WO2016073669A1 - Combination therapy of hsp90 inhibitory compounds with mtor inhibitors - Google Patents

Abstract

The present invention features methods of treating or preventing a sarcoma in a subject, comprising administering a therapeutically effective amount of an Hsp90 inhibitor in combination with an mTOR inhibitor, wherein the subject is selected as being of 16 years of age or less. The present invention also features compositions comprising an Hsp90 inhibitor and an mTOR inhibitor, and kits comprising the same.

Description

COMBINATION THERAPY OF HSP90 INHBITORY COMPOUNDS WITH mTOR INHIBITORS

RELATED APPLICATIONS

[0001] This application claims the benefit of priority to U.S. Provisional Patent Application No. 62/075,581, filed on November 5, 2014, the entire contents of which are hereby incorporated herein by reference.

BACKGROUND OF THE INVENTION

[0002] Heat shock proteins (HSPs) are a class of chaperone proteins that are up- regulated in response to elevated temperature and other environmental stresses, such as ultraviolet light, nutrient deprivation and oxygen deprivation. HSPs act as chaperones to other cellular proteins (called client proteins), facilitate their proper folding and repair and aid in the refolding of misfolded client proteins. There are several known families of HSPs, each having its own set of client proteins. The Hsp90 family is one of the most abundant HSP families accounting for about 1-2% of proteins in a cell that is not under stress, increasing to about 4-6% in a cell under stress. Inhibition of Hsp90 results in the degradation of its client proteins via the ubiquitin proteasome pathway. Unlike other chaperone proteins, the client proteins of Hsp90 are mostly protein kinases or transcription factors involved in signal transduction, and a number of its client proteins have been shown to be involved in the progression of cancer.

[0003] Ganetespib is a selective inhibitor of Hsp90. Many solid and hematologic tumors are dependent on Hsp90 client proteins including proteins involved in "oncogene addiction" (ALK, HER2, mutant BRAF and EGFR, androgen receptor, estrogen receptor, and JAK2); proteins involved in resistance to chemotherapy and radiation therapy (ATR, BCL2, BRCAl/2, CDKl/4, CHK1, survivin, and WEE1); proteins involved in angiogenesis (HIF-lalpha, VEGFR, PDFGR, and VEGF); and proteins involved in metastasis (MET, RAF, AKT, MMPs, HIF-lalpha, and IGF-1R). In preclinical models, inhibition of Hsp90 by ganetespib results in the inactivation, destabilization, and eventual degradation of these cancer-promoting proteins.

Ganetespib is being evaluated in trials in lung cancer, breast cancer, and other tumor types.

[0004] Malignant peripheral nerve sheath tumors (MPNSTs) are soft tissue sarcomas arising from peripheral nerve or show nerve sheath differentiation and are associated with a high risk of local recurrence and metastasis. MPNSTs account for 10% of all soft tissue sarcomas, and carry the highest risk for sarcoma specific death among all the soft tissue sarcoma histologies . At present, complete surgical resection is the only curative treatment for MPNST. MPNSTs are at high risk for local recurrence (32-65%) 5 and metastasis (40%). The most frequent sites of metastasis of MPNSTs are lung, liver, brain, soft tissue, bone, lymph nodes, and retroperitoneum . The outcome for unresectable, recurrent, or metastatic MPNST is dismal. Outcomes for unresectable, recurrent, or metastatic MPNSTs are very poor, underscoring an urgent need for new therapeutic options. Accordingly, there is a need in the art for new therapeutic strategies to treat this disease.

SUMMARY OF THE INVENTION

[0005] Hsp90 inhibitors, in particular Ganetespib, in combination with mTOR inhibitors, in particular sirolimus (rapamycin), are demonstrated herein to be particularly effective in specific dosing regimens for treating pediatric patients with sarcomas. It is also demonstrated herein that those Hsp90 inhibitors in combination with mTOR inhibitors, in particular sirolimus, are particularly effective in treating refractory sarcomas, including malignant peripheral nerve sheath tumors (MPNSTs). It is a surprising and unexpected finding of the present invention that drugs that target Hsp90 and mTOR show synergistic activity in treating sarcomas, and in particular MPNSTs. The particular dosing regimens disclosed herein demonstrate potency against these certain specific types of cancer, while showing minimal side effects.

[0006] Accordingly, in a first aspect, the invention provides methods of treating or preventing a sarcoma in a subject, comprising administering a therapeutically effective amount of an Hsp90 inhibitor in combination with an mTOR inhibitor, wherein the subject is selected as being of 16 years of age or less.

[0007] In another embodiment, the invention provides methods of reducing or inhibiting the progression of a sarcoma in a subject, comprising administering a

therapeutically effective amount of an Hsp90 inhibitor in combination with an mTOR inhibitor, wherein the subject is selected as being of 16 years of age or less.

[0008] In related embodiments, the Hsp90 inhibitor is ganetespib (5-[2,4-dihydroxy-5-(l- methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lH-indol-5-yl)-3H-l,2,4-triazole-3-one), or a pharmaceutically acceptable salt or a tautomer thereof.

[0009] In related embodiments, the mTOR inhibitor is sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)-9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a- hexadecahydro-9,27-dihydroxy-3-[(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l- methylethyl]-10,21-dimethoxy-6,8,12,14,20,26-hexamethyl-23,27-epoxy-3H-pyrido[2,l-c][l,4] oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone.

[0010] In one embodiment, the sarcoma is a refractory sarcoma, e.g., a bone or soft tissue sarcoma. In another embodiment, the sarcoma is a malignant peripheral nerve sheath tumor (MPNST), e.g., an unresectable or metastatic sporadic or neurofibromatosis type-1 associated MPNST.

[0011] In related embodiments, the sarcoma was previously treated and not responsive. In another embodiment, the the sarcoma is a recurrent sarcoma.

[0012] In one embodiment, the amount of the Hsp90 inhibitor administered is from 100- 200 mg/m². In another embodiment, the amount of the Hsp90 inhibitor administered is from 150-200 mg/m². [0013] In one embodiment, the Hsp90 inhibitor is administered intravenously. In another embodiment, the Hsp90 inhibitor is infused over 60 minutes.

[0014] In one embodiment, the amount of mTOR inhibitor administered is 2-12 mg. In another embodiment, the amount of mTOR inhibitor administered is 2-4 mg.

[0015] In one embodiment, the mTOR inhibitor is administered orally.

[0016] In one embodiment, the Hsp90 inhibitor and mTOR inhibitor are administered over a 28 day cycle. In a related embodiment, the 28 day cycle is repeated one or more times.

[0017] In one embodiment, the Hsp90 inhibitor is administered on days 1, 8 and 15.

[0018] In one embodiment, the mTOR inhibitor is administered on days 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27 and 28.

[0019] In one aspect, the invention provides methods of treating or preventing a sarcoma in a subject, comprising administering a therapeutically effective amount of ganetespib (5-[2,4-dihydroxy-5-(l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lH-indol-5- yl)-3H-l,2,4-triazole-3-one), or a pharmaceutically acceptable salt or a tautomer thereof, in combination with sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)-9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27-dihydroxy-3-[(lR)- 2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21-dimethoxy- 6,8,12,14,20,26-hexamethyl-23,27-epoxy-3H-pyrido[2,l-c][l,4] oxaazacyclohentriacontine- 1,5,11,28,29 (4H,6H ,31H)-pentone, wherein the subject is selected as being of 16 years of age or less.

[0020] In one aspect, the invention provides methods of reducing or inhibiting the progression of a sarcoma in a subject, comprising administering a therapeutically effective amount of ganetespib (5-[2,4-dihydroxy-5-(l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl- lH-indol-5-yl)-3H-l,2,4-triazole-3-one), or a pharmaceutically acceptable salt or a tautomer thereof, in combination with sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)-9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27- dihydroxy-3-[(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21- dimethoxy-6,8,12,14,20,26-hexamethyl-23,27-epoxy-3H-pyrido[2,l-c][l,4] oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone, wherein the subject is selected as being of 16 years of age or less.

[0021] In one aspect, the invention provides kits to practice any one of the methods described herein.

[0022] In one embodiment, the invention provides compositions comprising an Hsp90 inhibitor and an mTOR inhibitor. In one embodiment, the Hsp90 inhibitor is ganetespib (5- [2,4-dihydroxy-5-(l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lH-indol-5-yl)-3H-l,2,4- triazole-3-one), or a pharmaceutically acceptable salt or a tautomer thereof. In another embodiment, the mTOR inhibitor is sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)-9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27- dihydroxy-3-[(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21- dimethoxy-6,8,12,14,20,26-hexamethyl-23,27-epoxy-3H-pyrido[2,l-c][l,4]

oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone.

[0023] In one embodiment, the amount of the Hsp90 inhibitor is between 100-500 mg/m². In another embodiment, the amount of the Hsp90 inhibitor is between 100-200 mg/m², preferably 150-200 mg/m².

[0024] In one embodiment, the amount of mTOR inhibitor is between 2-12 mg. In another embodiment, the amount of mTOR inhibitor is between 2-4 mg.

[0025] In one embodiment, the compostion comprising the combination of the Hsp90 inhibitor and the mTOR inhibitor exhibits therapeutic synergy.

DETAILED DESCRIPTION OF THE INVENTION

[0026] The present invention provides novel combinations of drugs that target Hsp90 and mTOR to treat refractory sarcomas, and in particular MPNSTs, in pediatric patients. It is a surprising and unexpected finding of the present invention that drugs that target Hsp90 and mTOR show synergistic activity in treating sarcomas, and in particular MPNSTs. DEFINITIONS

[0027] Unless otherwise specified, the below terms used herein are defined as follows:

[0028] he articles "a", "an" and "the" are used herein to refer to one or to more than one (i.e. to at least one) of the grammatical object of the article unless otherwise clearly indicated by contrast. By way of example, "an element" means one element or more than one element.

[0029] The term "including" is used herein to mean, and is used interchangeably with, the phrase "including but not limited to".

[0030] The term "or" is used herein to mean, and is used interchangeably with, the term "and/or," unless context clearly indicates otherwise.

[0031] The term "such as" is used herein to mean, and is used interchangeably, with the phrase "such as but not limited to".

[0032] Unless specifically stated or obvious from context, as used herein, the term "about" is understood as within a range of normal tolerance in the art, for example within 2 standard deviations of the mean. About can be understood as within 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1 %, 0.05%, or 0.01% of the stated value. Unless otherwise clear from context, all numerical values provided herein can be modified by the term about.

[0033] Any compositions or methods provided herein can be combined with one or more of any of the other compositions and methods provided herein.

[0034] As used herein, the terms "administer," "administering" or

"administration" include any method of delivery of a pharmaceutical composition or agent into a subject's system or to a particular region in or on a subject. In certain embodiments of the invention, an agent is administered intravenously,

intramuscularly, subcutaneously, intradermally, intranasally, orally,

transcutaneously, or mucosally. In a preferred embodiment, an agent is administered intravenously. Administering an agent can be performed by a number of people working in concert. Administering an agent includes, for example, prescribing an agent to be administered to a subject and/or providing instructions, directly or through another, to take a specific agent, either by self-delivery, e.g., as by oral delivery, subcutaneous delivery, intravenous delivery through a central line, etc.; or for delivery by a trained professional, e.g., intravenous delivery,

intramuscular delivery, intratumoral delivery, etc.

[0035] The term "agent" refers to any active agent that affects any biological process. Active agents that are considered drugs for purposes of this application are agents that exhibit a pharmacological activity. Examples of agents include active agents that are used in the prevention, diagnosis, alleviation, treatment or cure of a disease condition. In an exemplary embodiment, an agent is a chemotherapeutic agent.

[0036] As used herein, the term "in combination" refers to the use of more than one therapeutic agent (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, or more). The use of the term "in combination" does not restrict the order in which the therapeutic agents are administered to a subject afflicted with cancer. A first therapeutic agent, such as a compound described herein, can be administered prior to (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks before), concomitantly with, or subsequent to (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks after) the administration of a second

therapeutic agent or treatment, such as an anti-cancer agent, to a subject with cancer. In certain embodiments, one agent may be administered more frequently than the other agent such that multiple doses of one agent are administered for each dose of the other agent(s). [0037] As used herein, "diagnosing" and the like, is meant to refer to a clinical or other assessment of the condition of a subject based on observation, testing, or circumstances for identifying a subject having a disease, disorder, or condition based on the presence of at least one indicator, such as a sign or symptom of the disease, disorder, or condition. Typically, diagnosing using the method of the invention includes the observation of the subject for multiple indicators of the disease, disorder, or condition in conjunction with the methods provided herein. Diagnostic methods provide an indicator that a disease is or is not present. A single diagnostic test typically does not provide a definitive conclusion regarding the disease state of the subject being tested.

