Patent Description:
The present disclosure is related to compounds for inhibition of indoleamine-<NUM>,<NUM>-dioxygenase pathway, in particular salts of indoximod with enhanced pharmacokinetic properties relative to indoximod.

Tryptophan degradation into kynurenine is mediated by indoleamine-<NUM>,<NUM>-dioxygenase (IDO1) expressed by plasmacytoid dendritic cells, placental, epithelial and tumor cells and by tryptophan-<NUM>,<NUM>-dioxygenase (TDO2) expressed mainly by the liver and tumor cells.

IDO1 plays an important role in the regulation of immune responses by triggering anergy on reactive effector T cells and by modulating differentiation and activation of regulatory T cells (Tregs). From a more general viewpoint, the IDO enzyme is involved in pathway that comprises all proteins that directly or indirectly contribute to modulate the immunosuppressive functions dependent on IDO activity, including proteins that mediate induction of IDO expression, activation of enzymatic activity by reductases, post-translational modifications that regulate activity, protein degradation, and the interpretation and transmission of the signals elicited by low concentrations of Trp and the presence of Trp catabolites [collectively known as kynurenines (Kyns)] including catabolic stress sensors integrated into the General Control Nonrepressed-<NUM> (GCN2) pathway, the Aryl Hydrocarbon Receptor (AhR) pathway, and the mammalian Target Of Rapamycin (mTOR) pathways. This concept of integrated downstream regulatory pathways with IDO at the center has emerged from studies on multiple model systems by many research groups and this notion may be critically important for understanding how the IDO pathway is induced, how IDO exerts downstream effects, and the mechanism of action of IDO pathway inhibitors that target IDO directly or target other components of the IDO pathway [<NUM>, <NUM>].

Therefore, direct pharmacological inhibition of IDO1 enzymatic activity or inhibition of the upstream factors that activate IDO1 enzyme or inhibition of the downstream effects of IDO1 enzymatic activity should stimulate an immune response by multiple mechanisms that may involve preventing anergy of effector T cells, reactivating anergic effector T cells, preventing the activation of regulatory T cells, promoting phenotypic conversion of Tregs to pro-inflammatory TH17 cells and promoting phenotypic reprogramming of immunosuppressive dendritic cells into immunostimulatory dendritic cells.

For these reasons, numerous enzymatic inhibitors of IDO have been described and are being developed to treat or prevent IDO related diseases such as cancer and infectious diseases. Numerous molecules that inhibit IDO enzymatic activity either as competitive or non-competitive inhibitors have been described in the literature, for example in patent applications <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>, <CIT>.

The general use of pharmaceutically acceptable salts is described by <NPL>, <NPL>, and <NPL>.

One of the first IDO pathway inhibitors studied in preclinical models has been <NUM>-methyl-DL-tryptophan (1mT), a racemic mixture of enantiomers, which was shown to mediate immune dependent rejection of allogeneic fetuses in mice [<NUM>] and immune dependent enhancement of antitumor activity of chemotherapy and radiotherapy [<NUM>]. Each one of these enantiomers shows different biological properties. <NUM>-methyl-L-tryptophan (L1mT) has been shown to inhibit IDO1 enzymatic activity (Ki=<NUM>, [<NUM>]) in cell-free assays using purified recombinant IDO1 enzyme, and in tumor cells treated with INFγ or in tumor cell lines transfected with expression vectors that encode IDO1 under the control of an heterologous promoter, while the D isomer (indoximod) does not inhibit enzymatic activity in these type of assays [<NUM>]. Nonetheless, both isomers are capable of restoring T cell proliferation in an MLR assay with IDO+ dendritic cells as the stimulator cells, or in syngeneic antigen-dependent T cell proliferation assays using IDO+ DCs isolated from tumor draining lymph nodes [<NUM>]. In this type of assay, where IDO+ DCs are present, T cells do not proliferate. However, inhibition of the IDO pathway by these inhibitors restores the proliferative capacity of T cells. Interestingly, both isomers show different potency in this assay, with indoximod being more potent (EC50=<NUM>) than L1mT (EC50= <NUM>-<NUM>) or the racemic mixture (<NUM>-<NUM>) [<NUM>]. Moreover, despite the fact that indoximod does not show inhibition of enzymatic activity in other types of assays, it shows inhibition of enzymatic activity in this co-culture assay, as seen by reduced Trp degradation and Kyn synthesis.

A somewhat puzzling issue has been the fact that indoximod does not show inhibition of IDO1 enzymatic activity in vitro, but somehow mimics the biological consequences of IDO1 inhibition in vivo or in cell based assays. Experimental evidence from a number of research laboratories points to the conclusion that indoximod is participating in the inhibition of the IDO1 pathway. Several possible mechanisms by which this could be taking place are: <NUM>) inhibition of isoforms of IDO1, <NUM>) inhibition of IDO2, <NUM>) alternative formation of indoximod -derived metabolites, <NUM>) racemization of indoximod into L1mT, <NUM>) inhibition of Trp transport, <NUM>) inhibition of the GCN2 pathway by formation of indoximod-tRNA complexes, <NUM>) inhibition of enzymes involved in Trp sensing such as WARS1 or WARS2, <NUM>) alteration of autophagy under conditions of amino acid deprivation induced stress or <NUM>) bypassing mechanisms that inactivate mTOR under conditions of amino acid deficiency [<NUM>]. These mechanisms are not necessarily mutually exclusive, and so far are compatible with the current experimental data. Further investigations are needed to elucidate which of these biochemical mechanisms is responsible for the biological activity of indoximod.

The biological activity of indoximod to relieve immunosuppression in vivo and in vitro is supported by studies performed in several laboratories in murine preclinical models. Indoximod has demonstrated activity in the following biological assays:.

For these reasons, indoximod is being investigated in human clinical trials for cancer indications. Indoximod is being studied in several cancer indications in combination with different chemotherapeutic and biological immunotherapeutic agents, such as docetaxel, paclitaxel, gemcitabine, Nab-paclitaxel, temozolomide, ipilimumab, sipuleucel-T, or vaccines.

Indoximod is orally bioavailable with a favorable pharmacokinetic (PK) profile (Tmax: ~ <NUM>; half-life: ~<NUM>) and an excellent safety profile. Pharmacokinetic studies in patients have demonstrated that indoximod shows a linear PK profile at doses of up to <NUM>/dose, with maximum plasma concentration (Cmax) of <NUM> and drug exposure (AUC(<NUM>-last)) levels of ~<NUM>. However, increasing doses above <NUM>/dose up to <NUM>/dose, does not result in a linear or proportional increase in Cmax or drug exposure, thus potentially limiting the therapeutic activity of this investigational drug.

Mixed-lymphocyte response (MLR) T cell proliferation assay show that T cells that are in an IDO+ environment restore ~<NUM>% of their proliferative capacity at concentrations of indoximod higher than <NUM>. Murine antitumor experiments show that biological effects of indoximod are observed when mice are dosed with indoximod in the drinking water at <NUM>/mL (~<NUM>/kg/day), or dosed orally at <NUM>/kg bid, which results in Cmax higher than <NUM> and exposures greater than <NUM>. For these reasons, it is desirable to increase the Cmax and exposure to indoximod in human clinical trials so they may reach the levels necessary for therapeutic activity. However, the non-linear pharmacokinetic profile of this drug makes it unlikely that this could be solved by increasing the dose given to patients.

For the above mentioned reasons we investigated whether different formulation of indoximod such as spray dry dispersions or salts or indoximod prodrugs in different salt forms would increase solubility and absorption rate or reduce blood clearance to levels that increase the maximum concentration and exposure to indoximod. Moreover, we looked for prodrugs and its salts that could result in increases parameters of exposure when dosed orally and in pill (capsule or tablet) dosage formulation.

The results of these investigations showed that a few selected prodrugs resulted in increases in parameters of exposure; and that increases in in vitro solubility and in vivo exposure could be achieved by a few salts of indoximod upon oral administration.

In one aspect the invention describes salts and pharmaceutical compositions comprising salts according to Formula 1a
<CHM>
Wherein A-pn is an anion selected from the group consisting of HPO<NUM><NUM>- (hemi-phosphate), HSO<NUM>- (hydrogen sulfate), and CH<NUM>S(O<NUM>)O- (mesylate), wherein the ionization state -p is -<NUM> or -<NUM> and the stoichiometric ratio n is <NUM> or ½, respectively, such that the stoichiometric conditions of charge neutrality are satisfied.

In another aspect, the present disclosure provides the above-described salt for use in a method of treating cancer, comprising orally administering a therapeutically effective amount of the salt to a subject in need thereof.

In another aspect, the present disclosure provides the above-described pharmaceutical composition for use in a method of treating cancer in a subject in need thereof, comprising orally administering a therapeutically effective amount of the salt in an appropriate pharmaceutical form or vehicle.

In another aspect, the present disclosure provides the above-described salt or pharmaceutical composition:.

Further embodiments of the invention are set out in the appended claims.

Indoximod (<NUM>-methyl-D-tryptophan, D1mT) is an investigational inhibitor of the indoleamine-<NUM>,<NUM>-dioxygenase (IDO) pathway that is being tested in several human clinical trials for multiple cancer indications, in combination with standard and experimental chemotherapeutic and immunomodulatory agents and active immunotherapies.

In the presence of IDO+ dendritic cells, CD8+ effector T cells become anergic and unable to proliferate. Moreover, regulatory T cells (CD4+ CD25+ FoxP3+) are activated in the presence of IDO+ DCs and become able to mediate systemic immunosuppression to tumor or viral antigens. Indoximod is capable to revert these processes, allowing effector T cells to proliferate and directing reprogramming of Tregs to a TH17 helper-like phenotype. In in vitro assays, these effects are mediated by indoximod with an EC50 of ~ <NUM> [<NUM>]. In preclinical murine tumor models, antitumor effects, stimulation of effector T cells and reprogramming of Tregs in the draining lymph nodes requires daily doses of ~ <NUM>/kg, with exposures > <NUM>.

Human pharmacokinetic experiments at oral doses that range between <NUM> to <NUM>/dose have shown that the pharmacokinetic parameters Cmax and exposure (AUC<NUM>-inf) increase linearly with dose, up to a range of ~ <NUM>/dose. At these doses, Cmax in plasma reaches an average of ~<NUM> and AUC<NUM>-inf reaches ~ <NUM>. The Cmax and AUC parameters do not significantly increase above those values at higher doses of up to <NUM>/dose. Therefore, in order to achieve indoximod concentration and exposure levels that are comparable to those that produce immunomodulatory and antitumor therapeutic effects in murine models it would be useful to increase the Cmax and exposure levels of indoximod.

The present disclosure describes compounds of formula 1a, 1b and formula <NUM> that produce a higher exposure and maximum serum concentration of indoximod upon oral administration, compared to oral administration of equivalent molar doses of indoximod.

In one example, a salt of indoximod is disclosed. In one example, the salt has a structure according to Formula 1a:
<CHM>
wherein A-pn is an inorganic or organic anion in an ionization state -p. In one example, the anion is present at a stoichiometric ratio n that ensures molecular charge neutrality.

In one example, the anion A-pn is selected from the group consisting of chloride, phosphate, sulfate, mesylate, besylate, acetate, ascorbate, aspartate, glutamate, glutarate, lactate, maleate, malonate, oxalate, succinate, fumarate, tartrate and citrate. In one example, the anion is presented at a stoichiometric ratio n such that the resulting salt is charge neutral. Accordingly, in one example, the anion has an ionization state p of -<NUM>, -<NUM> or -<NUM> and is presented at a stoichiometric ratio n of <NUM>, <NUM>/<NUM> or <NUM>/<NUM>, respectively, such that the stoichiometric conditions of charge neutrality are satisfied. In one embodiment, the phosphate is HPO<NUM>-<NUM>, and the HPO<NUM>-<NUM> is present at a stoichiometric ratio n of <NUM>. In one example, the phosphate is HPO<NUM>-, and the HPO<NUM>- is present at a stoichiometric ratio n of <NUM>. In one example, the sulfate is SO<NUM>-<NUM>, and the SO<NUM>-<NUM> is present at a stoichiometric ratio n of <NUM>. In one embodiment, the mesylate is CH<NUM>SO<NUM>-, and the CH<NUM>SO<NUM>- present at a stoichiometric ratio n of <NUM>.

In another example the anion A-pn is Cl- at a stoichiometric ratio n of <NUM>. In another preferred example the anion A-pn is Cl- at a stoichiometric ratio n of <NUM> and the crystalline form is an anhydrous isoform of Form <NUM>.

In one example, the salt has a structure according to Formula 1b:
<CHM>
wherein C+pm is a cation in an ionization state +p. In one example, the cation is present at a stoichiometric ratio m that ensures molecular charge neutrality. In one example, the C+pm is selected from the group consisting of Li+, Na+, K+, Mg+<NUM> and Ca+<NUM>. In one example, when p is +<NUM>, m is <NUM>, and when p is +<NUM>, m is ½.

In one example, a prodrug of indoximod is disclosed. In one example, the structure of the prodrug, in free base or salt form, is provided in Formula <NUM>:
<CHM>.

