Use of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] to ameliorate proteasome inhibitor resistance

Abstract

Methods and corresponding uses are provided for treating disease in patients involving the use of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] to improve drug resistance, including resistance to proteasome inhibitors and immunomodulatory imide drugs. The disease may be, for example, relapsed / refractory multiple myeloma.

Description

[Technical field]

[0001] The present invention is in the field of therapeutics, including the combination of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] and a proteasome inhibitor, for example in the treatment of cancer. [Background technology]

[0002] Sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is a coordination complex of ruthenium with anticancer activity (also known as BOLD-100, KP1339, NKP-1339, IT-139, and Na[RuIIICl4(Hind)2]). Methods for preparing alkali metal salts of trans-[tetrachlorobis(1H-indazole)ruthenate(III)] are described, for example, in PCT Publication WO 2018204930, and such compounds have the formula I: [ka] where M is an alkali metal cation, such as the sodium salt: [ka] It is.

[0003] Myeloma is a lymphoid malignancy with plasma cells present primarily in the bone marrow, although malignant plasma cells can also be found in peripheral blood, soft tissues, and organs. Myeloma is thus a plasma cell disorder that manifests in forms that can be differentiated by the site of involvement, with several different compartments affected in multiple myeloma, only one site affected in plasmacytoma, adjacent sites affected in localized myeloma, and involvement of tissues other than the bone marrow in extramedullary myeloma. As used herein, the term "myeloma" thus refers to a spectrum of diseases recognized in the art as such. Within this spectrum of diseases, relapsed MM is generally considered as a recurrence of disease after a previous response, typically based on objective clinical criteria, and relapsed / refractory MM (RRMM) is generally defined as disease that becomes non-responsive or progressive to therapy or within 60 days of previous treatment in patients who have achieved a minimal response (MR) or better response in previous therapy. Treatment of RRMM presents unique challenges.

[0004] Immunomodulatory imide drugs (IMiDs) have been used in the treatment of myeloma, including the IMiD drug thalidomide and its analogs, including lenalidomide, pomalidomide, and iverdimide. These drugs are understood to act as modulators of the protein cereblon.

[0005] The proteasome is a protease complex that mediates the selective hydrolysis of proteins in cooperation with ubiquitin, which acts as a marker for regulated protein hydrolysis, in both transformed and normal eukaryotic cells. The use of proteasome inhibitors (PIs) has been transformative in the treatment of multiple myeloma (MM), mantle cell lymphoma (MCL), and amyloidosis. PIs are understood to have anti-proliferative and anti-tumor activity mediated by inhibiting proteasomal degradation of proteins by targeting various proteasome components, and a distinction has been recognized between inhibitors of the constitutive proteasome expressed in most cell types and the immunoproteasome expressed in immune cells. Clinical activity of PIs has also been reported in Waldenstrom's macroglobulinemia, T-cell lymphoma amyloidosis, and other lymphoproliferative disorders, such as lymphoid malignancies. Three proteasome inhibitors, bortezomib, carfilzomib, and ixazomib citrate, are used for the treatment of multiple myeloma and are being tested for additional uses. Marizomib, oprozomib, and delanzomib are PIs being used in clinical trials for MM and other indications. Immunoproteasome inhibitors include ONX-0914 and KZR-616, and have been described, for example, in WO 2006099261, WO 2006092326, WO 2006045066, WO 2007149512, WO 2010108172, WO 2014152134, WO 2014152127, WO 2011109355, WO 2014056954, WO 2014056748. With the increasing success of PIs in treating a number of diseases, resistance to proteasome inhibitors has become a growing problem. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] International Publication No. 2006099261 [Patent Document 2] International Publication No. 2006092326 [Patent Document 3] International Publication No. 2006045066 [Patent Document 4] International Publication No. 2007149512 [Patent Document 5] International Publication No. 2010108172 [Patent Document 6] International Publication No. 2014152134 [Patent Document 7] International Publication No. 2014152127 [Patent Document 8] International Publication No. 2011109355 [Patent Document 9] International Publication No. 2014056954 [Patent Document 10] International Publication No. 2014056748 Summary of the Invention

[0007] Methods and corresponding uses are provided for improving therapeutic resistance to proteasome inhibitors (PIs) in patients in need thereof, comprising administering an effective amount of trans-[tetrachloridobis(1H-indazole)ruthenate(III)]sodium (BOLD-100). Methods for treating cancer or lymphoproliferative disorders in patients in need thereof are also provided, comprising administering an effective amount of BOLD-100 in combination with a combined effective amount of a proteasome inhibitor (PI). A combined effective amount of PI is a PI dosage or regimen in which the therapeutic benefit of the PI is improved by combined treatment with BOLD-100. Diseases or disorders suitable for combined treatment with BOLD-100 and PIs may be, for example, diseases that are prone to develop resistance to treatment with PIs in the absence of treatment with BOLD-100. The use of BOLD-100 to improve therapeutic resistance to proteasome inhibitors may therefore be prophylactic, in the sense of improving the transition to a PI-resistant disease state.

[0008] The effective amount of BOLD-100 and the combined effective amount of a PI may, for example, be additively or synergistically effective, with synergy being assessed by any one of a variety of art-recognized scales. The disease may be resistant to treatment with a PI alone. The disease may be multiple myeloma (MM), for example, relapsed refractory MM (RRMM). Selected embodiments thus involve treatment of RRMM, where RRMM has acquired resistance to a PI, and the treatment includes administration of a PI in combination with BOLD-100, where BOLD-100 is used in an amount effective to overcome or ameliorate the acquired resistance to the PI. BOLD-100 and the PI may be administered sequentially in any order, or may be administered in combination, in a co-formulation or separately.

[0009] Also provided is a method of treating a myeloma resistant to a therapeutic agent, comprising administering to a patient an effective amount of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)], wherein the therapeutic agent is effective to treat a myeloma that is not resistant. The therapeutic agent may be, for example, a PI, an immunomodulatory imide drug, a cereblon modulator drug, thalidomide, or a thalidomide analog (e.g., lenalidomide, pomalidomide, or iverdimide). [Brief description of the drawings]

