Methods for treating immunorefractory lung cancer

Abstract

This disclosure features methods for treating immunorefractory lung cancer using RAS inhibitors. This disclosure also features combination therapies for treating immunorefractory lung cancer.

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority to U.S. Provisional Application No. 63 / 317,649, filed March 8, 2022, the entire disclosure of which is incorporated herein by reference as if set forth in its entirety.

[0002] Sequence Listing This application contains a Sequence Listing that has been submitted electronically in XML format and is incorporated herein by reference in its entirety. The XML copy was created on March 3, 2023, has the file name "51432-025WO2_Sequence_Listing_3_3_23", and is 1,785 bytes in size. [Background technology]

[0003] Cancer remains one of the most deadly threats to human health. In the United States, cancer affects approximately 1.3 million new cases each year and is the second leading cause of death after heart disease, accounting for approximately one in four deaths.

[0004] The development of immune checkpoint inhibitors (ICIs) has dramatically improved the treatment of various solid tumors. However, initial or acquired resistance to ICI treatment often hampers sustained antitumor activity. Current response biomarkers for anti-PD-1 or anti-PD-L1 treatment include tumor mutation burden, programmed cell death ligand-1 (PD-L1) expression, and T cell density. ICI-induced antitumor immunity relies on lymphocyte infiltration into the tumor core, with "T cell-inflammed" tumors demonstrating the best response. In contrast, "cold tumors," also known as immunorefractory or immune-evasive tumors, may be defined in part by a lack of T cell infiltration and a low IFN-g gene signature and are poorly responsive to immune checkpoint inhibition (Bonaventura et al., Front. Immunol. 2019). Novel compositions and methods for treating immunorefractory tumors are needed. Summary of the Invention

[0005] The present disclosure provides compositions and uses thereof for treating immunorefractory lung cancer. The disclosure is based, at least in part, on the observation that treating immunorefractory lung cancer with a RAS inhibitor (e.g., a RAS(ON) inhibitor) compound that inhibits a mutant RAS G12C protein sensitizes the cancer to treatment with an immunotherapeutic agent. In some embodiments, the compound inhibits a RAS that has an oncogenic G12C mutation. In some embodiments, the RAS inhibitor is, for example, a covalent inhibitor that can form a covalent bond with an oncogenic RAS G12C mutant at the G12C position. In some embodiments, treatment with a RAS inhibitor sensitizes the cancer to treatment with an immune checkpoint inhibitor or an SHP2 inhibitor. In some embodiments, a compound or combination of compounds described herein is administered to a subject who has previously failed immunotherapy treatment, such as immunotherapy treatment with an immune checkpoint inhibitor.

[0006] In one aspect, the present disclosure provides a method for administering to a subject a RAS G12CA method of treating immunorefractory lung cancer in a subject by administering an (ON) inhibitor is provided.

[0007] In another aspect, the present disclosure provides a method for administering a RAS inhibitor to a subject in need thereof. G12C This paper provides a method for immunologically altering the tumor microenvironment of cold lung cancer by administering an (ON) inhibitor.

[0008] In some embodiments, the RAS G12C (ON) inhibitors are tri-complex RAS G12C (ON) inhibitor.

[0009] In another aspect, the disclosure provides a method of treating immunorefractory lung cancer in a subject, comprising administering to the subject a RAS inhibitor of Formula I: [ka] or a pharmaceutically acceptable salt thereof wherein the dotted lines represent 0, 1, 2, 3, or 4 non-adjacent double bonds; A is —N(H or CH3)C(O)—(CH2)—, where the amino nitrogen is —CH(R 10 )-, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene; B is absent or -CH(R 9 )-,>C=CR 9 R 9’ , or >CR 9 R 9’ wherein carbon is —N(R 11 )C(O)—; optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or 5- to 6-membered heteroarylene; G is optionally substituted C1-C4 alkylene, optionally substituted C1-C4 alkenylene, optionally substituted C1-C4 heteroalkylene; [ka] [In the formula, [ka] -C(R 7 R 8 )-bonded to ], [ka] [In the formula, [ka] -C(R 7 R 8 )-; optionally substituted C1-C4 heteroalkylene, or 3- to 8-membered heteroarylene; L is a linker, said linker being acyclic or comprising a monocyclic, fused bicyclic, fused polycyclic, bridged bicyclic, or bridged polycyclic group; W is a bridging group comprising a vinyl ketone, vinyl sulfone, ynone, haloacetyl, or alkynyl sulfone; X 1 is an optionally substituted C1-C2 alkylene, NR, O, or S(O) n and X 2 is O or NH, X 3 is N or CH, n is 0, 1, or 2; R is hydrogen, cyano, optionally substituted C1-C4 alkyl, optionally substituted C2-C4 alkenyl, optionally substituted C2-C4 alkynyl, C(O)R', C(O)OR', C(O)N(R')2, S(O)R', S(O)2R', or S(O)2N(R')2; Each R’ are independently H or optionally substituted C1-C4 alkyl; Y 1 is C, CH, or N, Y 2 , Y 3 , Y 4 , and Y 7 are independently C or N, Y 5 is CH, CH2, or N, Y 6 is C(O), CH, CH2, or N, R 1 is cyano, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 6-membered cycloalkenyl, optionally substituted 3- to 6-membered heterocycloalkyl, optionally substituted 6- to 10-membered aryl, or optionally substituted 5- to 10-membered heteroaryl; R 2 is absent, hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 7-membered heterocycloalkyl, optionally substituted 6-membered aryl, optionally substituted 5- or 6-membered heteroaryl; R 3 does not exist, or R 1 and R 2 combine with the atom to which they are attached to form an optionally substituted 3- to 14-membered heterocycloalkyl, or R 2 and R 3 combine with the atom to which they are attached to form an optionally substituted 3- to 8-membered cycloalkyl or an optionally substituted 3- to 14-membered heterocycloalkyl; R 4 is absent, hydrogen, halogen, cyano, or methyl optionally substituted with 1 to 3 halogens; R 5is hydrogen; C1-C4 alkyl optionally substituted with halogen, cyano, hydroxy, or C1-C4 alkoxy; cyclopropyl, or cyclobutyl; R 6 is hydrogen or methyl, and R 7 is hydrogen, halogen, or optionally substituted C1-C3 alkyl, or R 6 and R 7 combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R 8 is hydrogen, halogen, hydroxy, cyano, optionally substituted C1-C3 alkoxy, optionally substituted C1-C3 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 8-membered cycloalkyl, optionally substituted 3- to 14-membered heterocycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 6- to 10-membered aryl, or R 7 and R 8 combine with the carbon atom to which they are attached, forming C=CR 7’ R 8’ forming C=N(OH), C=N(O-C1-C3 alkyl), C=O, C=S, C=NH, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted 3- to 7-membered heterocycloalkyl; R 7a and R 8a are independently hydrogen, halo, optionally substituted C1-C3 alkyl, or combined with the carbon to which they are attached to form a carbonyl; R 7’ is hydrogen, halogen, or optionally substituted C1-C3 alkyl, and R 8’is hydrogen, halogen, hydroxy, cyano, optionally substituted C1-C3 alkoxy, optionally substituted C1-C3 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 8-membered cycloalkyl, optionally substituted 3- to 14-membered heterocycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 6- to 10-membered aryl, or R 7’ and R 8’ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R 9 is H, F, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 7-membered heterocycloalkyl; or R 9 and L, combined with the atom to which they are attached, form an optionally substituted 3- to 14-membered heterocycloalkyl; R 9’ is hydrogen or optionally substituted C1-C6 alkyl, or R 9 and R 9’ combine with the atom to which they are attached to form a 3- to 6-membered cycloalkyl or a 3- to 6-membered heterocycloalkyl; R 10 is hydrogen, halo, hydroxy, C1-C3 alkoxy, or C1-C3 alkyl; R 10a is hydrogen or halo, R 11 is hydrogen or C1-C3 alkyl, and R 21 is H or C1-C3 alkyl.

