Nitroxoline for use in the treatment or prevention of malignant peripheral nerve sheath tumors

Abstract

The present invention relates to a composition comprising nitroxoline or a pharmaceutically acceptable salt thereof for use in the treatment or prevention of MPNST.

Description

[Technical Field]

[0001] The present invention relates to a new use of nitroxoline. [Background technology]

[0002] Nitroxoline has been used as an antibiotic in humans and, although not widely used, has been commercially available since the 1960s. It is used to treat or prevent biofilm infections, such as urinary tract infections. It is particularly effective at disrupting biofilms, and this effect is thought to be due to its metal cation chelating properties. Nitroxoline is metabolized in the liver to its corresponding sulfate and glucuronide metabolites. There is evidence that both of these metabolites share antibacterial activity. It is also used in anticancer therapy via its antiproliferative effects. Nitroxoline has the systematic name 5-nitroquinolin-8-ol.

[0003] Malignant peripheral nerve sheath tumor (MPNST) is a form of cancer of the connective tissue that surrounds nerves. A sarcoma is defined as an MPNST if it meets at least one of the following criteria: it arises from a peripheral nerve, arises from a pre-existing benign nerve sheath tumor (neurofibroma), or shows Schwann cell differentiation on histological examination. MPNST is considered aggressive and is associated with poor survival rates.

[0004] MPNSTs usually present as an expanding, palpable mass. Pain is a variable complaint. Rapid expansion occurs more frequently in the setting of NF1, raising concern for malignant degeneration of the neurofibroma. MPNSTs arising from peripheral nerves can result in a variety of clinical patterns, including radicular pain, paresthesia, and motor dysfunction.

[0005] Many MPNSTs arise from plexiform neurofibromas (benign nerve sheath tumors), which themselves arise from cutaneous nerves during early development or from more internal nerve bundles such as cranial nerves or proximal large peripheral nerve sheaths. Plexiform neurofibromas have a 10-15% lifetime incidence of transformation to MPNSTs.

[0006] Many people with MPNST are also found to have neurofibromatosis type 1 (NF1), an autosomal dominant inherited disorder. NF1 is caused by germline mutations in the NF1 tumor suppressor gene, which encodes a protein called neurofibromin. Neurofibromin functions as a GTPase-activating (GAP) protein, inactivating the intracellular signaling protein Ras by converting its active GTP-bound form to its inactive GDP-bound form. This, in turn, leads to downregulation of Ras activity. Loss of neurofibromin activity increases Ras activity, which, in turn, promotes the transcription of numerous genes required for cell growth and proliferation.

[0007] Longitudinal imaging studies of patients with plexiform neurofibromas have identified a subset of distinct nodular lesions that emerge within preexisting plexiform neurofibromas, grow rapidly relative to the surrounding tumor, and show uptake on FDG-PET [Evans, DG, et al., J Med Genet, 2002. 39(5): pp. 311-314]. Biopsies often reveal atypical neurofibromatous neoplasms (ANNUBPs) of uncertain biological potential, which share overlapping histopathological features with MPNSTs and have been implicated as potential MPNST precursors [Miettinen, MM, et al., Hum Pathol, 2017. 67: pp. 1-10]. 9p21.3 deletions, encoding the entire CDKN2A / B locus (INK4 / ARF locus), have been identified as a solitary, highly recurrent genetic abnormality in the majority (94%, n=15 / 16) of human ANNUBPs. Furthermore, CDKN2A(p16 INK4A ) and its alternative reading frame p14 ARF Haploinsufficiency or homozygous loss of α-glucan has been identified in 60-80% of MPNSTs in two recent independent studies [Brohl, AS, et al., Sci Rep, 2017.7(1):p.14992, Lee, W., et al., Nat Genet, 2014.46(11):p.1227-32].

[0008] The Ink4a / Arf tumor suppressor is crucial for suppressing plexiform neurofibroma progression by inducing broad features of senescence. Mice carrying conditional ablation of Nf1 and Ink4a / Arf in the embryonic neural crest spontaneously develop tumors histopathologically indistinguishable from human ANNUBP, which then progress to MPNST with high penetrance [Rhodes, SD, et al., Hum Mol Genet, 2019]. This tissue lineage-specific model is the first to recapitulate the malignant transformation of pre-existing plexiform neurofibromas and ANNUBP precursor lesions, as seen in human patients.