[0010] [Figure 1] 1 is a line graph illustrating the anticancer activity of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] (BOLD-100), showing inhibition of proliferation of various multiple myeloma (MM) cancer cell lines. [Figure 2A] Included is a line graph showing the growth curves of cell lines MM.1S and MM.1S BR in the presence of increasing doses of BOLD-100 over a 24 hour period, demonstrating the greater sensitivity of the PI-resistant lines to BOLD-100 compared to the parental MM.1S cell line (BOLD-100 IC50: MM.1S = 83 μM; MM.1S BR = 23 μM). [Figure 2B] 1 includes a line graph showing the growth curves of cell lines OPM2, OPM2 LR (lenalidomide resistant) and OPM2 PR (pomalidomide resistant) in the presence of increasing doses of BOLD-100 over a 24 hour period, demonstrating the higher sensitivity of lenalidomide and pomalidomide resistant lines to BOLD-100 (BOLD-100 IC50: OPM2=120 μM; OPM2 PR=51 μM). [Figure 3A] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3B] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3C] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3D]Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3E] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3F] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3G] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3H]Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3I] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3J] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3K] Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3L]Included are bar graphs and scatter plots, showing that BOLD-100 overcomes PI resistance in vitro. The bar graphs show the survival percentage normalized to vehicle control for the indicated treatments. The scatter plots show the corresponding combination index and fraction effect size for the treatments, with dotted lines indicating the areas of antagonism, additive effect, and synergy from top to bottom, respectively. [Figure 3M] The bar graphs show the same data as shown in the other panels of FIG. 3, except that all cells in each data set were treated with 10 nM bortezomib (left hand bars) or 10 nM carfilzomib (right hand bars). [Figure 4A] 1 includes bar graphs and photographs showing the efficacy of BOLD-100 in combination with bortezomib (Btz) in mediating robust cytotoxic effects in PI-sensitive (MM.1S) cell lines in clonogenic assays. [Figure 4B] 1 includes bar graphs and photographs showing the efficacy of BOLD-100 in combination with bortezomib (Btz) in mediating robust cytotoxic effects in resistant (MM.1 BR) cell lines in a clonogenic assay. [Diagram 5] 1 is a line graph showing that BOLD-100 negatively affects survival of three MM.1S cell lines, but has minimal impact in the myeloid cell line HS-5. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0011] As disclosed herein, BOLD-100 in combination with a PI has the ability to mediate an additive therapeutic effect in PI-sensitive cancer cells and overcome resistance in PI-resistant cancer cells. Thus, when the therapeutic efficacy of a PI is reduced, typically due to acquired drug resistance from prolonged exposure, adding BOLD-100 to a treatment regimen restores the efficacy of the PI. Additionally, the combination of BOLD-100 and a PI promotes a durable PI therapy in the sense that the additive combination of BOLD-100 and a PI is further complemented by a synergistic effect on any PI-resistant cells that arise during the course of treatment. In this way, BOLD-100 can be used to inhibit the progression of a PI-sensitive disease to PI resistance. These forms of durable PI combination therapy with BOLD-100 may therefore have unique utility in cancers or lymphoproliferative disorders that are prone to develop resistance to treatment with PIs. This is the case, for example, in the treatment of MM, particularly relapsed / refractory MM, or in the prevention of the progression of MM to RRMM.

[0012] An embodiment of the present invention involves a method for preparing a drug product containing the sodium salt of trans-[tetrachlorobis(1H-indazole)ruthenic acid (III)] (i.e., BOLD-100), as described below.

[0013] One embodiment of the present invention provides a method for preparing a sterile, lyophilized drug product containing sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)]. The formulation is suitable for administration to a patient. The formulation includes sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], a pH buffer, and a cryoprotectant. A general method for providing the formulation includes the steps of preparing an aqueous buffer solution, preparing an aqueous cryoprotectant solution, dissolving sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] in the buffer solution, adding the cryoprotectant solution, sterile filtration (e.g., aseptic filtration), filling into vials under sterile conditions, and lyophilizing under sterile conditions. Suitable buffers include, but are not limited to, citric acid (salts), TRIS, acetic acid (salts), EDTA, HEPES, tricine, and imidazole. The use of phosphate buffers is possible but not preferred. A preferred embodiment of the present invention is the use of citric acid / sodium citrate buffer. Suitable cryoprotectants include, but are not limited to, sugars, monosaccharides, disaccharides, polyalcohols, mannitol, sorbitol, sucrose, trehalose, dextran, and dextrose. A preferred embodiment of the present invention is the use of mannitol as a cryoprotectant.

[0014] As described herein above, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] can decompose to compound A in water (Scheme II). Those skilled in the art will recognize that limiting this decomposition reaction is advantageous in order to obtain the highest purity product. It has been found that cooling the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] solution during the formulation process has been found to greatly reduce the amount of compound A present in the lyophilized product. In one embodiment of the invention, the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] solution is cooled to 4° C. during the formulation process. In another embodiment of the invention, the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] solution is cooled to 2-8° C. during the formulation process. In another embodiment of the invention, the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] solution is cooled to 2-15° C. during the formulation process.

[0015] One embodiment of the present invention provides a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], a suitable buffer, and mannitol. In some embodiments, the suitable buffer comprises a citrate buffer. For example, in some embodiments, the citrate buffer comprises sodium citrate and citric acid. An alternative embodiment of the present invention provides a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, and mannitol. An alternative embodiment of the present invention provides a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, and mer,trans-[RuIIICl3(Hind)2(H2O)]. An alternative embodiment of the present invention provides a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, mer, trans-[RuIIICl3(Hind)2(H2O)], and a cesium salt. An alternative embodiment of the present invention provides a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, and mannitol, wherein the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is amorphous. An alternative embodiment of the present invention provides a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, and mer, trans-[RuIIICl3(Hind)2(H2O)], wherein the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is amorphous.An alternative embodiment of the present invention provides a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, mer,trans-[RuIIICl3(Hind)2(H2O)], and a cesium salt, wherein the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is amorphous.An alternative embodiment of the present invention provides a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, mer,trans-[RuIIICl3(Hind)2(H2O)], and a cesium salt, wherein the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is amorphous. mer,trans-[Ru III Cl3(Hind)2(H2O)] is about 0.01 to about 0.4 weight percent of the composition; and cesium is about 0.00001 to about 0.01 weight percent of the composition. A composition is provided.

[0016] Alternative embodiments of the present invention include sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, mer,trans-[Ru III Cl3(Hind)2(H2O)] is about 0.01 to about 0.4 weight percent of the composition; and cesium is about 0.00001 to about 0.01 weight percent of the composition. A composition is provided.

[0017] Alternative embodiments of the present invention include sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, mer,trans-[Ru III Cl3(Hind)2(H2O)] is about 0.01 to about 0.2 weight percent of the composition; and cesium is about 0.00001 to about 0.01 weight percent of the composition. A composition is provided.

[0018] An alternative embodiment of the present invention is sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, mer,trans-[Ru III Cl3(Hind)2(H2O)] is from about 0.01 to about 0.40 weight percent of the composition; and cesium is about 0.00001 to about 0.01 weight percent of the composition. A composition is provided.

[0019] An alternative embodiment of the present invention is sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, The composition is a lyophilized powder, III Cl3(Hind)2(H2O)] is from about 0.01 to about 0.40 weight percent of the composition; and cesium is about 0.00001 to about 0.01 weight percent of the composition. A composition is provided.

[0020] An alternative embodiment of the present invention is sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, The composition is a lyophilized powder, IIICl3(Hind)2(H2O)] is about 0.01 to about 0.3 weight percent of the composition; and cesium is about 0.00001 to about 0.1 weight percent of the composition. A composition is provided.

[0021] Alternative embodiments of the present invention include sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, mer,trans-[Ru III Cl3(Hind)2(H2O)] is about 0.01 to about 0.3 weight percent of the composition; and cesium is about 0.00001 to about 0.1 weight percent of the composition. A composition is provided.

