administered to a patient. Water is a useful excipient for intravenous administration of any of the agents described herein. Saline, aqueous dextrose, and aqueous glycerol solutions can also be used as liquid excipients, particularly for injectable solutions. Suitable pharmaceutical excipients also include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glyceryl monostearate, talc, sodium chloride, dried skim milk powder, glycerin, propylene, glycol, water, ethanol, etc. Any agent described herein can also contain minor amounts of wetting agents, emulsifying agents, or pH buffering agents, if necessary. Other examples of suitable pharmaceutical excipients are described in Remington's Pharmaceutical Sciences 1447-1676 (Alfonso R. Gennaro eds., 19th ed. 1995), which is incorporated herein by reference.

[0150] The present disclosure encompasses the described pharmaceutical compositions (and / or additional therapeutic agents) in a variety of formulations. Any pharmaceutical composition (and / or additional therapeutic agent) described herein may take the form of a solution, suspension, emulsion, drops, tablets, pills, pellets, capsules, capsules containing liquid, gelatin capsules, powders, sustained release formulations, suppositories, emulsions, aerosols, sprays, suspensions, lyophilized powders, freeze suspensions, dry powders, or any other form suitable for use. In some embodiments, the composition is in the form of a capsule. In some embodiments, the composition is in the form of a tablet. In yet other embodiments, the pharmaceutical composition is formulated in the form of a softgel capsule. In some embodiments, the pharmaceutical composition is formulated in the form of a gelatin capsule. In yet other embodiments, the pharmaceutical composition is formulated as a liquid.

[0151] If necessary, the pharmaceutical composition of the present invention (and / or additional therapeutic agent) can also include a solubilizing agent. The agent can also be delivered using a suitable vehicle or delivery device known in the art. The combination therapy outlined herein can be co-delivered in a single delivery vehicle or delivery device.

[0152] The formulations containing the pharmaceutical composition of the present disclosure (and / or additional therapeutic agent) may be presented in convenient unit dosage form and may be prepared by any of the methods well known in the art of pharmacy. Such methods generally include the step of bringing into association a therapeutic agent with a carrier which constitutes one or more accessory ingredients. Typically, the formulations are prepared by uniformly and intimately bringing into association a therapeutic agent with liquid carriers, finely divided solid carriers, or both, and then, if necessary, shaping the product into the desired formulation (e.g., wet or dry granulation, powder blending, etc., followed by tableting, etc., using conventional methods known in the art).

[0153] In some embodiments, any pharmaceutical composition described herein (and / or additional therapeutic agent) is formulated in accordance with routine procedures into a composition compatible with the modes of administration described herein.

[0154] Routes of administration include, for example, topical, oral, intradermal, transdermal, subcutaneous, intramuscular, intraperitoneal, intravenous, intranasal, epidural, sublingual, intranasal, intracerebral, intravaginal, intrarectal, or by inhalation. Administration can be local or systemic. In some embodiments, administration is by the intravenous route. The method of administration is at the discretion of the physician and will depend in part on the site of the medical condition. In most cases, administration will result in the release of any agent described herein onto or into the affected area.

[0155] In some embodiments, GM-CSF (and / or the additional therapeutic agent) is administered via an intravenous route.

[0156] In some embodiments, the pharmaceutical compositions (and / or additional therapeutic agents) described herein are formulated according to routine procedures into compositions suitable for administration. For intravenous administration, suitable carriers include physiological saline, bacteriostatic water, Cremophor ELTM (BASF, Parsippany, NJ), or phosphate buffered saline (PBS). The carrier must be stable under the conditions of manufacture and storage and must be preserved against microorganisms. The carrier can be, for example, a solvent or dispersion medium, such as water, ethanol, polyol (e.g., glycerol, propylene glycol, and liquid polyethylene glycol), and suitable mixtures thereof.

[0157] Dosage forms suitable for parenteral administration (e.g., intravenous, intramuscular, intraperitoneal, subcutaneous, and intraarticular injection and infusion) include, for example, solutions, suspensions, dispersions, emulsions, and the like. They can also be prepared in the form of sterile solid compositions (e.g., lyophilized compositions) that can be dissolved or suspended in a sterile injectable medium immediately before use. These may, for example, contain suspending or dispersing agents known in the art. Formulation components suitable for parenteral administration include sterile diluents such as water for injection, saline, fixed oils, polyethylene glycols, glycerin, propylene glycol, or other synthetic solvents; antibacterial agents such as benzyl alcohol or methylparabens; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as EDTA; buffers such as acetate, citrate, and phosphate; and agents for adjusting tonicity such as sodium chloride or dextrose.

[0158] The composition for oral delivery can be in the form of, for example, tablets, lozenges, aqueous or oily suspensions, granules, powders, emulsions, capsules, syrups, or elixirs.The composition for oral administration can contain one or more agents, such as sweeteners, such as fructose, aspartame, or saccharin, flavorings, such as peppermint, wintergreen oil, or cherry, coloring agents, and preservatives, for example, to provide a palatable preparation.