[0010] In another aspect, the present disclosure provides a method for altering the tumor microenvironment of immunologically cold lung cancer in a subject in need thereof by administering to the subject a RAS inhibitor of Formula I or a subformula thereof as described herein. In some embodiments, the subject is resistant to immune checkpoint inhibitors prior to altering the tumor microenvironment. In some embodiments, administration of the RAS inhibitor alters the tumor microenvironment, thereby sensitizing the cancer to treatment with an immune checkpoint inhibitor.

[0011] In each of the preceding embodiments, the method may further include administering to the subject an SHP2 inhibitor.

[0012] In another aspect, the present disclosure provides a method for administering to a subject a RAS G12C A method of treating immunorefractory lung cancer in a subject by administering an (ON) inhibitor, an SHP2 inhibitor, and an immune checkpoint inhibitor is provided.

[0013] In yet another aspect, the present disclosure provides a method for administering a RAS inhibitor to a subject in need thereof. G12C A method for immunologically transforming the tumor microenvironment of cold lung cancer is provided by administering an (ON) inhibitor, an SHP2 inhibitor, and an immune checkpoint inhibitor.

[0014] In some embodiments of any of the methods described herein, the Ras inhibitor is a compound of Formula I: [ka] or a pharmaceutically acceptable salt thereof wherein the dotted lines represent 0, 1, 2, 3, or 4 non-adjacent double bonds; A is —N(H or CH3)C(O)—(CH2)—, where the amino nitrogen is —CH(R 10)-, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene; B is absent or -CH(R 9 )-,>C=CR 9 R 9’ , or >CR 9 R 9’ wherein carbon is —N(R 11 )C(O)—; optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or 5- to 6-membered heteroarylene; G is optionally substituted C1-C4 alkylene, optionally substituted C1-C4 alkenylene, optionally substituted C1-C4 heteroalkylene; [ka] [In the formula, [ka] -C(R 7 R 8 )-bonded to ], [ka] [In the formula, [ka] -C(R 7 R 8 )-; optionally substituted C1-C4 heteroalkylene, or 3- to 8-membered heteroarylene; L is a linker, said linker being acyclic or comprising a monocyclic, fused bicyclic, fused polycyclic, bridged bicyclic, or bridged polycyclic group; W is a bridging group comprising a vinyl ketone, vinyl sulfone, ynone, haloacetyl, or alkynyl sulfone; X 1 is an optionally substituted C1-C2 alkylene, NR, O, or S(O) n and X 2 is O or NH, X 3 is N or CH, n is 0, 1, or 2; R is hydrogen, cyano, optionally substituted C1-C4 alkyl, optionally substituted C2-C4 alkenyl, optionally substituted C2-C4 alkynyl, C(O)R', C(O)OR', C(O)N(R')2, S(O)R', S(O)2R', or S(O)2N(R')2; Each R ’ are independently H or optionally substituted C1-C4 alkyl; Y 1 is C, CH, or N, Y 2 , Y 3 , Y 4 , and Y 7 are independently C or N, Y 5 is CH, CH2, or N, Y 6 is C(O), CH, CH2, or N, R 1 is cyano, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 6-membered cycloalkenyl, optionally substituted 3- to 6-membered heterocycloalkyl, optionally substituted 6- to 10-membered aryl, or optionally substituted 5- to 10-membered heteroaryl; R 2is absent, hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 7-membered heterocycloalkyl, optionally substituted 6-membered aryl, optionally substituted 5- or 6-membered heteroaryl; R 3 does not exist, or R 1 and R 2 combine with the atom to which they are attached to form an optionally substituted 3- to 14-membered heterocycloalkyl, or R 2 and R 3 combine with the atom to which they are attached to form an optionally substituted 3- to 8-membered cycloalkyl; or R 4 is absent, hydrogen, halogen, cyano, or methyl optionally substituted with 1 to 3 halogens; R 5 is hydrogen; C1-C4 alkyl optionally substituted with halogen, cyano, hydroxy, or C1-C4 alkoxy; cyclopropyl, or cyclobutyl; R 6 is hydrogen or methyl, and R 7 is hydrogen, halogen, or optionally substituted C1-C3 alkyl, or R 6 and R 7 combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R 8 is hydrogen, halogen, hydroxy, cyano, optionally substituted C1-C3 alkoxy, optionally substituted C1-C3 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 8-membered cycloalkyl, optionally substituted 3- to 14-membered heterocycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 6- to 10-membered aryl, or R 7 and R8 combine with the carbon atom to which they are attached, forming