[0009] Current MPNST treatments, including surgery and chemoradiotherapy, are debilitating and largely ineffective. After 10 years, overall survival is only about 50% for patients with NF-1-associated MPNST. This highlights the need for new treatments. Summary of the Invention

[0010] The present inventors found that nitroxoline inhibits cell proliferation and increases apoptosis in stem cells that recapitulate transformed Nf1 MPNST in vitro, and thus nitroxoline is expected to alleviate, treat, and prevent MPNST.

[0011] Thus, the present invention is a composition comprising nitroxoline or a pharmaceutically acceptable salt thereof for use in treating or preventing MPNST.

[0012] A first aspect of the present invention is a composition comprising nitroxoline or a pharmaceutically acceptable salt thereof for use in the treatment or prevention of MPNST.

[0013] A second aspect of the invention is the use of nitroxoline or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for use in the treatment or prevention of MPNST.

[0014] A third aspect of the present invention provides a method for treating or preventing MPNST, comprising administering to a patient a composition comprising nitroxoline or a pharmaceutically acceptable salt thereof. [Brief explanation of the drawings]

[0015] [Figure 1] 1 shows the dose response of nitroxoline on proliferation and apoptosis of Nf1 − / − and Nf1 − / −;Ink4a / Arf+ / − DNSCs in an in vitro assay. DETAILED DESCRIPTION OF THE INVENTION

[0016] In the present invention, as demonstrated by the data herein, nitroxoline inhibits cell proliferation and increases apoptosis in stem cells that recapitulate transformed Nf1 MPNST in vitro, and is therefore an effective treatment for MPNST. Preferably, nitroxoline is used to treat or prevent MPNST in subjects with neurofibromatosis type 1.

[0017] As used herein, the term "treatment" or "treating" refers to therapeutic (curative) treatment, including reducing the size of MPNST. A biopsy may be used to diagnose MPNST. As used herein, the term "prevention" or "preventing" refers to "prophylactic" treatment, including administering nitroxoline to a patient, e.g., a patient with plexiform neurofibroma but who has not yet developed MPNST, to avoid the development of MPNST. Plexiform neurofibromas may have begun to grow, e.g., grow rapidly.

[0018] "Patient" and "subject" are used interchangeably and refer to a subject receiving nitroxoline. Preferably, the subject is a human. Suitably, the subject has neurofibromatosis, preferably neurofibromatosis type I.

[0019] In one embodiment, nitroxoline is used to treat or prevent MPNST and the patient has undergone or will undergo surgery to remove some or all of the MPNST. This may be particularly advantageous when the MPNST is large and / or has spread beyond tissue boundaries, making it difficult to remove all of it by surgery and / or where rapid removal of at least some of it is desirable / beneficial.

[0020] In one embodiment, nitroxoline is used to treat or prevent MPNST and the patient has undergone or will undergo surgery to remove some or all of a plexiform neurofibroma. Typically, in this embodiment, the plexiform neurofibroma has not evolved into a MPNST.

[0021] The term "surgery" has its usual meaning in the art. Surgery is an invasive technique based on physical intervention into an organ / organ system / tissue for diagnostic or therapeutic reasons.

[0022] In one embodiment, the patient has a 9p21.3 deletion encoding the entire CDKN2A / B locus (INK4 / ARF locus), or a CDKN2A(p16 INK4A ) and its alternative reading frame p14 ARF These individuals have genetic abnormalities, such as haploinsufficiency or homozygous loss of ANNUBP, which have been identified as isolated and highly recurrent in the majority of human ANNUBPs associated with MPNST.

[0023] As used herein, a pharmaceutically acceptable salt is a salt with a pharmaceutically acceptable acid or base. Pharmaceutically acceptable acids include both inorganic acids such as hydrochloric acid, sulfuric acid, phosphoric acid, diphosphoric acid, hydrobromic acid, or nitric acid, and organic acids such as citric acid, fumaric acid, maleic acid, malic acid, ascorbic acid, succinic acid, tartaric acid, benzoic acid, acetic acid, methanesulfonic acid, ethanesulfonic acid, salicylic acid, stearic acid, benzenesulfonic acid, or p-toluenesulfonic acid. Pharmaceutically acceptable bases include hydroxides of alkali metals (e.g., sodium or potassium) and alkaline earth metals (e.g., calcium or magnesium), as well as organic bases such as alkylamines, arylamines, or heterocyclic amines.

[0024] The present invention is directed to a composition comprising nitroxoline or a pharmaceutically acceptable salt thereof for use in the treatment or prevention of MPNST.