[0159] Compositions for topical delivery can be in the form of, for example, creams, gels, ointments, lotions, sprays, aqueous or oily suspensions, powders, or emulsions. Increased skin permeability and penetration can be achieved by non-invasive methods, for example, by the use of any nanocarrier in combination with any pharmaceutical composition (and / or additional therapeutic agent) described herein. The skin can act as a reservoir and can be used to deliver the compositions (and / or additional therapeutic agent) described herein in a sustained manner over a longer period of time.

[0160] Any pharmaceutical composition (and / or additional therapeutic agent) described herein can be administered by controlled or sustained release means or by delivery devices known to those skilled in the art. Examples include, but are not limited to, those described in U.S. Patent Nos. 3,845,770; 3,916,899; 3,536,809; 3,598,123; 4,008,719; 5,674,533; 5,059,595; 5,591,767; 5,120,548; 5,073,543; 5,639,476; 5,354,556; and 5,733,556, each of which is incorporated herein by reference in its entirety. Such dosage forms may be useful for providing controlled or sustained release of one or more active ingredients, for example, using hydropropylcellulose, hydropropylmethylcellulose, polyvinylpyrrolidone, other polymer matrices, gels, permeable membranes, osmotic systems, multi-layer coatings, microparticles, liposomes, microspheres, or combinations thereof to provide desired release profiles at various ratios. Suitable controlled or sustained release formulations known to those skilled in the art, including those described herein, can be easily selected for use with the active ingredients of the medicaments described herein. Thus, the present disclosure provides single unit dosage forms suitable for oral administration, such as, but not limited to, tablets, capsules, gelcaps, and caplets adapted for controlled or sustained release.

[0161] Controlled or sustained release of an active ingredient can be stimulated by various conditions including, but not limited to, changes in pH, changes in temperature, stimulation with light of appropriate wavelengths, enzyme concentration or availability, water concentration or availability, or other physiological conditions or compounds.

[0162] In other embodiments, a controlled release system can be placed in the vicinity of the therapeutic target area, thereby requiring only a fraction of the systemic dose (see, e.g., Goodson, in Medical Applications of Controlled Release, supra, vol. 2, pp. 115-138 (1984)). Other controlled release systems discussed in the review by Langer, 1990, Science 249:1527-1533 may also be used.

[0163] The pharmaceutical preparation is preferably sterile. Sterilization can be accomplished, for example, by filtration through sterile filtration membranes. If the composition is lyophilized, filter sterilization can be performed prior to or after lyophilization and reconstitution.

[0164] Usage / Dosage It will be understood that the actual dose of the composition administered according to the present disclosure will vary depending on the specific dosage form and mode of administration. Those skilled in the art can take into account many factors that may modify the action of the composition (e.g., body weight, sex, diet, administration time, administration route, excretion rate, patient condition, drug combination, genetic disposition, and reaction sensitivity). Administration can be performed continuously or in one or more separate doses within the maximum tolerated dose range. The optimal administration rate for a given group of conditions can be ascertained by those skilled in the art using conventional dose administration tests.

[0165] In some embodiments, GM-CSF is administered in a total dose of about 125 μg, about 150 μg, or about 200 μg, or about 250 μg, or about 300 μg, or about 350 μg. In some embodiments, GM-CSF is administered in a total dose of about 250 μg.

[0166] In some embodiments, GM-CSF is administered at a dose of about 125 μg, about 150 μg, or about 200 μg, or about 250 μg, or about 300 μg, or about 350 μg.

[0167] In some embodiments, GM-CSF is administered twice daily.

[0168] In some embodiments, the GM-CSF is sargramostim and is administered at a dose of about 125 μg twice daily.

[0169] Combination and additional therapeutic agents In some embodiments, the pharmaceutical compositions of the present disclosure are co-administered in combination with an additional agent, such as a neurological agent and / or an additional therapeutic agent. Co-administration can be simultaneous or sequential.

[0170] In some embodiments, the additional neurological agent and / or additional therapeutic agent of the present disclosure and GM-CSF are administered to the patient simultaneously. As used herein, the term "simultaneously" means that the neurological agent and / or additional therapeutic agent and GM-CSF are administered at a time interval of about 60 minutes or less, e.g., about 30 minutes or less, about 20 minutes or less, about 10 minutes or less, about 5 minutes or less, or about 1 minute or less. The administration of the neurological agent and / or additional therapeutic agent and GM-CSF can be by simultaneous administration of a single formulation (e.g., a formulation including the additional therapeutic agent and a GM-CSF composition) or separate formulations (e.g., a first formulation including the neurological agent and / or additional therapeutic agent and a second formulation including a GM-CSF composition).