[0022] Alternative embodiments of the present invention include sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, the composition is a lyophilized powder; sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 11.5 to about 14.0 weight percent of the composition; citric acid is about 43.9 to about 53.7 weight percent of the composition; sodium citrate is about 25.7 to about 23.1 weight percent of the composition; mannitol is about 11.5 to about 14.0 weight percent of the composition; mer,trans-[Ru III Cl3(Hind)2(H2O)] is between about 0.01 and about 0.3 weight percent of the composition; and cesium is about 0.00001 to about 0.1 weight percent of the composition. A composition is provided.

[0023] Alternative embodiments of the present invention include sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], sodium citrate, citric acid, mannitol, mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, the composition is a lyophilized powder; sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 10.2 to about 15.3 weight percent of the composition; citric acid is about 39.0 to about 58.5 weight percent of the composition; sodium citrate is about 20.5 to about 30.8 weight percent of the composition; mannitol is about 10.2 to about 15.3 weight percent of the composition; mer,trans-[Ru III Cl3(Hind)2(H2O)] is between about 0.01 and about 0.3 weight percent of the composition; and cesium is about 0.00001 to about 0.1 weight percent of the composition. A composition is provided.

[0024] An alternative embodiment of the present invention is sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mer,trans-[Ru III Cl3(Hind)2(H2O)], and a cesium salt, the composition is a lyophilized powder, and the sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 10.2 to about 15.3 weight percent of the composition; mer,trans-[Ru III Cl3(Hind)2(H2O)] is about 0.01 and about 0.3 weight percent, and cesium is about 0.00001 to about 0.1 weight percent of the composition. A composition is provided.

[0025] An alternative embodiment of the present invention is a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, and sodium citrate, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 49.86 weight percent of the composition; mannitol is about 49.86 weight percent of the composition; citric acid is about 0.187 weight percent of the composition; and sodium citrate is about 0.093 weight percent of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0026] An alternative embodiment of the present invention is a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, and sodium citrate, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 40 to about 60 weight percent of the composition; Mannitol is about 40 to about 60 weight percent of the composition; citric acid is about 0.01 to about 0.5 weight percent of the composition; and sodium citrate is about 0.001 to about 0.25 weight percent of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0027] An alternative embodiment of the present invention is a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, and sodium citrate, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 30 to about 70 weight percent of the composition; Mannitol is about 30 to about 70 weight percent of the composition; citric acid is about 0.001 to about 1 weight percent of the composition; and sodium citrate is about 0.0001 to about 1 weight percent of the composition. A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0028] An alternative embodiment of the present invention is a method for preparing a ruthenate comprising the steps of: sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, and Ru III A composition comprising Cl(Hind)(H2O), sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 49.86 weight percent of the composition; mannitol is about 49.86 weight percent of the composition; citric acid is about 0.187 weight percent of the composition; sodium citrate is about 0.093 weight percent of the composition; And Ru III Cl3(Hind)2(H2O) is less than or equal to 0.5 weight percent of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0029] An alternative embodiment of the present invention is a method for preparing a ruthenate comprising the steps of: sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, and Ru III A composition comprising Cl(Hind)(H2O), sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 40 to about 60 weight percent of the composition; Mannitol is about 40 to about 60 weight percent of the composition; citric acid is about 0.01 to about 0.5 weight percent of the composition; sodium citrate is about 0.001 to about 0.25 weight percent of the composition; And Ru III Cl3(Hind)2(H2O) is about 0 to about 0.5 weight percent of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0030] An alternative embodiment of the present invention is a mixture of sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, Ru III A composition comprising Cl3(Hind)2(H2O) and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 30 to about 70 weight percent of the composition; Mannitol is about 30 to about 70 weight percent of the composition; citric acid is about 0.001 to about 1 weight percent of the composition; sodium citrate is about 0.0001 to about 1 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is less than or equal to 0.5 weight percent of the composition; and cesium is 0.25 weight percent or less of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0031] An alternative embodiment of the present invention is a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 49.61 weight percent of the composition; mannitol is about 49.86 weight percent of the composition; citric acid is about 0.187 weight percent of the composition; sodium citrate is about 0.093 weight percent of the composition; and the cesium is about 0.25 weight percent of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0032] An alternative embodiment of the present invention is a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 40 to about 60 weight percent of the composition; Mannitol is about 40 to about 60 weight percent of the composition; citric acid is about 0.01 to about 0.5 weight percent of the composition; sodium citrate is about 0.001 to about 0.25 weight percent of the composition; and cesium is about 0.1 to about 0.5 weight percent of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0033] An alternative embodiment of the present invention is a composition comprising sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 30 to about 70 weight percent of the composition; Mannitol is about 30 to about 70 weight percent of the composition; citric acid is about 0.001 to about 1 weight percent of the composition; sodium citrate is about 0.0001 to about 1 weight percent of the composition; and cesium is about 0.01 to about 1 weight percent of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0034] An alternative embodiment of the present invention is a mixture of sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), Ru III A composition comprising Cl3(Hind) (HN=C(Me)ind) and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 46.61 weight percent of the composition; mannitol is about 49.86 weight percent of the composition; citric acid is about 0.187 weight percent of the composition; sodium citrate is about 0.093 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is less than or equal to 0.5 weight percent of the composition; Ru III Cl3(Hind)2(CH3CN) is less than or equal to 1.25 weight percent of the composition; Ru III Cl3(Hind) (HN=C(Me)ind) is less than or equal to 1.0 weight percent of the composition; and cesium is 0.25 weight percent or less of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0035] An alternative embodiment of the present invention is a mixture of sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), Ru III A composition comprising Cl3(Hind) (HN=C(Me)ind) and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 46.61 weight percent of the composition; mannitol is about 49.86 weight percent of the composition; citric acid is about 0.187 weight percent of the composition; sodium citrate is about 0.093 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is less than or equal to 0.5 weight percent of the composition; Ru III Cl3(Hind)2(CH3CN) is less than or equal to 1.25 weight percent of the composition; Ru III Cl3(Hind) (HN=C(Me)ind) is less than or equal to 1.0 weight percent of the composition; and cesium is 0.25 weight percent or less of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0036] An alternative embodiment of the present invention is a mixture of sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), Ru III A composition comprising Cl3(Hind) (HN=C(Me)ind) and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 40 to about 60 weight percent of the composition; Mannitol is about 40 to about 60 weight percent of the composition; citric acid is about 0.01 to about 0.5 weight percent of the composition; sodium citrate is about 0.001 to about 0.25 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is about 0.5 weight percent or less of the composition; Ru III Cl3(Hind)2(CH3CN) is less than or equal to about 1.25 weight percent of the composition; Ru III Cl3(Hind) (HN=C(Me)ind) is less than or equal to about 1.0 weight percent of the composition; and cesium is 0.25 percent or less of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0037] An alternative embodiment of the present invention is a mixture of sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), Ru III A composition comprising Cl3(Hind) (HN=C(Me)ind) and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 30 to about 70 weight percent of the composition; Mannitol is about 30 to about 70 weight percent of the composition; citric acid is about 0.001 to about 1 weight percent of the composition; sodium citrate is about 0.0001 to about 1 weight percent of the composition; Ru IIICl3(Hind)2(H2O) is about 0.5 weight percent or less of the composition; Ru III Cl3(Hind)2(CH3CN) is less than or equal to about 1.25 weight percent of the composition; Ru III Cl3(Hind) (HN=C(Me)ind) is less than or equal to about 1.0 weight percent of the composition; and cesium is 0.25 percent or less of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0038] An alternative embodiment of the present invention is a mixture of sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), Ru III A composition comprising Cl3(Hind) (HN=C(Me)ind) and cesium, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)] is about 20 to about 80 weight percent of the composition; Mannitol is about 20 to about 80 weight percent of the composition; citric acid is about 0.0001 to about 5 weight percent of the composition; sodium citrate is about 0.00001 to about 5 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is about 0.5 weight percent or less of the composition; Ru III Cl3(Hind)2(CH3CN) is less than or equal to about 1.25 weight percent of the composition; Ru III Cl3(Hind) (HN=C(Me)ind) is less than or equal to about 1.0 weight percent of the composition; and cesium is 0.25 percent or less of the composition; A composition is provided. In some such embodiments, the composition is a lyophilized powder.