[0038] As used herein, the terms "identify" or "select" refer to a choice in preference to another. In other words, to identify a subject or select a subject is to perform the active step of picking out that particular subject from a group and confirming the identity of the subject by name or other distinguishing feature.

[0039] As used herein, the term "recur" refers to the re-growth of tumor or cancerous cells in a subject in whom primary treatment for the tumor has been administered. The tumor may recur in the original site or at another site.

[0040] As used herein, the term "refractory" cancer or tumor is understood as a malignancy which is either initially unresponsive to chemo- or radiation therapy, or which becomes unresponsive over time. A cancer refractory to on intervention may not be refractory to all interventions. A refractory cancer is typically not amenable to treatment with surgical interventions. Exemplary refractory cancers or tumors of the invention include refractory sarcomas, for example refractory bone and soft tissue sarcomas, and in particular MPNSTs.

[0041] As used herein, the term "sarcoma" is meant to refer to a broad group of cancers that includes tumors that form in the bones and in the soft tissues. In certain embodiments, the sarcoma is a refractory sarcoma, which may be a bone or soft tissue sarcoma. In other embodiments, the sarcoma is a malignant peripheral nerve sheath tumor (MPNST).

[0042] As used herein, terms "subject", "patient" and "mammal" are used interchangeably. The terms "subject" and "patient" refer to an animal (e.g., a bird such as a chicken, quail or turkey, or a mammal), preferably a mammal including a non- primate (e.g., a cow, pig, horse, sheep, rabbit, guinea pig, rat, cat, dog, and mouse) and a primate (e.g., a monkey, chimpanzee and a human), and more preferably a human. In one embodiment, the subject is a non-human animal such as a farm animal (e.g., a horse, cow, pig or sheep), or a pet (e.g., a dog, cat, guinea pig or rabbit). In another embodiment, the subject is a human.

[0043] As used herein, the term "synergistic" or "synergy" refers to a combination of compounds described herein, which, when taken together, is more effective than the additive effects of the individual therapies. A synergistic effect of a combination of therapies {e.g., a combination of therapeutic agents) permits the use of lower dosages of one or more of the therapeutic agent(s) and/or less frequent

administration of the agent(s) to a subject with a disease or disorder, e.g., a proliferative disorder. The ability to utilize lower the dosage of one or more therapeutic agent and/or to administer the therapeutic agent less frequently reduces the toxicity associated with the administration of the agent to a subject without reducing the efficacy of the therapy in the treatment of a disease or disorder. In addition, a synergistic effect can result in improved efficacy of agents in the prevention, management or treatment of a disease or disorder, e.g. cancer. Finally, a synergistic effect of a combination of therapies may avoid or reduce adverse or unwanted side effects associated with the use of either therapeutic agent alone.

[0044] As used herein, a "therapeutically effective amount" is that amount sufficient to treat a disease in a subject. A therapeutically effective amount can be administered in one or more administrations. [0045] As used herein, the terms "treat," "treating" or "treatment" refer, preferably, to an action to obtain a beneficial or desired clinical result including, but not limited to, alleviation or amelioration of one or more signs or symptoms of a disease or condition, diminishing the extent of disease, stability (i.e., not worsening) state of disease, amelioration or palliation of the disease state, diminishing rate of or time to progression, and remission (whether partial or total), whether detectable or undetectable. "Treatment" can also mean prolonging survival as compared to expected survival in the absence of treatment. Treatment does not need to be curative.

[0046] The recitation of an embodiment for a variable or aspect herein includes that embodiment as any single embodiment or in combination with any other embodiments or portions thereof.

[0047] Any compositions or methods provided herein can be combined with one or more of any of the other compositions and methods provided herein.

Sarcomas

Malignant peripheral nerve sheath tumors

[0048] Malignant peripheral nerve sheath tumors (MPNSTs) are soft tissue sarcomas arising from peripheral nerve or show nerve sheath differentiation and are associated with a high risk of local recurrence and metastasis (1). MPNSTs account for 10% of all soft tissue sarcomas, and carry the highest risk for sarcoma specific death among all the soft tissue sarcoma histologies (2). At present, complete surgical resection is the only curative treatment for MPNST (3,4). MPNSTs are at high risk for local recurrence (32-65%) (5) and metastasis (40%) (6). The most frequent sites of metastasis of MPNSTs are lung, liver, brain, soft tissue, bone, lymph nodes, and retroperitoneum (7). The outcome for unresectable, recurrent, or metastatic MPNST is dismal. Neurofibromatosis Type 1 and MPNST

[0049] Approximately half of all MPNSTs arise from individuals with NF1( 8). NF1 is a common autosomal dominant tumor predisposition syndrome. The gene responsible for NF1 encodes for a protein called neurofibromin, which includes a GTPase activating protein that regulates hydrolysis of Ras-GTP to Ras-GDP (9,10). Patients with NF1 have decreased levels of neurofibromin, which can lead to dysregulated Ras and tumorigenesis (11). MPNSTs are the most common NF1- associated malignancy. The lifetime risk of MPNST in NF1 is 8-13% compared to 0.001% in the general population (7,8). The majority of NFl-associated MPNSTs arise within pre-existing plexiform neurofibromas (12). This may lead to difficulty and delay in diagnosing MPNSTs in patients with NF1 because the clinical indicators of malignancy such as mass, pain, and edema may also be features of active, benign plexiform neurofibromas. NF1 associated MPNSTs are frequently more located in the trunk (13,14) as opposed to extremity location seen more commonly in sporadic MPNSTs, tend to be large (>5cm) (15), and may have a greater propensity to metastasize ( 7). These may be potential reasons why NFl-associated MPNSTs appear to have a worse outcome than sporadic MPNSTs. Gene expression profiling of NF1 associated (n=25) and sporadic (n=17) MPNSTs did not identify a molecular signature that could reliably distinguish between both groups (16,17).

[0050] The only known curative approach to MPNSTs is complete surgical resection with wide negative margins (3,4). The role of chemotherapy for MPNSTs has not been defined to date.

Refractory bone and soft tissue sarcomas

[0051] The prognosis in advanced and metastatic sarcomas remains poor.

Continued progress while minimizing acute and late effects of current therapy is dependent on the development of novel therapeutic approaches for malignant sarcomas. Hsp90 inhibitors have demonstrated benefit in a variety preclinical of bone and soft tissue sarcomas, including synovial sarcoma (21, 32) Ewing Sarcoma (33), osteosarcoma (34, 35), and rhabdomyosarcoma (36, 37). Disruptions of the PI3K-Akt-mTOR signaling pathway are associated with different sarcoma types (38). Rapamycin and other mTOR inhibitors have demonstrated inhibition of growth in a variety of in vitro and in vivo sarcoma preclinical models (39-43). Early clinical studies of single agent mTOR inhibition have demonstrated potential benefit in a subset of sarcoma patients who have failed standard therapy (variety of soft tissue sarcoma, GIST, Ewing Sarcoma, and Osteosarcoma), however the activity of this drug has been primarily cytostatic (38), and activation of feedback mechanisms have been of concern (44). Thus rationale combination of agents with mTOR inhibitors may result in improved responses in these difficult to treat patients. mTOR signaling has also been implicated in ER stress pathway (45). The present invention shows that ganetespib and sirolimus, two agents that work on separate and synergistic pathways, will not only be beneficial to MPNSTs, but other refractory bone and soft tissue sarcomas. mTOR and NF1

[0052] Signaling intermediates downstream of Ras are hyperactivated as a result of NF1 gene inactivation and these specific proteins are critical for transmitting the Ras growth signal and for the development of neoplasia in patients with NF1 (11). One of these downstream proteins is the mTOR molecule. Studies have

demonstrated that the NF1 tumor suppressor regulates mTOR pathway activation. mTOR was found to be activated in both NF1 deficient primary human and mouse cells as well as in human and genetically-engineered Nf 1 mouse tumor models. This aberrant activation was dependent on Ras and PI3 kinase/AKT signaling (64,65). In this regard, Nf 1 loss in mouse embryonic fibroblasts (65) and primary mouse astrocytes (64) was shown to result in Ras- and PI3K-dependent mTOR pathway activation, which could be inhibited with sirolimus. Moreover, Nf 1-/- astrocytes are highly sensitive to sirolimus treatments that have no effect on normal astrocyte growth (64). The increased proliferation associated with loss of neurofibromin expression in human MPNST cell lines was dramatically reduced by treatment with sirolimus (65). Sirolimus treatment of optic gliomas developing in a genetically- engineered Nfl mouse model resulted in attenuated mTOR signaling in vivo (58) as well as tumor growth in vivo (66). While tumors rapidly ceased to proliferate, there was no evidence of apoptosis or senescence, and sirolimus had no early effect on microvasculature in either preclinical model. mTOR and MPNST

[0053] A transgenic mouse model (67) carrying compound mutations in the Nfl and p53 tumor suppressors on the same chromosome develop aggressive MPNSTs that are histologically indistinguishable from human MPNSTs. Tumors grow with consistent and rapid kinetics, and on average mice only survive 10.7 days after the tumor is detected. This model was used to test the role of mTOR in tumorigenesis in vivo and assess the therapeutic utility of rapamycin (27). Animals with palpable tumors (approximately 300 mm³) were injected LP. with 5 mg/kg rapamycin per day. Control mice died on average in 12.2 days, and tumors grew 9.7-fold. In contrast, rapamycin potently suppressed MPNST growth, and allowed the animals to survive. Inhibition of S6 phosphorylation was observed in tumor and non-tumor tissue, demonstrating that rapamycin was effectively suppressing the mTOR pathway in vivo. Moreover rapamycin mediated its anti-tumor effects within 24 hours by potently suppressing proliferation, as assessed by BrDU incorporation in control and rapamycin treated tumors. Consistent with in vitro observations, apoptotic and senescent cells were not detected. As such, tumor growth was dependent on continued exposure to rapamycin, as tumors re-exhibited S6 phosphorylation and resumed growing at a rate comparable to control treated tumors following

rapamycin removal. Although the response to rapamycin was potent, effects were cytostatic. This model has been used to develop more effective mTOR-inhibitor based combination therapies (30). Hsp90 and mTOR combination

[0054] Tumor cells often exhibit specific stress-related phenotypes caused by insults such as excessive DNA damage as well as replicative, metabolic, and proteotoxic stress (68). Agents that further enhance or sensitize cancer cells to these stresses could thus be developed as potential anti-cancer therapies (69,70).

Proteotoxic or endoplasmic reticulum (ER) stress is induced when unfolded proteins accumulate in the ER (28). Cancer cells frequently exhibit high levels of ER stress caused by a multitude of factors (69,70). Oncogenic RAS also causes ER stress (29). Aneuploidy in particular has been shown to induce proteotoxic stress in cells (71). ER stress activates a signal transduction pathway called the unfolded protein response (UPR) (28). The UPR is an initially protective mechanism to reduce protein accumulation, however, when ER stress levels become insurmountable, cell death ensues (28). Hsp90 maintains protein homeostasis by folding newly synthesized and misfolded proteins, assembling and dissembling protein complexes and resolving protein aggregates (47). Hsp90 also directly stabilizes two key stress sensing components of UPR: IREl and pPERK/PERK 72. Therefore Hsp90 inhibitors would expect to promote ER stress in cancer cells directly by impairing global protein folding in already compromised tumor cells and inactivating subsequent adaptive responses of UPR.

[0055] MPNSTs are highly aneuploid and are driven by constitutive activation of Ras. They were demonstrated to have much higher levels of ER stress when compared to normal peripheral nerve as confirmed by three markers of UPR activation (BiP upregulation, eukaryotic translational initiation factor 2a (eIF2a) phosphorylation, and accumulation of the spliced active form of XBP-1 (sXBP-1). In addition, agents that induced ER stress triggered cell death at concentrations that did not affect the viability of normal cells. [0056] To date, no previous targeted agents have been shown capable of causing tumor regression in the highly aggressive genetically engineered MPNST model or in human tumors (see Table 1).

[0057] Table 1. Completed and select ongoing clinical trials with targeted for refractory MPNST.

[0058] Pre-clinical data presented demonstrate strong scientific rationale to study this combination in a clinical trial. Ganetespib is a potent, next generation Hsp90 inhibitor that has shown superior activity to first generation agents in preclinical studies. It has a favorable safety profile and promising anti-tumor activity over a broad range of tumor types in early clinical trials. Sirolimus is commercially available, oral, and relatively inexpensive. It has a long safety record, demonstrated efficacy in preclinical cancer models, and was used in the transgenic MPNST mouse model. The present application proposes a multi-institutional open label phase I/II trial of ganetespib in combination with sirolimus in patients with refractory MPNST, and other sarcoma types. Pre-clinical work with this combination of agents in other sarcomas is currently underway, which will aid in the further development of this combination for other sarcomas. The results of the trial described herein will provide valuable toxicity, tolerability, and pharmacokinetic information for a drug combination with potential uses other Ras driven tumors. Importantly, if the combination proves as effective as hypothesized in MPNSTs, it will provide a therapeutic strategy for this highly refractory and aggressive malignancy.