[0171] Co-administration does not require that the therapeutic agents be administered simultaneously, provided that the timing of administration is such that the pharmacological activity of the neurological agent and / or additional therapeutic agent and GM-CSF overlaps in time, thereby providing a combined therapeutic effect. For example, the neurological agent and / or additional therapeutic agent and the targeting moiety, GM-CSF composition can be administered sequentially. As used herein, the term "sequentially" means that the neurological agent and / or additional therapeutic agent and GM-CSF are administered at a time interval of more than about 60 minutes. For example, the time between the sequential administration of the neurological agent and / or additional therapeutic agent and GM-CSF can be more than about 60 minutes, more than about 2 hours, more than about 5 hours, more than about 10 hours, or more than about 1 day, more than about 2 days, more than about 3 days, more than about 1 week, more than about 2 weeks, or more than about 1 month. The optimal administration time will depend on the rate of metabolism, excretion, and / or pharmacodynamic activity of the additional therapeutic agent and GM-CSF administered. Either the neurological agent and / or additional therapeutic agent, or the GM-CSF composition may be administered first.

[0172] Co-administration also does not require that the therapeutic agents be administered to a patient by the same route of administration. Rather, each therapeutic agent can be administered by any suitable route, which may or may not be parenteral.

[0173] In some embodiments, the GM-CSF described herein acts synergistically when co-administered with a neurological agent and / or additional therapeutic agent. In such embodiments, the GM-CSF composition and the neurological agent and / or additional therapeutic agent may be administered at lower doses than would be used if the agent were used in a monotherapy setting.

[0174] Biological samples In some embodiments, the biological sample is selected from a biopsy specimen, a tissue, and / or a bodily fluid.

[0175] In some embodiments, the biological sample is selected from blood, a skin or tissue sample, a tissue biopsy specimen, a formalin-fixed or paraffin-embedded tissue specimen, a cytological sample, cultured cells, plasma, serum, pus, urine, sweat, tears, mucus, sputum, saliva, and / or other bodily fluids.

[0176] In some embodiments, the biological sample is selected from blood, a skin or tissue sample, plasma, serum, pus, urine, sweat, tears, mucus, sputum, saliva, cerebrospinal fluid (CSF) and / or other bodily fluids.

[0177] In some embodiments, the biological sample is or comprises monocytes, hi some embodiments, the biological sample is or comprises a monocyte population.

[0178] In some embodiments, the biological sample is peripheral blood lymphocytes (PBLs), for example, peripheral blood lymphocytes (PBLs) isolated by leukapheresis and centrifugal elutriation.

[0179] In some embodiments, the biological sample is a lymphocyte population, for example a lymphocyte population isolated by leukapheresis and centrifugal elutriation.

[0180] kit The present disclosure also provides kits that can detect the presence or absence of the biomarkers described herein.

[0181] Exemplary kits of the invention include various reagents, including, for example, agents for detecting the biomarkers described herein. In some embodiments, the kits include one or more detection reagents, including those useful in the various detection methods described herein. In some embodiments, the kits include materials necessary for the evaluation, including, for example, plates with wells, syringes, etc. In some embodiments, the kits further include a label or printed instructions directing the use of the described reagents.

[0182] In some embodiments, kits are provided for measuring the present biomarkers in biological samples. In some embodiments, the kits include a multi-well sample plate coated with immobilized capture antibodies that bind to the biomarkers; a detection antibody that is covalently linked to an enzyme and also binds to the biomarker; a colored or fluorescent product catalyzed by the enzyme bound to the detection antibody; and an appropriate buffer. In some embodiments, the kits have an ELISA plate specific for detecting one of the biomarkers. In some embodiments, the kits detect one or more of LRRK2, HMOX1, TLR2, TLR8, RELA, ATG7, and GABARAPL2, for example, about 1, or about 2, or about 3, or about 4, or about 5.

[0183] In some embodiments, a companion diagnostic, complementary diagnostic, or co-diagnostic test kit is provided that includes: (a) an array of nucleic acids or proteins suitable for detection of one or more, e.g., about 1, or about 2, or about 3, or about 4, or about 5, about 10, about 15, or about 20, of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA, ATG3, ATG7, and GABARAPL2; and (b) instructions for use.

[0184] In some embodiments, companion, complementary, or co-diagnostic test kits are provided that contain reagents and instructions for use in one or more of the methods described herein.