[0039] In some embodiments, the present invention provides a unit dosage form comprising the formulation or composition described herein.The expression "unit dosage form" as used herein refers to a physically separate unit of the formulation provided that is suitable for the subject to be treated.However, it is understood that the total daily dose of the formulation provided is determined by the attending physician within the scope of sound medical judgment.The specific effective dose level for any specific subject or organism depends on various factors, including the disorder being treated and the severity of the disorder; the activity of the specific active agent used; the specific formulation used; the age, weight, general health, sex and diet of the subject; the time of administration and the rate of excretion of the specific active agent used; the duration of treatment; the drugs and / or additional therapies used in combination or simultaneously with the specific compound(s) used, and similar factors well known in the medical field.

[0040] The composition of the present invention may be provided as a unit dosage form. In some embodiments, a vial containing sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, and sodium citrate is the unit dosage form.

[0041] In some embodiments, the vial of the present invention containing sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, and cesium is a unit dosage form.

[0042] In some embodiments, sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, sodium citrate, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), Ru IIIThe vials of the present invention containing Cl3(Hind) (HN=C(Me)ind), and cesium are in unit dosage form.

[0043] Still further encompassed by the present invention are pharmaceutical packs and / or kits comprising a composition described herein, or a unit dosage form comprising a provided composition, and a container (e.g., a foil or plastic package, or other suitable container). Instructions for use may additionally be provided in such kits.

[0044] In some embodiments, the present invention may be provided as a unit dosage form. Indeed, a vial containing sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, and sodium citrate is the unit dosage form depicted in Table 3. [Table 1] In some embodiments, the pharmaceutical components described in Table 3 further comprise cesium; The cesium is not more than 0.25 weight percent of the composition.

[0045] In some embodiments, the pharmaceutical components listed in Table 3 include cesium, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), and Ru III Further containing Cl3(Hind) (HN=C(Me)ind); the cesium is less than or equal to about 0.25 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is less than or equal to about 0.5 weight percent of the composition; Ru III Cl3(Hind)2(CH3CN) is less than or equal to about 1.25 weight percent of the composition; And Ru IIICl3(Hind) (HN=C(Me)ind) is less than or equal to about 1.0 weight percent of the composition.

[0046] In some embodiments, the pharmaceutical composition is selected from those in Table 4. [Table 2] In some embodiments, the pharmaceutical components described in Table 4 further comprise cesium; The cesium is not more than 0.25 weight percent of the composition.

[0047] In some embodiments, the pharmaceutical components listed in Table 4 include cesium, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), and Ru III Further containing Cl3(Hind) (HN=C(Me)ind); the cesium is less than or equal to about 0.25 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is less than or equal to about 0.5 weight percent of the composition; Ru III Cl3(Hind)2(CH3CN) is less than or equal to about 1.25 weight percent of the composition; And Ru III Cl3(Hind) (HN=C(Me)ind) is less than or equal to about 1.0 weight percent of the composition.

[0048] In some embodiments, the present invention may be provided as a unit dosage form. Indeed, a vial containing sodium trans-[tetrachlorobis(1H-indazole)ruthenate(III)], mannitol, citric acid, and sodium citrate is the unit dosage form depicted in Table 5. [Table 3] In some embodiments, the pharmaceutical components described in Table 5 further comprise cesium; The cesium is not more than 0.25 weight percent of the composition.

[0049] In some embodiments, the pharmaceutical components listed in Table 5 include cesium, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), and Ru III Further containing Cl3(Hind) (HN=C(Me)ind); the cesium is less than or equal to about 0.25 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is less than or equal to about 0.5 weight percent of the composition; Ru III Cl3(Hind)2(CH3CN) is less than or equal to about 1.25 weight percent of the composition; And Ru III Cl3(Hind) (HN=C(Me)ind) is less than or equal to about 1.0 weight percent of the composition.

[0050] In some embodiments, the pharmaceutical composition is selected from those in Table 6. [Table 4] In some embodiments, the pharmaceutical components described in Table 6 further comprise cesium; The cesium is not more than 0.25 weight percent of the composition.

[0051] In some embodiments, the pharmaceutical components listed in Table 6 include cesium, Ru III Cl3(Hind)2(H2O), Ru III Cl3(Hind)2(CH3CN), and Ru III Further containing Cl3(Hind) (HN=C(Me)ind); the cesium is less than or equal to about 0.25 weight percent of the composition; Ru III Cl3(Hind)2(H2O) is less than or equal to about 0.5 weight percent of the composition; RuIII Cl3(Hind)2(CH3CN) is less than or equal to about 1.25 weight percent of the composition; And Ru III Cl3(Hind) (HN=C(Me)ind) is less than or equal to about 1.0 weight percent of the composition.

[0052] In some embodiments, the pharmaceutical components are as set forth in any of Tables 3-6 and further comprise cesium. In some embodiments, the cesium is present in an amount of about 0.001, 0.002, 0.003, 0.004, 0.005, 0.006, 0.007, 0.008, 0.009, 0.010, 0.015, 0.020, 0.025, 0.030, 0.035, 0.040, 0.045, 0.050, 0.055, 0.060, 0.065, present in an amount of 0.070, 0.075, 0.080, 0.085, 0.090, 0.095, 0.10, 0.15, 0.20, 0.25, 0.30, 0.35, 0.40, 0.45, 0.50, 0.55, 0.60, 0.65, 0.70, 0.75, 0.80, 0.85, 0.90, 0.95, or 1.0 weight percent.

[0053] In some embodiments, an effective amount of BOLD-100, alone or in combination, for example in combination with a PI, may be effective to reduce the amount of GRP78 in a cell following administration of BOLD-100.

[0054] In certain embodiments, BOLD-100 is administered after the PI, e.g., at least 1, 2, 3, 4, 5, 6, or 7 days, or 1-7 days, after the PI. Alternatively, BOLD-100 may be administered at least approximately simultaneously with the PI, e.g., within 1, 2, 3, 4, 5, or 6 hours of the PI, or within 20, 21, 22, 23, 24, 25, 26, 27, or 28 hours of each other. Alternatively, BOLD-100 may be administered before the PI, e.g., at least about 8, 9, 10, 11, 12, 13, 14, 15, or 16 hours before the PI.