C=CR 7’ R 8’ forming C=N(OH), C=N(O-C1-C3 alkyl), C=O, C=S, C=NH, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted 3- to 7-membered heterocycloalkyl; R 7a and R 8a are independently hydrogen, halo, optionally substituted C1-C3 alkyl, or combined with the carbon to which they are attached to form a carbonyl; R 7’ is hydrogen, halogen, or optionally substituted C1-C3 alkyl, and R 8’ is hydrogen, halogen, hydroxy, cyano, optionally substituted C1-C3 alkoxy, optionally substituted C1-C3 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 8-membered cycloalkyl, optionally substituted 3- to 14-membered heterocycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 6- to 10-membered aryl, or R 7’ and R 8’ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R 9 is H, F, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 7-membered heterocycloalkyl; or R 9 and L, combined with the atom to which they are attached, form an optionally substituted 3- to 14-membered heterocycloalkyl; R 9’ is hydrogen or optionally substituted C1-C6 alkyl, or R 9 and R 9’combine with the atom to which they are attached to form a 3- to 6-membered cycloalkyl or a 3- to 6-membered heterocycloalkyl; R 10 is hydrogen, halo, hydroxy, C1-C3 alkoxy, or C1-C3 alkyl; R 10a is hydrogen or halo, R 11 is hydrogen or C1-C3 alkyl, and R 21 is H or C1-C3 alkyl.

[0015] In some embodiments, the RAS inhibitor is a compound of formula II: [ka] or a pharmaceutically acceptable salt thereof.

[0016] In some embodiments, the RAS inhibitor is a compound of formula III: [ka] or a pharmaceutically acceptable salt thereof.

[0017] In some embodiments, the RAS inhibitor is a compound of formula IV: [ka] or a pharmaceutically acceptable salt thereof.

[0018] In some embodiments, the RAS inhibitor is a compound of formula V: [ka] or a pharmaceutically acceptable salt thereof.

[0019] In some embodiments, the RAS inhibitor is a compound of formula VI: [ka] or a pharmaceutically acceptable salt thereof [In the formula, X e and X f are independently N or CH, and R 12 is an optionally substituted C1-C6 alkyl, an optionally substituted C1-C6 heteroalkyl, or an optionally substituted 3- to 7-membered heterocycloalkyl.

[0020] In some embodiments of the compound of Formula I, R 7 is methyl, or R 8 is methyl. In some embodiments of the compound of Formula I, R 7 is methyl, or R 8 is methyl.

[0021] In some embodiments, the RAS inhibitor is a compound of formula VII: [ka] or a pharmaceutically acceptable salt thereof [In the formula, R 13 is hydrogen, an optionally substituted 3-10 membered heterocycloalkyl, or an optionally substituted C1-C6 heteroalkyl.

[0022] In some embodiments, R 2 is an optionally substituted C1-C6 alkyl or an optionally substituted 3- to 6-membered cycloalkyl.

[0023] In some embodiments, L is acyclic. In some embodiments, L is monocyclic.

[0024] In some embodiments, A is an optionally substituted 6-membered arylene. In some embodiments, A is an optionally substituted 5-6-membered heteroarylene. In some embodiments, A is an optionally substituted C1-C4 heteroalkylene. In some embodiments, A is an optionally substituted 3-6-membered heterocycloalkylene.

[0025] In some embodiments, B is —CHR 9 In some embodiments, R 9 is F, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 7-membered heterocycloalkyl. In some embodiments, B is an optionally substituted 6-membered arylene. In some embodiments, B is a 6-membered arylene.

[0026] In some embodiments, W is a bridging group comprising a vinyl ketone, a vinyl sulfone, an ynone, or an alkynyl sulfone.

[0027] In some embodiments, W is a bridging group comprising a vinyl ketone. In some embodiments, W has the structure of Formula VIIIa: [ka] [In the formula, R 16a , R 16b , and R 16c are independently hydrogen, —CN, halogen, or —C1-C3 alkyl optionally substituted with one or more substituents independently selected from —OH, —O—C1-C3 alkyl, —NH2, —NH(C1-C3 alkyl), —N(C1-C3 alkyl)2, or 4- to 7-membered saturated heterocycloalkyl.

[0028] In some embodiments, W is an ynone-containing bridging group. In some embodiments, W has the structure of Formula VIIIb: [ka] [In the formula, R 17 is hydrogen; -C1-C3 alkyl optionally substituted with one or more substituents independently selected from -OH, -O-C1-C3 alkyl, -NH2, -NH(C1-C3 alkyl), -N(C1-C3 alkyl)2, or 4- to 7-membered saturated cycloalkyl; or 4- to 7-membered saturated heterocycloalkyl.