Compositions

[0059] The present invention features in certain aspects, compositions comprising an Hsp90 inhibitor and an mTOR inhibitor. It is a surprising finding of the present invention that the combination of a Hsp90 inhibitor and an mTOR inhibitor show synergistic activity.

[0060] In one embodiment, the Hsp90 inhibitor is ganetespib (5-[2,4-dihydroxy-5-

(l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lH-indol-5-yl)-3H-l,2,4-triazole-3- one).

[0061] Ganetespib (formerly called STA-9090) is a novel, injectable resorcinolic triazolone small molecule inhibitor of Hsp90 (46). Hsp90 is a molecular chaperone that regulates posttranslational folding, stability, and function of its client proteins, many of which play critical roles in cell growth, differentiation, and survival (47,48). Ganetespib is shown as Formula (I) below:

[0062] When a disclosed compound is named or depicted by structure, it is to be understood that solvates (e.g., hydrates) of the compound or a pharmaceutically acceptable salt thereof is also included. "Solvates" refer to crystalline forms wherein solvent molecules are incorporated into the crystal lattice during crystallization.

[0063] Solvates may include water or nonaqueous solvents such as ethanol, isopropanol, DMSO, acetic acid, ethanolamine and ethyl acetate. When water is the solvent molecule incorporated into the crystal lattice of a solvate, it is typically referred to as a "hydrate". Hydrates include stoichiometric hydrates as well as compositions containing variable amounts of water.

[0064] Ganetespib inhibits Hsp90 chaperone activity by binding to its N-terminal adenosine triphosphate (ATP) pocket. Hsp90 inhibition causes its client proteins to adopt aberrant conformations, which are then targeted for ubiquination and degradation by the proteosome (49). Hsp90 client proteins include wild type and mutated forms of many important signaling proteins associated with cancer, such as BCR-ABL, BRAF, CDK4, KIT, c-MET, c-SRC, EGFR, LCK, HER2 and VEGFR.

Ganetespib is currently being investigated in adults with a broad range of tumor types. It is being explored in both once weekly and twice weekly schedule, and has been generally well tolerated. Most frequent adverse events were gastrointestinal or constitutional in nature, and were generally mild to moderate in severity.

Preliminary signals of anti-tumor activity have been observed on the ongoing clinical trials in a broad range of tumor types.

[0065] In another embodiment, the mTOR inhibitor is sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)-

9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27-dihydroxy-3- [(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21- dimethoxy-6,8, 12, 14,20,26-hexamethyl-23,27-epoxy-3H-pyrido [2, 1-c] [1,4]

oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone. [0066] Sirolimus (rapamycin) is a mammalian target of rapamycin (mTOR) kinase inhibitor that has been FDA approved for immunosuppression following kidney transplantation (52). mTOR is a serine/threonine kinase, which belongs to

phosphatidylinositol-3 kinase (PI3K) related kinases (PIKKs) family.

[0067] Sirolimus inhibited tumor growth in preclinical models by inducing cell cycle arrest and apoptosis, leading to recognition of the mTOR pathway as a target for cancer therapy (53,54). Rapamycin analogs, such as temsirolimus (55), an intravenous soluble ester (pro drug) of sirolimus and everolimus (56), and oral mTOR inhibitor, have been FDA approved for the treatment of advanced renal cell carcinoma. At this time, it is unclear whether one compound will have advantage over the others in a particular tumor type.

[0068] In one embodiment, the amount of the Hsp90 inhibitor is between 100-500 mg/m², between 100-200 mg/m², and preferably between 150-200 mg/m².

[0069] In another embodiment, the amount of mTOR inhibitor is between 2-12 mg, preferably between 2-4 mg.

[0070] It is a surprising and unexpected finding of the present invention that drugs that target Hsp90 and mTOR show synergistic activity.

Methods of Treatment

[0071] The present invention relates to methods of treating or preventing a sarcoma in a subject, comprising administering a therapeutically effective amount of an Hsp90 inhibitor in combination with an mTOR inhibitor, wherein the subject is selected as being of 16 years of age or less.

[0072] The invention also relates to methods of reducing or inhibiting the progression of a sarcoma in a subject, comprising administering a therapeutically effective amount of an Hsp90 inhibitor in combination with an mTOR inhibitor, wherein the subject is selected as being of 16 years of age or less. [0073] In certain embodiments, the Hsp90 inhibitor is ganetespib (5-[2,4- dihydroxy-5-(l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lii-indol-5-yl)-3ii- l,2,4-triazole-3-one). In other embodiments, the mTOR inhibitor is sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)- 9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27-dihydroxy-3- [(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21- dimethoxy-6,8, 12, 14,20,26-hexamethyl-23,27-epoxy-3H-pyrido [2, 1-c] [1,4]

oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone.

[0074] Sarcomas were described supra. In embodiments of the invention, the sarcoma is a refractory sarcoma. The refractory sarcoma may be a bone or soft tissue sarcoma. In certain exemplary embodiments, the sarcoma is a malignant peripheral nerve sheath tumor (MPNST). The MPNST may be an unresectable or metastatic sporadic or neurofibromatosis type-1 associated MPNST.

[0075] The sarcoma may be a sarcoma that was previously treated and not responsive, or the sarcoma may be a recurrent sarcoma.

[0076] Thus, in various aspects, the invention includes methods of administering the Hsp90 inhibitor and mTOR inhibitor disclosed herein for therapeutic purposes for any sarcoma. As such, the invention provides methods of treating an individual afflicted with a sarcoma.

[0077] The compositions described herein are formulated to be compatible with its intended route of administration. Examples of routes of administration include parenteral, e.g., intravenous, intradermal, subcutaneous, oral, intranasal {e.g., inhalation), transdermal (topical), transmucosal, and rectal.

[0078] Effective dosages and schedules for administering the compositions of the invention may be determined empirically, and making such determinations is within the skill in the art. The dosage ranges for the administration of the compositions are those large enough to produce the desired effect in which the symptoms/disorder are/is affected. The dosage should not be so large as to cause adverse side effects, such as unwanted cross-reactions, anaphylactic reactions, and the like. Generally, the dosage will vary with the age, condition, sex and extent of the disease in the patient, route of administration, or whether other drugs are included in the regimen, and can be determined by one of skill in the art. The dosage can be adjusted by the individual physician in the event of any counterindications. Dosage can vary, and can be administered in one or more dose administrations daily, for one or several days. Guidance can be found in the literature for appropriate dosages for given classes of pharmaceutical products. For example, guidance in selecting appropriate doses for antibodies can be found in the literature on therapeutic uses of antibodies, e.g., Handbook of Monoclonal Antibodies, Ferrone et al., eds., Noges Publications, Park Ridge, N.J., (1985) ch. 22 and pp. 303-357; Smith et al., Antibodies in Human

Diagnosis and Therapy, Haber et al., eds., Raven Press, New York (1977) pp. 365-389.

[0079] According to certain embodiments, the amount of the Hsp90 inhibitor administered is from 100-200 mg/m². In further embodiments, the amount of the Hsp90 inhibitor administered is from 150-200 mg/m². The Hsp90 is preferably administered intravenously, over a period of 60 minutes.

[0080] In other embodiments, the amount of mTOR inhibitor administered is 2-12 mg. In further embodiments, the amount of mTOR inhibitor administered is 2-4 mg. Preferably ,the mTOR inhibitor is administered orally.

[0081] Preferably, the Hsp90 inhibitor is administered on days 1, 8 and 15. The mTOR inhibitor is preferably administered on days 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27 and 28.

[0082] The Hsp90 inhibitor and mTOR inhibitor may be administered over a cycle of days, for example a cycle of 28 days. The 28 day cycle may be repeated one or more times. [0083] The therapeutic agents of the combination therapies described herein can be administered sequentially or concurrently. In one embodiment, the

administration of the Hsp90 inhibitor and the mTOR inhibitor are done

concurrently. In another embodiment, the administration of the Hsp90 inhibitor and the mTOR inhibitor are done separately. In another embodiment, the administration of the Hsp90 inhibitor and the mTOR inhibitor are done sequentially. In one embodiment, the administration of the Hsp90 inhibitor and the mTOR inhibitor are done until the sarcoma is cured or stabilized or improved.

[0084] Different dosing regimens may be used as appropriate.

[0085] Following administration of the Hsp90 and mTOR inhibitors for treating or preventing a sarcoma, the efficacy of the therapeutic peptide can be assessed in various ways well known to the skilled practitioner.

[0086] The compositions disclosed herein may be also administered

prophylactically to patients or subjects who are at risk for a sarcoma.

Kits and/or Pharmaceutical Packages

[0087] The present invention also contemplates kits and pharmaceutical packages that are drawn to reagents or components that can be used in practicing the methods disclosed herein. The kits can include any material or combination of materials discussed herein or that would be understood to be required or beneficial in the practice of the disclosed methods. For example, the kits could include a peptide of the invention, or one or more additional active agents. In addition, a kit can include a set of instructions for using the components of the kit for its therapeutic and/or diagnostic purposes.

[0088] Throughout this application, various publications are referenced. The disclosures of these publications in their entireties are hereby incorporated by reference into this application in order to more fully describe the state of the art to which this invention pertains. The references disclosed are also individually and specifically incorporated by reference herein for the material contained in them that is discussed in the sentence in which the reference is relied upon.

[0089] It will be apparent to those skilled in the art that various modifications and variations can be made in the present invention without departing from the scope or spirit of the invention. Other embodiments of the invention will be apparent to those skilled in the art from consideration of the specification and practice of the invention disclosed herein. It is intended that the specification and examples be considered as exemplary only, with a true scope and spirit of the invention being indicated by the following claims.

[0090] The following examples further demonstrate several embodiments of this invention. While the examples illustrate the invention, they are not intended to limit it.

EXAMPLES

Example 1-- A Phase I/II Trial of ganetespib in combination with the mTOR inhibitor sirolimus for pediatric patients with unresectable or metastatic malignant peripheral nerve sheath tumors (MPNSTs)

[0091] Previously, no targeted agents have been able to cause tumor regression in a genetically engineered MPNST mouse model or human MPNST. Ganetespib is a novel, injectable, small molecule inhibitor of Hsp90 and is currently being

investigated in adults with a broad range of tumor types with a favorable safety profile and promising early results. Ganetespib has been studied in preclinical in vivo models with a variety of targeted agents with no marked apparent

pharmacological interactions. Sirolimus is a commercially available orally administered mTOR inhibitor and is the active metabolite of temsirolimus, which is FDA approved agent for advanced metastatic renal cell carcinoma. Sirolimus has been studied and tolerated in combination with multiple cytotoxic and targeted agents in a variety of tumor types.

[0092] It is a surprising and unexpected finding of the present invention that drugs that target Hsp90 and mTOR show synergistic activity in treating sarcomas, and in particular MPNSTs. The examples described herein are meant to demonstrate that ganetespib in combination with sirolimus will cause tumor regression in patients, and in a particular in a specific sub-set of patients identified as being 16 years of age or less, with refractory MPNSTs.

Objectives

Patient Population

[0093] The design and protocol described herein will be applied to a pediatric patient population. As used herein, a pediatric patient population is meant to refer to a particular sub -population of subjects less than 16 years of age.

Primary Objective

[0094] Phase I: To assess the safety, tolerability, and maximum tolerated dose (MTD)/ recommended dose of ganetespib when administered in combination with sirolimus in patients aged 16 years of less with refractory or relapsed sarcomas including unresectable or metastatic sporadic or neurofibromatosis type 1 (NF1) associated MPNST.

[0095] Phase II: To determine the clinical benefit of ganetespib in combination with sirolimus for patients aged 16 years or less with unresectable or metastatic sporadic or NF1 associated MPNST.

Secondary Objectives

[0096] Phase I: To describe the plasma pharmacokinetic profile of ganetespib and sirolimus when administered in combination therapy [0097] Phase I/II: To determine changes in pharmacodynamic parameters including phospho-S6, phosphorylated eIF2 alpha, Akt Phosphorylation, Hsp70, and G6PD in tumor tissue and peripheral blood mononuclear cells at baseline and during treatment and correlate with changes in clinical or radiologic outcome.