In one example, R<NUM> is -OH, -OC<NUM>-<NUM>alkyl, -OCH<NUM>CH(OH)CH<NUM>OH, -O(CH<NUM>)<NUM>N(CH<NUM>)<NUM>, - OC<NUM>-<NUM>alkyl-R<NUM>, -NHC(S)HR<NUM>(COOH), -NHC(R)HR<NUM>(COOH), -OC<NUM>-<NUM>alkylR<NUM>, -OC<NUM>-<NUM>alkyl-C(S)H(NH<NUM>)(COOH), or -OC<NUM>-<NUM>alkyl-C(R)H(NH2)(COOH). In one example, R<NUM> is - NHC(S)HR<NUM>(COOCH<NUM>) or -NHC(R)HR<NUM>(COOCH<NUM>).

In one example, R<NUM> is -H, -C(O)C(S)H(NH<NUM>)R<NUM>, -C(O)C(R)H(NH<NUM>)R<NUM>,-C(O)CH<NUM>C(S)H(NH<NUM>)-C(O)OCH<NUM>, -C(O)OR<NUM>, or -C(O)NHR<NUM>.

In one example, R<NUM> is tetrahydropyran or
<CHM>.

In one example, R<NUM> is -H, -C<NUM>-<NUM>alkyl, -(CH<NUM>)<NUM>-<NUM>SH, -C<NUM>-<NUM>alkylSC<NUM>-<NUM>alkyl, -C<NUM>-<NUM>alkylOC<NUM>-<NUM>alkyl, -CH<NUM>-R<NUM>, -CH<NUM>OH, -CH(OH)CH<NUM>, -(CH<NUM>)<NUM>-<NUM>C(O)NH<NUM>,-(CH<NUM>)<NUM>-<NUM>C(O)OH, -(CH<NUM>)<NUM>-<NUM>NH<NUM>, or - (CH<NUM>)<NUM>-<NUM>NC(=NH<NUM>)NH<NUM>.

In one example, when R<NUM> is not -H, C(S) and C(R) are carbons with the S or R stereochemistry, respectively.

In one example, R<NUM> is -H, C<NUM>-<NUM>alkylR<NUM>, or R<NUM>. In one example, R<NUM> is selected from the group consisting of -H, aryl, alkylaryl, heteroaryl, cycloalkyl, and heterocycloalkyl, wherein the aryl, alkylaryl, heteroaryl, cycloalkyl or heterocycloalkyl is optionally substituted with one two or three R<NUM> groups.

In one example, each R<NUM> is independently halogen, cyano, nitro, -OR, -N(R)<NUM>, -SR, -C(O)OR, C<NUM>-<NUM>alkyl, C<NUM>-<NUM>haloalkyl, -C(O)N(R)<NUM>, -C(O)R, -S(O)R, -S(O)OR, - S(O)N(R)<NUM>, -S(O)<NUM>R, -S(O)<NUM>OR, -S(O)<NUM>N(R)<NUM>, -OC(O)R, -OC(O)OR, -OC(O)N(R)<NUM>, -N(R)C(O)R, -N(R)C(O)OR, or -N(R)C(O)N(R)<NUM>, wherein R is H or C<NUM>-<NUM>alkyl.

In some examples of the prodrug of Formula <NUM>, R<NUM> cannot be -OH when R<NUM> is H.

Furthermore, in all examples, the prodrug cannot be Nα-tert-butoxycarbonyl-<NUM>-methyl-D-tryptophan, ethyl Nα-benzyl-<NUM>-methyl-D-tryptophanate, or benzyl Nα- (tert-butoxycarbonyl)-<NUM>-methyl-D-tryptophanate.

In one example, HAn is an acid. In one example, the acid HAn is selected from the group consisting of PO<NUM>H<NUM> (phosphoric acid), SO<NUM>H<NUM> (sulfuric acid), HCl (hydrochloric acid), HSO<NUM>CH<NUM> (methyl sulfonic acid), C<NUM>H<NUM>SO<NUM>H (benzyl sulfonic acid), acetic acid, ascorbic acid, aspartic acid, glutamic acid, glutaric acid, lactic acid, maleic acid, malonic acid, oxalic acid, succinic acid, fumaric acid, tartaric acid and citric acid.

In one example, the acid HAn is present at a stoichiometric ratio n such that the resulting prodrug is charge neutral. Accordingly, in one example, the stoichiometric ratio n of the acid HAn is <NUM>, <NUM>, <NUM> or <NUM> such that the prodrug is charge neutral.

The disclosure also provides prodrugs of indoximod, in their free base or salt form. In one example, the prodrugs of indoximod are represented by compounds of Formula <NUM>,
<CHM>
wherein.

In another example, the disclosure provides prodrugs of indoximod, in their free base or salt form, as represented by compounds of Formula <NUM>,
<CHM>.

In a preferred example, the disclosure provides prodrugs of indoximod, in their free base or salt form, as represented by compounds of Formula <NUM>,
<CHM>
wherein.

In another preferred example, the disclosure provides prodrugs of indoximod, in their free base or salt form, as represented by compounds of Formula <NUM>,
<CHM>
wherein.

In a most preferred example, the disclosure provides prodrugs of indoximod, in their free base or salt form, as represented by compounds of Formula <NUM>,
<CHM>
wherein.

In a preferred example, the disclosure provides prodrugs of indoximod, in their free base or as a pharmaceutically appropriate salt form, as represented by compounds of Formula <NUM> represented in Table <NUM>.

In one example, the prodrug substantially includes at least one of the following compounds: (i) ethyl Nα-(L-leucyl)-<NUM>-methyl-D-tryptophanate; (ii) <NUM>,<NUM>-dihydroxypropyl <NUM>-methyl-D-tryptophanate; (iii) Nα-(L-leucyl)-<NUM>-methyl-D-tryptophan; (iv) ethyl Nα-(L-isoleucyl)-<NUM>-methyl-D-tryptophanate; (v) Nα-(L-glycyl)-<NUM>-methyl-D-tryptophan; (vi) (S)-<NUM>-amino-<NUM>-(((R)-<NUM>-carboxy-<NUM>-(<NUM>-methyl-<NUM>H-indol-<NUM>-yl)ethyl)amino)-<NUM>-oxohexanoic acid; (vii) Nα-(L-lysyl)-<NUM>-methyl-D-tryptophan; (viii) Nα-(L-phenylalanyl)-<NUM>-methyl-D-tryptophan; (ix) ethyl Nα-(L-glutaminyl)-<NUM>-methyl-D-tryptophanate; (x) <NUM>-(dimethylamino)ethyl <NUM>-methyl-D-tryptophanate; (xi) (<NUM>-ethoxy-<NUM>-oxido-<NUM>,<NUM>,<NUM>-dioxaphospholan-<NUM>-yl)methyl <NUM>-methyl-D-tryptophanate; (xii) <NUM>-(tetrahydro-<NUM>H-pyran-<NUM>-yl)ethyl <NUM>-methyl-D-tryptophanate; (xiii) ethyl <NUM>-methyl-D-tryptophanate; (xiv) isopropyl <NUM>-methyl-D-tryptophanate; (xv) Nα-(L-methionyl)-<NUM>-methyl-D-tryptophan; or (xvi) ethyl Nα-(L-methionyl)-<NUM>-methyl-D-tryptophanate.

In one example, the disclosure provides a pharmaceutical composition comprising salts of indoximod, as represented by compounds of Formula 1a and 1b,
<CHM>
wherein A-n is an inorganic or organic anion and C+pm is an inorganic cation in an ionization state and at a stoichiometric ratio that ensures molecular charge neutrality.

In a further example, the disclosure provides a pharmaceutical composition comprising salts of indoximod, as represented by compounds of Formula 1a, wherein A-pn is an anion selected from the group consisting of chloride, phosphate, sulfate, mesylate, besylate, acetate, ascorbate, aspartate, glutamate, glutarate, lactate, maleate, malonate, oxalate, succinate, fumarate, tartrate and citrate, wherein negative charge p is -<NUM>, -<NUM> or -<NUM> at stoichiometric ratio n of <NUM>, ½ or <NUM>/<NUM>, respectively, so that it satisfies stoichiometric conditions of charge neutrality.

In a further example, the disclosure provides a pharmaceutical composition comprising salts of indoximod, as represented by compounds of Formula 1b, wherein C+pm is an cation selected from the group of Li+, Na+, K+, Mg+<NUM> or Ca+<NUM>, wherein positive charge p is +<NUM> or +<NUM> at stoichiometric ratio m of <NUM> or ½, respectively, so that it satisfies stoichiometric conditions of charge neutrality.

In a further example, the disclosure provides a pharmaceutical composition comprising salts of indoximod, as represented by compounds of Formula 1a, wherein A-pn is an anion selected from the group consisting of HPO<NUM>-<NUM> (phosphate), SO<NUM>-<NUM> (sulfate), H<NUM>PO<NUM>- (phosphate), Cl-, and CH<NUM>SO<NUM>- (mesylate), at stoichiometric ratio n of <NUM>, <NUM>, <NUM> or <NUM>, respectively.

In a further example, the disclosure provides a pharmaceutical composition comprising salts of indoximod, as represented by compounds of Formula 1a, wherein A-pn is Cl- at a stoichiometric ratio n of <NUM>.

In a further example, the disclosure provides a pharmaceutical composition comprising salts of indoximod, as represented by compounds of Formula 1a, wherein A-pn is Cl- at a stoichiometric ratio n of <NUM> and the crystalline form is an anhydrous isoform of Form <NUM>. In an example, the disclosure provides a pharmaceutical composition comprising prodrugs of indoximod, in their free base or salt form. In one example, the prodrugs of indoximod are represented by compounds of Formula <NUM>,
<CHM>
wherein.

In another example, the disclosure provides a pharmaceutical composition comprising prodrugs of indoximod, in their free base or salt form, as represented by compounds of Formula <NUM>,
<CHM>.

In a preferred example, the disclosure provides a pharmaceutical composition comprising prodrugs of indoximod, in their free base or salt form, as represented by compounds of Formula <NUM>,
<CHM>
wherein.

In a most preferred example, the disclosure provides a pharmaceutical composition comprising prodrugs of indoximod, in their free base or salt form, as represented by compounds of Formula <NUM>,
<CHM>
wherein.

In a most preferred example, the disclosure provides a pharmaceutical composition comprising prodrugs of indoximod, in their free base or salt form, as represented by compounds of Formula <NUM>,
wherein
<CHM>.

In a preferred example, the disclosure provides a pharmaceutical composition comprising prodrugs of indoximod, in their free base or as a pharmaceutically appropriate salt form, as represented by compounds of Formula <NUM> represented in Table <NUM>.

In another example, the disclosure provides methods of use of compositions of formulas <NUM> and <NUM>, to modulate the activity of indoleamine-<NUM>,<NUM>-dioxygenase pathway in a subject in need thereof, comprising the oral administration of therapeutically effective amounts such compositions to such subject in an appropriate pharmaceutical form or vehicle.

In another example, the disclosure provides methods of use of compositions of formulas 1a, 1b and <NUM>, for the treatment of cancer in a subject in need thereof, comprising the oral administration of therapeutically effective amounts of such compositions to such subject in an appropriate pharmaceutical form or vehicle.

In another example, the disclosure provides methods of use of compositions of formulas 1a, 1b and <NUM>, for the treatment of tumor-specific immunosuppression associated with cancer, in a subject in need thereof, comprising the oral administration of sufficient amounts such compositions to such subject in an appropriate pharmaceutical form or vehicle.

In another example, the disclosure provides methods of use of compositions of formulas 1a, 1b and <NUM>, to treat immunosuppression associated with infectious diseases (e. g HIV-<NUM> infection, influenza), in a subject in need thereof, comprising the oral administration of sufficient amounts such compositions to such subject in an appropriate pharmaceutical form or vehicle.

In one embodiment, a salt and/or a prodrug of indoximod is included in a pharmaceutical composition, and the composition is included in a solid capsule, gelatin capsule, tablet or pill. In one embodiment, the salt and/or the prodrug is included in a dissolvable capsule.

In specific embodiments, the compositions of the present invention may additionally contain other adjunct components conventionally found in pharmaceutical compositions, at their art-established usage levels. Thus, for example, the compositions may contain additional materials useful in physically formulating various dosage forms of the compositions of the present invention, such as dyes, flavoring agents, preservatives, antioxidants, opacifiers, thickening agents and stabilizers. The formulations can be sterilized and, if desired, mixed with auxiliary agents, e.g., lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, colorings, flavorings and/or aromatic substances and the like which do not deleteriously interact with the oligonucleotide(s) of the formulation.

In certain embodiments, pharmaceutical compositions of the present invention comprise one or more excipients. In certain such embodiments, excipients are selected from water, salt solutions, alcohol, polyethylene glycols, gelatin, lactose, lactose monohydrate, amylase, magnesium stearate, talc, silicic acid, viscous paraffin, hydroxymethylcellulose, microcrystalline cellulose and polyvinylpyrrolidone.