[0029] In some embodiments, W is a bridging group comprising a vinyl sulfone. In some embodiments, W has the structure of Formula VIIIc: [ka] [In the formula, R 18a , R 18b , and R 18c are independently hydrogen, —CN, or —C1-C3 alkyl optionally substituted with one or more substituents independently selected from —OH, —O—C1-C3 alkyl, —NH2, —NH(C1-C3 alkyl), —N(C1-C3 alkyl)2, or 4- to 7-membered saturated heterocycloalkyl.

[0030] In some embodiments, W is a bridging group comprising an alkynyl sulfone. In some embodiments, W has the structure of formula VIIId: [ka] [In the formula, R 19 is hydrogen; -C1-C3 alkyl optionally substituted with one or more substituents independently selected from -OH, -O-C1-C3 alkyl, -NH2, -NH(C1-C3 alkyl), -N(C1-C3 alkyl)2, or 4- to 7-membered saturated heterocycloalkyl; or 4- to 7-membered saturated heterocycloalkyl.

[0031] In some embodiments, W has the structure of Formula VIIe: [ka] [In the formula, X e is a halogen, and R 20 is hydrogen; -OH, -O-C1-C3 alkyl, -NH2, -NH(C1-C3 alkyl), -N(C1-C3 alkyl)2, or -C1-C3 alkyl optionally substituted with one or more substituents independently selected from 4- to 7-membered saturated heterocycloalkyl.

[0032] In some embodiments, the RAS inhibitor is a compound set forth in Table 1 or a pharmaceutically acceptable salt thereof.

[0033] In some embodiments, the RAS inhibitor is [ka] or a pharmaceutically acceptable salt thereof.

[0034] In some embodiments, the RAS inhibitor is a compound of formula IX: [ka] or a pharmaceutically acceptable salt thereof wherein A is a 6-membered heterocycloalkyl optionally substituted with methyl, —OH, or ═O; A' is a 5-6 membered saturated heterocycloalkyl or a 5-6 membered heteroaryl, each of which is optionally substituted with methyl, methoxy, or halogen; R 2 is methyl or halomethyl, R 9’ and R 9’’ are each methyl, or R 9’ and R 9’’ together form an unsubstituted saturated C3-C6 cycloalkyl, and R17 is hydrogen; -C1-C3 alkyl optionally substituted with one or more substituents independently selected from -OH, -O-C1-C3 alkyl, -NH2, -NH(C1-C3 alkyl), -N(C1-C3 alkyl)2, or 4- to 7-membered saturated cycloalkyl; or 4- to 7-membered saturated heterocycloalkyl.

[0035] In some embodiments, the RAS inhibitor is a compound of formula X: [ka] or a pharmaceutically acceptable salt thereof [Wherein A is [ka] and B is CH(R 9 )[where R 9 teeth, [ka] ] and L is [ka] and W is [ka] [ka] It is.]

[0036] In some embodiments of any of the methods described herein, the method further comprises administering to the subject an immune checkpoint inhibitor.

[0037] In another aspect, the disclosure provides a method of treating immunorefractory lung cancer in a subject by administering to the subject a RAS inhibitor, a SHP2 inhibitor, and an immune checkpoint inhibitor, wherein the RAS inhibitor: [ka] or a pharmaceutically acceptable salt thereof; The above SHP2 inhibitors [ka] or a pharmaceutically acceptable salt thereof.

[0038] In some embodiments of the methods described herein, the subject is administered an immune checkpoint inhibitor that is a PD-1 inhibitor.

[0039] In another aspect, the present disclosure provides a method of increasing susceptibility to immunorefractory lung cancer in a subject, comprising administering to the subject a RAS inhibitor of Formula I or a subformula thereof as described herein.

[0040] In some embodiments, the subject has previously been administered an immune checkpoint inhibitor. In some embodiments, the subject is refractory to treatment with an immune checkpoint inhibitor. In some embodiments, the subject has acquired resistance to treatment with an immune checkpoint inhibitor.

[0041] In some embodiments, administration of a RAS inhibitor sensitizes the cancer to treatment with an immune checkpoint inhibitor.

[0042] In some embodiments, the combination of inhibitors is administered simultaneously or sequentially, hi some embodiments, the inhibitors are administered as a single formulation or as separate formulations.

[0043] In some embodiments, the subject has one or more tumors with low tumor mutational burden. In some embodiments, the subject has one or more tumors that are microsatellite stable. In some embodiments, the subject has one or more tumors with low microsatellite instability. In some embodiments, the subject has one or more tumors with low tumor immune infiltrate.

[0044] In some embodiments, administration of a RAS inhibitor or a combination of inhibitors disclosed herein alters the tumor immune infiltrate compared to the tumor immune infiltrate in the absence of the inhibitor(s). In some embodiments, the tumor immune infiltrate comprises antigen-presenting cells, myeloid cells, or lymphoid cells. In some embodiments, administration of a RAS inhibitor or a combination of inhibitors disclosed herein alters the anti-tumor immune response compared to the tumor immune infiltrate in the absence of the inhibitor(s). In some embodiments, administration of a RAS inhibitor or a combination of inhibitors disclosed herein alters the tumor microenvironment compared to the tumor immune infiltrate in the absence of the inhibitor(s). In some embodiments, administration of a RAS inhibitor or a combination of inhibitors disclosed herein converts an immunologically cold tumor to an immunologically hot tumor. In some embodiments, the method reduces tumor size or inhibits tumor growth.