[0098] Phase I/II: To assess patient-reported pain severity and the impact of pain on daily activities before and during treatment with ganetespib and sirolimus and to correlate with changes in clinical or radiologic outcome.

[0099] Phase I/II: To evaluate the utility of three-dimensional MRI (3D-MRI) analysis in comparison to 1-dimensional and 2-dimensional measurements as a method to more sensitively monitor response.

TRIAL DESIGN

[00100] The study will be a multi-institutional open label phase I/II trial of ganetespib in combination with sirolimus in patients with refractory sarcoma including MPNST. The primary objective of the initial phase I component is to evaluate the safety and tolerability of this combination, and to determine the maximum tolerated dose (MTD)/recommend dose for this combination.

[00101] As discussed supra, the present trials are designed for a particular subset of patients aged 16 years or less, with sarcomas, in particular refractory sarcoma including MPNSTs.

[00102] The phase I component will be open to the identified patient population with refractory sarcomas. Secondary objectives of this portion will be to describe the plasma pharmacokinetic profile of ganetespib and sirolimus when administered in this combination therapy. The starting dose and escalation schema are as described below. Dose Ganetespib Sirolimus (mg)

Level (mg/m ) Loading dose Maintenance

IV days 1, 8, 15 CI Dl only PO once daily

every 28 days continuous

-2 100 6 2

-1 150 6 2

1* 150 12 4

2 200 12 4

* Starting dose

Each cycle is considered 28 days

[00103] The recommended phase II dose of weekly single agent ganetespib is 200 mg/m²/dose IV over one hour x 3 weeks; 1 week off. The starting dose of Ganetespib selected is a 25% dose decrease and used previously in the docetaxel combination trials (49). The starting dose for sirolimus was selected using similar doses derived from earlier phase I single and combination studies (59, 61, 62) with sirolimus trough levels ranging from 5-10 ng/mL. There will be no plans to exceed the recommended doses of either agent (Dose level 2). Three to six patients will be entered per dose level using standard 3+3 design. Patients will be closely evaluated for the

development of toxicity. Toxicities observed during the first cycle will be used to define the MTD/Recommended dose.

[00104] Once the doses are determined in the phase I portion, the phase II component will open. The primary objective of the phase II portion will be to determine the clinical benefit rate (CR, PR, or stable disease > 4 months using WHO criteria) of ganetespib in combination with sirolimus for patients with refractory sporadic or NF1 associated MPNST. Additional sarcoma strata may be considered directed by preclinical findings and rationale. We propose an open label Simon's optimal two-stage design with a target benefit rate of 25%. Ten patients will be enrolled on the first stage, with no further accrual if no response (CR, PR, or stable disease > 4 months) is observed. If at least one response is observe, accrual will continue until a total of 20 patients have been enrolled. If > 3 of 20 evaluable patients responds, this regimen will be considered active and could provide a therapeutic upfront strategy for this malignancy. Sites of measureable disease will be evaluated within 4 weeks prior to starting therapy and then every 2 cycles. Patients will be able to remain on treatment as long as they do not experience progressive disease or unacceptable toxicity up to a maximum of 13 cycles (1 year). Response will be determined by WHO criteria. MPNSTs are typically complex, non-spherical tumors and 2-D may thus reflect changes in tumor size better than 1-D (RECIST). In addition, previous and ongoing trials of refractory phase II MPNST trials (erlotinib and bevacizumab/everolimus) have used WHO criteria and will allow for direct comparisons. Secondary objectives will be to determine changes in

pharmacodynamic parameters including phospho-S6, AKT phosphorylation, pEIF2cc, Hsp70, and G6PD in tumor tissues from diagnosis/recurrence, and on treatment if feasible. These markers will also be evaluated in peripheral blood mononuclear cells at baseline and during treatment. Changes in patient reported pain and impact of pain on daily activities will also be assessed before and during treatment.

[00105] Eligibility Criteria

[00106] Patients must have baseline evaluations performed prior to the first dose of study drug and must meet all inclusion / exclusion criteria. Results of all baseline evaluations, which assure that all inclusion and exclusion criteria have been satisfied must be reviewed by the Principal Investigator or his/her designee prior to enrollment of the patient. In addition, the patient must be thoroughly informed about all aspects of the study, including the study visit schedule and required evaluations and all regulatory requirements for informed consent. The written informed consent must be obtained from the patient prior to enrollment. The following criteria apply to all patients enrolled onto the study unless otherwise specified. Inclusion Criteria

[00107] Age 16 years or less.

[00108] Phase I: Patients with unresectable, recurrent, or metastatic histologically confirmed soft tissue or bone sarcoma of one of the following subtypes:

Leiomyosarcoma*

Liposarcoma*

MFH/pleomorphic undifferentiated sarcoma* Rhabdomyosarcoma*

Malignant peripheral nerve sheath tumor (MPNST)*

Osteosarcoma*

Ewings*

Alveolar soft part sarcoma Malignant giant cell tumor of bone Desmoplastic small round blue cell tumor Synovial Sarcoma Undifferentiated sarcoma

[00109] *Patients with leiomyosarcoma, liposarcoma, osteosarcoma, Ewings, MPNST, rhabdomyosarcoma or MFH must have documentation that they have received, not been eligible for, or refused at least one prior chemotherapy regimen prior to enrollment.

[00110] Phase II: Patients with unresectable or metastatic histologically confirmed sporadic or NF1 associated high grade MPNST who have experienced progression after one or more prior regimens of cytotoxic chemotherapy. Patients who have refused cytotoxic chemotherapy or for whom treatment on this protocol prior to receiving cytotoxic chemotherapy is felt to be in the best interest for the patient by the local investigator will also be eligible.

[00111] Diagnostic criteria for NF1 are (NIH Consensus Conference 1987):

Presence of 2 or more of the following criteria:

[00112] Six or more cafe-au-lait spots (>0.5 cm in prepubertal subjects or >1.5 cm in postpubertal subjects)

[00113] > 2 neurofibromas or 1 plexiform neurofibroma

[00114] 3. Freckling in the axilla or groin

[00115] 4. Optic glioma

[00116] 5. Two or more Lisch nodules

[00117] 6. A distinctive bony lesion (dysplasia of the sphenoid bone or dysplasia or thinning of long bone cortex)

[00118] 7. A first degree relative with NF1

[00119] ECOG Performance Status of < 2.

[00120] Must be able to swallow whole pills.

[00121] Patients must have measurable disease, defined as at least one tumor that is measurable (defined as those that can be accurately measured in at least two dimensions (longest diameter > 20 mm with conventional techniques or > 10 mm using spiral CT scan) in two dimensions on CT or MRI scan. Baseline radiologic scans must be performed within 4 weeks of starting treatment.

[00122] Adequate organ function within 2 weeks of Day 1 of study defined as:

[00123] Adequate hematologic function as shown by: ANC > 1.0 x 10⁹/L, Platelets > 75,000 x 10⁹/L, Hgb > 9 g/dL (transfusion of packed red blood cells allowed).

[00124] Adequate liver function as shown by: serum bilirubin < 1.5 x ULN, ALT and AST < 3.0 x ULN (< 5x ULN in patients with liver metastases). [00125] Fasting serum cholesterol < 300 mg/dL OR < 7.75 mmol/L AND fasting triglycerides < 300 mg/dL OR < 3.42 mmol/L. NOTE: In the case one or both of these thresholds are exceeded, the patient can only be included after initiation of appropriate lipid lowering medication.

[00126] Serum creatinine < ULN or creatinine clearance > 60 ml/min/1.73 m² [00127] Adequate Cardiac Function defined as:

[00128] No history of congenital prolonged QTc syndrome, NYHA Class III or IV congestive heart failure (CHF)

[00129] No clinically significant cardiac arrhythmias, stroke or myocardial infarction within 6 months prior to enrollment

[00130] Second or third degree atrioventricular block unless treated with a permanent pacemaker

[00131] Complete left bundle branch block

[00132] QTcF < 480 ms. Note: Patients with Grade 1 prolonged QTcF (450-480 msec) at the time of study enrollment should have correctable causes of prolonged QTc addressed if possible (i.e. electrolytes, medications).

[00133] Use of medications that have been linked to the occurrence of torsades de pointes within the last 7 days.

[00134] Prior therapy: Patients must have fully recovered from the acute toxic effects of all prior anti-cancer therapy. Recovery is defined as a toxicity < grade 2 (CTCAE v 4.0), unless otherwise specified in the inclusion/exclusion criteria.

[00135] Myelosuppressive chemotherapy: Patients must have not received myelosuppressive chemotherapy within 3 weeks of enrollment onto this study.

[00136] Biologic agent: At least 7 days after the last dose of biologic agent. For agents that have known adverse events occurring beyond 7 days after administration, this period must be extended beyond the time which adverse events are known to occur.

[00137] Immunotherapy: At least 42 days after the completion of any type of immunotherapy, e.g. tumor vaccines.

[00138] Monoclonal antibodies: Patients may not have received monoclonal antibodies within 3 weeks of enrollment onto this study.

[00139] Radiation: At least 14 days after local palliative XRT (small port); or at least 4 weeks otherwise.

[00140] Stem Cell Transplant: No evidence of graft versus host disease, and > 2 months must have elapsed since transplant.

[00141] Hematopoietic growth factors: At least 7 days must have elapsed since completion of therapy with a growth factor. At least 14 days must have elapsed after receiving pegfilgrastim.

[00142] Fertile men and women of childbearing potential must agree to use an effective method of birth control from Day 1 of study and for 120 days after last study drug administration in both sexes. Effective methods of birth control includes: surgically sterile, barrier device (condom, diaphragm), contraceptive coil, abstinence, or oral contraception.

Exclusion Criteria

[00143] Patients currently receiving other anti-cancer agents are not eligible.

[00144] Patients receiving chronic, systemic treatment with corticosteroids or another immunosuppressive agent (for example, cyclosporine). Topical or inhaled corticosteroids are allowed.

[00145] Patients should not receive immunization with attenuated live vaccines within one week of study entry or during study period. [00146] Uncontrolled brain or leptomeningeal metastases, including patients who continue to require glucocorticoids for brain or leptomeningeal metastases.

[00147] Other malignancies within the past 3 years except for adequately treated carcinoma of the cervix or basal or squamous cell carcinomas of the skin. Stable NF1 related tumors, such as optic pathway tumors, which do not require treatment at time of study enrollment, will not be considered an exclusion criterion.

[00148] Patients who have any known severe and/or uncontrolled medical conditions or other conditions that could affect their participation in the study such as:

[00149] Severely impaired lung function defined as spirometry and DLCO that is 50% of the normal predicted value corrected for hemoglobin and alveolar volume and/or O2 saturation that is 88% or less at rest on room air. For patients who do not have respiratory symptoms (e.g. dyspnea at rest, known requirement for

supplemental oxygen), pulmonary function tests are not required.

[00150] Significant vascular disease (e.g. aortic aneurysm, symptomatic peripheral vascular disease) within 6 months prior to enrollment

[00151] Uncontrolled diabetes as defined by fasting serum glucose >1.5 x ULN

[00152] Active (acute or chronic) or uncontrolled severe infectious hepatitis.

[00153] Note: A detailed assessment of Hepatitis B/C medical history and risk factors must be done at screening for all patients. HBV DNA and HCV RNA PCR testing are required at screening for all patients with a positive medical history based on risk factors and/or confirmation of prior HBV/HCV infection.

[00154] A known history of HIV seropositivity, as immune deficiency increases the risk for opportunistic infection.

[00155] Impairment of gastrointestinal function or gastrointestinal disease that may significantly alter the absorption of sirolimus (e.g., ulcerative disease, uncontrolled nausea, vomiting, diarrhea, malabsorption syndrome or small bowel resection).

[00156] Patients with an active, bleeding diathesis or significant coagulopathy (in absence of therapeutic anticoagulation).

[00157] Female patients who are pregnant or breast feeding, or adults of reproductive potential who are not using effective birth control methods. (Women of childbearing potential must have a negative urine or serum pregnancy test within 7 days prior to administration of study drugs)

[00158] Patients with a known hypersensitivity to rapamycins (sirolimus, temsirolimus, everolimus) or to its excipients. Excipients: Tablets:

butylhyroxytoluene/butylated hydroxytoluene (BHT), magnesium stearate, lactose monohydrate, hypromellose/hydroxypropyl methylcellulose, crospovidone, lactose anhydrous. The excipients comply with the requirements of the applicable

compendial monographs (Ph. Eur., USP/NF).

[00159] Patients unwilling or unable to comply with the protocol.

[00160] CYP3A4/CYP2C19 substrates: Patients should not have received these medications within 1 week of entry and is not allowed while on study.