In certain embodiments, a pharmaceutical composition of the present invention is prepared using known techniques, including, but not limited to mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or tableting processes.

Additional embodiments relate to the pharmaceutical formulations wherein the formulation is selected from the group consisting of a solid, powder, liquid and a gel. In certain embodiments, a pharmaceutical composition of the present invention is a liquid (e.g., a suspension, elixir and/or solution). In certain of such embodiments, a liquid pharmaceutical composition is prepared using ingredients known in the art, including, but not limited to, water, glycols, oils, alcohols, flavoring agents, preservatives, and coloring agents.

In certain embodiments, a pharmaceutical composition of the present invention is a solid (e.g., a powder, tablet, and/or capsule). In certain of such embodiments, a solid pharmaceutical composition comprising one or more ingredients known in the art, including, but not limited to, starches, sugars, diluents, granulating agents, lubricants, binders, and disintegrating agents.

In certain embodiments, a pharmaceutical composition of the present invention comprises a delivery system. Examples of delivery systems include, but are not limited to, liposomes and emulsions. Certain delivery systems are useful for preparing certain pharmaceutical compositions including those comprising hydrophobic compounds. In certain embodiments, certain organic solvents such as dimethylsulfoxide are used.

In certain embodiments, a pharmaceutical composition of the present invention comprises a co-solvent system. Certain of such co-solvent systems comprise, for example, benzyl alcohol, a nonpolar surfactant, a water-miscible organic polymer, and an aqueous phase. In certain embodiments, such co-solvent systems are used for hydrophobic compounds. A non-limiting example of such a co-solvent system is the VPD co-solvent system, which is a solution of absolute ethanol comprising <NUM>% w/v benzyl alcohol, <NUM>% w/v of the nonpolar surfactant Polysorbate <NUM> and <NUM>% w/v polyethylene glycol <NUM>. The proportions of such co-solvent systems may be varied considerably without significantly altering their solubility and toxicity characteristics. Furthermore, the identity of co-solvent components may be varied: for example, other surfactants may be used instead of Polysorbate <NUM>; the fraction size of polyethylene glycol may be varied; other biocompatible polymers may replace polyethylene glycol, e.g., polyvinyl pyrrolidone; and other sugars or polysaccharides may substitute for dextrose.

In certain embodiments, a pharmaceutical composition of the present invention comprises a sustained-release system. A non-limiting example of such a sustained-release system is a semi-permeable matrix of solid hydrophobic polymers. In certain embodiments, sustained-release systems may, depending on their chemical nature, release pharmaceutical agents over a period of hours, days, weeks or months.

In certain embodiments, a pharmaceutical composition of the present invention is prepared for oral administration. In certain of such embodiments, a pharmaceutical composition is formulated by combining one or more agents and pharmaceutically acceptable carriers. Certain of such carriers enable pharmaceutical compositions to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral ingestion by a subject. Suitable excipients include, but are not limited to, fillers, such as sugars, including lactose, lactose monohydrate, sucrose, mannitol, or sorbitol; cellulose preparations such as, for example, maize starch, wheat starch, rice starch, potato starch, gelatin, gum tragacanth, methyl cellulose, hydroxypropylmethyl-cellulose, sodium carboxymethylcellulose, microcrystalline cellulose, and/or polyvinylpyrrolidone (PVP). In certain embodiments, such a mixture is optionally ground and auxiliaries are optionally added. In certain embodiments, pharmaceutical compositions are formed to obtain tablets or dragee cores. In certain embodiments, disintegrating agents (e.g., cross-linked carboxymethyl cellulose, such as croscarmellose sodium, cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof, such as sodium alginate) are added.

In certain embodiments, dragee cores are provided with coatings. In certain such embodiments, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures. Dyestuffs or pigments may be added to tablets or dragee coatings.

In certain embodiments, pharmaceutical compositions for oral administration are push-fit capsules made of gelatin. Certain of such push-fit capsules comprise one or more pharmaceutical agents of the present invention in admixture with one or more filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In certain embodiments, pharmaceutical compositions for oral administration are soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. In certain soft capsules, one or more pharmaceutical agents of the present invention are be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In addition, stabilizers may be added.

In certain embodiments, pharmaceutical compositions are prepared for buccal administration. Certain of such pharmaceutical compositions are tablets or lozenges formulated in conventional manner.

In certain embodiments, a pharmaceutical composition is prepared for administration by injection (e.g., intravenous, subcutaneous, intramuscular, etc.). In certain of such embodiments, a pharmaceutical composition comprises a carrier and is formulated in aqueous solution, such as water or physiologically compatible buffers such as Hanks's solution, Ringer's solution, or physiological saline buffer. In certain embodiments, other ingredients are included (e.g., ingredients that aid in solubility or serve as preservatives). In certain embodiments, injectable suspensions are prepared using appropriate liquid carriers, suspending agents and the like. Certain pharmaceutical compositions for injection are presented in unit dosage form, e.g., in ampoules or in multi-dose containers. Certain pharmaceutical compositions for injection are suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. Certain solvents suitable for use in pharmaceutical compositions for injection include, but are not limited to, lipophilic solvents and fatty oils, such as sesame oil, synthetic fatty acid esters, such as ethyl oleate or triglycerides, and liposomes. Aqueous injection suspensions may contain substances that increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, such suspensions may also contain suitable stabilizers or agents that increase the solubility of the pharmaceutical agents to allow for the preparation of highly concentrated solutions.

In certain embodiments, a pharmaceutical composition of the present invention may be an effervescent tablet or granulate. Effervescent tablets most commonly consist of a soluble acid source and a carbonate source to produce carbon dioxide gas, the latter serving as disintegrant. The acidity needed for the effervescent reaction can be derived from food acids, acid anhydrides and acid salts. The food acid can for example be citric acid, tartaric acid, malic acid, fumaric acid, adipic acid or succinic acid. The acid anhydride may be succinic anhydride or citric anhydride or the like. The acid salts may be e.g. sodium dihydrogen phosphate (monosodium phosphate), disodium dihydrogen pyrophosphate (sodium acid pyrophosphate), acid citric salts (sodium dihydrogen citrate and disodium hydrogen citrate), sodium acid sulfite (sodium bisulfite). Suitable carbonate sources are for example sodium bicarbonate, sodium carbonate, potassium bicarbonate, potassium carbonate, sodium sesquicarbonate (mixture of equal molar amounts of sodium carbonate and sodium bicarbonate), glycine carbonate, L-lysine carbonate, arginine carbonate, calcium carbonate.

Effervescence may also be induced by the formation of other gases such as oxygen, e.g. released from sodium perborate or from a combination of e.g. a peroxygen compound that yields active oxygen on mixture with water (e.g. sodium perborate monohydrate or sodium percarbonate) and a chlorine compound that liberates hypochlorite on contact with water (e.g. sodium dichloroisocyanurate or calcium hypochlorite).

The pharmaceutical composition of the present invention can be manufactured according to standard methods known in the art. Granulates and effervescent tablets according to the invention can be obtained by dry compaction or wet granulation. These granulates can subsequently be mixed with e.g. suitable disintegrating agents, glidants and lubricants and be compressed into tablets or filled into e.g. sachets of suitable size. Effervescent tablets can also be obtained by direct compression of a suitable powder mixture, i.e. without any preceding granulation of the excipients.

Suitable powder or granulate mixtures according to the invention are also obtainable by spray drying (e.g., by hot process spray drying or by basic spray drying) , lyophilization, melt extrusion, pellet layering, coating of the active pharmaceutical ingredient or any other suitable method. Preferably, the conditions are chosen such as to prevent amorphization of the active pharmaceutical ingredient. The so obtained powders or granulates can be mixed with one or more suitable ingredients and the resulting mixtures can either be compressed to form effervescent tablets or filled into sachets.

Terms used herein may be preceded and/or followed by a single dash, "- ", or a double dash, "=", to indicate the bond order of the bond between the named substituent and its parent moiety; a single dash indicates a single bond and a double dash indicates a double bond or a pair of single bonds in the case of a spiro-substituent. In the absence of a single or double dash it is understood that a single bond is formed between the substituent and its parent moiety; further, substituents are intended to be read "left to right" unless a dash indicates otherwise. For example, C<NUM>-<NUM>alkoxycarbonyloxy and -OC(O)C<NUM>-<NUM>alkyl indicate the same functionality; similarly arylalkyl, arylalkyl-, and -alkylaryl indicate the same functionality.

Further, certain terms herein may be used as both monovalent and divalent linking radicals as would be familiar to those skilled in the art, and by their presentation linking between two other moieties. For example, an alkyl group can be both a monovalent radical or divalent radical; in the latter case, it would be apparent to one skilled in the art that an additional hydrogen atom is removed from a monovalent alkyl radical to provide a suitable divalent moiety.

The term "alkenyl" as used herein, means a straight or branched chain hydrocarbon containing from <NUM> to <NUM> carbons, unless otherwise specified, and containing at least one carbon-carbon double bond. Representative examples of alkenyl include, but are not limited to, ethenyl, <NUM>-propenyl, <NUM>-methyl-<NUM>-propenyl, <NUM>-butenyl, <NUM>-pentenyl, <NUM>-hexenyl, <NUM>-heptenyl, <NUM>-methyl-<NUM>-heptenyl, <NUM>-decenyl, and <NUM>,<NUM>-dimethylocta <NUM>,<NUM>-dienyl.

The term "alkoxy" as used herein, means an alkyl group, as defined herein, appended to the parent molecular moiety through an oxygen atom. Representative examples of alkoxy include, but are not limited to, methoxy, ethoxy, propoxy, <NUM>-propoxy, butoxy, tert-butoxy, pentyloxy, and hexyloxy.

The term "alkyl" as used herein, means a straight or branched chain hydrocarbon containing from <NUM> to <NUM> carbon atoms, unless otherwise specified. Representative examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, n-hexyl, <NUM>-methylhexyl, <NUM>,<NUM>-dimethylpentyl, <NUM>,<NUM>-dimethylpentyl, n-heptyl, n-octyl, n-nonyl, and n-decyl. When an "alkyl" group is a linking group between two other moieties, then it may also be a straight or branched chain; examples include, but are not limited to -CH<NUM>-, -CH<NUM>CH<NUM>-, -CH<NUM>CH<NUM>CHC(CH<NUM>)-, -CH<NUM>CH(CH<NUM>CH<NUM>)CH<NUM>-.

The term C<NUM>-<NUM>alkyl refers to a linear or branched alkyl of <NUM> to <NUM> carbon atoms.