[0055] In certain embodiments, treatment with BOLD-100 may be part of first-line or induction treatment, or alternatively, BOLD-100 may be used in the treatment of relapsed or refractory disease.For example, in MM or RRMM, BOLD-100 may be combined with one or more of PI (e.g., bortezomib, carfilzomib or ixazomib), lenalidomide, pomalidomide, dexamethasone, prednisone, cyclophosphamide, thalidomide, daratumumab, elotuzumab, isatuximab, selinexor, melphalan, cyclophosphamide, doxorubicin, liposomal doxorubicin and panobinostat.MM treatment with BOLD-100 may precede, for example, autologous stem cell transplantation.

[0056] Titratable dosages may be adapted, for example, to allow patients to take a medicine in a dose smaller than a unit dose, with a "unit dose" being defined as the maximum dose of a medicine that can be taken at any one time or within a particular dosing period. Since not all patients require the same dose to achieve the same benefit, dose titration allows different patients to incrementally increase the dose until they feel the medicine is effective. A person with a larger body or a faster metabolism may require a larger dose to achieve the same effect as another person with a smaller body or a slower metabolism. Thus, titratable dosages have an advantage over standard dosage forms.

[0057] In select embodiments, the formulation may be adapted to be delivered in a manner that targets one or more of the following: sublingual, buccal, oral, rectal, nasal, parenteral, and via the pulmonary system. The formulation may be, for example, in one or more of the following forms: gel, gel spray, tablet, liquid, capsule, by injection, or for vaporization.

[0058] Conventional pharmaceutical practice may be used to provide suitable formulations or compositions for administering the formulation to a subject.Routes of administration may include, for example, parenteral, intravenous, intradermal, subcutaneous, intramuscular, intracranial, intraorbital, intraocular, intraventricular, intracapsular, intraspinal, intrathecal, intracisternal, intraperitoneal, intranasal, inhalation, aerosol, topical, sublingual or oral administration.The therapeutic formulation may be in the form of a liquid solution or suspension; for oral administration, the formulation may be in the form of a tablet or capsule; for intranasal formulation, the formulation may be in the form of a powder, nasal drops or aerosol; for sublingual formulation, the formulation may be in the form of drops, aerosol or tablet.

[0059] Methods well known in the art for preparing formulations can be found, for example, in "Remington: The Science and Practice of Pharmacy" (21st edition), ed. David Troy, 2006, Lippincott Williams & Wilkins. Formulations for parenteral administration may contain, for example, excipients, sterile water, or saline, polyalkylene glycols, such as polyethylene glycol, oils of vegetable origin, or hydrogenated naphthalenes. Biocompatible biodegradable lactide polymers, lactide / glycolide copolymers, or polyoxyethylene-polyoxypropylene copolymers may be used to control the release of the compounds. Other potentially useful parenteral delivery systems include ethylene-vinyl acetate copolymer particles, osmotic pumps, implantable infusion systems, and liposomes. Formulations for inhalation may contain excipients, such as lactose, or may be aqueous solutions, such as polyoxyethylene-9-lauryl ether, glycocholate and deoxycholate, or may be oily solutions for administration in the form of nasal drops or as a gel.

[0060] The pharmaceutical composition of the present invention may be in any form that allows the composition to be administered to a patient.For example, the composition may be in the form of a solid, liquid or gas (aerosol).The pharmaceutical composition of the present invention is formulated so that the active ingredient contained therein is bioavailable upon administration of the composition to a patient.The composition administered to a patient may take the form of one or more dosage units, for example, a tablet, capsule or cachet may be a single dosage unit, and a container of the compound in aerosol form may hold multiple dosage units.

[0061] The materials used in the preparation of pharmaceutical compositions should be pharma- ceutical pure and non-toxic in the amount used. The compositions of the present invention may contain one or more compounds (active ingredients) known for a particular desired effect. It is clear to those skilled in the art that the optimal dosage of the active ingredient(s) in the pharmaceutical composition depends on various factors. Relevant factors include, but are not limited to, the type of subject (e.g., human), the specific form of the active ingredient, the mode of administration, and the composition used.

[0062] Generally, pharmaceutical compositions comprise the formulation of the present invention described herein in admixture with one or more carriers.The carrier(s) may be particulate, so that the composition is, for example, in tablet or powder form.The carrier(s) may be liquid, so that the composition is, for example, an oral syrup or an injectable liquid.Additionally, the carrier(s) may be gaseous, so as to provide an aerosol composition, for example, useful in inhalation administration.

[0063] When intended for oral administration, the compositions are preferably in either solid or liquid form, with semi-solid, semi-liquid, suspension and gel forms being included within the forms considered herein as either solid or liquid.

[0064] As a solid dosage form for oral administration, the composition may be formulated into a form such as a powder, granules, compressed tablets, pills, capsules, cachets, chewing gum, wafers, or lozenges, etc. Such solid compositions typically contain one or more inert diluents or edible carriers. Additionally, one or more of the following adjuvants may be present: binders, such as syrup, acacia, sorbitol, polyvinylpyrrolidone, carboxymethylcellulose, ethylcellulose, microcrystalline cellulose, gum tragacanth or gelatin, and mixtures thereof; excipients, such as starch, lactose or dextrin, disintegrants, such as alginic acid, sodium alginate, Primogel, and corn starch; lubricants, such as magnesium stearate or Sterotex; fillers, such as lactose, mannitol, starch, calcium phosphate, sorbitol, methylcellulose, and mixtures thereof; lubricants, such as magnesium stearate, high molecular weight polymers, such as polyethylene glycol, high molecular weight fatty acids, such as stearic acid, silica, wetting agents, such as sodium lauryl sulfate, glidants, such as colloidal silicon dioxide; sweeteners, such as sucrose or saccharin, flavorings, such as peppermint, methyl salicylate or orange flavoring, and coloring agents. When the composition is in the form of a capsule, for example, a gelatin capsule, it may contain, in addition to materials of the above type, a liquid carrier such as polyethylene glycol or a fatty oil.

[0065] The formulation may be in the form of a liquid, such as an elixir, syrup, solution, aqueous or oily emulsion or suspension, or even a dry powder that can be reconstituted with water and / or other liquid media prior to use. The liquid may be for oral administration or for delivery by injection, as two examples. When intended for oral administration, a preferred composition contains, in addition to the compound of the present invention, one or more of a sweetener, thickener, preservative (e.g., alkyl p-hydroxybenzoate), dye / colorant, and flavor enhancer (flavoring). In a composition intended to be administered by injection, one or more of a surfactant, a preservative (e.g., alkyl p-hydroxybenzoate), a wetting agent, a dispersing agent, a suspending agent (e.g., sorbitol, glucose, or other sugar syrup), a buffer, a stabilizer, and an isotonic agent may be included. The emulsifier may be selected from lecithin or sorbitol monooleate.