[00161] Seville orange, star fruit, grapefruit and their juices, and St. John's Wort use are not allowed while on study.

[00162] Enzyme inducing anticonvulsants: Patients may not be taking enzyme - inducing anticonvulsants, and may not have received these medications within 1 week of entry, as these patients may experience different drug disposition. These medications include:

Carbamazepine (Tegretol)

Felbamate (Felbtol)

Phenobarbitol Phenytoin (Dilantin) Primidone (Mysoline) Oxcarbazepine (Trileptal)

Inclusion of Women and Minorities

[00163] Subjects of both genders and from all racial and ethnic groups are eligible for this trial if they meet the criteria. To date, there is no information that suggests differences in absorption, metabolism, or disposition or disease response among racial or ethnic groups or between the genders, indicating that results of the trial will be applicable to all groups. Efforts will be made to extend the accrual to a

representative population, but in a phase I/II trial which will accrue a limited number of patients, a balance must be struck between patient safety considerations and limitations on the number of individuals exposed to potentially toxic or ineffective treatments on the one hand and the need to explore gender, racial, and ethnic aspects of clinical research on the other. If differences in outcome that correlate to gender, racial, or ethnic identity are noted, accrual may be expanded or additional studies may be performed to investigate those differences more fully

TREATMENT PLAN Agent Administration

[00164] Ganetespib is commercially available. Ganetespib will be diluted to appropriate dose in D5W prior to administration. Ganetespib will be given at hour 0 on days 1, 8, and 15 intravenously over 1 hour. Loperamide 2 mg should be given 1 to 2 hours prior to ganetespib administration and then every 4 hours for 12 hours post infusion to all patients as prophylactic diarrhea management. *A11 efforts should be made to adhere to this schedule. However, to provide some flexibility to changes in patients' schedules and holidays for long-term treatment, +/- 1 day in changes will not be considered a protocol deviation and will not require reporting to the NCI, IRB, or FDA.

[00165] Sirolimus will be purchased commercially (2 mg tablets) and administered once daily. On cycle 1, on day 1 a loading dose of sirolimus will be given orally x 1. Subsequent doses of sirolimus will be given once daily continuously with no breaks in between cycles. On days when sirolimus and ganetespib are given together, they should begin at hour 0 together. Sirolimus should be taken at approximately the same time every day, preferably in the morning, consistently with or without food. If a patient misses a dose, the dose may be taken within 6 hours of missed dose. Otherwise, patient must wait until the next day's dose. If a patient vomits after a dose of sirolimus it will not be repeated. All doses prescribed and dispensed to the patient and all dose changes during the study will be recorded in a patient diary.

[00166] A cycle of therapy is considered to be 28 days.

[00167] Ganetespib drug doses should be adjusted based on BSA determined prior to the beginning of each cycle.

Criteria for starting subsequent cycles

[00168] A cycle may be repeated every 28 days if the patient has at least stable disease, has not experienced dose-limiting toxicity, and has recovered from the prior cycle as evidenced by return to baseline eligibility. A cycle can be extended by an additional 14 days (to day 42) to allow for recovery without modifying the dose of the drug on subsequent cycles if toxicity did not meet dose limiting criteria. Patient must not met one of the off protocol therapy or off study criteria defined in section 5.8 and 5.9. A cycle may be administered a maximum of 13 times.

Dose Escalation Schema [Phase I component]

Inter-patient Escalation Table 2.

Criteria for Dose Escalation

[00169] Cohorts of 3 to 6 patients will be treated with drug combination at each dose level. When a minimum of three patients who are evaluable for toxicity have completed one cycle of therapy at a dose level without evidence of dose-limiting toxicity (DLT), subsequent patients may be enrolled at the next higher dose level.

[00170] If DLT is observed in 1 patient from the initial cohort of 3 patients at a given dose level, an additional 3 patients will be entered at that dose level. If none of these additional patients experiences a DLT (1/6 with DLT), the dose will be escalated. If > 1 of the additional patients experience a DLT (> 2/6 with DLT), the MTD has been exceeded, and the next lower dose level will be considered the MTD.

[00171] If the MTD has been exceeded at the starting dose level, then the

subsequent cohort of patients will be de-escalated per schema above. Definition of Dose Limiting Toxicity (DLT) (Criteria for entire study)

[00172] Toxicity will be graded using the NCI Common Terminology Criteria for Adverse Events (CTCAE) version 4.0. A copy of the CTCAE version 4.0 can be downloaded from the CTEP website (http://ctep.cancer.gov). Any suspected or confirmed dose-limiting toxicity should be reported within 24 hours to the overall study Principal Investigator.

[00173] DLT will be defined as any of the following events that are possibly, probably, or definitely attributable to ganetespib or sirolimus. The DLT observation period for the purposes of dose escalation will be the first cycle of therapy. Dose limiting hematological and non-hematological toxicities are defined below:

Non-hematological DLT

Any Grade > 4 non-hematological toxicity

Any Grade 3 non-hematological toxicity with the specific exclusion of Grade 3 nausea and vomiting of < 3 day duration. Grade 3 Diarrhea < 3 days duration

Grade 3 ALT/ AST that returns to meet initial eligibility criteria within 7 days of study drug interruption and that do not recur upon study re-challenge.

Grade 3 fever or infection < 5 days duration.

Grade 3 electrolyte imbalances that respond to oral or intravenous supplementation.

Allergic reactions that necessitate discontinuation of study drug will not be considered dose-limiting.

Grade 2 non-hematological toxicity that persists for > 7 days and is considered sufficiently medically significant or intolerable by patients that it requires treatment interruption. Any adverse event requiring interruption of study drug for > 7 days or which recurs upon drug challenge.

Hematological DLT

Grade 4 Thrombocytopenia

Grade 4 Neutropenia

Grade 4 Anemia

Intra-Patient Escalation

[00174] Intra-patient dose escalation is not allowed.

Definition of Maximum Tolerated Dose (MTD)

[00175] The MTD is defined as the dose level immediately below the dose at which > 33% of patients in a cohort experience a DLT.

[00176] In order to escalate a dose level, < 33% of patients in a cohort should have a DLT.

[00177] At least 3 patients in a cohort must be evaluable for the definition of the MTD in order to escalate.

[00178] If 1 out of 3 patients in a cohort experience a DLT, then at least 6 patients must be enrolled and evaluable for the definition of MTD prior to dose escalation.

[00179] To determine extended tolerability, toxicities observed during the first treatment cycle will be used to define the MTD. A patient will be considered evaluable for definition of the MTD if at least 70% of the prescribed sirolimus dose has been administered to the patient during the first treatment cycle based on diary review and pill count of returned drug, and patient must have received all 3 doses of ganetespib in the cycle. If a discrepancy occurs, pill count will be used for adherence measurement. If a patient has less than 70% adherence, the patient will be replaced in the cohort. In addition, anyone who receives one or more doses and experiences a DLT will be considered evaluable for definition of MTD. Dosing for Phase II component

[00180] Patients will be treated at the recommend dose from the Phase I component of this trial.

General Concomitant Medication and Supportive Care Guidelines

[00181] Patients must be instructed not to take any additional medications

(including over-the-counter products) during the trial without prior consultation with the investigator. All medications taken at the time of screening should be recorded. If concomitant therapy must be added or changed, the reason and name of the drug/therapy should be recorded.

[00182] In general, the use of any concomitant medication/therapies deemed necessary for the care of the patient are allowed, including drugs given

prophylactically (e.g. antiemetics), with the following exceptions:

[00183] No other investigational therapy should be given to patients

[00184] No chronic treatment with systemic steroids or another

immunosuppressive agent (for example, cyclosporine) with the exception of patients with endocrine deficiencies who are allowed to received physiologic or stress doses of steroids if necessary.

[00185] Certain CYP3A4/ CYP2C19 substrates land enzyme inducing

anticonvulsants are prohibited on this trial. Exercise caution when using sirolimus with other drugs or agents that are modulators of CYP3A4.

[00186] Use of medications that have been linked to the occurrence of torsades de pointes are prohibited.

[00187] Patients should not receive immunization with attenuated live vaccines during the study period. Close contact with those who have received attenuated live vaccines should be avoided. Examples of live vaccines include intranasal influenza, measles, mumps, rubella, oral polio, BCG, yellow fever, varicella and TY21a typhoid

[00188] Concomitant Cancer and other Therapy

[00189] Concurrent cancer therapy, including chemotherapy, radiation therapy, immunotherapy, or biologic therapy may not be administered.

Supportive Care

[00190] Appropriate antibiotics, blood products, antiemetics (except systemic steroids and aprepitant), fluids, electrolytes and general supportive care are to be used as necessary. Platelets should be transfused for thrombocytopenia following institutional guidelines. All blood products will be administered following institutional guidelines to prevent graft-versus-host disease. Corticosteroids are permissible as premedication for blood product transfusions, or as treatment for an acute allergic reaction.

[00191] Diarrhea management (refer to Operations Manual):

[00192] All patients will receive prophylactic loperamide 2 mg prior to and every 4 hours during the first 12 hours post infusion of ganetespib.

[00193] For patients with Grade 1 or 2 Diarrhea: Loperamide as an initial 4 mg dose followed by 2 mg doses every 4 hours (Do not exceed 16 mg in 24 hours).

Patient should continue until free of diarrhea for 12 hours.

[00194] For Grade 3 or 4 Diarrhea: Consider hospitalization with IVF if clinically indicated and antibiotics as appropriate.

[00195] Patients must take some form of PCP prophylaxis while on sirolimus.

For example: trimethoprim/sulfamethoxazole as per institutional guidelines, or inhaled pentamidine. [00196] Good oral hygiene and mouth care are encouraged, as mucositis is one of the toxicities of sirolimus.

[00197] Growth Factors that support platelet or white cell number or function can only be administered for culture proven bacteremia, clinical sepsis, or invasive fungal infection with neutropenia. ASCO guidelines and regulatory authority labeling for providing growth factor support are recommended.

[00198] Vaccinations: Patients receiving immunosuppressants, including sirolimus, should not be administered live vaccines. In addition, the response to vaccines (non-live) administered while the patient is immunosuppressed can be variable, and clinicians should check titers following for response if a non-live vaccine must be administered during this time.

[00199] Patients with positive hepatitis screen:

[00200] It is mandatory that patients positive for HBV-DNA or HBsAg are treated prophylactically with an antiviral for at least 1 week prior to receiving study drug.

[00201] The antiviral treatment should continue throughout the entire study period and for at least 4 weeks after the last dose of study drug.

[00202] Patients on antiviral prophylaxis treatment or positive HBV antibodies should be tested for HBV-DNA according to study visit schedule.

Management for Surgery

[00203] Patients undergoing minor surgery should hold study drugs for 2 weeks prior to procedure, if feasible, and for 2 weeks after. Patients will be considered evaluable for toxicity and/or response as long as they meet criteria as defined in Section 11.1.1.

Duration of Therapy

[00204] Treatment may continue for a maximum of 1 year (13 cycles) in the absence of disease progression or unacceptable adverse events. [00205] Duration of Follow Up

[00206] Patients will be followed until 30 days after the last dose of ganetespib and sirolimus or longer if the patient is removed from treatment with ganetespib and sirolimus for unacceptable adverse events. Adverse events will be followed until resolution or stabilization of the adverse event to < grade 1 using the CTCAE v 4 as detailed herein.

Criteria for Removal from Study Treatment [00207] Disease progression

[00208] Intercurrent illness that prevents further administration of treatment

[00209] Unacceptable adverse event(s), including significant irreversible grade 4 toxicity attributed to sirolimus or ganetespib

[00210] Patients with Grade 4 QTcF prolongation and torsades de pointes or polymorphic ventricular tachycardia or signs and symptoms of serious arrhythmia OR repeated grade 3 or higher QTcF prolongation must discontinue treatment with ganetespib

[00211] Greater than 3 weeks have elapsed since the last dose of sirolimus or greater than 8 weeks since the last dose of ganetespib

[00212] Patient decides to withdraw from the study

[00213] General or specific changes in the patient's condition render the patient unacceptable for further treatment in the judgment of the investigator

Off Study Criteria

[00214] Thirty days after the last dose of investigational agent

[00215] Death

[00216] Lost to follow-up

[00217] Withdrawal of consent for any further data submission [00218] Entry onto another therapeutic study

DOSING MODIFICATIONS/ Management for Specific Side Effects

[00219] Dosing changes for ganetespib related toxicities are described herein. If toxicity cannot be clearly attributed to either agent alone, the toxicity will be attributed to both agents, and modifications will be made to both agents. Should a patient require permanent discontinuation of either ganetespib or sirolimus, the patient can continue on study receiving the agent, which is tolerated for as long as no other off treatment criteria are met. The study Pis should be contacted to discuss questions regarding toxicity attribution and to discuss patients who may meet criteria to continue treatment with either agent alone.