[0025] In an alternative embodiment, the present invention is directed to a composition comprising nitroxoline or a pharmaceutically acceptable salt thereof for use in treating or preventing MPNST, wherein nitroxoline is the only active agent in the composition, meaning that the composition does not contain other ingredients that may be used in the treatment or prevention of MPNST.

[0026] Alternatively, the composition comprising nitroxoline or a pharmaceutically acceptable salt thereof may also comprise one or more active agents, preferably one active agent, in the composition. The additional active agent(s) may be an active agent for treating or preventing MPNST.

[0027] The composition of the present invention may contain a pharmaceutically acceptable carrier. "Pharmaceutically acceptable carrier" refers to any diluent or excipient, such as a filler or binder, that is compatible with other components of the composition and is not harmful to the recipient. Pharmaceutically acceptable carriers can be selected according to standard pharmaceutical practice based on the desired route of administration.

[0028] In the present invention, the composition can be administered in various dosage forms. In one embodiment, the composition can be formulated in a form suitable for oral, rectal, parenteral, intranasal, or transdermal administration, or for administration by inhalation or suppository.

[0029] The compositions can be orally administered, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders, or granules. Preferably, the compositions are formulated to be suitable for oral administration, such as tablets and capsules. Tablets and capsules can be prepared using binders such as syrup, acacia, gelatin, sorbitol, tragacanth, cellulose, or polyvinylpyrrolidone; fillers such as lactose, sucrose, corn starch, calcium phosphate, sorbitol, or glycine; lubricants such as magnesium stearate, talc, polyethylene glycol, or silica; and surfactants such as sodium lauryl sulfate. Liquid compositions can contain conventional additives, such as suspending agents, for example, sorbitol syrup, methylcellulose, sugar syrup, gelatin, carboxymethylcellulose, or edible fats; emulsifiers and surfactants such as lecithin or acacia; vegetable oils such as almond oil, coconut oil, cod liver oil, or peanut oil; and preservatives such as butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT). Liquid compositions can be encapsulated in, for example, gelatin to provide a unit dosage form.

[0030] The compositions can also be administered parenterally, subcutaneously, intravenously, intramuscularly, intrasternally, transdermally or by infusion techniques.

[0031] Composition can also be administered by inhalation.The advantage of inhalation medicine is that compared with many drugs that are taken orally, it is delivered directly to the area with abundant blood supply.Therefore, because the surface area of ​​the alveoli is huge and the blood supply is abundant, and first-pass metabolism is avoided, absorption is very rapid.

[0032] The present invention also provides an inhalation device comprising a composition of the invention, typically a metered dose inhaler (MDI) that includes a pharmaceutically acceptable chemical propellant to force the medicament out of the inhaler.

[0033] The composition can also be administered intranasally. The highly permeable tissue of the nasal cavity is highly receptive to drugs, absorbing them quickly and efficiently. Nasal drug delivery is less painful and invasive than injections, causing less anxiety among patients. This method results in very rapid absorption and generally avoids first-pass metabolism, thus reducing patient-to-patient variability. Furthermore, the present invention also provides an intranasal device comprising the composition of the present invention.

[0034] The composition can also be administered transdermally. For topical delivery, transdermal and transmucosal patches, creams, ointments, jellies, solutions, or suspensions can be employed. Thus, the present invention also provides a transdermal patch containing the composition.

[0035] The composition may also be administered by sublingual administration. Accordingly, the present invention also provides a sublingual tablet comprising the composition.

[0036] The compositions may also be formulated with agents that reduce degradation of the substance by processes other than the patient's normal metabolism, such as antibacterial agents or inhibitors of protease enzymes that may be present in the patient or in commensal or parasitic organisms living on or in the patient and that can degrade the compound.

[0037] Liquid dispersions for oral administration can be syrups, emulsions, and suspensions.

[0038] Suspensions and emulsions may contain as a carrier, for example, a natural gum, agar, sodium alginate, pectin, methylcellulose, carboxymethylcellulose, or polyvinyl alcohol. Suspensions or solutions for intramuscular injection may contain, together with the active compound, a pharmaceutically acceptable carrier, for example, sterile water, olive oil, ethyl oleate, glycols such as propylene glycol, and, if desired, an appropriate amount of lidocaine hydrochloride.

[0039] The solutions for injection or infusion may contain as carrier, for example, sterile water or preferably they may be in the form of sterile aqueous isotonic saline solutions.