[0045] In some embodiments, the immunorefractory lung cancer is non-small cell lung cancer or small cell lung cancer.In some embodiments, the immunorefractory lung cancer comprises Ras mutation.In some embodiments, the Ras mutation is K-Ras G12C, H-Ras C12C or N-Ras G12C.In some embodiments, the Ras mutation is K-Ras G12C.

[0046] It is specifically contemplated that any limitation discussed with respect to one embodiment of the present disclosure may apply to any other embodiment of the present disclosure. Furthermore, any compound or composition of the present disclosure may be used in any method of the present disclosure, and any method of the present disclosure may be used to produce or utilize any compound or composition of the present disclosure. [Brief explanation of the drawings]

[0047] [Figure 1] Figure 1 shows the tumor immune profile of mouse syngeneic Lewis lung (eLL2) KRASWT / G12C NRAS- / - A2 tumors. The composition of eLL2 KRASWT / G12C NRAS- / - A2 tumors is represented by an average of 2.37% T cells (CD8+, CD4+, and gdT cells), 0.35% B cells (CD19+), 1.38% NK cells (NKp46+), 3.35% dendritic cells (CD11c+ / MHC IIhi), 39.72% myeloid cells (Ly6G+ and Ly6C+), 8.5% macrophages (F4 / 80+), 6.52% other CD45+ cells, and 37.79% CD45- cells. [Figure 2A] Representative immunohistochemical staining of CD8+ cells in eLL2 KRASWT / G12C NRAS- / - A2 tumors. Arrows indicate positive staining, and the scale bar represents 100 μm. [Figure 2B] Quantification of four tumors exhibiting an immune desert tumor microenvironment with an average of 0.225% cytotoxic T cells infiltrating the tumor is shown. [Figure 3A] Figure 1 shows in vivo tumor cell growth in the murine syngeneic eLL2 KRASWT / G12C NRAS- / -A2 model in mice treated with vehicle and isotype control. [Figure 3B] 1 shows in vivo tumor cell growth in the murine syngeneic eLL2 KRASWT / G12C NRAS− / −A2 model in mice treated with Compound A and isotype control. [Figure 3C]1 shows in vivo tumor cell growth in the murine syngeneic eLL2 KRASWT / G12C NRAS− / −A2 model in mice treated with RMC-4550 and isotype control. [Figure 3D] 1 shows in vivo tumor cell growth in the murine syngeneic eLL2 KRASWT / G12C NRAS− / −A2 model in mice treated with Compound A, RMC-4550, and isotype control. [Figure 3E] Figure 1 shows in vivo tumor cell growth in the murine syngeneic eLL2 KRASWT / G12C NRAS- / -A2 model in vehicle and anti-PD-1 treated mice. [Figure 3F] Figure 1 shows in vivo tumor cell growth in the murine syngeneic eLL2 KRASWT / G12C NRAS- / -A2 model in mice treated with Compound A and anti-PD-1. [Figure 3G] Figure 1 shows in vivo tumor cell growth in the murine syngeneic eLL2 KRASWT / G12C NRAS- / -A2 model in mice treated with RMC-4550 and anti-PD-1. [Figure 3H] 1 shows in vivo tumor cell growth in the murine syngeneic eLL2 KRASWT / G12C NRAS− / − A2 model in mice treated with Compound A, RMC-4550, and anti-PD-1. [Figure 3I] The percentage of tumors that increased less than 2-fold from baseline volume over time after treatment with RMC-4550, Compound A, or both is shown. [Figure 3J] Graph showing the percentage of tumors that increased less than 2-fold from baseline volume over time after treatment with RMC-4550, Compound A, anti-PD-1, or a combination thereof. [Figure 3K] Shown is the percent change in body weight over time after tumor implantation and treatment with RMC-4550, Compound A, or both. Treatment was well tolerated, as measured by body weight. [Figure 3L]Percent weight change over time after tumor implantation and treatment with RMC-4550, Compound A, anti-PD-1, or a combination thereof is shown. Treatment was well tolerated, as measured by body weight. [Figure 4A] 1 graphically depicts that Compound A, dual combinations with RMC-4550 or anti-PD-1, and triple combinations significantly increased CD8+ T cell infiltration. [Figure 4B] 1 shows that Compound A, dual combinations with RMC-4550 or anti-PD-1, and triple combinations significantly increased CD4+ T cell infiltration. [Figure 4C] 1 is a graph showing that monotherapy and combination therapy with Compound A and RMC-4550 significantly reduces Ly6G+ myeloid suppressor cells. [Figure 5A] 10 graphically depicts the increase in the percentage of granzyme B-secreting CD8+ T cells with the dual combination of Compound A and RMC-4550, or the triple combination with anti-PD-1. [Figure 5B] 1 shows that the proportion of CD107a+CD8+ T cells was increased by the dual combination of Compound A and RMC-4550, or the triple combination with anti-PD-1. [Figure 5C] 1 is a graph showing that the dual combination of Compound A and RMC-4550, or the triple combination with anti-PD-1, increased the percentage of TNFalpha+CD8+ T cells. [Figure 6A] 1 is a graphical representation of IHC quantification of T cell infiltration after 4 days of treatment with Compound A and RMC-4550, in combination with anti-PD-1, or in the triple combination, showing a significant increase in CD8+ T cells. [Figure 6B] IHC quantification of T cell infiltration after 4 days of treatment with Compound A and RMC-4550, combination with anti-PD-1, or triple combination, showing a significant increase in CD4+ T cells. DETAILED DESCRIPTION OF THE INVENTION

[0048] The present disclosure generally relates to compositions and methods for treating immunorefractory lung cancer. The disclosure is based, at least in part, on the observation that treating immunorefractory lung cancer with a compound that inhibits a mutant RAS G12C protein sensitizes the cancer to treatment with an immunotherapeutic agent. In some embodiments, the compound inhibits a RAS that has an oncogenic G12C mutation. In some embodiments, the RAS inhibitor is, for example, a covalent inhibitor that can form a covalent bond with an oncogenic RAS G12C mutation at the G12C position. In some embodiments, treatment with a RAS inhibitor sensitizes the cancer to treatment with an immune checkpoint inhibitor or an SHP2 inhibitor. In some embodiments, a compound or combination of compounds described herein is administered to a subject who has previously failed immunotherapy treatment, such as treatment with an immune checkpoint inhibitor.