The term "aryl," as used herein, means a phenyl (i.e., monocyclic aryl), or a bicyclic ring system containing at least one phenyl ring or an aromatic bicyclic ring containing only carbon atoms in the aromatic bicyclic ring system. The bicyclic aryl can be azulenyl, naphthyl, or a phenyl fused to a monocyclic cycloalkyl, a monocyclic cycloalkenyl, or a monocyclic heterocyclyl. The bicyclic aryl is attached to the parent molecular moiety through any carbon atom contained within the phenyl portion of the bicyclic system, or any carbon atom with the napthyl or azulenyl ring. The fused monocyclic cycloalkyl or monocyclic heterocyclyl portions of the bicyclic aryl are optionally substituted with one or two oxo and/or thia groups. Representative examples of the bicyclic aryls include, but are not limited to, azulenyl, naphthyl, dihydroinden-<NUM>-yl, dihydroinden-<NUM>-yl, dihydroinden-<NUM>-yl, dihydroinden-<NUM>-yl, <NUM>,<NUM>-dihydroindol-<NUM>-yl, <NUM>,<NUM>-dihydroindol-<NUM>-yl, <NUM>,<NUM>-dihydroindol-<NUM>-yl, <NUM>,<NUM>-dihydroindol-<NUM>-yl, inden-<NUM>-yl, inden-<NUM>-yl, inden-<NUM>-yl, inden-<NUM>-yl, dihydronaphthalen-<NUM>-yl, dihydronaphthalen-<NUM>-yl, dihydronaphthalen-<NUM>-yl, dihydronaphthalen-<NUM>-yl, <NUM>,<NUM>,<NUM>,<NUM>-tetrahydronaphthalen-<NUM>-yl, <NUM>,<NUM>,<NUM>,<NUM>-tetrahydronaphthalen-<NUM>-yl, <NUM>,<NUM>-dihydrobenzofuran-<NUM>-yl, <NUM>,<NUM>-dihydrobenzofuran-<NUM>-yl, <NUM>,<NUM>-dihydrobenzofuran-<NUM>-yl, <NUM>,<NUM>-dihydrobenzofuran-<NUM>-yl, benzo[d][<NUM>,<NUM>]dioxol-<NUM>-yl, benzo[d][<NUM>,<NUM>]dioxol-<NUM>-yl, <NUM>-chromen-<NUM>-on-<NUM>-yl, <NUM>-chromen-<NUM>-on-<NUM>-yl, <NUM>-chromen-<NUM>-on-<NUM>-yl, <NUM>-chromen-<NUM>-on-<NUM>-yl, isoindoline-<NUM>,<NUM>-dion-<NUM>-yl, isoindoline-<NUM>,<NUM>-dion-<NUM>-yl, inden-<NUM>-on-<NUM>-yl, inden-<NUM>-on-<NUM>-yl, inden-<NUM>-on-<NUM>-yl, inden-<NUM>-on-<NUM>-yl, <NUM>,<NUM>-dihydrobenzo[b][<NUM>,<NUM>]dioxin-<NUM>-yl, <NUM>,<NUM>-dihydrobenzo[b][<NUM>,<NUM>]dioxin-<NUM>-yl, <NUM>-benzo[b][<NUM>,<NUM>]oxazin3(<NUM>)-on-<NUM>-yl, 2Hbenzo[b][<NUM>,<NUM>]oxazin3(<NUM>)-on-<NUM>-yl, <NUM> benzo[b][<NUM>,<NUM>]oxazin3(<NUM>)-on-<NUM>-yl, 2Hbenzo[b][<NUM>,<NUM>]oxazin3(<NUM>)-on-<NUM>-yl, benzo[d]oxazin-<NUM>(<NUM>)-on-<NUM>-yl, benzo[d]oxazin-<NUM>(<NUM>)-on-<NUM>-yl, benzo[d]oxazin-<NUM>(<NUM>)-on-<NUM>-yl, benzo[d]oxazin-<NUM>(<NUM>)-on-<NUM>-yl, quinazolin-<NUM>(<NUM>)-on-<NUM>-yl, quinazolin-<NUM>(<NUM>)-on-<NUM>-yl, quinazolin-<NUM>(<NUM>)-on-<NUM>-yl, quinazolin-<NUM>(<NUM>)-on-<NUM>-yl, quinoxalin-<NUM>(<NUM>)-on-<NUM>-yl, quinoxalin-<NUM>(<NUM>)-on-<NUM>-yl, quinoxalin-<NUM>(<NUM>)-on-<NUM>-yl, quinoxalin-<NUM>(<NUM>)-on-<NUM>-yl, benzo[d]thiazol-<NUM>(<NUM>)-on-<NUM>-yl, benzo[d]thiazol-<NUM>(<NUM>)-on-<NUM>-yl, benzo[d]thiazol-<NUM>(<NUM>)-on-<NUM>-yl, and, benzo[d]thiazol-<NUM>(<NUM>)-on-<NUM>-yl. In certain examples, the bicyclic aryl is (i) naphthyl or (ii) a phenyl ring fused to either a <NUM> or <NUM> membered monocyclic cycloalkyl, a <NUM> or <NUM> membered monocyclic cycloalkenyl, or a <NUM> or <NUM> membered monocyclic heterocyclyl, wherein the fused cycloalkyl, cycloalkenyl, and heterocyclyl groups are optionally substituted with one or two groups which are independently oxo or thia.

The term "arylalkyl," " alkylaryl," and "arylalkyl-" as used herein, means an aryl group, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of arylalkyl include, but are not limited to, benzyl, <NUM>-phenylethyl, <NUM>-phenylpropyl, and <NUM>-naphth-<NUM>-ylethyl.

The terms "cyano" and "nitrile" as used herein, mean a -CN group.

The term "cycloalkyl" as used herein, means a monocyclic or a bicyclic cycloalkyl ring system. Monocyclic ring systems are cyclic hydrocarbon groups containing from <NUM> to <NUM> carbon atoms, where such groups can be saturated or unsaturated, but not aromatic. In certain examples, cycloalkyl groups are fully saturated. Examples of monocyclic cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptyl, and cyclooctyl. Bicyclic cycloalkyl ring systems are bridged monocyclic rings or fused bicyclic rings. Bridged monocyclic rings contain a monocyclic cycloalkyl ring where two non-adjacent carbon atoms of the monocyclic ring are linked by an alkylene bridge of between one and three additional carbon atoms (i.e., a bridging group of the form -(CH<NUM>)w-, where w is <NUM>, <NUM>, or <NUM>). Representative examples of bicyclic ring systems include, but are not limited to, bicyclo[<NUM>. <NUM>]heptane, bicyclo[<NUM>. <NUM>]heptane, bicyclo[<NUM>. <NUM>]octane, bicyclo[<NUM>. <NUM>]nonane, bicyclo[<NUM>. <NUM>]nonane, and bicyclo[<NUM>. <NUM>]nonane. Fused bicyclic cycloalkyl ring systems contain a monocyclic cycloalkyl ring fused to either a phenyl, a monocyclic cycloalkyl, a monocyclic cycloalkenyl, a monocyclic heterocyclyl, or a monocyclic heteroaryl. The bridged or fused bicyclic cycloalkyl is attached to the parent molecular moiety through any carbon atom contained within the monocyclic cycloalkyl ring. Cycloalkyl groups are optionally substituted with one or two groups which are independently oxo or thia. In certain examples, the fused bicyclic cycloalkyl is a <NUM> or <NUM> membered monocyclic cycloalkyl ring fused to either a phenyl ring, a <NUM> or <NUM> membered monocyclic cycloalkyl, a <NUM> or <NUM> membered monocyclic cycloalkenyl, a <NUM> or <NUM> membered monocyclic heterocyclyl, or a <NUM> or <NUM> membered monocyclic heteroaryl, wherein the fused bicyclic cycloalkyl is optionally substituted by one or two groups which are independently oxo or thia.

"Cycloalkenyl" as used herein refers to a monocyclic or a bicyclic cycloalkenyl ring system. Monocyclic ring systems are cyclic hydrocarbon groups containing from <NUM> to <NUM> carbon atoms, where such groups are unsaturated (i.e., containing at least one annular carbon-carbon double bond), but not aromatic. Examples of monocyclic ring systems include cyclopentenyl and cyclohexenyl. Bicyclic cycloalkenyl rings are bridged monocyclic rings or a fused bicyclic rings. Bridged monocyclic rings contain a monocyclic cycloalkenyl ring where two non-adjacent carbon atoms of the monocyclic ring are linked by an alkylene bridge of between one and three additional carbon atoms (i.e., a bridging group of the form -(CH<NUM>)w- , where w is <NUM>, <NUM>, or <NUM>). Representative examples of bicyclic cycloalkenyls include, but are not limited to, norbornenyl and bicyclo[<NUM>. <NUM>]oct-<NUM>-enyl. Fused bicyclic cycloalkenyl ring systems contain a monocyclic cycloalkenyl ring fused to either a phenyl, a monocyclic cycloalkyl, a monocyclic cycloalkenyl, a monocyclic heterocyclyl, or a monocyclic heteroaryl. The bridged or fused bicyclic cycloalkenyl is attached to the parent molecular moiety through any carbon atom contained within the monocyclic cycloalkenyl ring. Cycloalkenyl groups are optionally substituted with one or two groups which are independently oxo or thia.

The term "halo" or "halogen" as used herein, means Cl, Br, I or F.

The term "haloalkyl" as used herein, means at least one halogen, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of haloalkyl include, but are not limited to, chloromethyl, <NUM>-fluoroethyl, trifluoromethyl, pentafluoroethyl, and <NUM>-chloro-<NUM>-fluoropentyl.

The term "heteroaryl," as used herein, means a monocyclic heteroaryl or a bicyclic ring system containing at least one heteroaromatic ring. The monocyclic heteroaryl can be a <NUM> or <NUM> membered ring. The <NUM> membered ring consists of two double bonds and one, two, three or four nitrogen atoms and optionally one oxygen or sulfur atom. The <NUM> membered ring consists of three double bonds and one, two, three or four nitrogen atoms. The <NUM> or <NUM> membered heteroaryl is connected to the parent molecular moiety through any carbon atom or any nitrogen atom contained within the heteroaryl. Representative examples of monocyclic heteroaryl include, but are not limited to, furyl, imidazolyl, indolyl, <NUM>-methyl-indolyl, isoxazolyl, isothiazolyl, oxadiazolyl, oxazolyl, pyridinyl, pyridazinyl, pyrimidinyl, pyrazinyl, pyrazolyl, pyrrolyl, tetrazolyl, thiadiazolyl, thiazolyl, thienyl, triazolyl, and triazinyl. The bicyclic heteroaryl consists of a monocyclic heteroaryl fused to a phenyl, a monocyclic cycloalkyl, a monocyclic cycloalkenyl, a monocyclic heterocyclyl, or a monocyclic heteroaryl. The fused cycloalkyl or heterocyclyl portion of the bicyclic heteroaryl group is optionally substituted with one or two groups which are independently oxo or thia. When the bicyclic heteroaryl contains a fused cycloalkyl, cycloalkenyl, or heterocyclyl ring, then the bicyclic heteroaryl group is connected to the parent molecular moiety through any carbon or nitrogen atom contained within the monocyclic heteroaryl portion of the bicyclic ring system. When the bicyclic heteroaryl is a monocyclic heteroaryl fused to a phenyl ring or a monocyclic heteroaryl, then the bicyclic heteroaryl group is connected to the parent molecular moiety through any carbon atom or nitrogen atom within the bicyclic ring system. Representative examples of bicyclic heteroaryl include, but are not limited to, benzimidazolyl, benzofuranyl, benzothienyl, benzoxadiazolyl, benzoxathiadiazolyl, benzothiazolyl, cinnolinyl, <NUM>,<NUM>-dihydroquinolin-<NUM>-yl, <NUM>,<NUM>-dihydroisoquinolin-<NUM>-yl, furopyridinyl, indazolyl, indolyl, isoquinolinyl, naphthyridinyl, quinolinyl, purinyl, <NUM>,<NUM>,<NUM>,<NUM>-tetrahydroquinolin-<NUM>-yl, <NUM>,<NUM>,<NUM>,<NUM>-tetrahydroquinolin-<NUM>-yl, <NUM>,<NUM>,<NUM>,<NUM>-tetrahydroquinolin-<NUM>-yl, <NUM>,<NUM>,<NUM>,<NUM>-tetrahydroisoquinolin-<NUM>-yl, thienopyridinyl, <NUM>,<NUM>,<NUM>,<NUM>-tetrahydrobenzo[c][<NUM>,<NUM>,<NUM>]oxadiazolyl, and <NUM>,<NUM>-dihydrobenzo[c][<NUM>,<NUM>,<NUM>]oxadiazol-<NUM>(<NUM>)-onyl. In certain examples, the fused bicyclic heteroaryl is a <NUM> or <NUM> membered monocyclic heteroaryl ring fused to either a phenyl ring, a <NUM> or <NUM> membered monocyclic cycloalkyl, a <NUM> or <NUM> membered monocyclic cycloalkenyl, a <NUM> or <NUM> membered monocyclic heterocyclyl, or a <NUM> or <NUM> membered monocyclic heteroaryl, wherein the fused cycloalkyl, cycloalkenyl, and heterocyclyl groups are optionally substituted with one or two groups which are independently oxo or thia.

The term "heteroarylalkyl" and " alkylheteroaryl" as used herein, means a heteroaryl, as defined herein, appended to the parent molecular moiety through an alkyl group, as defined herein. Representative examples of heteroarylalkyl include, but are not limited to, fur-<NUM>-ylmethyl, <NUM>-imidazol-<NUM>-ylmethyl, <NUM>-imidazol-<NUM>-ylmethyl, <NUM>-(pyridine-<NUM>-yl)ethyl, pyridine-<NUM>-ylmethyl, pyridine-<NUM>-ylmethyl, pyrimidin-<NUM>-ylmethyl, <NUM>-(pyrimidin-<NUM>-yl)propyl, thien-<NUM>-ylmethyl, and thien-<NUM>-ylmethyl.