Dose modifications for non-hematological toxicity

[00220] If a patient experiences a non-hematological dose limiting toxicity, the attributable treatment will be withheld. If the toxicity resolves to meet on study parameters within 14 days of drug discontinuation, the patient may resume treatment at a reduced dose. Doses reduced for toxicity will not be re-escalated, even if there is minimal to no toxicity with the reduced dose.

[00221] If toxicity does not resolve to meet on study entry parameters within 14 days of drug discontinuation, the patient must discontinue the protocol therapy.

[00222] If dose-limiting toxicity recurs in a patient who has resumed treatment at a reduced dose, the patient must discontinue the protocol therapy.

[00223] Patients with reported Grade 3 severity at any of the specified ECG time points (QTcF > 500ms), ganetespib should be withheld. Ganetespib may be restarted at dose reduction (Section 6.3) when QTcF is < Grade 1. While the QTcF

prolongation is Grade 3 or higher, patients should have additional ECG monitoring until QTcF prolongation returns to < Grade 1. Dose modifications for hematological toxicity

[00224] If a patient experiences hematological dose-limiting toxicity, the

attributable treatment will be withheld. Counts should be checked twice weekly during this time. When toxicity resolves to meet study parameters within 14 days of drug discontinuation, the patient may resume treatment at the reduced Doses reduced for toxicity will not be re-escalated, even if there is minimal to no toxicity with the reduced dose.

[00225] If toxicity does not resolve to meet on study entry parameters within 14 days of drug discontinuation, the patient must discontinue protocol therapy.

[00226] If dose-limiting toxicity recurs in a patient who has resumed treatment at a reduced dose, the patient must protocol therapy.

Ganetespib dose level modifications

[00227] A single dose reduction of 30% is allowed for toxicities outlined supra. Sirolimus dose level modification

[00228] A single dose reduction of 50% is allowed for toxicities outlined supra.

Management of Specific Side Effects

Management of hypersensitivity reactions to ganetespib infusions

[00229] If an infusion hypersensitivity reaction to ganetespib is suspected, the following management is provided as guidance.

[00230] Mild or moderate symptoms:

[00231] Stop ganetespib infusion

[00232] Administer IV dexamethasone and diphenhydramine HC1 or therapeutic equivalent

[00233] After recovery from symptoms, resume ganetespib infusion at reduced rate [00234] In subsequent cycle, consider premedication with steroids and diphenhydramine HC1

[00235] Severe Symptoms (such as hypotension, angioedema, respiratory distress, or generalized uticaria):

[00236] Stop ganetespib infusion

[00237] Administer IV dexamethasone, diphenhydramine HC1 or therapeutic equivalent

[00238] Consider epinephrine and/or bronchodialators as indicated

[00239] In subsequent cycles, give prophylactic premedication regimen with steroids, diphenydramine HC1 and reduce rate of ganetespib

[00240] The following is an example of infusion premedication regimen:

[00241] Dexamethasone 12 mg orally and diphenhydramine HC1 25-50 mg orally approximately 12 to 24 hours prior to next dose of study

[00242] Repeat dexamethasone 12 mg orally and diphenhydramine HC1 25-50 mg orally approximately 4-6 hours prior to the re-challenge.

[00243] If severe symptoms recur with optimal premedication, treatment with ganetespib must be discontinued.

Management of stomatitis/ oral mucositis/mouth ulcers

[00244] Stomatitis/oral mucositis/mouth ulcers due to sirolimus should be treated using local supportive care. Please note that investigators in earlier trials have described the oral toxicities associated with sirolimus as mouth ulcers, rather than mucositis or stomatitis. If your examination reveals mouth ulcers rather than a more general inflammation of the mouth, please classify the adverse event as such. Please follow the paradigm below for treatment of stomatitis/oral mucositis/mouth ulcers: [00245] For mild toxicity (Grade 1), use conservative measures such as nonalcoholic mouth wash or salt water (0.9%) mouth wash several times a day until resolution.

[00246] For more severe toxicity (Grade 2 in which case patients have pain but are able to maintain adequate oral alimentation, or Grade 3 in which case patients cannot maintain adequate oral alimentation), the suggested treatments are topical analgesic mouth treatments (i.e., local anesthetics such as benzocaine, butyl aminobenzoate, tetracaine hydrochloride, menthol, or phenol) with or without topical corticosteroids, such as triamcinolone oral paste 0.1% (Kenalog in Orabase®). For intolerable grade > 2 stomatitis/oral mucositis/mouth ulcers, sirolimus will be held, and restarted with a dose reduction by 1 dose level after recovery from toxicity to grade 1 or less.

[00247] Agents containing hydrogen peroxide, iodine, and thyme derivatives may tend to worsen mouth ulcers. It is preferable to avoid these agents.

[00248] Antifungal agents must be avoided unless a fungal infection is diagnosed. In particular, systemic imidazole antifungal agents (ketoconazole, fluconazole, itraconazole, etc.) should be avoided in all patients due to their strong inhibition of sirolimus metabolism, thereby leading to higher sirolimus exposures. Therefore, topical antifungal agents are preferred if an infection is diagnosed. Similarly, antiviral agents such as acyclovir should be avoided unless a viral infection is diagnosed.

[00249] Note: Stomatitis/oral mucositis should be appropriately graded using the functional grading given on the NCI-CTC for adverse events, version 4.

Management of hyperlipidemia and hyperglycemia

[00250] Treatment of hyperlipidemia should take into account the pre-treatment status and dietary habits. Blood tests to monitor hyperlipidemia must be taken in the fasting state. Grade 2 or greater hypercholesterolemia (> 300 mg/dL or 7.75 mmol/L) or Grade 2 or greater hypertriglyceridemia (> 300 mg/dL or > 3.42 mmol/L) should be treated with a 3-hydroxy-3-methyl-glutaryl (HMG)-CoA reductase inhibitor (e.g., atorvastatin, pravastatin) or appropriate lipid-lowering medication, in addition to diet. Patients should be monitored clinically and through serum biochemistry for the development of rhabdomyolysis and other adverse events as required in the product label/data sheets for HMG-CoA reductase inhibitors. Note: Concomitant therapy with fibrates and an HMG-CoA reductase inhibitor is associated with an increased risk of a rare but serious skeletal muscle toxicity manifested by rhabdomyolysis, markedly elevated creatinine kinase (CPK) levels and myoglobinuria, acute renal failure and sometimes death. The risk versus benefit of using this therapy should be determined for individual patients based on their risk of cardiovascular

complications of hyperlipidemia. Grade 4 hypercholesterolemia (> 500 mg/dL or > 12.92 mmol/L) hold sirolimus until cholesterol is < Grade 4. If triglycerides > 500- 1,000, and HDL is low, consider fibrate or niacin. For triglycerides > 1000 mg/dL or >11.4mmol/L, hold sirolimus while instituting fibrate or niacin therapy until triglycerides are < Grade 4.

[00251] Grade 3 hyperglycemia has been observed in patients receiving sirolimus therapy. In many cases the affected patients had an abnormal fasting glucose at baseline. Based on this finding, it is suggested that optimal glucose control should be achieved before starting a patient on sirolimus and should be monitored during sirolimus therapy.

Management of non-infectious pneumonitis

[00252] Non-infectious pneumonitis is a class effect of rapamycin derivatives. Cases of non-infectious pneumonitis (including interstitial lung disease) have also been described in patients taking sirolimus. Some of these have been severe and on rare occasions, a fatal outcome was observed.

[00253] A diagnosis of non-infectious pneumonitis should be considered in patients presenting with non-specific respiratory signs and symptoms such as hypoxia, pleural effusion, cough or dyspnea, and in whom infectious, neoplastic and other non-medicinal causes have been excluded by means of appropriate

investigations. Patients should be advised to report promptly any new or worsening respiratory symptoms.

[00254] Patients who develop radiological changes suggestive of non-infectious pneumonitis and have few or no symptoms may continue sirolimus therapy without dose alteration. Dose modifications and retreatment are described in Table 3 below.

[00255] Table 3. Management of non-infectious pneumonitis

Worst Grade Required Investigations Management of Sirolimus Dose Pneumonitis Pneumonitis Adjustment

Grade 1 CT scans with lung No specific therapy is Administer 100% of windows and pulmonary required sirolimus dose.

function testing including:

spirometry, DLCO, and

room air 02 saturation at

rest. Repeat chest x-ray/CT

scan every 2 Cycles until

return to baseline.

Grade 2 CT scan with lung Symptomatic only. Reduce sirolimus dose windows and pulmonary Prescribe to 50% lower dose than function testing including: corticosteroids if cough previously

spirometry, DLCO, and is troublesome. administered until room air 02 saturation at recovery to < Grade 1. rest. Repeat each Sirolimus may also be subsequent Cycle until interrupted if return to baseline. symptoms are

Consider bronchoscopy * troublesome. Patients will be withdrawn from protocol treatment if they fail to recover to < Grade 1 within 3 weeks.

Grade 3 CT scan with lung Prescribe Hold treatment until windows and pulmonary corticosteroids if recovery to < Grade 1. function testing including: infective origin is ruled May restart protocol spirometry, DLCO, and out. Taper as medically treatment within 2 room air 02 saturation at indicated. weeks at a reduced dose rest; Repeat each if evidence of clinical subsequent Cycle until benefit.

return to baseline. Patients will be Bronchoscopy is withdrawn from the recommended * study if they fail to recover to < Grade 1 within 2 weeks.

Grade 4 CT scan with lung Prescribe Discontinue treatment.

windows and required corticosteroids if

pulmonary function testing infective origin is ruled

includes: spirometry, out. Taper as medically

DLCO, and room air 02 indicated.

saturation at rest. Repeat

each subsequent Cycle

until return to baseline.

Bronchoscopy is

recommended *.

* A bronchoscopy with biopsy and/or bronchoalveolar lavage should be considered (grade 2) or is recommended (grade 3 or 4). For any grade infection should be ruled out prior to prescribing corticosteroids. PHARMACEUTICAL INFORMATION Ganetespib

[00256] Ganetespib, chemical name: 5-[2,4-dihydroxy-5-(l-methylethyl)phenyl]-

2,4-dihydro-4-(l-methyl-lH-indol-5-yl)-3H-l,2,4-triazole-3-one, is a novel triazolone heterocyclic compound. Its molecular formula is C20H20N4O3. Ganetespib is a white to off-white powder with a molecular weight of

364.40 g/mol.

Preparation and Administration

[00257] The current ganetespib investigational product is a concentrate for solution for infusion provided in a single-use vial containing 300 mg /vial of ganetespib, as described in the Pharmacy Manual. The concentration of ganetespib is 25 mg/mL in a polyethylene glycol 300 (PEG 300), polysorbate 80 (Tween-80) and dehydrated alcohol non-aqueous solvent system. The drug product is a clear, colorless-to-pale-yellow solution, essentially free of visible particles.

[00258] Ganetespib Drug Product, 25 mg/mL, 300 mg/vial (identified with a dark blue color cap and applicable product label): Each vial contains 12 mL of deliverable volume (12.84 mL total including an overage per USP requirements) equivalent to 300 mg of ganetespib at a concentration of 25 mg/mL in a PEG 300, polysorbate 80, and dehydrated alcohol non-aqueous solvent system. The drug product, as noted, is a clear, colorless-to-pale-yellow solution. [00259] The amount of ganetespib administered will depend upon the patient's body surface area. The drug product is diluted before infusion.

[00260] Ganetespib must be diluted prior to administration. The appropriate drug administration instructions per the preparation guidelines must be carefully followed prior to use. Refer to the Pharmacy Manual for detailed ganetespib preparation guidelines.

[00261] Based on preclinical data, use of vascular access devices (VADs) (such as ports and peripherally-inserted central catheters [PICCS]) containing silicone catheters for ganetespib administration is permitted. Use of VADs made of any other material is not permitted. Following ganetespib administration through a VAD, care should be taken to flush the line after each dose of study drug.

Formulation, Packaging and Labeling

[00262] The ganetespib drug product is provided in a 30 mL type I amber glass vial fitted with a 20 millimeter stopper and sealed with an aluminum crimp and flip-off cap..

[00263] Starting with the 300 mg drug product, the storage condition will be changed to 20-25°C (68°F to 77°F) with excursions allowed between 15°C and 30°C (59°F and 86°F).