[0049] Additional aspects of the present disclosure are described below.

[0050] General method The practice of the present disclosure will employ, unless otherwise indicated, conventional techniques of cell culture, molecular biology (including recombinant techniques), microbiology, cell biology, biochemistry, and immunology, which are within the skill of the art. Such techniques are described in Molecular Cloning: A Laboratory Manual, third edition (Sambrook et al., 2001); Cold Spring Harbor Press; Oligonucleotide Synthesis (P. Herdewijn, ed., 2004); Animal Cell Culture (RIFreshney), ed., 1987); CCBlackwell, eds.); Gene transfer Vectors for Mammalian Cells (JMMiller & MPCalos, eds., 1987); Current Protocols in Molecular Biology (FMAusubel et al., eds., 1987); PCR: The Polymerase Chain Reaction, (Mullis et al., eds., 1994); Current Protocols in Immunology (JEColigan et al. al., eds., 1991);Short Protocols in Molecular Biology(Wiley and Sons, 1999); Manual of Clinical Laboratory Immunology (B. Detrick, NRRose, and JDFolds eds., 2006); Immunochemical Protocols (J. Pound, ed., 2003); Lab Manual in Biochemistry: Immunology and Biotechnology (A. Nigam and A. Ayyagari, eds.2007); Immunology Methods Manual: The Comprehensive Sourcebook of Techniques (Ivan Lefkovits, ed., 1996); Using Antibodies: A Laboratory Manual (E. Harlow and D. Lane, eds., 1988); and others.

[0051] definition In this application, unless otherwise clear from the context, (i) the term "a" means "one or more," (ii) is used to mean "and / or" unless expressly indicated to refer to alternatives only or that the alternatives are mutually exclusive, but the present disclosure supports definitions that refer to alternatives only and to "and / or," (iii) the terms "comprising" and "including" are understood to encompass the itemized components or steps, whether presented by themselves or with one or more additional components or steps, and (iv) when ranges are presented, the endpoints are included.

[0052] As used herein, the term "about" is used to indicate that a value includes the standard deviation of error for the device or method being used to determine the value. In certain embodiments, the term "about" refers to a range of values ​​that is included by 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% or less in either direction (above or below) the stated value, unless otherwise stated or otherwise apparent from the context (e.g., where such number may exceed 100% of the possible values).

[0053] As used herein, the term "adjacent" in the context of describing adjacent atoms means divalent atoms that are directly joined by a covalent bond.

[0054] As used herein, "compounds of the invention" and similar terms refer to the Ras inhibitors described herein, including compounds of Formula I and subformulas thereof, and compounds in Table 1 and Table 2, as well as salts (e.g., pharmaceutically acceptable salts), solvates, hydrates, stereoisomers (including atropisomers), and tautomers thereof, whether or not explicitly stated.

[0055] Those of skill in the art will understand that certain compounds described herein can exist in one or more different isomeric (e.g., stereoisomers, geometric isomers, atropisomers, tautomers) or isotopic (e.g., one or more atoms replaced with a different isotope of that atom, such as hydrogen replaced with deuterium) forms. Unless otherwise specified, or apparent from context, the depicted structures can be understood to represent any such isomeric or isotopic forms, individually or in combination.

[0056] The compounds described herein can be asymmetric (e.g., having one or more stereocenters). All stereoisomers, such as enantiomers and diastereomers, are intended unless otherwise specified. Compounds of the present disclosure containing asymmetrically substituted carbon atoms can be isolated in optically active or racemic forms. Methods for preparing optically active forms from optically active starting materials, such as by resolution of racemic mixtures or by stereoselective synthesis, are known in the art. Many geometric isomers of olefins, C=N double bonds, and the like, can also be present in the compounds described herein, and all such stable isomers are contemplated in the present disclosure. Cis and trans geometric isomers of the compounds of the present disclosure are described and can be isolated as a mixture of isomers or as separated isomeric forms.

[0057] In some embodiments, one or more compounds described herein can exist in different tautomeric forms. Unless explicitly excluded, as is clear from the context, reference to such a compound encompasses all such tautomeric forms. In some embodiments, a tautomeric form results from the exchange of a single bond with an adjacent double bond and the accompanying migration of a proton. In certain embodiments, a tautomeric form can be a prototropic tautomer, which is an isomeric protonation state having the same empirical formula and total charge as the referenced form. Examples of moieties having prototropic tautomeric forms are ketone-enol pairs, amide-imidic acid pairs, lactam-lactim pairs, amide-imidic acid pairs, enamine-imine pairs, and cyclic forms in which protons can occupy more than one position in a heterocyclic ring system, such as 1H- and 3H-imidazole, 1H-, 2H-, and 4H-1,2,4-triazole, 1H- and 2H-isoindole, and 1H- and 2H-pyrazole. In some embodiments, the tautomeric forms can be in equilibrium or sterically locked into one form by appropriate substitution. In certain embodiments, the tautomeric forms arise from acetal interconversion.