The terms "heterocyclyl" or "heterocycloalkyl" as used herein, means a monocyclic heterocycle or a bicyclic heterocycle. The monocyclic heterocycle is a <NUM>, <NUM>, <NUM>, <NUM> or <NUM> membered ring containing at least one heteroatom independently selected from the group consisting of O, N, and S where the ring is saturated or unsaturated, but not aromatic. The <NUM> or <NUM> membered ring contains <NUM> heteroatom selected from the group consisting of O, N and S. The <NUM> membered ring can contain zero or one double bond and one, two or three heteroatoms selected from the group consisting of O, N and S. The <NUM> or <NUM> membered ring contains zero, one or two double bonds and one, two or three heteroatoms selected from the group consisting of O, N and S. The monocyclic heterocycle is connected to the parent molecular moiety through any carbon atom or any nitrogen atom contained within the monocyclic heterocycle. Representative examples of monocyclic heterocycle include, but are not limited to, azetidinyl, azepanyl, aziridinyl, diazepanyl, <NUM>,<NUM>-dioxanyl, <NUM>,<NUM>-dioxolanyl, <NUM>,<NUM>-dithiolanyl, <NUM>,<NUM>-dithianyl, imidazolinyl, imidazolidinyl, isothiazolinyl, isothiazolidinyl, isoxazolinyl, isoxazolidinyl, morpholinyl, oxadiazolinyl, oxadiazolidinyl, oxazolinyl, oxazolidinyl, piperazinyl, piperidinyl, pyranyl, pyrazolinyl, pyrazolidinyl, pyrrolinyl, pyrrolidinyl, tetrahydrofuranyl, tetrahydrothienyl, thiadiazolinyl, thiadiazolidinyl, thiazolinyl, thiazolidinyl, thiomorpholinyl, <NUM>,<NUM>-dioxidothiomorpholinyl (thiomorpholine sulfone), thiopyranyl, and trithianyl. The bicyclic heterocycle is a monocyclic heterocycle fused to either a phenyl, a monocyclic cycloalkyl, a monocyclic cycloalkenyl, a monocyclic heterocycle, or a monocyclic heteroaryl. The bicyclic heterocycle is connected to the parent molecular moiety through any carbon atom or any nitrogen atom contained within the monocyclic heterocycle portion of the bicyclic ring system. Representative examples of bicyclic heterocyclyls include, but are not limited to, <NUM>,<NUM>-dihydrobenzofuran-<NUM>-yl, <NUM>,<NUM>-dihydrobenzofuran-<NUM>-yl, indolin-<NUM>-yl, indolin-<NUM>-yl, indolin-<NUM>-yl, <NUM>,<NUM>-dihydrobenzothien-<NUM>-yl, decahydroquinolinyl, decahydroisoquinolinyl, octahydro-<NUM>-indolyl, and octahydrobenzofuranyl. Heterocyclyl groups are optionally substituted with one or two groups which are independently oxo or thia. In certain examples, the bicyclic heterocyclyl is a <NUM> or <NUM> membered monocyclic heterocyclyl ring fused to phenyl ring, a <NUM> or <NUM> membered monocyclic cycloalkyl, a <NUM> or <NUM> membered monocyclic cycloalkenyl, a <NUM> or <NUM> membered monocyclic heterocyclyl, or a <NUM> or <NUM> membered monocyclic heteroaryl, wherein the bicyclic heterocyclyl is optionally substituted by one or two groups which are independently oxo or thia.

The term "hydroxy" as used herein, means an -OH group.

The term "nitro" as used herein, means a -NO<NUM> group.

The term "oxo" as used herein means a =O group.

The term "thia" as used herein means a -S- group.

The term "saturated" as used herein means the referenced chemical structure does not contain any multiple carbon-carbon bonds. For example, a saturated cycloalkyl group as defined herein includes cyclohexyl, cyclopropyl, and the like.

The term "unsaturated" as used herein means the referenced chemical structure contains at least one multiple carbon-carbon bond, but is not aromatic. For example, a unsaturated cycloalkyl group as defined herein includes cyclohexenyl, cyclopentenyl, cyclohexadienyl, and the like.

As used herein, the term "individual" or "patient," used interchangeably, refers to any animal, including mammals, preferably mice, rats, other rodents, rabbits, dogs, cats, swine, cattle, sheep, horses, or primates, and most preferably humans.

As used herein, the phrase "therapeutically effective amount" refers to the amount of active compound or pharmaceutical agent that elicits the biological or medicinal response that is being sought in a tissue, system, animal, individual or human by a researcher, veterinarian, medical doctor or other clinician.

In certain embodiments, a therapeutically effective amount can be an amount suitable for.

As used here, the terms "treatment" and "treating" means (i) ameliorating the referenced disease state, for example, ameliorating a disease, condition or disorder in an individual who is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder (i.e., reversing or improving the pathology and/or symptomatology) such as decreasing the severity of disease; or (ii) eliciting the referenced biological effect (e.g., IDO modulation or tryptophan degradation inhibition).

Manifestation of amelioration of a disease condition with underlying IDO-mediated immunosuppression may require the concomitant or sequential administration of additional therapeutic agents, such as antineoplastic agents in the case of cancer, or antiretroviral agents in the case of viral diseases. For example, administration of IDO inhibitors for the treatment of cancer does not always produce a direct antitumor effect when used as a single agent. However, when combined with chemotherapeutic drugs (antineoplastic) the antitumor effect observed is higher than the sum of effects of each agent alone.

As used herein, the terms "catalytic pocket", "catalytic site", "active site" collectively and indistinctly refer to a region of the enzyme that contains amino acid residues responsible for the substrate binding (charge, hydrophobicity, steric hindrance) and catalytic amino acid residues which act as proton donors or acceptors or are responsible for binding a cofactor and participate in the catalysis of a chemical reaction.

As used herein, the phrase "pharmaceutically acceptable salt" refers to both pharmaceutically acceptable acid and base addition salts and solvates. Such pharmaceutically acceptable salts include salts of acids such as hydrochloric, phosphoric, hydrobromic, sulfuric, sulfinic, formic, toluenesulfonic, methanesulfonic, nitric, benzoic, citric, tartaric, maleic, hydroiodic, alkanoic such as acetic, HOOC-(CH<NUM>)n-COOH where n is <NUM>-<NUM>, and the like. Non-toxic pharmaceutical base addition salts include salts of bases such as sodium, potassium, calcium, ammonium, and the like. Those skilled in the art will recognize a wide variety of non-toxic pharmaceutically acceptable addition salts.

As used herein, the term "indoximod" refers to <NUM>-methyl-D-tryptophan, also referred to as D-1MT or D1mT.

As used herein, the term "prodrug of indoximod" refers to any substance that after in vivo administration is metabolized to produce indoximod as one of the main metabolites.

All reagents and solvents were purchased from commercial sources. All commercial reagents and solvents were used as received without further purification. The reactions were monitored using analytical thin layer chromatography (TLC) with <NUM> EM Science silica gel plates (60F-<NUM>). The developed TLC plates were visualized by short wave UV light (<NUM>) or immersion in potassium permanganate solution followed by heating on a hot plate. Flash chromatography was performed with Selecto Scientific silica gel, <NUM>-<NUM> particle sizes. All reactions were performed in flame or oven-dried glassware under a nitrogen atmosphere. All reactions were stirred magnetically at ambient temperature unless otherwise indicated. <NUM>H NMR spectra were obtained with a Bruker DRX400, Varian VXR400 or VXR300. <NUM>H NMR spectra were reported in parts per million (δ) relative to TMS (<NUM>), DMSO-d<NUM> (<NUM>) or CD<NUM>OD (<NUM>) as an internal reference. All <NUM>H NMR spectra were taken in CDCl<NUM> unless otherwise indicated. Comparative compounds and salts are marked *.

To a suspension of D-1MT (<NUM>, <NUM> mmol) in ethanol (<NUM>) at <NUM> was added SOCl<NUM> (<NUM>, <NUM> mmol) and the mixture was stirred at <NUM> overnight. After cooling to rt, the solvent was distilled-off and the crude was diluted with diethyl ether (<NUM>), the white solid was filtered-off and washed with dry ether to afford the desired product (<NUM>, <NUM> %).

To a suspension of D-1MT (<NUM>, <NUM> mmol) in isopropanol (<NUM>) at <NUM> rt, was added SOCl<NUM> (<NUM>, <NUM> mmol) and the mixture was stirred at <NUM> overnight. After cooling to rt, the solvent was distilled-off and the crude was basified with <NUM> % aq NaHCO<NUM> (<NUM>), the product was extracted with CH<NUM>Cl<NUM>, the combined organic extract was dried over Na<NUM>SO<NUM> and the solvent was distilled-off under reduced pressure. The free base was converted to its HCl salt by adding dry HCl in dioxane, the solvent was removed under reduced pressure to afford the desired product as white solid (<NUM>, <NUM>%).

To a stirred solution of D-1MT (<NUM>, <NUM> mmol) in <NUM>:<NUM> THF/<NUM> NaHCO<NUM> (<NUM>, <NUM> mmol) was added the appropriate chloroformate dropwise. The mixture was allowed to stir for <NUM>. and the solution was diluted with water and extracted with ether 2x. The aqueous layer was cooled to <NUM> oC and conc HCl solution was added to adjust the pH to ~<NUM>. The cold aqueous layer was immediately extracted with ethyl acetate and the combined organic layers were washed with water, brine and dried. The solvent was removed under reduced pressure to afford crude the carbamate. The crude was purified by column chromatography and treated with activated charcoal to afford the pure carbamate.

To a mixture of D-1MT (<NUM>, <NUM> mmol) in dioxane (<NUM>) at <NUM> was added NaOH (<NUM> dissolved in <NUM> DI water), followed by the addition of Boc<NUM>O. The reaction was stirred at <NUM> for <NUM> and stirred overnight at rt. The solution was concentrated under reduced pressure to approx. one third the original volume. The reaction was acidified with 1N HCl at <NUM> and the product was extracted with EtOAc. The organic extract was washed with brine and dried over Na<NUM>SO<NUM>, the solvent was evaporated under reduced pressure to afford the product that was used directly in the next step without further purification (<NUM>, <NUM>%).

In <NUM> of DMF was dissolved Nα-(tert-butoxycarbonyl)-<NUM>-methyl-D-tryptophan (<NUM>, <NUM> mmol) to which Cs<NUM>CO<NUM> (<NUM>, <NUM> mmol) and benzyl bromide (<NUM>, <NUM> mmol) was added. The resulting suspension was allowed to stir at room temperature for <NUM> hours. After the end of reaction (TLC), the DMF was removed under reduced pressure followed by suspending the residue in toluene/ethyl acetate before washing with distilled water (<NUM> x <NUM>) and brine. The organic layer was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by column chromatography on silica gel (<NUM>, <NUM>%).

Ethyl acetate (<NUM>) and MeOH (<NUM>) in a RB flask equipped with a septum and a needle vent were cooled in an ice bath with stirring. Acetyl chloride (<NUM>) was added slowly. The resulting solution was stirred at <NUM> for <NUM> minutes and MeOH (<NUM>) was added. A flask containing benzyl Nα-(tert-butoxycarbonyl)-<NUM>-methyl-D-tryptophanate (<NUM>, <NUM> mmol) was placed in an ice bath and the cold, freshly prepared HCl (<NUM> in EtOAc) was poured into the flask containing benzyl Nα-(tert-butoxycarbonyl)-<NUM>-methyl-D-tryptophanate slowly. The solution was stirred vigorously at <NUM> for <NUM> where the formation of a white suspension was observed and the flask was removed from the ice bath. The suspension was allowed to stir vigorously for <NUM>. The solution was cooled in an ice bath diluted with ether (<NUM>) and the suspension was filtered and the solid cake washed with cold ether. The solid was allowed to dry under high vacuum and the desired product was isolated as a colorless solid (<NUM>, <NUM>%). <NUM>H NMR (d<NUM>-dmdso); <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (s, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>).

To a solution of N-(tert-butoxycarbonyl)-<NUM>-methyl-D-tryptophan (<NUM> mmol), appropriate alcohol or amine (<NUM> mmol) and HATU (<NUM> mmol) in acetonitrile (<NUM>) at <NUM> was added DIPEA (<NUM> mmol) and the solution was allowed to warm to rt. After stirring overnight (<NUM>), the reaction was diluted with water (<NUM>) and the product was extracted with CH<NUM>Cl<NUM> (<NUM> x <NUM>). The combined organic extract was washed with water (<NUM> mLx1), brine (25mlx1) dried over Na<NUM>SO<NUM> and concentrated under reduced pressure to afford the crude. Chromatographic purification afforded the desired product.

To a solution of NLG-<NUM>-A-E43 (<NUM>, <NUM> mmol) in THF (<NUM>) was added water (<NUM>) and lithium monohydrate (<NUM>, <NUM> mmol) and the mixture stirred under ambient temperature for <NUM>. The mixture was neutralized with <NUM> HCl (at <NUM>) and poured into ice cold water (<NUM>). The aqueous layer was extracted with EtOAc (<NUM> x <NUM>). The combined organic layers were dried over Na<NUM>SO<NUM> and concentrated. The crude product was purified by flash column chromatography to afford the desired product as white solid (<NUM>, <NUM> %). <NUM>H NMR: <NUM> and <NUM> (two s, <NUM>). <NUM>-<NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dt, <NUM>, J = <NUM>)
<CHM>.

To a mixture of tBoc protected amine (<NUM> mmol) in dioxane (<NUM>) at rt was added HCl (<NUM>, <NUM> solution in dioxane). After stirring for <NUM>, the solvent was distilled-off under reduced pressure. The residue was stirred with methyl tert-butyl ether (<NUM>), the solid was filtered and dried under reduced pressure to afford the desired product.

The following compounds were synthesized following procedures described in the above sections.

To a solution of NLG-<NUM>-B. <NUM>-E15 (<NUM>, <NUM> mmol) in CH<NUM>Cl<NUM> (<NUM>) was added HCl (<NUM>, <NUM> solution in dioxane) at <NUM> and the solution was allowed to warm to rt. After stirring for <NUM>, the solvent was evaporated and the reaction was diluted with trifluoroacetic acid (<NUM>) and the solution was stirred for <NUM> at rt. After evaporating trifluoroacetic acid the reaction was diluted with dry HCl solution (<NUM>, <NUM> solution in dioxane) and the mixture was stirred for <NUM>. The solvent was evaporated under reduced pressure, the product was triturated with ethanol:ether (<NUM>:<NUM>, <NUM>) and the product was filtered and washed with dry ether (<NUM>). The product was dried under reduced pressure (<NUM>, <NUM>%). <NUM>H NMR (<NUM>, CD<NUM>OD): <NUM>-<NUM> (m, <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>).