[0066] The liquid pharmaceutical formulations of the present invention, whether in solution, suspension or other similar form, may contain one or more of the following adjuvants: sterile diluents, such as water for injection, saline solution, preferably saline, Ringer's solution, isotonic sodium chloride, fixed oils, such as synthetic mono- or diglycerides, polyethylene glycols, glycerin, propylene glycol or other solvents that can serve as solvents or suspending media; antibacterial agents, such as benzyl alcohol or methylparabens; antioxidants, such as ascorbic acid or sodium bisulfite; chelating agents, such as ethylenediaminetetraacetic acid; buffers, such as acetate (salts), citrate (salts) or phosphate (salts) and agents for adjusting tonicity, such as sodium chloride or dextrose. Parenteral preparations may be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic. Saline is the preferred adjuvant. Injectable pharmaceutical compositions are preferably sterile.

[0067] Pharmaceutical preparations may be intended for external administration, in which case carriers may suitably comprise solution, emulsion, ointment, cream or gel bases.Bases may comprise, for example, one or more of the following: petrolatum, lanolin, polyethylene glycol, beeswax, mineral oil, diluents such as water and alcohol, and emulsifiers and stabilizers.Thickening agents may be present in pharmaceutical compositions for external administration.When intended for transdermal administration, compositions may comprise transdermal patch or iontophoresis device.

[0068] The preparation may be intended for rectal administration, for example, in the form of a suppository that melts in the rectum and releases the drug. The composition for rectal administration may contain an oily base as a suitable non-irritating excipient. Such bases include, but are not limited to, lanolin, cocoa butter and polyethylene glycol. Low melting waxes are preferred for the preparation of suppositories, and mixtures of fatty acid glycerides and / or cocoa butter are suitable waxes. The wax may be melted and the aminocyclohexyl ether compound is dispersed homogeneously therein by stirring. The molten homogeneous mixture is then poured into conveniently sized molds, cooled and thereby solidified.

[0069] Preparations may contain various materials that modify the physical form of solid or liquid dosage units.For example, compositions may contain materials that form coating shells around active ingredients.The materials that form coating shells are typically inert, and may be selected from, for example, sugar, shellac, and other enteric coating agents.Alternatively, active ingredients may be placed in gelatin capsules or cachets.

[0070] The pharmaceutical preparation may be composed of a gaseous dosage unit, for example, in the form of an aerosol. The term aerosol is used to describe a variety of systems ranging from those of colloidal nature to systems composed of pressurized packages. Delivery may be by liquefied or compressed gas or by a suitable pump system that dispenses the active ingredient. The aerosol of the compound of the present invention may be delivered in a single-phase, two-phase, or three-phase system to deliver the active ingredient(s). The delivery of the aerosol includes the necessary container, activator, valve, and subcontainer, etc., which may be taken together to form a kit.

[0071] Some biologically active compounds may be in the form of a free base or a pharma- ceutically acceptable salt, such as hydrochloride, sulfate, phosphate, citrate, fumarate, methanesulfonate, acetate, tartrate, maleate, lactate, mandelate, salicylate, succinate, and other salts known in the art. Appropriate salts are selected to enhance the bioavailability or stability of the compound for the appropriate mode (e.g., oral or parenteral route of administration) used.

[0072] The present invention also provides a kit containing the pharmaceutical preparation together with instructions for the use of the preparation.Preferably, the commercial package contains one or more unit doses of the preparation.Light and / or air sensitive preparations may require special packaging and / or formulation.For example, packaging that is light-tight and / or sealed from contact with ambient air and / or constructed with suitable coating or excipients may be used.

[0073] The formulation of the present invention can be provided alone or in combination with other compounds (e.g., small molecules, nucleic acid molecules, peptides, or peptide analogs) in the presence of carriers or any pharma- ceutically or biologically acceptable carriers.As used herein, "pharma-ceutically acceptable carriers" or "excipients" include any and all physiologically compatible solvents, dispersion media, coatings, antibacterial and antifungal agents, and isotonic and absorption retardants, etc.Carriers can be suitable for any suitable mode of administration.Pharmaceutically acceptable carriers generally include sterile aqueous solutions or dispersions and sterile powders.Auxiliary active compounds can also be incorporated into the formulation.

[0074] An "effective amount" of a formulation according to the present invention includes a therapeutically effective amount or a prophylactically effective amount. A "therapeutically effective amount" refers to an amount effective, at the dosage and time period required, to achieve a desired therapeutic result. A therapeutically effective amount of a formulation may vary according to factors such as the disease state, age, sex, and weight of the individual, and the ability of the compound to elicit a desired response in the individual. Dosage regimens may be adjusted to provide optimal therapeutic success. A therapeutically effective amount may also be an amount in which the therapeutically beneficial effects outweigh any toxic or adverse effects of the formulation or active compound. A "prophylactically effective amount" refers to an amount effective, at the dosage and time period required, to achieve a desired prophylactic result. Typically, a prophylactic dose is used prior to or at an early stage of disease in a subject, such that the prophylactically effective amount may be lower than the therapeutically effective amount. For any particular subject, the timing and dosage of treatment may be adjusted over time (e.g., timing may be daily, every other day, weekly, monthly) according to the individual's need and the professional judgment of the person administering or supervising the administration of the composition.

[0075] In therapeutic applications, synergy between active ingredients occurs when the observed combined therapeutic effect is greater than the sum of the therapeutic effects of the individual active ingredients, or when new therapeutic effects are generated that the active ingredients could not generate alone. Thus, when the components of a formulation are present in synergistically effective amounts, the formulation produces a therapeutic effect that is greater than that achieved by the individual active ingredients administered alone in equivalent dosages. In this context, the enhancement of therapeutic effect may take the form of increased efficacy or potency and / or reduced adverse effects. The synergistic effect may be mediated in whole or in part by the pharmacokinetics and / or pharmacodynamics of the active ingredients in the subject, so that the amounts and ratios of the ingredients in the formulation may be synergistic in vivo. This in vivo synergy may be produced using a formulation that contains the active ingredients in amounts and ratios that are also synergistic in in vitro assays of efficacy. As used herein, the term "synergistically effective amount" thus refers to an amount that is synergistic in vivo and / or in vitro.

[0076] Synergy can be calculated by multiple drug effect analysis according to the methodology described by Chou and Talalay using Compusyn software, version 1.0, and can be numerically reflected in compound interactions performed by the median equation principle (see Chou TC. “Drug combination studies and their synergy quantification using the Chou-Talalay method.” Cancer Res. 2010 Jan 15;70(2):440-6). The CI value indicates the synergistic, additive or antagonistic behavior of drug combinations. CI<1, =1, and >1 indicate synergy, additive effect, and antagonism, respectively.

[0077] An alternative numerical quantification of synergy is often expressed as the fractional inhibitory concentration index (FICI), which represents the sum of the fractional inhibitory concentration (FIC) of each drug tested, and is determined for each drug by dividing the minimum inhibitory concentration (MIC; the lowest concentration of drug that prevents visible growth of bacteria in a standard in vitro assay - a standard colorimetric assay based on resazurin) of each drug when used in combination by the MIC of each drug when used alone. In a very general sense, an FICI lower or higher than 1 indicates positively correlated activity (at least additive synergy) or the absence of a positive interaction, respectively. More definitively, synergy of two compounds can be conservatively defined as a FICI of ≦0.5 (see Odds, 2003; additivity or additive synergy corresponds to a FICI of >0.5 and ≦1; no interaction (indifference) corresponds to a FICI of >1 and ≦4; antagonism corresponds to a FICI of >>4). Synergy of three compounds has been defined as a FICI of ≦1.0. (Berenbaum, 1978; Yu et al., 1980).