Sirolimus

[00264] Sirolimus is a macrocyclic lactone produced by Streptomyces

hygroscopicus. The chemical name of sirolimus (also known as rapamycin) is (3S,6R ,7E,9R ,10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)- 9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27-dihydroxy-3- [(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21- dimethoxy-6,8, 12, 14,20,26-hexamethyl-23,27-epoxy-3H-pyrido [2, 1-c] [1,4] oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone. Its molecular formula is C51H79N013 and its molecular weight is 914.2

Preparation and Administration

[00265] For this trial 2 mg tablets will be provided.

[00266] Food effects: In 22 healthy volunteers receiving Rapamune Oral Solution, a high-fat meal (861.8 kcal, 54.9% kcal from fat) altered the bioavailability

characteristics of sirolimus. Compared with fasting, a 34% decrease in the peak blood sirolimus concentration (Cmax), a 3.5-fold increase in the time-to-peak concentration (tmax), and a 35% increase in total exposure (AUC) was observed. After administration of Rapamune Tablets and a high-fat meal in 24 healthy volunteers, Cmax, tmax, and AUC showed increases of 65%, 32%, and 23%, respectively. To minimize variability, tablets should be taken consistently with or without food.

Formulation, Packaging and Labelling

[00267] Tablets should be stored at 20° to 25°C (USP Controlled Room

Temperature) (68° to 77°F). Use cartons to protect blister cards and strips from light. Dispense in a tight, light-resistant container as defined in the USP.

MEASUREMENT OF EFFECTS Antitumor Effect - Solid Tumors Definitions

[00268] Evaluable for toxicity for Phase I component: A patient will be considered evaluable for definition of the MTD if at least 70% of the prescribed sirolimus dose has been administered to the patient during the first treatment cycle based on diary review and pill count of returned drug [if a discrepancy occurs, pill count will be used for adherence measurement] and all 3 doses of ganetespib in the first cycle. If a patient has less than 70% adherence, the patient will be replaced in the cohort. In addition, anyone who receives one or more doses and experiences a DLT will be considered evaluable for definition of MTD.

[00269] In the phase II component: Patients who have received at least one dose of study drug will be considered evaluable for toxicity from the time of their first dose of ganetespib and sirolimus until the last evaluation on trial.

[00270] Evaluable for objective response:

[00271] Measurable disease: Measurable lesions are defined as those that can be accurately measured in at least two dimensions (longest diameter > 20mm with conventional techniques or >10 mm using spiral CT scan). All tumor measurements must be recorded in millimeters (or decimal fractions of centimeters).

[00272] Non-measurable disease: All other lesions (or sites of disease), including small lesions (longest diameter < 20mm with conventional techniques or <10 mm using spiral CT scan), are considered non-measurable disease. Bone lesions, leptomeningeal disease, ascites, pleural/pericardial effusions, lymphangitis cutis/pulmonis, inflammatory breast disease, abdominal masses (not followed by CT or MRI), and cystic lesions are all non-measurable.

[00273] Any patient who is enrolled and receives at least one dose of sirolimus and ganetespib will be considered evaluable for response provided: (1) the patient demonstrates progressive disease or death while on protocol therapy; (2) the patient is observed on protocol therapy for at least one cycle and the tumor is not removed surgically prior to the time complete response or partial response is confirmed; or (3) the patient demonstrates a complete or partial response or stable disease as confirmed according to protocol criteria. Patients who electively terminate therapy before receiving all ganetespib doses and > 80% of the required sirolimus doses during the first treatment cycle and do not expire within 28 days from start of treatment will be replaced. Disease Parameters

[00274] Measurable, bidimensional: Malignant disease measurable in two

dimensions by ruler with surface area determined by multiplying the longest diameter by the greatest perpendicular diameter.

[00275] Index Lesions: Index lesions should be selected on the basis of their measurability in two dimensions and their suitability for accurate repeated measurements (by imaging techniques CT or MRI). A sum of the product(s) of the longest diameter (LD) and greatest perpendicular diameter of all index lesions will be calculated and reported as the baseline sum. The baseline sum will be used as a reference by which to characterize the objective tumor response.

[00276] Non-index Lesions: All other lesions (or sites of disease) including any measurable lesions over and above the index lesions should be identified as non- index lesions and should also be recorded at baseline. Measurement of these lesions is not required, but the presence or absence of each should be noted throughout follow-up.

Methods for Evaluation of Measurable Disease

[00277] All measurements should be taken and recorded in metric notation using a ruler or calipers. All baseline evaluations should be performed as closely as possible to the beginning of treatment and never more than 4 weeks before the beginning of the treatment.

[00278] The same method of assessment and the same technique should be used to characterize each identified and reported lesion at baseline and during follow-up. Imaging-based evaluation is preferred to evaluation by clinical examination when both methods have been used to assess the antitumor effect of a treatment.

[00279] Conventional CT and MRI: These techniques should be performed with cuts of 10 mm or less in slice thickness contiguously. Spiral CT should be performed using a 5 mm contiguous reconstruction algorithm. This applies to tumors of the chest, abdomen, and pelvis. Head and neck tumors and those of extremities may require specific protocols.

[00280] WHO response criteria

[00281] WHO will be used for this trial for several reasons:

[00282] 1) MPNSTs are typically complex non spherical tumors, and 2- dimensional measurements may thus better reflect changes in tumor size than 1- dimensional measurements (RECIST).

[00283] 2) The phase 2 trial of erlotinib and everolimus/bevacizumab, which will be used as a historical control for determination of time to progression, used WHO criteria. In order to allow for the closest comparison, this trial will therefore use WHO response criteria.

Evaluation of Index Lesions

[00284] Complete Response (CR): Disappearance of all known disease, determined by two consecutive observations not less than 4 weeks apart.

[00285] Partial Response (PR): A > 50% decrease in the total tumor load of the lesions that have been measured to determine the effect of therapy not less than four weeks apart. The observations must be consecutive.

[00286] Bidimensionally measurable: single lesion, > 50% decrease in tumor area (multiplication of longest diameter by the greatest perpendicular diameter); multiple lesions, a 50% decrease in the sum of the products of the perpendicular diameters of the multiple lesions.

[00287] In addition there can be no appearance of new lesions or progression of any lesion.

[00288] Stable Disease (SD): A 50% decrease in total tumor area cannot be established nor has a 25% increase in the size of one or more measurable lesions been demonstrated. [00289] Progressive Disease (PD): A > 25% increase in the area of one or more measurable lesions or the appearance of new lesions.

Evaluation of Non-Index Lesions

[00290] Complete Response (CR): Complete disappearance of all known disease for at least four weeks.

[00291] Partial Response (PR): Estimated decrease in tumor area of > 50% for at least four weeks.

[00292] Stable Disease (SD): No significant change for at least four weeks. This includes stable disease, estimated decrease of < 50%, and lesions with estimated increase of < 25%.

[00293] Progressive Disease (PD): Appearance of any new lesions not previously identified or an estimated increase of > 25% in existent lesions.

[00294] Although a clear progression of "non-index" lesions only is exceptional, the opinion of the treating physician should prevail in such circumstances, and the progression status should be confirmed at a later time by the Principal Investigator.

Evaluation of Best Overall Response

Duration of Response

[00295] Duration of overall response: The duration of overall response is measured from the time measurement criteria are met for CR or PR (whichever is first recorded) until the first date that recurrent or progressive disease is objectively documented (taking as reference for progressive disease the smallest measurements recorded since the treatment started, best response scan).

[00296] The duration of overall CR is measured from the time measurement criteria are first met for CR until the first date that recurrent disease is objectively documented.

[00297] Duration of stable disease: Stable disease is measured from the start of the treatment until the criteria for progression are met, taking as reference the smallest measurements recorded since the treatment started.

Progression-Free Survival

[00298] PFS is defined as the duration of time from start of treatment to time of objective progression or death.

STATISTICAL CONSIDERATIONS Study Design/Endpoints

Phase I

[00299] In the phase I component, a conventional dose-escalation design is used. The initial starting dose of ganetespib is approximately 1 dose level below the recommended phase 2 weekly dose in combination with the recommended adult dose of sirolimus of 4 mg once daily. This will be followed by one dose escalation to the ganetespib weekly recommended phase 2 dose and sirolimus recommended adult dose. The total number of dose levels will likely only be 2, with the possibility of 2 dose de-escalations. Cohorts of 3 to 6 patients will be treated per dose level. [00300] The MTD/Recommended dose will be defined as the dose level immediately below the level at which >33% of patients in a cohort experience a DLT based on toxicities observed in the first treatment cycle. In the phase 1 portion, this study is largely exploratory, precluding formal statistical comparisons of treatment groups. Toxicity and tolerability will be measured using CTCAE-4 and summarized by dose level. Grade >3 toxicities attributable to agents will be presented in frequency tables for each dose level.

Phase II

[00301] In the phase II component, the primary endpoint will be clinical benefit rate, which will be defined as a CR, PR, or stable disease >4 months. A completed trial of erlotinib in patients with refractory MPNST demonstrated a median time to progression of 48 days and progressive disease in the first response evaluation after completion of 2 treatment cycles in 19/20 patients— . Thus stable disease > 4 months can be considered potentially beneficial and worthy of further investigation. An evaluable patient will be classified a responder (success) for the primary endpoint if the patient achieves a PR, CR or stable disease at > 4 months. The target clinical benefit rate will be 25%, and a clinical benefit rate < 5% will be considered

uninteresting. Using Simon's optimal two-stage phase II design, the first stage will require 10 patients, with no further accrual if 0 of 10 respond. If >1/10 patients respond, accrual will continue until a total of 20 patients have been enrolled. If >3/20 patients respond, this combination will be considered of sufficient activity.

Assuming the number of successes is binomially distributed, this design has a one sided alpha of 0.07 and a power of 88% for detecting a true success probability of at least 25% versus the null hypothesis success rate of 5% or less. Definitions

Evaluable for Adverse Effects

[00302] Any patient who experiences DLT at any time during protocol therapy is considered evaluable for adverse effects. Patient without DLT who receive at least 85% of the prescribed sirolimus dose per protocols guidelines and had the appropriate toxicity monitoring performed are also considered evaluable for adverse effects. Patients who are not evaluable for adverse effects at a given dose level will be replaced.

Evaluable for Response

[00303] Any eligible patient who is enrolled and receives at least one dose of drug will be considered evaluable for response provided: 1) the patient demonstrates progressive disease or death while on protocol therapy or 2) the patient is observed on protocol therapy for at least one cycle and the tumor is not removed surgically prior to the time complete response or partial response or stable disease is confirmed, or 3) patient demonstrates a complete or partial response or stable disease per protocol criteria. The evaluation period for determination of best response will be 6 treatment cycles. All other patients will be considered non- responders

Sample Size/Accrual Rate

[00304] Phase I: 3 to 6 patients per cohort with 1 dose escalations (potential for 2 de-escalation). Thus a minimum of 6 patients to a maximum of 18 patients will be enrolled.

[00305] Phase II: 10 patients in first stage with an additional 10 patients in the second stage for a total of 20 patients. The maximum number of patients for entire study will be 38. It is expected that 15-25 patients be enrolled per year, and enrollment is expected to be completed in approximately 2.5 years. Analysis of Secondary Endpoints

[00306] Summary statistics will be used to describe the study population and baseline characteristics.

[00307] Pharmacokinetic analysis will be conducted using non-compartmental methods and estimated pharmacokinetic parameters including AUC, clearance, half- life and volume of distribution presented for each dose level using summary statistics (mean, standard deviation, median and range).

[00308] Analysis of secondary endpoints will be predominantly descriptive using primarily non-parametric analyses and will be interpreted as being exploratory and hypothesis generating. Results will be summarized with graphical analysis and correlations will be made if feasible.

[00309] Pharmacodynamic parameters including phospho-S6, phospho-S6, phosphorylated eIF2 alpha, Akt Phosphorylation, Hsp70, and G6PD will be evaluated in surrogate tissue (peripheral blood mononuclear cells) and tumor tissue (when feasible) at baseline and during treatment. Changes in these parameters will be correlated to radiographic response using logistic regression analysis.

[00310] Patient reported pain severity and impact of pain on activities of daily living will be summarized using descriptive statistics. Analysis of change in pain severity and pain interference over time may involve individual t-tests between 2 time points (or non-parametric Wilcoxon-Mann- Whitney tests) and repeated measures Analysis of Variance when appropriate. Correlations and chi-square analyses to compare pain severity and impact with objective tumor response and adverse events will be made if feasible. CORRELATIVE STUDIES Pharmacokinetics

[00311] The plasma pharmacokinetic profile of ganetespib and sirolimus will be described when administered in combination therapy. Pre-therapy levels will be drawn at baseline. Pharmacokinetic analysis will occur on cycle 1, day 15 as to capture steady state sirolimus levels. Ganetespib has previously demonstrated no accumulation. On day 15: Ganetespib will be collected at hours 0, 1 hr (at the end of infusion), 2, 4, 6, 8, and 24 hours. Sirolimus will be collected at hours 0, 1, 2, 4, and 24 hours. All patients enrolled in the phase I portion will be required to participate in pharmacokinetic testing. In the phase II component, up to 10 patients will have optional pharmacokinetic analysis performed for broader experience at the

MTD/recommended dose.