Dioxane (<NUM>) and MeOH (<NUM>, <NUM> mmol) in a RB flask equipped with a septum and a needle vent were cooled in an ice bath with stirring. Acetyl chloride (<NUM>, <NUM> mmol) was added slowly. The resulting solution was stirred at <NUM> for <NUM> minutes and MeOH (<NUM>) was added. A flask containing NLG-<NUM>-A-E22 (<NUM>, <NUM> mmol) was placed in an ice bath and the cold, freshly prepared HCl (<NUM> in dioxane) was poured into the flask containing NLG-<NUM>-A-E22 slowly. The solution was allowed to warm to RT and stirred vigorously for <NUM>. The solvent was removed using rotary evaporator to afford pure white solid (<NUM>, <NUM>%). (DMSO-d6) <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (s, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (br s, <NUM>), <NUM> (d, <NUM>, J = <NUM>).

A solution of NLG1558-A-E23 (<NUM>, <NUM> mmol ) in THF (<NUM>) at <NUM> was added TFA (<NUM>, <NUM> mmol) and water (<NUM>, <NUM> mmol) and the cooling bath was removed, the mixture was stirred at rt for <NUM>. HCl (<NUM>, <NUM> mmol; <NUM> solution in dioxane) was added and continued stirring for <NUM>. The reaction was stirred at <NUM> for <NUM> minutes. The precipitated white solid was filtered and washed with MTBE to afford the hydrochloride salt (<NUM>, <NUM>%). <NUM>H NMR (<NUM>, DMSO-d<NUM>): <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (s, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>).

Appropriate D-tryptophanate hydrochloride ester (<NUM>, <NUM> mmol) and appropriate acid (<NUM> mmol) were stirred in acetonitrile (<NUM>) at <NUM>. HATU (<NUM>, <NUM> mmol) and iPr<NUM>NEt (<NUM>, <NUM> mmol) were added and the reaction stirred overnight at room temperature. The solvent was removed under reduced pressure and the crude was diluted with water (<NUM>) and dichloromethane (<NUM>). The organic layer was separated and the aqueous layer was extracted with dichloromethane (<NUM> x <NUM>). The combined organic layer was washed with brine (<NUM>), dried over Na<NUM>SO<NUM>, and concentrated under reduced pressure. The crude product was purified by flash column chromatography to afford the desired product.

tert-Butyl(S)-<NUM>-(((R)-<NUM>-(benzyloxy)-<NUM>-(<NUM>-methyl-<NUM>-indol-<NUM>-yl)-<NUM>-oxopropan-<NUM>-yl)amino)-<NUM>-((tert-butoxycarbonyl)amino)-<NUM>-oxopentanoate (<NUM>, <NUM> mmol) was suspended in MeOH (<NUM>) and THF (<NUM>). After cooling to <NUM>, NaOH sol'n (<NUM>, <NUM>) was added and the reaction stirred for <NUM>. The solution was acidified with <NUM> HCl to pH = <NUM> and the solvents were concentrated under reduced pressure (<NUM>). The solution was partitioned between water and DCM in a separatory funnel and the organic layer was collected. The aqueous layer was extracted with DCM (<NUM> x <NUM>) and the combined organic layer was washed with water and brine. Chromatographic purification afforded the desired product (<NUM>, <NUM>%). <NUM>H NMR(Chloroform-d, <NUM>): δ = <NUM> (s, <NUM>), <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (s, <NUM>), <NUM> (dt, <NUM>, J = <NUM>, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (apparent d overlapped with CDCl<NUM>, <NUM>,), <NUM> (d, <NUM>, J = <NUM>).

To Nα-((S)-<NUM>-(tert-butoxy)-<NUM>-((tert-butoxycarbonyl)amino)-<NUM>-oxopentanoyl)-<NUM>-methyl-D-tryptophan (<NUM>, <NUM> mmol) was added HCl (<NUM> in dioxane) (<NUM>). The resulting solution was allowed to stir at room temperature for <NUM> hours. The solution was concentrated and the solid was dissolved in MeOH and treated with activated charcoal and heated to <NUM> for <NUM>. The solution was filtered through celite and the filtrate concentrated to afford the desired product as a beige solid (<NUM>, <NUM> %). <NUM>H NMR (DMSO-d<NUM>, <NUM>): (mixture of rotamers) <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (q, <NUM>, J = <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (tt, <NUM>, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dt, <NUM>, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (br s, <NUM>), <NUM> - <NUM> (m, <NUM>). <CHM>
<CHM>.

To a solution of appropriate amide (<NUM> mmol) in THF (<NUM>) was added water (<NUM>) and lithium monohydrate (<NUM>, <NUM> mmol) and the mixture stirred under ambient temperature for <NUM>. The mixture was neutralized with <NUM> HCl (at <NUM>) and poured into ice cold water (<NUM>). The aqueous layer was extracted with EtOAc (<NUM> x <NUM>). The combined organic layers were dried over Na<NUM>SO<NUM> and concentrated. The crude product was purified by flash column chromatography to afford the desired product.

To a solution of appropriate tBoc protected amine (<NUM> mmol) in dioxane (<NUM>) was added HCl solution (<NUM>, <NUM> solution in dioxane) at <NUM>. The solution was allowed to warm to rt and stirred vigorously for <NUM>-<NUM>. The solvent was removed using rotary evaporator. The solid was diluted with dry ether (<NUM>) and the product was filtered to afford the crude product. The crude was dried under high vacuum to afford the desired product.

To a solution NLG-<NUM> free base (<NUM>, <NUM> mmol) in acetonitrile (<NUM>) at <NUM> was added Boc<NUM>O (<NUM>, <NUM> mmol) and the reaction was allowed to warm to RT and stirred for <NUM>. The solvent was removed under reduced pressure and the crude was purified by column chromatography to afford the desired product (<NUM>, <NUM>%). <NUM>H NMR: <NUM> (s, <NUM>), <NUM>-<NUM> (m, <NUM>, <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>).

To a solution of NLG-<NUM>-A-E24 (<NUM>, <NUM> mmol) in dry pyridine (<NUM>) at <NUM> was added POCl<NUM> and the solution was allowed to warm to rt. After stirring overnight (<NUM>), ethanol (<NUM>) was added and the reaction continued for <NUM>. The solvent was removed under reduced pressure and the crude was purified by column chromatography (<NUM>, <NUM>%). <NUM>H NMR: <NUM> (t, <NUM>, J = <NUM>), <NUM> (s, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> and <NUM> (two s, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>).

To a solution NLG-<NUM>-B-E24 (<NUM>, <NUM> mmol) in dry CH<NUM>Cl<NUM> (<NUM>) at <NUM> was added anhydrous HCl (<NUM>, <NUM> solution in dioxane) and the mixture was allowed to warm to rt. After stirring for <NUM>, the solvent was removed under reduced pressure and the crude was washed with dry ether (<NUM> x <NUM>). The white solid was filtered and the product was dried under reduced pressure (<NUM>, <NUM> %). (CD3OD-d<NUM>) <NUM> (td, <NUM>, J = <NUM>, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (s, <NUM>), <NUM> (h, <NUM>, J = <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>).

To an ice cold aqueous HCl (<NUM>, <NUM> mmol; <NUM>) solution was added D1MT (<NUM>, <NUM> mmol). After stirring for <NUM> minutes, the clear solution was evaporated under reduced pressure and the crude was evaporated thrice with Ethanol (<NUM>). The crude was stirred in Ethanol and tert-butylmethylether and filtered to afford the desired product (<NUM>, <NUM>%).

An alternative method was developed where ~ <NUM> of D-1MT was suspended in <NUM> glass bottle with <NUM> of acetonitrile. <NUM> HCl solution pre-dissolved in acetonitrile (<NUM>/mL) was added into the D-1MT free form solution according to <NUM>:<NUM> molar ratio to free base:acid, and then kept shaking at room temperature overnight to form salt. The filtered solid was dried under vacuum at <NUM> overnight. A white powder (<NUM>) was obtained by the above process, and characterized by XRPD, DSC and TGA (<FIG>). The purity was <NUM>% area based on the HPLC analysis, and the stoichiometry was analyzed by ELSD, the calculated molar ratio (API:HCl acid) were <NUM>:<NUM>. The powder was crystalline as assessed by polarized light microscopy (PLM) and by X-ray powder dispersion spectrometry (XRPD, <FIG>). The salt was anhydrous as assessed by thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) (<FIG>).

To a stirred solution methane sulfonic acid (<NUM>, <NUM> mmol) in DI water (<NUM>) was added D-1MT (<NUM>, <NUM> mmol) in <NUM> portions. The solution was stirred vigorously for <NUM> at <NUM> until the solution was homogeneous. The solution was concentrated under reduced pressure and the solid collected (<NUM>, <NUM>%). <NUM>H NMR(Methanol-d<NUM>, <NUM>): δ = <NUM> (s, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (ddd, <NUM>, J = <NUM>, <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (ddd, <NUM>, J = <NUM>, <NUM>, <NUM>), <NUM> (dd, <NUM>, J=<NUM>, <NUM>), <NUM> (dt, <NUM>, J = <NUM>, <NUM>).

To the solution of phosphoric acid (<NUM>, <NUM>. 87mmol) in deionized water (<NUM>) at <NUM>, was added D-1MT (<NUM>, <NUM>) portion wise and the mixture was stirred at <NUM> overnight. Solution was then concentrated to half of its original volume and allowed to stand at room temperature overnight. Resulting precipitate was filtered, washed with cold ethanol, and dried to yield NLG-<NUM> as white solid (<NUM>, <NUM>%). <NUM>H NMR (<NUM>, DMSO-d<NUM>) δ <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (ddd, <NUM>, J = <NUM>, <NUM>, <NUM>), <NUM> - <NUM> (m, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>).

An alternative method was developed where ~ <NUM> of D-1MT was suspended in <NUM> glass bottle with <NUM> of THF. <NUM> of H<NUM>PO<NUM> solution pre-dissolved in THF (<NUM>/mL) was added into the D-1MT free form solution according to <NUM>:<NUM> molar ratio to free base:acid, and then kept shaking at room temperature overnight to form salt. The filtered solid was dried under vacuum at <NUM> overnight, checked by XRPD, DSC, TGA and ELSD. A white powder (<NUM>) was obtained, which showed to be crystalline by PLM and XRPD pattern (<FIG>). The salt was anhydrous based on DSC and TGA data (<FIG>). The purity was <NUM>%, and the stoichiometry was analyzed by ELSD, the calculated molar ratio (free base:phosphoric acid) were <NUM>:<NUM>.

To a suspension of D-1MT (<NUM>, <NUM> mmol) in water/THF (<NUM>:<NUM>, <NUM>) at rt, was added <NUM> H<NUM>SO<NUM> (<NUM>, <NUM> mmol) and the mixture was stirred at rt overnight. The white solid was filtered-off and washed with cold THF to afford the sulfate salt of D-1MT (<NUM>, <NUM>%). (DMSO-d<NUM>) <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (dd, <NUM>, J = <NUM>, <NUM>), <NUM> (s, <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM> (t, <NUM>, J = <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM> (br s, <NUM>).

To a solution of free base (<NUM> mmol) in EtOH (<NUM>) at <NUM> was added phosphoric acid (<NUM> mmol; a solution in EtOH <NUM>) or (<NUM> mmol in case of diamine) and the mixture was allowed to warm to RT and stirred for <NUM>-<NUM>. The solvent was removed under reduced pressure and the residue was diluted with methyl tert-butylether (<NUM>), after stirring for <NUM>-<NUM> the solid was filtered and dried under reduced pressure to afford the desired product. For NLG-<NUM>-<NUM>, the free base was generated from NLG-<NUM>-<NUM> using ion-exchange resin.

To a solution of free base (<NUM>, <NUM>. 723mmol) in ethanol (<NUM>) at rt, was added methanesulfonic or benzenesulfonic acid (<NUM> mmol or <NUM> mmol in case of diamines) and the mixture was stirred at rt overnight. Ethanol was evaporated and the crude product was stirred in methyl tert-butyl ether for <NUM>-<NUM>. The precipitate was filtered and dried to yield the corresponding methanesulfonate or benzenesulfonate salt.

To a solution of free base (<NUM> mmol) in dry THF (<NUM>) at <NUM> was added sulfuric acid (<NUM> mmol or <NUM> mmol) as a solution in THF (<NUM>) and the solution was allowed to warm to rt. After stirring for <NUM>-<NUM>, the solvent was distilled-off and the crude was stirred with methyl tert-butyl ether, the solid was filtered and dried under vacuum to yield the desired product.