[0185] array SEQ ID NO:1 is the amino acid sequence of wild-type GM-CSF:

[0186] APARSPSPSTQPWEHVNAIQEARRLLNLSRDTAAEMNETVEVISEMFDLQEPTCLQTRLELYKQGLRGSLTKLKGPLTMMASHYKQHCPPTPETSCATQIITFESFKENLKDFLLVIPFDCWEPVQE

[0187] SEQ ID NO:2 is the amino acid sequence of sargramostim:

[0188] APARSPSPSTQPWEHVNAIQEALRLLNLSRDTAAEMNETVEVISEMFDLQEPTCLQTRLELYKQGLRGSLTKLKGPLTMMASHYKQHCPPTPETSCATQIITFESFKENLKDFLLVIPFDCWEPVQE

[0189] SEQ ID NO:3 is the amino acid sequence of molgramostim:

[0190] APARSPSPSTQPWEHVNAIQEARRLLNLSRDTAAEMNETVEVISEMFDLQEPTCLQTRLELYKQGLRGSLTKLKGPLTMMASHYKQHCPPTPETSCATQIITFESFKENLKDFLLVIPFDCWEPVQE

[0191] SEQ ID NO:4 is the amino acid sequence of regramostim:

[0192] APARSPSPSTQPWEHVNAIQEARRLLNLSRDTAAEMNETVEVISEMFDLQEPTCLQTRLELYKQGLRGSLTKLKGPLTMMASHYKQHCPPTPETSCATQTTFESFKENLKDFLLVIPFDCWEPVQE

[0193] definition The following definitions are used in connection with the invention disclosed herein: Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0194] An "effective amount," when used in connection with an agent effective to treat a coronavirus infection, is an amount effective to treat or ameliorate a coronavirus infection.

[0195] As used herein, "a," "an," or "the" can mean one or more. Additionally, the term "about," when used in connection with a referenced numerical indication, means the referenced numerical indication plus or minus up to 10% of the referenced numerical indication. For example, the language "about 50" covers a range from 45 to 55.

[0196] As referred to herein, all composition percentages are by weight of the total composition unless otherwise specified. As used herein, the word "include" and its variations are intended to be open-ended and that the description of items in a list does not exclude other items that may be useful in the materials, compositions, devices, and methods of this technology. Similarly, the words "can" and "may" and its variations are intended to be open-ended and that a description that an embodiment can or may include certain elements or features does not exclude other embodiments of this technology that do not include those elements or features.

[0197] Although the open-ended term "comprising" is used herein to describe and claim the present invention as a synonym for terms such as including, containing, or having, the present invention or embodiments thereof may alternatively be described using alternative terms such as "consisting of" or "essentially consisting of." EXAMPLES

[0198] Example 1: Monocyte proteomic profile following sargramostim treatment To understand in detail the changes in monocyte proteomic profile after sargramostim treatment, functional enrichment and pathway enrichment analyses of differentially regulated proteins were performed after 2 and 6 months of treatment. At 2 months, sargramostim treatment enriched multiple immune processes, including myeloid leukocyte-mediated immunity (p=1.33E-08), myeloid cell activation associated with immune response (p=1.30E-07), and leukocyte activation associated with immune response (p=7.25E-06) (Figure 1A). Kyoto Encyclopedia of Genes and Genomes (KEGG) and Reactome analyses showed enrichment of phagosome (p=2.74E-04) and innate immune system (p=1.29E-07), respectively (Figure 1A). In addition, enrichment of inflammatory processes was observed, including regulation of interleukin-8 (IL-8) (p=3.64E-02) and tumor necrosis factor (TNF) production pathways (p=1.73E-02) (Figure 1A). Furthermore, Ingenuity Pathway Analysis (IPA) revealed inhibition of neuroinflammatory signaling pathways (p=1.43E-04), IL-8 pathway (p=8.8E-04), integrin-linked kinase (ILK) pathway (p=2.8E-02), and nitric oxide and reactive oxygen species (ROS) pathways (p=3.35E-02) (Figure 1A). Sargramostim also affected endosome (p=3.14E-02), Golgi vesicle (p=1.60E-03), endoplasmic reticulum-Golgi vesicle-mediated (p=1.02E-02), and lysosome targeting of proteins (p=2.78E-02), as well as regulation of late endosome-to-lysosome transport (p=3.88E-03) (Figure 1A). Cellular component and reactome analysis showed enrichment of secretory vesicles (p=1.14E-13), endocytic vesicles (p=2.37E-05), transport vesicles (p=1.18E-03), phagosomes (p=5.16E-03), Golgi-associated vesicles (p=9.24E-03), lysosomes (p=2.59E-09), and trans-Golgi network vesicle budding (p=4.19E-02) (Figure 1A).Sargramostim treatment also induced changes in biological processes related to RNA processing, such as regulation of mRNA processing (p=2.16E-05) and regulation of RNA splicing (p=3.96E-04) (Figure 1A). Similarly, KEGG and Reactome tests showed pathways affecting RNA processing (including RNA transport, p=3.70E-02), mRNA splicing (major and minor pathways, p=3.54E-05 and p=4.31E-02, respectively), and spliceosome (1.24E-03) (Figure 1A).