[0040] In an embodiment of the invention, the composition is administered in an amount effective to treat or prevent MPNST. The effective dose will be apparent to one skilled in the art and will depend on many factors, including age, sex, and weight, and a physician will be able to determine the effective amount.

[0041] In a preferred embodiment, the composition comprises 30 mg to 600 mg, preferably 50 mg to 500 mg, more preferably 100 mg to 400 mg, even more preferably 150 mg to 350 mg, and most preferably 200 mg to 300 mg of nitroxoline.

[0042] The composition may be administered once daily, twice daily, three times daily, or four times daily.

[0043] In an embodiment of the present invention, the composition is administered at least once a day. Preferably, it is administered as a single daily dose. Preferably, the single daily dose is 90 mg to 1800 mg, preferably 150 mg to 1500 mg, more preferably 300 mg to 1200 mg, even more preferably 450 mg to 1050 mg, and most preferably 600 mg to 900 mg of nitroxoline.

[0044] In an embodiment of the invention, the composition is administered twice daily, preferably with each dose being 45 mg to 900 mg, preferably 75 mg to 750 mg, more preferably 150 mg to 600 mg, even more preferably 225 mg to 525 mg, and most preferably 300 mg to 450 mg of nitroxoline.

[0045] In an embodiment of the invention, the composition is administered three times daily, preferably with each dose being 30 mg to 600 mg, preferably 50 mg to 500 mg, more preferably 100 mg to 400 mg, even more preferably 150 mg to 350 mg, and most preferably 200 mg to 300 mg of nitroxoline.

[0046] In an embodiment of the invention, the composition is administered four times daily, preferably with each dose being 15 mg to 500 mg, preferably 50 mg to 400 mg, more preferably 100 mg to 300 mg, even more preferably 125 mg to 225 mg, and most preferably 150 mg to 200 mg of nitroxoline.

[0047] Preferably, the dosing regimen is such that the total daily dose of nitroxoline does not exceed 1500 mg.

[0048] Suitably, an effective dose of nitroxoline results in an intracellular concentration of 1 to 150 μM, preferably 10 to 100 μM, more preferably 25 to 50 μM.

[0049] To treat or prevent MPNST, compositions containing nitroxoline are used in a chronic dosing regimen, i.e., chronic long-term treatment, preferably lasting at least 1 month, preferably at least 2 months, e.g., at least 3 months.

[0050] The present invention also relates to the use of nitroxoline or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for use in the treatment or prevention of MPNST. This embodiment of the invention may have any of the preferred features described above.

[0051] The present invention also relates to a method for treating or preventing MPNST, comprising administering to a patient a composition comprising nitroxoline or a pharmaceutically acceptable salt thereof. This embodiment of the invention may have any of the preferred features described above. The method of administration may be according to any of the routes described above.

[0052] For the avoidance of doubt, the present invention also encompasses pro-drugs which react in vivo to yield compounds of the present invention.

[0053] Experimental Section In vitro drug screening using Nf1 and Nf1-Ink4a / Arf mutant DNSCs Nf1-Ink4a / Arf-deficient cells, contained within embryonic dorsal root ganglia (DRG) / nerve roots (DNSCs), can be reimplanted into the neural microenvironment and generate tumors histologically indistinguishable from human ANNUBP within 6 weeks. These lesions further progress to high-grade MPNST with 100% penetrance by 3-4 months after implantation. By providing high efficiency of tumor development within a defined latency period, this approach provides a solid foundation for the evaluation of experimental therapeutics designed to treat, delay, and / or prevent the progression of ANNUBP and MPNST driven by the loss of Nf1 and Ink4 / Arf [Chen, Z., et al., Cancer Cell, 2014.26(5):pp.695-706].

[0054] Consistent with their in vivo phenotype, Nf1-Ink4a / Arf mutant DNSCs have increased cell cycle activity and proliferation in vitro. flox / flox Nf1 obtained by transient infection with - / - and Nf1 - / - ;Ink4a / Arf + / - Nf1 carrying mutant DNSC and adenovirus carrying Cre recombinase flox / flox ;Ink4a / Arf flox / + DNSC is used to enhance nitroxylon selectivity by Nf1 - / - and Nf1- / - ;Ink4a / Arf + / - Comparison was made between DNSCs.