[0058] Unless otherwise stated, structures depicted herein are also meant to include compounds that differ only in the presence of one or more isotopically enriched atoms. Exemplary isotopes that can be incorporated into the compounds of the present disclosure include: 2 H, 3 H, 11 C. 13 C. 14 C. 13 N, 15 N, 15 O. 17 O. 18 O. 32 P, 33 P, 35 S, 18 F, 36 Cl, 123 I, and 125 Isotopically labeled compounds (e.g.,3 H and 14 C) may be useful in compound or substrate tissue distribution assays. 3 H), and carbon-14 (i.e., 14 C) isotopes can be useful for their ease of preparation and detectability. Additionally, heavier isotopes, such as deuterium (i.e., 2 Substitution with, for example, H, may result in greater metabolic stability and may confer certain therapeutic advantages (e.g., longer in vivo half-life or reduced dosage requirements). In some embodiments, one or more hydrogen atoms may be 2 H or 3 H or one or more carbon atoms are replaced by 13 C or 14 It is replaced by C-enriched carbon. 15 O. 13 N, 11 C and 18 Positron-emitting isotopes, such as F, are useful in positron emission tomography (PET) studies to examine substrate receptor occupancy. The preparation of isotopically labeled compounds is known to those skilled in the art. For example, isotopically labeled compounds can generally be prepared by following procedures similar to those disclosed for the compounds of the present disclosure described herein, substituting an isotopically labeled reagent for a non-isotopically labeled reagent.

[0059] Non-limiting examples of moieties that may contain one or more deuterium substitutions in the compounds of the invention include: [ka] and any position "R" can be deuterium (D). Additional examples include: [ka] Parts such as, similar R 1 Deuteration of the R 1The definition of is found herein. Deuteration of moieties within the substituent W in the compounds of the invention, where W is defined herein (e.g., Formula I and subformulas thereof, as well as specific examples of W described herein, e.g., [ka] (See also [ka] Deuteration at any available position in any A moiety of the compounds of the formulae described herein is also contemplated, such as: [ka] may occur in the compounds of the present invention at linker positions such as:

[0060] In further embodiments, silylated substitutions are also contemplated, such as in linkers such as: [ka]

[0061] As is known in the art, many chemical compounds can exist in a variety of different solid forms, such as amorphous or crystalline forms (e.g., polymorphs, hydrates, solvates), etc. In some embodiments, the compounds of the present disclosure can be utilized in any such form, including any solid form. In some embodiments, the compounds described or illustrated herein can be provided or utilized in hydrate or solvate form.

[0062] At various places herein, substituents for compounds of the present disclosure are disclosed in groups or in ranges. It is specifically intended that the present disclosure include each and every individual subcombination of the members of such groups and ranges. For example, the term "C1-C6 alkyl" is specifically intended to individually disclose methyl, ethyl, C3 alkyl, C4 alkyl, C5 alkyl, and C6 alkyl. Furthermore, when a compound contains multiple positions where a substituent is disclosed in a group or range, unless otherwise specified, the disclosure is intended to cover individual compounds containing each and every individual subcombination of the elements at each position, as well as groups (e.g., genera and subgenera) of compounds.

[0063] The term "optionally substituted X" (e.g., optionally substituted alkyl) is intended to be equivalent to "X, where X is optionally substituted" (e.g., "alkyl, where the alkyl is optionally substituted"). The feature "X" (e.g., alkyl) itself is not intended to imply optionality. As described herein, certain compounds of interest can contain one or more "optionally substituted" moieties. Generally, the term "substituted," whether preceded by the term "optionally" or not, means that one or more hydrogens of the specified moiety are replaced with a suitable substituent, e.g., any of the substituents or groups described herein. Unless otherwise specified, an "optionally substituted" group can have a suitable substituent at every substitutable position of the group, and when more than one position in any given structure can be substituted with more than one substituent selected from a particular group, the substituents can be the same or different at all positions. For example, in the term "optionally substituted C-C alkyl-C-C heteroaryl," the alkyl portion, the heteroaryl portion, or both can be optionally substituted. Combinations of substituents contemplated by this disclosure are preferably those that result in the formation of stable or chemically feasible compounds. The term "stable," as used herein, refers to compounds that remain substantially unchanged when subjected to conditions that foresee their production, detection, and, in certain embodiments, their recovery, purification, and use for one or more of the purposes disclosed herein.