[0199] Proteomic and scRNA-seq analysis after 6 months of sargramostim treatment also showed enrichment of immune processes including monocyte-macrophage mediated immunity (p=6.03E-34), activation (p=2.29E-35), and innate immune response (p=2.91E-27). In addition, regulation of leukocyte activation (p=5.05E-03), lymphocyte chemotaxis (p=5.02E-03), and regulation of lymphocyte activation (p=6.540E-04) were shown (Figure 2A). Similarly, proteomic Reactome analysis showed enrichment of innate immunity (p=3.66E-19) and phagosome formation (p=3.28E-02) (Figure 2A and Figure 2B). IPA of the scRNA-seq dataset after 6 months showed enrichment for chemokine signaling (p=1.84E-04), dendritic cell maturation (p=2.03E-04), and phagosome formation (p=3.73E-07) (Figure 2A and Figure 2B). In addition, enrichment for regulation of ROS metabolism (p=3.22E-03), oxidative stress (p=7.07E-04), and oxygen-containing compounds (p=1.16E-03) was also observed (Figure 2A). These data suggest an antioxidant effect after sargramostim treatment. Interestingly, IPA of the proteomics data after 6 months showed inhibition of oxidative phosphorylation (p=1.36E-06). This suggests that regulation of ROS production may occur after sargramostim treatment (Figure 2B). Furthermore, the proteomic data after 6 months showed enrichment of mitochondrial function (p=8.26E-40) as organelles responsible for the respiratory electron transport pathway (p=5.29E-15) enriched in the Reactome analysis (Figure 2A). In addition, cellular component analysis of both the proteomic and scRNA-seq datasets showed enrichment of secretory vesicles (p=1.59E-38) and cytoplasmic vesicles (p=6.56E-03) (Figure 2A). The proteomic dataset after 6 months showed enrichment of autophagy (p=4.63E-04) and macroautophagy (1.51E-05) (Biological Process Analysis), and activation of the sirtuin signaling pathway (IPA, p=2.19E-11) (Figure 2A and Figure 2B).After 6 months, scRNA-seq data also showed enrichment for pathways related to the control of inflammatory responses (p=5.87E-03), with IL-10 negatively regulating cell membrane-associated inflammatory mediators (p=9.67E-03) and the receptor ACKR2 binding to most inflammatory CC chemokines (p=3.16E-03) (Figure 2A).

[0200] Example 2: Gene and Protein Expression Following Sargramostim Treatment Gene and protein expression was evaluated as biomarkers for predicting disease progression and treatment response after sargramostim treatment using ddPCR and Western blotting. Individual subjects' baseline and treatment protein expression of LRRK2, HMOX1, TLR2, TLR8, RELA, ATG7, and GABARAPL2 were compared 2 and 6 months after initiation of sargramostim treatment. At 2 months after initiation of sargramostim, 4 / 5 patients showed significant reductions in LRRK2 and HMOX1 protein expression, and 3 / 5 patients showed significant reductions in TLR2 protein expression (Figure 3A). Additionally, 3 / 5 patients showed reductions in TLR8 and NF-κB protein expression, with 1 / 3 patients showing significant downregulation: subject 2005 for TLR8 and subject 2003 for NF-κB (Figure 3A). Furthermore, one patient showed increased expression of ATG7 protein (subject 2003), and in the same subject, GABARAPL2 protein expression was significantly increased (Figure 3A). At 6 months, 4 / 5 patients showed significant downregulation of LRRK2 and TLR2 (Figure 3B). Of note, 5 / 5 patients showed decreased protein expression of HMOX1. Said downregulation was significant in 3 / 5 patients (Figure 3B). Similarly, 5 / 5 patients showed decreased NF-κB expression, and said downregulation was significant in 4 / 5 patients (Figure 3B). Furthermore, downregulation of TLR8 protein was observed in 2 / 5 patients (Figure 3B). Only subject 2003 showed limited increased expression of ATG7 protein, while 3 / 5 patients showed upregulation of GABARAPL2, approaching significance in 2 / 3 patients (subjects 2005 and 2006) (Figure 3B). At the gene level, we compared individual patients' baseline and treatment gene expression of LRRK2, HMOX1, TLR2, TLR8, RELA, ATG7, and GABARAPL2 at 2 and 6 months after the start of sargramostim treatment. At 2 months, 5 / 5 patients showed significant decreases in LRRK2 and HMOX1 gene expression, and 3 / 5 patients showed significant decreases in TLR2 and TLR8 gene expression after sargramostim treatment (Figure 3C).Although 4 / 5 patients showed a significant decrease in gene expression of NF-κB, the downregulation did not exceed 15% compared to baseline (Figure 3C). Interestingly, 4 / 5 patients showed a significant upregulation of ATG7 gene, but in 2 / 4 subjects (subjects 2005 and 2006), the upregulation did not exceed 17% compared to baseline (Figure 3C). Similarly, 3 / 5 patients showed a significant upregulation of GABARAPL2 gene, but the upregulation did not exceed 17% compared to baseline in 3 subjects (Figure 3C). At 6 months, 5 / 5 patients showed a significant downregulation of HMOX1 and TLR2 genes (Figure 3D). 3 / 5 and 4 / 5 patients showed a significant downregulation of LRRK2 and TLR8 genes, respectively (Figure 3D). Only 2 / 5 patients showed a significant decrease in gene expression of NF-κB, with a downregulation not exceeding 5% compared to baseline (Figure 3D). Interestingly, 4 / 5 patients showed significant upregulation of the ATG7 gene (Figure 3D). According to ddPCR data at 2 months, 3 / 5 patients showed a significant increase in gene expression of GABARAPL2, although in 2 / 3 subjects the upregulation did not exceed 6% compared to baseline (subjects 2004 and 2005) (Figure 3D).