[0078] [Example] As shown in the Examples below, BOLD-100 reverses PI resistance.

[0079] Example 1: Sensitivity to BOLD-100 The anticancer activity of trans-[tetrachloridobis(1H-indazole)ruthenate(III)]sodium (BOLD-100) was demonstrated by inhibition of proliferation of various multiple myeloma (MM) cancer cell lines. Specifically, various MM cell lines were treated with BOLD-100 for 24 hours and subjected to a Cell Titer Glo assay, which provides a metabolic readout corresponding to the number of metabolically active viable cells in the culture. Luminescence readings obtained in the assay were used to measure the percentage of cell death after 24 hours of treatment with increasing doses of BOLD-100. Each cell line was treated in triplicate, and the average cell death was used to determine the IC in these cell lines. 50 or 50% maximal inhibitory concentration dose was calculated using a nonlinear regression fit. Figure 1 shows the growth curves of different cell lines in the presence of increasing doses of BOLD-100. Table 1 shows the IC 50 Indicates the value. [Table 5] Example 2: Drug-resistant cell lines are more sensitive to BOLD-100 Proteasome inhibitors (PIs) and immunomodulatory drugs (IMiDs) are used as standard of care treatments in MM disease in the clinic. BOLD-100 was found to be more potent in MM cell lines that had developed to acquire resistance to each of these classes of drugs compared to sensitive controls as demonstrated in in vitro CTG assays. Specifically, the MM.1S cell line was developed to acquire partial resistance to the proteasome inhibitors bortezomib (MM.1S BR) and carfilzomib (MM.1S CR) by treating the MM.1S cell line with increasing doses of the drugs for approximately 6 months, and the OPM2 cell line was developed to acquire partial resistance to the IMiDs lenalidomide (OPM2 LR) and pomalidomide (OPM2 PR) by treating the OPM2 cell line with increasing doses of the drugs for approximately 6 months. Once resistance developed, cells were tested for their ability to grow independent of the presence of drug and all drug-resistant cell lines were found to be able to grow in the absence of bortezomib / carfilzomib and lenalidomide / pomalidomide while maintaining their growth rate in the absence of drugs. Figure 2A shows the growth curves of MM.1S and MM.1S BR in the presence of increasing doses of BOLD-100 over a 24 h period, demonstrating the higher sensitivity of the PI-resistant lines to BOLD-100 compared to the parental MM.1S (BOLD-100 IC 50 MM.1S = 83 μM; MM.1S BR = 23 μM). Figure 2B shows the growth curves of OPM2, OPM2 LR and OPM2 PR in the presence of increasing doses of BOLD-100 over a 24-hour period, demonstrating the higher sensitivity of lenalidomide- and pomalidomide-resistant lines to BOLD-100 (BOLD-100 IC 50 :OPM2=120μM;OPM2 PR=51μM).

[0080] Example 3: BOLD-100 overcomes PI resistance in vitro Cell lines were subjected to BOLD-100+PI treatment to investigate to what extent BOLD-100 could affect the survival of these resistant cell lines and overcome their resistance to PI. PI-sensitive and PI-resistant cells were either treated simultaneously with three increasing doses of both drugs, i.e., BOLD-100 and the respective PI, bortezomib (Btx) or carfilzomib (Cfz), for 24 h and subjected to CTG assay to determine survival (Figures 3A-3F), or treated sequentially with drugs, where cells were first exposed to a proteasome inhibitor (bortezomib or carfilzomib) for 24 h, followed by the addition of BOLD-100 for another 24 h (Figures 3G-3L).

[0081] The survival percentage normalized to the vehicle control in each cell line was expressed as the proportion affected by the drug combination at the concentrations used ("effect size ratio"). Compound interactions were calculated as combination index (CI) by multiple drug effect analysis performed by the median equation principle according to the methodology described by Chou and Talalay using Compusyn software, version 1.0 (see Chou TC. "Drug combination studies and their synergy quantification using the Chou-Talalay method." Cancer Res. 2010 Jan 15;70(2):440-6). CI values ​​are one way to indicate synergistic, additive or antagonistic behavior of drug combinations. Using this method, CI<0.8, =0.8-1.2, and >1.2 indicate synergy, additive effects, and antagonism, respectively, as shown in the figure. FIG. 3 depicts data derived from growth curves of parental and resistant cells in the presence of the drug combinations listed below demonstrating a decrease in survival with increasing concentrations of both drugs, as well as respective CI vs. effect size ratio plots for each cell line indicating synergy, additivity or antagonism: Bortezomib + BOLD-100 B100-25uM / Btx 2.5nM B100-50uM / Btx 2.5nM B100-100uM / Btx 2.5nM B100-25uM / Btx 5nM B100-50uM / Btx 5nM B100-100uM / Btx 5nM B100-25uM / Btx 10nM B100-100uM / Btx 10nM B100-100uM / Btx 10nM Carfilzomib + BOLD-100 B100-25uM / Cfz 2.5nM B100-50uM / Cfz 2.5nM B100-100uM / Cfz 2.5nM B100-25uM / Cfz 5nM B100-50uM / Cfz 5nM B100-100uM / Cfz 5nM B100-25uM / Cfz 10nM B100-100uM / Cfz 10nM B100-100uM / Cfz 10nM (B100 is BOLD-100, Btx is bortezomib, and Cfz is carfilzomib).

[0082] Figures 3A and 3B show that the combination of BOLD-100 plus bortezomib or carfilzomib in PI-sensitive MM cell lines shows an additive effect of the combination, and Figures 3C and 3E show that BOLD-100 demonstrates a synergistic effect on cell survival when combined with either bortezomib or carfilzomib in matched resistant cell lines. For bortezomib-resistant cell lines, the combination of BOLD-100 with carfilzomib as shown in Figure 3D also shows a synergistic impact on cell survival, indicating cross-resistance, an effect not seen in carfilzomib-resistant cell lines when BOLD-100 is combined with bortezomib in Figure 3F. The effect of co-dosing shown in Figures 3A-3F is also evident in the corresponding sequential dosing data shown in Figures 3G-3L.

[0083] An alternative way of showing the effects shown in the data of FIG. 3 is provided in FIG. 3L, where all cells in each data set are treated with 10 nM bortezomib (left hand bars) or 10 nM carfilzomib (right hand bars). As shown, when no BOLD-100 is added (DMSO column), there is a lower response in the resistant cell line compared to the sensitive cell line. Importantly, as the concentration of BOLD-100 is increased, the relative efficacy of treatment is reversed between sensitive and resistant cells. For example, at 100 uM BOLD-100, there is roughly double the cell death in the resistant cell line compared to the sensitive cell line.