To a solution of <NUM>-(<NUM>-imidazol-<NUM>-yl)phenol (<NUM> mmol) (prepared according to <NPL>) in DMF (<NUM>) was added triethylamine (<NUM> mmol). After stirred for <NUM>, a solution of <NUM>,<NUM>'-Dimethoxytrityl chloride (<NUM> mmol) in DMF (<NUM>) was added dropwise. After stirred overnight under a nitrogen atmosphere, the reaction mixture was poured into ice water (<NUM>). The solid was filtered off, washed with cold water and dissolved in ethyl acetate. The organic layer was dried over Na2SO4 and concentrated the crude product was taken into next step without further purification. To a suspension of (R)-methyl <NUM>-amino-<NUM>-(<NUM>-methyl-<NUM>-indol-<NUM>-yl)propanoate (<NUM> mmol) (prepared as described by <NPL>) in DCM (<NUM>) was added triphosgene (<NUM> mmol) and Et3N (<NUM> mmol) at <NUM> oC. The solution was allowed to stir for <NUM> and was concentrated to dryness. The crude residue was used immediately in the next step without purification. The crude residue was dissolved in DCM (<NUM>), the phenyl imidazole derivative (<NUM> mmol) and DMAP (<NUM> mmol) were added. The resulting solution was allowed to stir at rt overnight. The solvent was removed under reduced pressure and the crude residue was filtered through a plug of silica gel and concentrated. To the residue was added MeOH (<NUM>) and AcOH (<NUM>) and the solution was stirred at rt for <NUM>. The solution was diluted with water and made basic with solid K2CO3 (pH ~ <NUM>-<NUM>). The aqueous was extracted with EtOAc and the combined organic layers were washed with water, brine and dried (Na2SO4). The crude residue was purified by column chromatography on silica gel afforded the compound (<NUM>% yield). <NUM> NMR: <NUM>-<NUM> (m, <NUM>), <NUM> (s,<NUM>), <NUM> (s,<NUM>), <NUM>-<NUM> (m, <NUM>), <NUM> (d, <NUM>, J = <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>), <NUM>-<NUM> (m, <NUM>).

D-1MT (HPLC purity <NUM>%) free base is a white powder and it displays birefringence, needle shape and crystalline appearance under the polarized light microscope (PLM) and by X-ray powder dispersion spectroscopy (XRPD) (<FIG>). It only shows single melt endothermic peak with onset at <NUM> by thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) and ~<NUM>% weight loss from <NUM>-<NUM>, indicating that is an anhydrate form. This crystalline form is non-hygroscopic (<NUM>% weight gain from <NUM>-<NUM> %RH), and does not show changes after dynamic vapor sorption method (DVS). Furthermore, stability studies of the solid powder form indicate that D-1MT is chemically stable at the tested conditions (<NUM>/<NUM>%RH, <NUM>, <NUM>/<NUM>%RH, <NUM> and <NUM>) for <NUM> weeks. Additionally, it is also stable in solution in <NUM> N HCl, and <NUM> phosphate buffers pH <NUM>-<NUM> at <NUM> for <NUM> hours, while it shows minor degradation (<NUM>%-<NUM>%) in pH <NUM> and pH <NUM> buffers with <NUM>% H<NUM>O<NUM> (the most impurity was RRT=<NUM>).

The solubility of indoximod as free base in buffered or un-buffered solutions, as well as in simulated biological fluids (SGF, FaSSIF or FeSSIF) is shown in <FIG> (open symbols). Solubility of indoximod in aqueous solutions of pH <NUM>-<NUM> is <NUM>-<NUM>/mL, with higher solubility at pH <<NUM> or ><NUM>. This low solubility at neutral pH range is likely due to the high molecular packing energy of indoximod in the crystal, which is reflected by the very high melting point of <NUM>. This low solubility of indoximod in the pH range corresponding to intestinal pH may in part explain the limiting dose absorption at doses higher than <NUM> in humans. Therefore, we studied whether salts or sprayed dry dispersions of indoximod could increase solubility and exposure after oral dosing.

Several salts of indoximod were manufactured and their physicochemical properties were evaluated (Table <NUM>). The hydrochloride, sulfate, phosphate, hemi-phosphate, mesylate and hemi-mesylate salts were solid white powders that showed crystalline properties by PLM and XRPD and were anhydrous by TGA. These salts showed lower melting point than the free base, suggesting increased solubility in water in the range of pH between ><NUM> and <<NUM>. Most of these salts showed increases of solubility to ~<NUM>-<NUM>/mL in water and <NUM>-<NUM>/mL in SGF, with the hydrochloride salt showing a very significant increase to ><NUM>/mL in water or SGF.

Another indoximod salt tested was the maleic acid salt, which showed low melting point of <NUM> and poor crystallinity by PLM and XRPD. This salt has the appearance of a sticky white powder of hydrate or solvate form (<NUM>% weight loss by TGA).

The tosylate salt shows the appearance of a brown oil, which may be advantageous as that could increase the intestinal absorption of the active ingredient.

Other salts had less favorable physico-chemical properties. For example, lactate and N-methyl glucamine did not form a salt with indoximod, and the crystal showed a mixture of indoximod free base crystals and N-methyl glucamine or lactate crystals.

The sodium salt did not show crystalline morphology, it was a hydrate or solvate with very low melting and multiple decomposition peaks by TGA or DSC and thus it was not further characterized.

A list of indoximod sprayed dry dispersion (SDD) formulations were made in order to assess whether any SDD formulation was able to increase the molecular absorption by generating and maintaining a supersaturated state of indoximod in gastrointestinal fluid so that its absorption could be enhanced. In this study, SDD formulations were made by two methods: hot process spray dry - formulation solution heated up to <NUM> before spraying dry, and basic spray dry - formulation pH raised up to ~ <NUM> (room temperature) before spraying dry. The performance of each SDD formulation was investigated by in-vitro dissolution test in simulated gastric buffer (GB) and simulated intestinal fluid (SIF). As shown in Table <NUM>, CmaxGB represented the maximum concentration of indoximod in solution when enough of the SDD formulation was dissolved in GB for <NUM>; Cmax90 represents the maximum indoximod concentration when the SDD was dissolved in SIF for <NUM>; UltraC<NUM> represents the concentration in SIF after <NUM> of dissolution followed by ultracentrifugation to remove any particulates and UltraC<NUM> represents the concentration in SIF after <NUM> of dissolution followed by ultracentrifugation to remove any particulates. It was expected that the enhanced concentrations of indoximod in GB and SIF increased the absorption of indoximod when the SDD formulation was dosed in animals as well as human beings. Another criterion to evaluate these SDD formulations was physical and chemical stability of indoximod in these formulations. It was found that SDD formulations made by hot process spray drug method were in general more stable than those made by basic process spray dry. In addition, higher drug load in the powder was preferred since it could decrease the dose amount of the final formulation. Based on all these criteria, two SDD formulations were selected for further in vivo PK studies in monkeys. The first one was <NUM>% indoximod/ <NUM>% PVPVA-<NUM>, which showed a <NUM>-fold increased predicted intestinal concentration than indoximod (UltraC<NUM> <NUM> ng/mL vs <NUM> ng/mL); and the second was <NUM>% indoximod/ <NUM>% Affinisol <NUM>, which showed a <NUM>-fold higher predicted intestinal concentration than indoximod (UltraC<NUM> <NUM> ng/mL vs <NUM> ng/mL). These SDDs were prepared by the hot process dry spray which showed better stability properties.

In order to determine whether salts or SDDs that show increase in solubility compared to indoximod free base result in an increase in the maximum concentration (Cmax) and total exposure (AUC<NUM>->∞) of indoximod, we carried out a comparative crossover pharmacokinetic study in cynomolgus monkeys, which is a common species used to predict human oral bioavailability. Two groups of <NUM> monkeys each (all males) were orally dosed at <NUM>µmol/kg (Group <NUM>) or <NUM>µmol/kg (Group <NUM>) with: <NUM>) indoximod free base capsules; <NUM>) indoximod hydrochloride capsules; <NUM>) indoximod hemi phosphate capsules; <NUM>) SDD1 suspension (indoximod <NUM>%/<NUM>%PVPVA-<NUM>, (w/w)) and <NUM>) SDD2 suspension (indoximod <NUM>%/Affinisol <NUM><NUM>% (w/w)). Each monkey was dosed with each of the <NUM> dose formulations once every <NUM> days, and blood samples were obtained at <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM> and <NUM>. Concentration of indoximod was determined from plasma by a validated LC-MS/MS analytical method. Cmax and AUC(<NUM>-<NUM>) was calculated by non-compartmental analysis using WinNonLin software (Certara). For indoximod in capsule formulation, animals in Group <NUM> were orally dosed with <NUM> capsules A and animals in Group <NUM> were dosed with <NUM> capsules B. Compositions of capsules A and B are shown in Table <NUM>. For indoximod in SDD formulation, animals in Group <NUM> were dosed with <NUM>/kg of a <NUM> indoximod/mL suspension and animals in Group <NUM> were dosed with <NUM>/kg of a <NUM> indoximod/mL suspension. The SDD suspension formulations were prepared in <NUM>% methylcellulose (Methocel).

The average Cmax and AUC(<NUM>-<NUM>) parameter values observed in each group obtained after dosing with each formulation of indoximod are shown in Table <NUM>. The percentage of increase in these values as well as the P value obtained for the comparison of each formulation against that of indoximod free base is shown in Table <NUM>. Dosing of indoximod HCl capsules results in a significant increase in Cmax (<NUM>-<NUM>%) and exposure (<NUM>-<NUM>%) at both dose levels tested compared to dosing of indoximod free base capsules. Similarly, indoximod hemi phosphate capsules produced a significant increase in Cmax (<NUM>-<NUM>%) and exposure (<NUM>-<NUM>%). On the contrary, indoximod in SDD1 or SDD2 formulation produced a significant increase in Cmax (<NUM>-<NUM>%) but failed to increase the overall exposure with respect to indoximod free base capsules. For these reasons, indoximod salts in their hydrocholoride, hemi-phosphate or phosphate salts are preferred over indoximod in its free base form, either in capsules or in spray dry dispersions.

This study shows that the hydrochloride and phosphate salts of indoximod can produce an increase in Cmax and AUC pharmacokinetic parameters with respect to the free base, in the range of doses between <NUM>-<NUM>µmol/kg.

In order to determine whether salt formation increased the maximum concentration (Cmax) and total exposure (AUC<NUM>->∞) of indoximod in rats, we tested the hydrochloride, phosphate, sulfate and mesylate salts of indoximod, and formulated these into capsules by mixing them with appropriate excipients. Three dose levels were investigated: <NUM>, <NUM> or <NUM>µmol/kg.

Gelatin capsules (Torpac, <NUM> capacity) were prepared containing <NUM>, <NUM> or <NUM>µmol/capsule of indoximod or its salts, with or without excipients consisting of microcrystalline cellulose, lactose monohydrate, croscarmellose sodium and magnesium stearate, in proportions shown in Table <NUM>-<NUM>. Capsules were manually filled and the composition uniformity of a representative sample of capsules from each batch was verified by weight and by LC-MS/MS to determine the average indoximod content.

To test the pharmacokinetic profile achieved by dosing indoximod in its free base or salt forms, rats were dosed by intra-stomach delivery with <NUM> capsule A, <NUM> capsules B or <NUM> capsules C to achieve dose levels of <NUM>, <NUM> and <NUM>µmol/kg (equivalent to <NUM>, <NUM> and <NUM>/kg of indoximod, respectively). Rats were fasted <NUM> prior to dosing to eliminate any confounding food effects, and food was returned <NUM> after dosing. Blood samples were obtained from each rat at <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM> and <NUM> after dosing. The concentration of indoximod in plasma was determined by LC-MS/MS, and pharmacokinetic parameters were calculated using the software WinNonLin (Certara).

The most relevant pharmacokinetic parameters that were evaluated were the maximum concentration of indoximod (Cmax) and total exposure (AUC<NUM>->∞). Tables <NUM>-<NUM> and <FIG> show a summary of the experimental results.

Indoximod hydrochloride salt form results in non-statistically significant decrease in Cmax at low dose level, a statistically significant increase at the intermediate dose and a statistically significant decrease at high level. The drug exposure (AUC) for the hydrochloride salt did not show a significant change at the low and high dose level but showed a significant increase at the intermediate level. The different behavior of indoximod hydrochloride in rodents compared to primates is unexpected based on the solubility and dissolution profile of this salt, and it does not follow a dose dependent trend, which highlights the importance of conducting species-specific and dose-dependent tests for the prediction of pharmacokinetic profiles in humans.

Indoximod phosphate and hemiphosphate showed a significant increase in Cmax and AUC at the low and intermediate dose levels but a significant decrease in Cmax and a non-statistically significant decrease in exposure at the highest dose level.

The dose-dependent correlation for Cmax and AUC for the free base, HCl and PO<NUM>H<NUM> forms of indoximod is shown in <FIG>. This figure shows an increase in Cmax for the HCl and PO<NUM>H<NUM> salts with respect to the free base at the low and intermediate dose levels but a saturation in the Cmax dose-response curve at the highest dose level, which is not seen for the free base. The dose-response curve for AUC shows a more linear increase of AUC with dose, except for the PO<NUM>H<NUM> salt which seems to increase less than dose proportional at the highest dose level tested.