[0201] Example 3: Integrated scRNA-seq and proteomics data following sargramostim treatment The overlapping genes between the scRNA-seq and proteomics datasets for subjects 2003, 2004, and 2005 are shown in a Venn diagram showing the number of genes identified in each dataset and the number of overlapping genes in both datasets (Figure 4A). The Pearson product-moment correlation coefficient (r=0.3582, p=8.9627E-77) between the overlapping genes in scRNA-seq and proteomics showed a significant moderate positive association between the overlapping genes in both datasets (Figure 4B). This indicates that the expression of multiple genes in both datasets was changed in the same direction (up- or down-regulated).

[0202] Example 4: Correlation of clinical MDS-UPDRSIII scores by Western blot and ddPCRCorrelation between the expression (gene or protein level) of potential biomarkers and the MDS-UPDRSIII score of PD patients may be used to identify the possibility of using clinical biomarkers to predict disease progression and treatment response after therapy with immune modulators such as sargramostim. Therefore, we performed a correlation analysis between the raw score and its change in the MDS-UPDRSIII score and the protein levels of selected biomarkers. A positive correlation was shown between the change in the MDS-UPDRSIII score and LRRK2 (r=0.3534, p=0.017), HMOX1 (r=0.0881, p=0.565), TLR8 (r=0.1531, p=0.315), and NF-κB (r=0.4258, p=0.004) (Figure 5A), suggesting that a decrease in the expression of any of these proteins improves motor function. Additionally, a negative correlation was shown between the change in MDS-UPDRSIII score and the protein levels of ATG7 (r = -0.2440, p = 0.106) and GABARAPL2 (r = -0.1376, p = 0.367) (Figure 5A). Similarly, a positive correlation was shown between the raw MDS-UPDRSIII scores and the protein levels of LRRK2 (r = 0.2659, p = 0.077), HMOX1 (r = 0.1109, p = 0.468), TLR2 (r = 0.3704, p = 0.012), and NFκB (r = 0.1847, p = 0.224) (Figure 5B), while a negative correlation was shown between the raw MDS-UPDRSIII scores and the protein levels of ATG7 (r = -0.3002, p = 0.045) and GABARAPL2 (r = -0.1110, p = 0.468) (Figure 5B). Both LRRK2 and HMOX1 were able to predict the change in MDS-UPDRSIII score with 12.5% ​​of the variance of the change in MDS-UPDRSIII score (Figure 5C). However, in this model, LRRK2 exerted a stronger influence than HMOX1 (β=0.3594, p=0.0222). The effect of LRRK2 and NF-κB was significant (p=0.0139), with 18.4% of the change in MDS-UPDRSIII value (Figure 5C).The effects of NF-κB and GABARAPL2 showed a significant effect on the change in MDS-UPDRSIII score (p=0.0128) and a 18.7% effect on the change in MDS-LIPDRSIII score (Figure 5C). The effects of TLR2 and TLR8 showed no significant effect on the change in MDS-UPDRSIII score (p=0.567) (Figure 5C), and TLR8 and ATG7 showed no effect on the change in MDS-UPDRSIII score (p=0.2403) (Figure 5C). Further analysis of the impact on raw MDS-UPDRSIII scores showed significant impacts by LRRK2 / HMOX1, LRRK2 / NF-κB, and NF-κB / GABARAPL2 (p=0.2093, 0.2123, and 0.4761, respectively) (Figure 5D), while the other two pairs of predictors (TLR2 / TLR8 and TLR8 / ATG7) showed significant impacts on raw MDS-UPDRSIII scores (p=0.0314 and p=0.0046, respectively) (Figure 5D).