[00312] There are no additional required sirolimus trough levels. However, sirolimus trough levels can be drawn if concerns for toxicity or any other clinical indication, and recommendations to be discussed with principal investigator.

Pharmacodynamics

[00313] Correlative studies evaluating pharmacodynamic parameters on Hsp inhibition, mTOR inhibition, UPR activation, and oxidative stress will be explored in tumor tissue and peripheral blood mononuclear cells. Hsp 70 levels increase when Hsp90 is effectively inhibited and has been used as a pharmacodynamic response to Hsp90 inhibitors (73). Phospho-S6 is the marker of choice in mTOR PD studies (59, 74) and recent data indicate that baseline expression could have predictive value (75). The limited clinical efficacy of mTOR inhibitors have been proposed to result from AKT activation that can occur via the negative feedback pathways (76). Sirolimus did not induce AKT activation in MPNSTs in vivo(27), and combined rapamycin/IPI-504 did not suppress AKT phosphorylation or expression levels in the mouse model, indicating that this combination is not more effective because it inhibits AKT (30). EIF2cc phosphorylation is a marker of UPR activation (28) and was observed in the preclinical model as a marker of ER stress (30). Hsp90 inhibition appears to stimulate ROS production, and sirolimus appears to enhance these effects by suppressing endogenous anti-oxidants (30). Glucose 6-phosphate dehydrogenase (G6PD) has a well-established role in protecting cells from oxidative stress via its effects on glutathione production, one of the most important endogenous cellular antioxidants (77-79). G6PD expression can be suppressed by mTOR inhibitors in vitro through inhibitory effects on the transcription factor SREBP1 (80). It has been shown that sirolimus potently suppressed G6PD mRNA levels in MPNST tumor tissue (30). Sirolimus and Hsp90 inhibition combined dramatically suppressed G6PD mRNA and protein expression MPNSTs in vivo.

[00314] Changes in pharmacodynamic parameters will be examined: Hsp70, Phospho-S6, AKT phosphorylation, EIF2cc, and G6PD in peripheral blood

mononuclear cells performed on day 1 prior to ganetespib and sirolimus

administration, and on day 15, 6 hours post drug administration. Archival tissue will be obtained at diagnosis and/or recurrence to evaluate these tests if available. For consenting patients with tumors accessible safely by percutaneous biopsy, on the phase II component only, tumor biopsy will be obtained at baseline and then on day 8 or 15 of cycle 1 at some time point within 12 hours after infusion of ganetespib.

Patient reported pain and impact of pain of daily activities

[00315] Pain associated with a mass was found to be greatest risk factor associated with the development of MPNSTs in NF1 (12). Pain may also serve a surrogate marker for tumor response and clinical benefit. Patient reported pain severity and the impact of pain on daily activities will be assessed prior to treatment and during treatment prior to cycles 3, 5, 9 and 13. The relationship in the change in pain with radiologic response will be examined. The patient-reported pain evaluation will consist of two validated scales. The Numerical Rating Scale-11 (NRS-11) will be used to assess pain severity, and the Pain Interference Scale from the Brief Pain Inventory will be used to assess the impact of pain on daily activities. These scales have been placed on a single page to simplify administration. Total administration time is less than 3 minutes.

[00316] The Numerical Rating Scale-11 (NRS-11) is a self-report segmented 11- point numeric scale that assesses pain severity(81). It consists of a horizontal line with 0 representing "no pain" at the right end of the line and 10 representing "worst pain you can imagine" at the left end. Patients are asked to circle the one number from 0 to 10 that best describes how much their "most important tumor pain" hurt during the past week. It takes less than 1 minute to complete.

[00317] The Brief Pain Inventory is a 7-item self-report questionnaire that measures the extent to which pain interferes with daily functioning (82). Patients are asked to indicate how much pain interfered with various activities (general activity, mood, walking, normal work, relations with other people, sleep, and enjoyment of life) in the past week, with scores ranging from 0 (does not interfere) to 10

(completely interferes). A total score is obtained by taking the mean of the scores for all 7 items; thus, the total pain interference score can range from 0 to 7. This scale takes less than 2 minutes to complete.

Volumetric Tumor measurements

[00318] Using volumetric 3-dimensional MRI analysis of MPNSTs, response to ganetespib and sirolimus may be more sensitively monitored as compared to conventional two-dimensional MRI and one-dimensional MRI data analysis.

Volumetric MRI analysis has become the standard method to evaluate the growth rate of plexiform neurofibromas on clinical trials (83, 84). This method may have utility for MPNSTs, which are typically large and have a complex shape. [00319] Volumetric MRI analysis will be performed centrally at the NIH as a secondary objective and compared to standard response evaluation with ID-and 2D- measurements. MRI studies performed for response evaluation will be used for volumetric analysis, and no MRI sequence other than STIR MRI (Short Tl inversion recovery), which is commonly used in the evaluation of sarcomas and does not require contrast administration, will be required.

Brief Statistical Design

[00320] In the phase I component, a conventional 3+3 dose-escalation design is used. The initial starting dose of ganetespib is 150 mg/m², approximately 1 dose level below the recommended phase 2 weekly dose, in combination with the recommended adult dose of sirolimus of 4 mg once daily. This will be followed by one dose escalation of the ganetespib to 200 mg/m² weekly (recommended phase 2 dose) and sirolimus recommended adult dose. The Maximum tolerated dose

(MTD)/Recommended dose will be defined as the dose level immediately below the level at which > 33% of patients in a cohort experience a dose-limiting toxicity (DLT) based on toxicities observed in the first treatment cycle.

[00321] In the phase II component, the primary endpoint will be clinical benefit rate, which will be defined as a CR, PR, or stable disease > 4 cycles. An evaluable patient will be classified as a responder (success) for the primary endpoint if the patient achieves a PR, CR or stable disease at > 4 months. The target clinical benefit rate will be 25%, and a clinical benefit rate < 5% will be considered uninteresting. Using a Simon's optimal two-stage phase II design, the first stage will require 10 patients, with no further accrual if 0 of 10 respond. If >_1/10 patients respond, accrual will continue until a total of 20 patients have been enrolled. If >_3/20 patients respond, this combination will be considered of sufficient activity. Assuming the number of successes is binomially distributed, this design has a one sided alpha of 0.07 and a power of 88% for detecting a true success probability of at least 25% versus the null hypothesis success rate of 5% or less.

[00322] All publications cited herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control. In addition, the materials, methods, and examples throughout the specification are illustrative only and not intended to be limiting in any way.

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Claims

CLAIMS What is claimed is:

1. A method of treating or preventing a sarcoma in a subject, comprising administering a therapeutically effective amount of an Hsp90 inhibitor in

combination with an mTOR inhibitor, wherein the subject is selected as being of 16 years of age or less.

2. A method of reducing or inhibiting the progression of a sarcoma in a subject, comprising administering a therapeutically effective amount of an Hsp90 inhibitor in combination with an mTOR inhibitor, wherein the subject is selected as being of 16 years of age or less.

3. The method of claim 1 or 2, wherein the Hsp90 inhibitor is ganetespib (5-[2,4- dihydroxy-5-(l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lH-indol-5-yl)-3ii- l,2,4-triazole-3-one), or a pharmaceutically acceptable salt or a tautomer thereof.

4. The method of claim 1 or 2, wherein the mTOR inhibitor is sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)- 9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27-dihydroxy-3- [(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21- dimethoxy-6,8, 12, 14,20,26-hexamethyl-23,27-epoxy-3H-pyrido [2, 1-c] [1,4]

oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone.

5. The method of claim 1 or 2, wherein the sarcoma is a refractory sarcoma.

6. The method of claim 5, wherein the refractory sarcoma is a bone or soft tissue sarcoma.

7. The method of claim 1 or 2, wherein the sarcoma is a malignant peripheral nerve sheath tumor (MPNST).

8. The method of claim 7, wherein the MPNST is an unresectable or metastatic sporadic or neurofibromatosis type-1 associated MPNST.

9. The method of claim 1 or 2, wherein the sarcoma was previously treated and not responsive.

10. The method of claim 1 or 2, wherein the sarcoma is a recurrent sarcoma.

11. The method of any one of claims 1 or 2, wherein the amount of the Hsp90 inhibitor administered is from 100-200 mg/m².

12. The method of claim 11, wherein the amount of the Hsp90 inhibitor administered is from 150-200 mg/m².

13. The method of claim 11, wherein the Hsp90 inhibitor is administered intravenously.

14. The method of claim 13, wherein the Hsp90 inhibitor is infused over 60 minutes.

15. The method of any one of claims 1 or 2, wherein the amount of mTOR inhibitor administered is 2-12 mg.

The method of claim 15, wherein the amount of mTOR inhibitor administered L mg.

17. The method of claim 15, wherein the mTOR inhibitor is administered orally.

18. The method of any one of claims 1 or 2, wherein the Hsp90 inhibitor and mTOR inhibitor are administered over a 28 day cycle.

19. The method of claim 18, wherein the 28 day cycle is repeated one or more times.

20. The method of any one of claims 1 or 2, wherein the Hsp90 inhibitor is administered on days 1, 8 and 15.

21. The method of any one of claims 1 or 2, wherein the mTOR inhibitor is administered on days 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21,

22. 23, 24, 25, 26, 27 and 28.

22. A method of treating or preventing a sarcoma in a subject, comprising administering a therapeutically effective amount of ganetespib (5-[2,4-dihydroxy-5- (l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lH-indol-5-yl)-3H-l,2,4-triazole-3- one), or a pharmaceutically acceptable salt or a tautomer thereof, in combination with sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)- 9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27-dihydroxy-3- [(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21- dimethoxy-6,8, 12, 14,20,26-hexamethyl-23,27-epoxy-3H-pyrido [2, 1-c] [1,4]

oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone, wherein the subject is selected as being of 16 years of age or less.

23. A method of reducing or inhibiting the progression of a sarcoma in a subject, comprising administering a therapeutically effective amount of ganetespib (5-[2,4- dihydroxy-5-(l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lH-indol-5-yl)-3H l,2,4-triazole-3-one), or a pharmaceutically acceptable salt or a tautomer thereof, in combination with sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S _/26R_/27R _/34aS)-9_/10,12,13,14,21_/22_/23,24_/25_/26_/27_/32_/33_/34_/ 34a-hexadecahydro-9_/27- dihydroxy-3-[(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-

10,21-dimethoxy-6,8,12,14,20,26-hexamethyl-23,27-epoxy-3H-pyrido[2,l-c][l,4] oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone, wherein the subject is selected as being of 16 years of age or less.

24. A kit to practice any one of the methods of claim 1-23.

25. A composition comprising an Hsp90 inhibitor and an mTOR inhibitor.

26. The composition of claim 25, wherein the Hsp90 inhibitor is ganetespib (5- [2,4-dihydroxy-5-(l-methylethyl)phenyl]-2,4-dihydro-4-(l-methyl-lH-indol-5-yl)-3H- l,2,4-triazole-3-one), or a pharmaceutically acceptable salt or a tautomer thereof,.

27. The composition of claim 25, wherein the mTOR inhibitor is sirolimus (3S,6R ,7E,9R, 10R,12R ,14S,15E ,17E,19E ,21S,23S ,26R,27R ,34aS)- 9,10,12,13,14,21,22,23,24,25,26,27,32,33,34, 34a-hexadecahydro-9,27-dihydroxy-3- [(lR)-2-[(lS,3R,4R)-4-hydroxy-3-methoxycyclohexyl]-l-methylethyl]-10,21- dimethoxy-6,8, 12, 14,20,26-hexamethyl-23,27-epoxy-3H-pyrido [2, 1-c] [1,4]

oxaazacyclohentriacontine-1,5, 11,28,29 (4H,6H ,31H)-pentone.

28. The composition of claim 25, wherein the amount of the Hsp90 inhibitor is between 100-500 mg/m².

29. The composition of claim 28, wherein the amount of the Hsp90 inhibitor is between 100-200 mg/m², preferably 150-200 mg/m².

30. The composition of claim 25, wherein the amount of mTOR inhibitor is between 2-12 mg.

31. The composition of claim 30, wherein the amount of mTOR inhibitor is between 2-4 mg.

32. The composition of any one of claims 25-31, wherein the composition wherein the combination of the Hsp90 inhibitor and the mTOR inhibitor exhibits therapeutic synergy.