Similarly, other salt forms of indoximod such as sulfate or mesylate increase the Cmax and AUC ~<NUM>-<NUM>% when tested at <NUM>µmol/kg.

These tests indicate that the hydrochloride and phosphate salts of indoximod have increased solubility with respect to the free base form and display increased Cmax and AUC parameter values.

The pharmacokinetic profile of indoximod obtained after oral administration of several indoximod prodrugs was tested in such a way that reflected only differences in intestinal permeability and conversion of prodrug to indoximod in vivo without reflecting differences in solid state form such as differences in polymorphic crystals or amorphous solids which may impact solubility or solubilization rate for the different prodrugs. Therefore, indoximod and each of its prodrugs was solubilized in appropriate vehicle which was either saline solution, Cremaphor®:ethanol:saline (<NUM>:<NUM>:<NUM>), or Chremaphor:EtOH:saline:HCl (<NUM>:<NUM>:<NUM>:<NUM>. Indoximod or its prodrugs were dissolved at a concentration of <NUM>/mL and dosed to rats by oral gavage at <NUM>/kg to achieve a final dose of <NUM>/kg; or dissolved at <NUM>/mL and dosed to rats by oral gavage at <NUM>/kg to achieve a final dose of <NUM>/kg; or dissolved at a concentration of <NUM>/mL and dosed orally to mice by oral gavage at <NUM>/kg to achieve a final dose of <NUM>/kg. Blood samples (<NUM>-<NUM>) were collected from the femoral artery port from rats or by retro-orbital bleeding from mice and plasma was immediately collected by centrifugation and stored on dry ice to avoid prodrug hydrolysis after plasma collection. Blood samples were collected at <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM> and <NUM> after dosing from rats or at <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM> and <NUM> after dosing from mice. The concentration of indoximod and of each prodrug in plasma was determined by LC-MS/MS, and pharmacokinetic parameters were calculated for indoximod and its prodrugs. The pharmacokinetic parameters reflect the average of individual parameter values obtained from each individual rat (n) or one common parameter from a single pharmacokinetic curve derived from blood samples obtained from a group of mice (n).

Tables <NUM> and <NUM> show the indoximod Cmax and AUC(<NUM>->∞) obtained after dosing either indoximod or each one of the test prodrugs. Since all rats were orally dosed at the same dose of <NUM>/kg, but each prodrug has different molecular weight, in order to compare the values of Cmax and AUC(<NUM>->∞) obtained after dosing each prodrug vs. dosing indoximod as a free base, the measured Cmax and AUC(<NUM>->∞) and were normalized by multiplying them by the ratio of MWProdrug/MWindoximod, thus assuming linear pharmacokinetics within a ~<NUM>-fold dose range.

Table <NUM> shows that some prodrugs result in an effective increase in either Cmax, AUC or both pharmacokinetic parameters. Since the prodrugs were administered in completely soluble form, this suggests that those prodrugs that show enhanced Cmax and/or AUC of indoximod in plasma do so by a mechanism that involves a combination of factors including enhanced permeability of the prodrug through the intestinal cell wall, reduced clearance of the prodrug with respect to indoximod and good rate of conversion of the prodrug to indoximod in vivo. Not every prodrug form of indoximod resulted in enhanced maximum concentration and exposure of indoximod compared to administration of indoximod. In particular, exposure (AUC) to indoximod seems to be enhanced when dosing NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, and NLG-<NUM>, while indoximod Cmax seems to be enhanced when dosing NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM>, NLG-<NUM> and NLG-<NUM>.

Table <NUM> shows prodrugs that did not result in an effective increase in indoximod Cmax nor indoximod exposure when dosed orally to rats at <NUM>/kg, indicating that some of these chemical substitutions may either decrease permeability, or the rate of conversion to indoximod or increase the rate of prodrug clearance by routes that do not result in conversion to indoximod, or a combination of those effects.

Table <NUM> shows prodrugs that were tested by oral dosing to rats at <NUM>/kg. NLG-<NUM> causes an increase in Cmax and AUC when dosed to rats at <NUM>/kg. However, this prodrug results in a decrease in Cmax and AUC when dosed to mice at <NUM>/kg. Conversely, the highly similar molecule NLG-<NUM> does not produce a significant increase in Cmax or AUC when dosed at <NUM>/kg to rats, but it does produce a significant increase in Cmax and AUC in mice, suggesting that different species have different rates of absorption, elimination and metabolization of these prodrugs and that minimal changes in molecular structure can affect the outcome in different species. A dose dependent PK was carried out in mice, which were dosed at <NUM>, <NUM> and <NUM>/kg of indoximod, or at similar doses for prodrug NLG-<NUM> or NLG-<NUM>. A caveat of the comparison between dosing prodrugs vs indoximod as a free base was that prodrugs were fully soluble in the dosing formulation, while indoximod was insoluble at doses of <NUM> and <NUM>/kg. This may result in a time-dependent controlled release effect for indoximod which could result in lower Cmax but higher AUCs than when dosed in fully soluble form. NLG-<NUM> and NLG-<NUM> resulted in a significant increase in indoximod Cmax compared to what is observed when dosing indoximod in suspension, at all doses tested. However, NLG-<NUM> showed a dose dependent increase AUC for indoximod, where the percentage of increase in AUC decreases at higher doses. Table <NUM> also indicates that formation of carbamates on the amino group of indoximod result in prodrugs with marked reduction in pharmacokinetic parameters for indoximod.

To test which prodrugs have the best combined set of pharmacological properties (solubilization rate, solubility, intestinal permeability, clearance rate and rate of metabolization to indoximod) needed to achieve greater plasma concentrations of indoximod and increased exposure to indoximod after oral dosing in a capsule formulation, the prodrugs that showed enhanced indoximod Cmax or exposure when dosed in solution were prepared in several salt forms and mixed with excipients to form a powder blend. These blends were formulated so that each capsule contained the same molar dose of each prodrug. Gelatin capsules (Torpac, <NUM> capacity) were prepared containing <NUM>µmol/capsule A, <NUM>µmol/capsule B or <NUM>µmol/capsule C of indoximod free base (<NUM>, <NUM> or <NUM>/capsule, respectively) or its prodrugs in diverse salt forms, in an excipient blend consisting of microcrystalline cellulose, lactose monohydrate, croscarmellose sodium and magnesium stearate, in proportions shown in Tables <NUM>. 1a and <NUM>. The composition and uniformity of a representative sample of capsules from each batch was verified by weight and by LC-MS/MS to determine the average indoximod or prodrug content.

To test the pharmacokinetic profile achieved by dosing indoximod prodrugs in different salt forms, <NUM> capsule A (<NUM>µmol/capsule) or <NUM> capsules B (<NUM>µmol/capsule) or <NUM> capsules C (<NUM>µmol/capsule) were dosed to rats by intra-stomach delivery. The dose levels tested were equivalent to <NUM>/kg (<NUM>µmol/kg) of indoximod equivalent when dosing <NUM> capsule A of <NUM>µmol/capsule, <NUM>/kg (<NUM>µmol/kg) of indoximod equivalent when dosing <NUM> capsules B of <NUM>µmol/capsule and <NUM>/kg (<NUM>µmol/kg) of indoximod equivalent when dosing <NUM> capsules C of <NUM>µmol/capsule. Rats were fasted <NUM> prior to dosing to eliminate any confounding food effects, and food was returned <NUM> after dosing. Blood samples were obtained from each rat at <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM> and <NUM> after dosing. The concentration of indoximod in plasma was determined by LC-MS/MS, and pharmacokinetic parameters were calculated using the software WinNonLin (Certara).

The most relevant evaluated pharmacokinetic parameters were the maximum concentration of indoximod (Cmax) and total indoximod exposure (AUC<NUM>->∞). Tables <NUM> and <NUM> show a summary of the experimental results.

The statistical comparison of pharmacokinetic parameters indicated that ethyl Nα -(L-leucyl)-<NUM>-methyl-D-tryptophanate in its hydrochloride (NLG-<NUM>), phosphate (NLG-<NUM>), mesylate (NLG-<NUM>) or besylate (NLG-<NUM>) salt forms dosed at <NUM>-<NUM>µmol/kg was able to significantly (p<<NUM>) increase exposure of indoximod by <NUM>-<NUM>%, while its sulfate salt (NLG-<NUM>) did not result in a significant increase in Cmax or AUC at those doses. Similarly, significant increases in Cmax were observed for NLG-<NUM>, NLG-<NUM> and NLG-<NUM>. At doses of <NUM>µmol/kg, NLG-<NUM> hydrochloride, showed a minor increase in Cmax and AUC compared to indoximod.

Table <NUM> shows that <NUM>,<NUM>-dihydroxypropyl <NUM>-methyl-D-tryptophanate in its phosphate (NLG-<NUM>) form resulted in significant increase in Cmax (<NUM>-<NUM>%) and AUC (<NUM>-<NUM>%), while its hydrocholoride (NLG-<NUM>), and sulfate (NLG-<NUM>) salts resulted in less significant increases in Cmax and AUC. Interestingly, the mesylate salt of <NUM>,<NUM>-dihydroxypropyl <NUM>-methyl-D-tryptophanate (NLG-<NUM>) resulted in a decrease in Cmax and AUC, thought this decrease was not statistically significant.

Table <NUM> also shows that ethyl Nα-(L-methionyl)-<NUM>-methyl-D-tryptophanate (HCl, and phosphate salts, NLG-<NUM>) show a statistically significant increase in Cmax and AUC at doses of <NUM>-<NUM>µmol/kg.

Other prodrugs that were studied included: a) ethyl Nα-(L-glutaminyl)-<NUM>-methyl-D-tryptophanate (free base, HCl, phosphate or mesylate salts), b) Nα-glycyl-<NUM>-methyl-D-tryptophan (HCl or phosphate salt), c) methyl N<NUM>-((R)-<NUM>-ethoxy-<NUM>-(<NUM>-methyl-<NUM>H-indol-<NUM>-yl)-<NUM>-oxopropan-<NUM>-yl)-L-asparaginate (HCl form) and d) Nα-(L-lysyl)-<NUM>-methyl-D-tryptophan (free base, HCl, sulfate or phosphate salts). These prodrugs resulted in minor and non-statistically significant variations in the Cmax or AUC for indoximod compared to an equivalent molar dose of indoximod (Table <NUM>).

Interestingly, piperidin-<NUM>-ylmethyl <NUM>-methyl-D-tryptophanate in its HCl or phosphate salt forms (NLG-<NUM> and NLG-<NUM>) resulted in a statistically significant decrease in Cmax (<NUM>-<NUM>%, p<<NUM>) and AUC (<NUM>-<NUM>%, p<<NUM>) for indoximod. Since this compound showed an increase in Cmax (<NUM>%) and AUC (<NUM>%) when administered via oral solution, the difference in solubilization rate or final solubility may account for the observed differences when administered in powder form.

Since the rat shows a non-saturable linear increase in exposure with doses of indoximod of up to <NUM>/kg, while humans show a saturable exposure above doses of <NUM>/kg, we decided to evaluate two of the prodrug in primates, which may constitute a better model to predict human pharmacokinetics than rats. Cynomolgous monkeys (<NUM>-<NUM>) were dosed with indoximod, NLG-<NUM> HCl or NLG-<NUM> HCl at doses of <NUM>, <NUM> or <NUM>µmol/kg in a crossover study design where each animal received the same molar dose of either indoximod, NLG1564 HCl or NLG-<NUM> HCl every <NUM> days. Capsules were prepared according to the formulation described in Table <NUM>. Monkeys were orally dosed with <NUM> or <NUM> capsules A (<NUM>µmol/capsule) or <NUM> capsules B (<NUM>µmol/capsule). Blood samples were collected at <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM> and <NUM> post-dose, and the concentrations of prodrug and indoximod were analyzed by validated LC-MSMS methods.

The data in Table <NUM> shows that NLG-<NUM> HCl increases the Cmax of indoximod from ~ <NUM>-<NUM>% and AUC from <NUM>-<NUM>% in a statistically significant manner. Similarly, NLG-<NUM> HCl increases the Cmax of indoximod from ~ <NUM>-<NUM>% and AUC from <NUM>-<NUM>% in a statistically significant manner. The increase in pharmacodynamics indicators in primates was unexpectedly superior from the results observed in rats, indicating that in primates, prodrugs of indoximod of the present disclosure can provide a significant improvement in the maximum concentration and exposure to indoximod and are expected to improve exposure to the drug and therapeutic efficacy in human patients.

Claim 1:
A pharmaceutically acceptable salt of indoximod, having the structure of Formula 1a:
<CHM>
wherein A-p is an anion selected from the group consisting of HPO<NUM><NUM>- (hemi-phosphate), HSO<NUM>- (hydrogen sulfate), and CH<NUM>S(O<NUM>)O- (mesylate), wherein the ionization state -p is -<NUM> or -<NUM> and the stoichiometric ratio n is <NUM> or ½, respectively, such that the stoichiometric conditions of charge neutrality are satisfied.