[0203] Similarly, correlations were obtained between the MDS-UPDRSIII score and gene expression of selected biomarkers. This included positive correlations between the change in MDS-UPDRS-III score and LRRK2 (r=0.2469, p=0.057), HMOX1 (r=0.3193, p=0.013), TLR2 (r=0.4388, p=0.000), and TLR8 (r=0.0385, p=0.770) (Figure 6A). Each showed a relationship between decreased gene expression and improved motor function. Negative correlations were found between the change in MDS-UPDRSIII score and ATG7 (r=-0.5662, p=0.000) and GABARAPL2 (r=-0.0360, p=0.785) (Figure 6A). Each showed a relationship between increased gene expression and improved motor function. Similarly, replicate correlations were found between raw MDS-UPDRSIII scores and LRRK2 (r = 0.3299, p = 0.010), HMOX1 (r = 0.1598, p = 0.223), and TLR2 (r = 0.3665, p = 0.004) (Figure 6B). Negative correlations were found between raw MDS-UPDRSIII scores and ATG7 (r = -0.5960, p = 0.000) and GABARAPL2 (r = -0.0115, p = 0.930) (Figure 6B). The data demonstrated that both predictors were significant (p = 0.0336) with 11.2% of the variance in MDS-UPDRSIII score change (Figure 6C). HMOX1 was a stronger predictor (β=0.2611, p=0.0747), although it did not reach significance. Each unit increase in ddPCR HMOX1 expression increased the change in MDS-UPDRSIII score by 0.0409 points. The effects of LRRK2 and TLR2 were significant (p=0.0012), with a change in MDS-UPDRSIII value of 21% (Figure 6C). TLR2 had a greater effect than LRRK2 (β=0.4013, p=0.0018). The model predicted that a unit increase in ddPCR TLR2 expression would increase the change in MDS-UPDRSIII score by 0.0503 points.On the other hand, LRRK2 and TLR8 did not show a significant effect on the change in MDS-UPDRSIII score (p=0.1449) (Figure 6C). However, LRRK2 and ATG7 were associated with the change in MDS-UPDRSIII score (p=1.12E-05), with a 33% effect on the score change (Figure 6C). ATG7 had a greater effect than LRRK2 (P=-0.6326, p=0.0000), with a 1-unit increase in ddPCR relative expression of ATG7 affecting the MDS-UPDRSIII score change by 0.0400 points. The effect of TLR2 and TLR8 was significant (p=0.0009), with a 21.8% change in the MDS-UPDRSIII score (Figure 6C). TLR2 had a more significant effect than TLR8 (β=0.5124, p=0.0002). The model predicts that a one unit increase in ddPCR relative expression of TLR2 will increase the change in MDS-UPDRSIII score by 0.0643 points. TLR2 and ATG7 were significantly associated with the change in MDS-UPDRSIII score (p=1.11E-05), with an influence of 33% of the score (Figure 6C). ATG7 had the largest influence on the change in MDS-UPDRSIII score over TLR2 (β=-0.4850, p=0.0012). Here, a one unit increase in ddPCR of ATG7 decreased the change in MDS-UPDRSIII score by 0.0307 points. TLR8 and ATG7 also showed a significant influence on the change in MDS-UPDRSIII score (p=3.8E-06), with an influence of 35.5% (Figure 6C).

[0204] Equivalent Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the embodiments specifically described herein which equivalents are intended to be encompassed by the following claims.

[0205] Incorporation by Reference All patents and publications referenced herein are incorporated by reference in their entirety.

[0206] As used herein, all headings are for organizational purposes only and are not intended to limit the disclosure in any way. The contents of the individual sections are equally applicable to all sections.

Claims

1. A reagent and instructions for use, The reagents include HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, A comprising a nucleic acid or protein suitable for the detection of one or more of TP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7 and GABARAPL2, optionally one or more of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2 and GABARAPL2, A companion, complementary, or co-diagnostic test kit for selecting patients for drug treatment for Parkinson's disease.

2. The kit described in claim 1, wherein the drug for Parkinson's disease is granulocyte-macrophage colony-stimulating factor (GM-CSF). (i) one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFB1, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, and SDHA, which are downregulated during or after treatment with GM-CSF; optionally, one or more of HMOX1, TLR2, TLR8, RELA, and LRRK2, which are downregulated during or after treatment with GM-CSF; and / or (ii) one or more of ATG3, ATG7, and GABARAPL2 that are upregulated during or after treatment with GM-CSF; optionally, one or more of ATG7 and GABARAPL2 that are upregulated during or after treatment with GM-CSF; 2. The kit of claim 1, suitable for detecting

4. 2. The kit of claim 1, wherein the patient has one or more of the following symptoms: oxidative stress, loss of neurite integrity, apoptosis, neuron loss, and / or inflammatory response, cognitive impairment, cognitive decline, behavioral and personality changes, tremors, bradykinesia, rigidity, impaired posture and balance, loss of automatic movements, decreased coordination, changes in speech, photophobia, difficulty controlling eye muscles, slowed saccadic eye movements, difficulty swallowing, blepharospasm, fainting or lightheadedness due to orthostatic hypotension, dizziness, bladder control problems, well-formed hallucinations and delusions, changes in memory, concentration, and judgment, memory loss, depression, irritability, anxiety, rapid eye movement (REM) sleep disorder, seizures, paresthesia, numbness or tingling, convulsions, difficulty chewing or swallowing, muscle spasms and weakness in the limbs, and / or prickling or tingling in the feet or hands.