[0055] Nf1 as described above - / - and Nf1 - / - ;Ink4a / Arf + / - DNSCs were treated with serially diluted nitroxoline starting at 105 μM and incubated for 48 hours. After the incubation period, proliferation, viability, and apoptosis assays were performed as described below. The proliferation and viability of DNSCs were assessed using the CellTiter-Glo assay (Promega), which measures ATP consumption. - / - and Nf1 - / - ;Ink4a / Arf + / - DNSCs were plated in triplicate at 5,000 cells / well in 96-well dishes in 100 μl of DNSC growth medium supplemented with FGF and EGF and placed in a humidified incubator at 37°C and 5% CO2 for 48 hours, with or without experimental compounds. After incubation, 100 μL of CellTiterGlo reagent was added to each well. After 10 minutes, plates were read using a 96-well microplate luminometer. To assess cell apoptosis, the Caspase-Glo 3 / 7 kit (Promega) was used according to the manufacturer's instructions. - / - and Nf1 - / - ;Ink4a / Arf + / - DNSCs were plated and treated with experimental compounds. After 48 hours, 100 μL of Caspase-Glo 3 / 7 reagent was added to each well. After 10 minutes, the plates were read using a 96-well luminometer to measure caspase 3 / 7 activity.

[0056] result Nitroxoline inhibited cell proliferation and increased apoptosis in a dose-response manner, as seen in Figure 1. Antiproliferative effects were observed at concentrations greater than 3 μM in Nf1- / - lnk4a / Arf- / - cells and greater than 10 μM in Nf1- / - cells. Induction of apoptosis was evident at concentrations greater than 10 μM in Nf1- / - lnk4a / Arf- / - cells and at concentrations between 30 and 100 μM in Nf1- / - cells.

[0057] conclusion Nitroxoline inhibits cell proliferation and increases apoptosis in stem cells that recapitulate transformed Nf1 MPNST in vitro, and therefore holds promise for alleviating, treating, and preventing MPNST.

Claims

1. A composition comprising nitroxoline or a pharmaceutically acceptable salt thereof for use in the treatment or prevention of malignant peripheral nerve sheath tumors (MPNST).

2. 10. The composition for use according to claim 1 in the treatment of MPNST.

3. The composition for use according to claim 1 or 2, wherein the subject of said treatment or prevention has neurofibromatosis, preferably neurofibromatosis type I.

4. 10. A composition for use according to any one of the preceding claims, wherein the subject of said treatment or prevention is a human.

5. 10. A composition for use according to any one of the preceding claims, wherein the composition comprises 30mg to 600mg, preferably 50mg to 500mg, more preferably 100mg to 400mg, even more preferably 150mg to 350mg, and most preferably 200mg to 300mg of nitroxoline.

6. 10. A composition for use according to any one of the preceding claims, wherein administration is by two doses per day.

7. 7. The composition for use according to claim 6, wherein the dose is 45 mg to 900 mg, preferably 75 mg to 750 mg, more preferably 150 mg to 600 mg, even more preferably 225 mg to 525 mg, and most preferably 300 mg to 450 mg of nitroxoline.

8. The composition for use according to any one of claims 1 to 4, wherein the administration is by three doses per day.

9. 9. The composition for use according to claim 8, wherein the dose is 30 mg to 600 mg, preferably 50 mg to 500 mg, more preferably 100 mg to 400 mg, even more preferably 150 mg to 350 mg, and most preferably 200 mg to 300 mg of nitroxoline.

10. The composition for use according to any one of claims 1 to 4, wherein the administration is by four doses per day.

11. 11. The composition for use according to claim 10, wherein the dose is 15 mg to 500 mg, preferably 50 mg to 400 mg, more preferably 100 mg to 300 mg, even more preferably 125 mg to 225 mg, and most preferably 150 mg to 200 mg of nitroxoline.

12. 10. A composition for use according to any one of the preceding claims, which is administered orally or intravenously.

13. The composition for use according to any one of claims 1 to 11, which is administered parenterally, transdermally, sublingually, rectally or by inhalation administration.

14. A composition comprising nitroxoline or a pharmaceutically acceptable salt thereof for use in the treatment or prevention of MPNST, wherein nitroxoline is the only active agent in said composition.

15. 1. Use of nitroxoline or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for use in the treatment or prevention of MPNST.

16. Use according to claim 15, with any of the additional features according to claims 2 to 14.

17. A method for treating or preventing MPNST, comprising administering to a patient a composition comprising nitroxoline or a pharmaceutically acceptable salt thereof.

18. The method according to claim 17, having any of the additional features according to claims 2 to 14.

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