[0084] Example 4: Efficacy of BOLD-100 in PI-resistant cells demonstrated in clonogenic assays The effect of BOLD-100 in combination with proteasome inhibitors was also demonstrated in a colony formation assay where BOLD-100+PI was tested in both PI-sensitive and PI-resistant cell lines in a clonogenic assay. The design follows standard methods used in hematopoietic cells. In particular, cells and drugs were mixed together with semi-solid medium and plated sparsely to allow individual cells to form colonies. Cells were then allowed to grow into colonies in semi-solid medium for one week, after which colonies were stained with crystal violet and images were captured as shown in Figure 4A and Figure 4B. Figure 4 shows that there was a strong negative impact of BOLD-100 in combination with PI on cell survival when measured over a longer period of time, and in this assay, prevention of colony regrowth was seen, demonstrating the strong cytotoxic effect of the drug combination in both sensitive and resistant cell lines.

[0085] Example 5: Efficacy of BOLD-100 maintained in co-culture with bone marrow-derived stromal cells It is understood that bone marrow stromal cells provide a protective environment for the cells, isolating the bone marrow cells from drug effects. This example shows that BOLD-100 maintains its efficacy in PI-sensitive and PI-resistant MM cell lines in the presence of bone marrow stromal cells, an environment that partially mimics the in vivo environment. To do this, a co-culture assay was performed in which MM cells were grown in co-culture with the bone marrow stromal cell line HS-5. The MM cell lines adhere to the stromal cells in the culture, and the stromal cells can provide a protective effect against drug-induced apoptosis / growth inhibition. Dose-response curves were generated for the PI-sensitive cell line MM.1 S, as well as the two PI-resistant lines MM.1S BR and MM.1S CR, in the presence of HS-5 by standard CTG assay. As shown in Figure 5, BOLD-100 was able to negatively affect the survival of all three MM.1S cell lines, but had minimal impact in the bone marrow cell line HS-5.

[0086] References Chou TC.Drug combination studies and their synergy quantification using the Chou-Talalay method.Cancer Res.2010Jan15;70(2):440-6. Fricker LD.Proteasome Inhibitor Drugs.Annu Rev Pharmacol Toxicol.2020 Jan 6;60:457-476. Manasanch, E., Orlowski, R. Proteasome inhibitors in cancer therapy. Nat Rev Clin Oncol14, 417-433 (2017). Robak, P., Robak, T. Bortezomib for the Treatment of Hematologic Malignancies:15Years Later.Drugs RD 19,73-92(2019). Burris,H.A.et al.Safety and activity of IT-139,a ruthenium-based compound,in patients with advanced solid tumours:a first-in-human,open-label,dose-escalation phase I study with expansion cohort.ESMO open.1,e000154(2016). Although various embodiments of the present invention are disclosed herein, many adaptations and modifications may be made within the scope of the present invention in accordance with the ordinary general knowledge of those skilled in the art. Such modifications include the substitution of any aspect of the present invention with a known equivalent to achieve the same result in substantially the same way. Terms such as "exemplary" or "exemplary" are used herein to mean "serving as an example, instance, or illustration." Any embodiment described herein as "exemplary" or "exemplary" should not therefore be construed as necessarily preferred or advantageous over other embodiments, and all such embodiments are independent embodiments. Unless otherwise stated, numerical ranges are inclusive of the numbers defining the range, and the numbers are necessarily approximate to the given order of magnitude. The term "comprising" is used herein as an open-ended term and is substantially equivalent to the phrase "including, but not limited to," with the word "comprises" having a corresponding meaning. As used herein, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, a reference to "a thing" includes more than one such thing. Reference to a reference herein is not an admission that such reference is prior art to the present invention. Any priority document(s) and all publications, including but not limited to patents and patent applications, referenced herein, and all documents referenced in such documents and publications, are hereby incorporated by reference as if each individual publication was specifically and individually indicated to be incorporated by reference herein, and as if fully set forth herein. The present invention includes all embodiments and variations substantially as described above and with reference to the examples and drawings.

[0087] In some embodiments, the present invention excludes steps involving medical or surgical procedures.

Claims

1. 1. A composition comprising sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] and / or a proteasome inhibitor (PI) for use in a method for treating a disease, which is cancer or a lymphoproliferative disorder, in a human patient in need thereof, the method comprising administering to the patient an effective amount of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] and, in combination, an effective amount of a proteasome inhibitor (PI).

2. 1. A composition comprising sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] and / or a proteasome inhibitor (PI) for use in a method for ameliorating therapeutic resistance to a proteasome inhibitor (PI) in a disease in a patient in need thereof, the method comprising administering to the patient an effective amount of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)].

3. The composition described in claim 2, wherein the method further comprises administering to the patient an effective amount of the PI in combination.

4. 4. The composition of claim 1 or 3, wherein the effective amount of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] and the combined effective amount of the PI are synergistically effective.

5. 3. The composition of claim 1 or 2, wherein the disease is resistant to treatment with the PI or the disease is prone to developing resistance to the PI.

6. 3. The composition of claim 1 or 2, wherein the PI is bortezomib, carfilzomib, ixazomib citrate, marizomib, oprozomib, delanzomib, ONX 0914 or KZR 616.

7. The composition of claim 1 or 2, wherein the PI is a constitutive proteasome inhibitor.

8. The composition of claim 1 or 2, wherein the PI is an inhibitor of the immunoproteasome.

9. The composition of claim 1 or 2, wherein the disease is myeloma.

10. The composition of claim 1 or 2, wherein the disease is multiple myeloma (MM).

11. 10. The composition of claim 9, wherein the MM is relapsed / refractory MM.

12. 3. The composition of claim 1 or 2, wherein the disease is an amyloidosis, a lymphoproliferative disorder, a plasma cell disorder or a lymphoid malignancy.

13. 3. The composition of claim 1 or 2, wherein the disease is mantle cell lymphoma, Waldenstrom's macroglobulinemia, or T-cell lymphoma amyloidosis.

14. 2. The composition of claim 1, wherein the cancer is relapsed / refractory multiple myeloma and the PI is bortezomib or carfilzomib.

15. 1. A composition comprising sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)] for use in a method for treating a myeloma that is resistant to a therapeutic agent, the method comprising administering to a patient an effective amount of sodium trans-[tetrachloridobis(1H-indazole)ruthenate(III)], wherein the therapeutic agent is effective to treat a myeloma that is not resistant.

16. 16. The composition of claim 15, wherein the therapeutic agent is a proteasome inhibitor (PI), an immunomodulatory imide drug, a cereblon modulator drug, thalidomide, or a thalidomide analog.

17. 17. The composition of claim 16, wherein the PI is bortezomib, carfilzomib, ixazomib citrate, marizomib, oprozomib, delanzomib, ONX 0914, or KZR 616.

18. The composition of claim 16 , wherein the PI is a constitutive proteasome inhibitor.

19. The composition of claim 16 , wherein the PI is an inhibitor of the immunoproteasome.

20. 17. The composition of claim 16, wherein the thalidomide analog is lenalidomide, pomalidomide, or iveldimide.

21. 16. The composition of claim 15, wherein the myeloma is multiple myeloma (MM).

22. 22. The composition of claim 21, wherein the MM is relapsed / refractory MM.

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