5. The kit described in claim 1, wherein the detection is detection of a protein and / or a nucleic acid.

6. The detection of HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS, wherein the detection is determined by one or more of ELISA, Luminex multiplex assay, immunohistochemical staining, Western blotting, in-cell Western, immunofluorescence staining, or fluorescence-activated cell sorting (FACS). 41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, optionally one or more of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2.

7. The detection is performed by droplet digital PCR (ddPCR), reverse transcriptase PCR analysis, quantitative real-time PCR, single-strand conformation polymorphism analysis (SSCP), mismatch cleavage detection, heteroduplex analysis, deoxyribonucleic acid (DNA) sequencing, ribonucleic acid (RNA) sequencing, Northern blot analysis, in HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1 as determined by one or more of in situ hybridization, array analysis, and restriction fragment length polymorphism analysis. , KRT10, ACAA1, PCK2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, optionally one or more of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2.

8. The detection of HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, K, is determined by single-cell RNA sequencing and / or next-generation sequencing (NGS).

2. The kit of claim 1, wherein the kit is for detecting the presence, absence, or amount of one or more of RT10, ACAA1, PCK2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, optionally one or more of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2.

9. The detection of HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, G, 9. The kit of claim 8, wherein the kit is for detecting the presence, absence, or amount of one or more of LB1, KRT10, ACAA1, PCK2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, optionally one or more of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2.

10. A kit as described in claim 1, suitable for detection in biological samples including blood, skin or tissue samples, plasma, serum, pus, urine, sweat, tears, mucus, sputum, saliva, cerebrospinal fluid (CSF), and / or other bodily fluids.

11. A kit as described in claim 1, suitable for detection in a biological sample containing monocytes.

12. 3. The kit of claim 2, wherein the GM-CSF has the amino acid sequence of SEQ ID NO:1 or a variant that is at least 90%, or at least 93%, or at least 95%, or at least 97%, or at least 98% identical to the amino acid sequence of SEQ ID NO:

1.

13. 3. The kit of claim 2, wherein the GM-CSF has the amino acid sequence of one of SEQ ID NO:2, SEQ ID NO:3, and SEQ ID NO:4, or a variant having at least 90%, or at least 93%, or at least 95%, or at least 97%, or at least 98% identity to the amino acid sequence of one of SEQ ID NO:2, SEQ ID NO:3, and SEQ ID NO:

4.

14. 13. The kit of claim 12, wherein the GM-CSF is one of molgramostim, sargramostim, and regramostim.

15. The kit of claim 14, wherein the GM-CSF is sargramostim.

16. 3. The kit of claim 2, wherein the GM-CSF is suitable for administration by subcutaneous administration.

17. HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, ATP5F1D, AT a multi-well sample plate coated with immobilized capture antibodies that bind to one or more of P5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, optionally one or more of HMOXl, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2; Covalently linked to the enzymes HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, and ACAA1 , PCK2, ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, optionally also binding to one or more of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2; a colored or fluorescent product catalyzed by the enzyme bound to the detection antibody; and / or Suitable buffer The kit according to any one of claims 1 to 15, comprising:

18. HMOX1, TLR2, TLR8, RELA, IKBGG, ATG3, ATG7, leucine-rich repeat serine / threonine protein kinase 2 (LRRK2), GABARAPL2, RCOR1, GGA3, ALDH1A1, RFC1, BTF3L4, WBP2, EEA1, NCBP2, PEA15, MCM5, CLTA, VPS41, SRSF4, H2AFX, CD9, RFLNB, GLB1, KRT10, ACAA1, PCK2, ATP5F1D, ATP5 PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, and optionally one of HMOX1, TLR2, TLR8, RELA, ATG7, LRRK2, and GABARAPL2.

19. The kit of claim 18, wherein the ELISA plate is specific for detecting one, two, three, four, or five of LRRK2, HMOX1, TLR2, TLR8, RELA, ATG7, and GABARAPL2.

20. (a) an array of nucleic acids or proteins suitable for detecting one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, IKBGG, and ATP5F1D, ATP5PB, ATP5PF, ATP5PO, COX5B, COX7C, NDUFA2, NDUFBl, NDUFB4, NDUFS2, NDUFS3, NDUFS6, NDUFS7, SDHA, ATG3, ATG7, and GABARAPL2, optionally one or more of HMOX1, TLR2, TLR8, RELA, LRRK2, ATG7, and GABARAPL2; (b) instructions for use; and A companion diagnostic, complementary diagnostic, or co-diagnostic test kit comprising:

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