[0064] Suitable monovalent substituents at a substitutable carbon atom of an "optionally substituted" group are, independently, deuterium, halogen, -(CH2)0-4R°, -(CH2)0-4OR°, -O(CH2)0-4R o, —O—(CH2)0-4C(O)OR°, —(CH2)0-4CH(OR°)2, —(CH2)0-4SR°, —(CH2)0-4Ph [optionally substituted with R°], —(CH2)0-4O(CH2)0-1Ph [optionally substituted with R°], —CH═CHPh [optionally substituted with R°], —(CH2)0-4O(CH2)0-1-pyridyl [optionally substituted with R°]. ], 4-8 membered saturated or unsaturated heterocycloalkyl (e.g., pyridyl), 3-8 membered saturated or unsaturated cycloalkyl (e.g., cyclopropyl, cyclobutyl, or cyclopentyl), -NO2, -CN, -N3, -(CH2)0-4N(R°)2, -(CH2)0-4N(R°)C(O)R°, -N(R°)C(S)R°, -(CH2)0-4N(R °)C(O)NR°2, -N(R°)C(S)NR°2, -(CH2)0-4N(R°)C(O)OR°, -N(R°)N(R°)C(O)R°, -N(R°)N(R°)C(O)N R°2, -N(R°)N(R°)C(O)OR°, -(CH2)0-4C(O)R°, -C(S)R°, -(CH2)0-4C(O)OR°, -(CH2)0-4-C(O)-N(R o )2, -(CH2)0-4-C(O)-N(R o )-S(O)2-R o, -C(NCN)NR°2, -(CH2)0-4C(O)SR°, -(CH2)0-4C(O)OSiR°3, -(CH2)0-4OC(O)R°, -OC(O)(CH2)0-4SR°, -SC(S)SR°, -(CH2)0- 4SC(O)R°, -(CH2)0-4C(O)NR°2, -C(S)NR°2, -C(S)SR°, -(CH2)0-4OC(O)NR°2, -C(O)N(OR°)R°, -C(O)C(O)R°, -C(O)CH2C(O)R °, -C(NOR°)R°, -(CH2)0-4SSR°, -(CH2)0-4S(O)2R°, -(CH2)0-4S(O)2OR°, -(CH2)0-4OS(O)2R°, -S(O)2NR°2, -(CH2)0-4S(O) R°, -N(R°)S(O)2NR°2, -N(R°)S(O)2R°, -N(OR°)R°, -C(NOR°)NR°2, -C(NH)NR°2, -P(O)2R°, -P(O)R°2, -P(O)(OR°)2, -OP(O)R °2, -OP(O)(OR°)2, -OP(O)(OR°)R°, -SiR°3, -(C1-4 straight or branched chain alkylene)ON(R°)2, or -(C1-4 straight or branched chain alkylene)C(O)ON(R°)2, where each R° is optionally substituted as defined below and independently represents hydrogen, -C1-6 aliphatic, -CH2Ph, -O(CH2)0-1Ph, -CH2- (5-6 membered heteroaryl ring), or nitrogen, oxygen, or or a 3-6 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from sulfur, or, notwithstanding the above definitions, two independently occurring R° together with the atom(s) between them form a 3-12 membered saturated, partially unsaturated, or aryl mono- or bicyclic ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, which may be substituted as defined below.

[0065] Suitable monovalent substituents on R° (or a ring formed by taking two independently occurring R° together with the atoms between them) are independently halogen, —(CH)O-R ● ,-(Halo R ●), -(CH2)0-2OH, -(CH2)0-2OR ● , -(CH2)0-2CH(OR ● )2;-O(HaloR ● ), -CN, -N3, -(CH2)0-2C(O)R ● , -(CH2)0-2C(O)OH, -(CH2)0-2C(O)OR ● , -(CH2)0-2SR ● , -(CH2)0-2SH, -(CH2)0-2NH2, -(CH2)0-2NHR ● , -(CH2)0-2NR ● 2, -NO2, -SiR ● 3. -OSiR ● 3. -C(O)SR ● , -(C1-4 straight or branched chain alkylene)C(O)OR ● , or -SSR ● wherein each R ● is unsubstituted or, where preceded by "halo," is substituted only with one or more halogens, independently selected from C aliphatic, -CHPh, -O(CH)Ph, or a 5-6 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. Suitable divalent substituents on a saturated carbon atom of R include =0 and =S.

[0066] Suitable divalent substituents on a saturated carbon atom of an "optionally substituted" group include the following: =O, =S, =NNR * 2, =NNHC(O)R * , =NNHC(O)OR * , =NNHS(O)2R * , =NR * , =NOR * , -O(C(R * 2))2-3O-, or -S(C(R * 2) 2-3S-, wherein each R *is selected from hydrogen, a C1-6 aliphatic, which may be substituted as defined below, or an unsubstituted 5-6 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. Preferred divalent substituents attached to a vicinal substitutable carbon of an "optionally substituted" group include -O(CR * 2) 2-3O-, wherein each R * is selected from hydrogen, a C1-6 aliphatic which may be substituted as defined below, or an unsubstituted 5-6 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.

[0067] R * Suitable substituents on the aliphatic group include -R ● ,-(Halo R ● ), -OH, -OR ● , -O(HaloR ● ), -CN, -C(O)OH, -C(O)OR ● , -NH2, -NHR ● , -NR ● 2, or -NO2, wherein each R ● is unsubstituted or, where preceded by "halo", substituted only with one or more halogens and is independently a C1-4 aliphatic, -CH2Ph, -O(CH2)0-1Ph, or a 5-6 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.

[0068] Suitable substituents on a substitutable nitrogen of an "optionally substituted" group include -R † , -NR † 2. -C(O)R † , -C(O)OR † , -C(O)C(O)R † , -C(O)CHC(O)R † , -S(O)2R † , -S(O)NR † 2. -C(S)NR †2. -C(NH)NR † 2, or -N(R † )S(O)2R † wherein each R † are independently hydrogen, a C1-6 aliphatic, unsubstituted -OPh which may be substituted as defined below, or an unsubstituted 3-6 membered, saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur; or, regardless of the above definition, two independently occurring R † together with the intervening atom(s), form an unsubstituted 3-12 membered saturated, partially unsaturated, or aryl mono- or bicyclic ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.

[0069] R † Suitable substituents on the aliphatic group are independently halogen, —R ● ,-(Halo R ● ), -OH, -OR ● , -O(HaloR ● ), -CN, -C(O)OH, -C(O)OR ● , -NH2, -NHR ● , -NR ● 2, or -NO2, where each R ● is unsubstituted or, where preceded by "halo", substituted only with one or more halogens, and is independently a C1-4 aliphatic, -CH2Ph, -O(CH2)0-1Ph, or a 5-6 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. R † Suitable divalent substituents on a saturated carbon atom of include ═O and ═S.

[0070] Those of skill in the art reading this disclosure will understand that certain compounds described herein can be provided or utilized in any of a variety of forms, such as, for example, salt forms, protected forms, prodrug forms, ester forms, isomeric forms (e.g., optical or structural isomers), isotopic forms, etc. In some embodiments, reference to a particular compound may refer to a particular form of that compound. In some embodiments, reference to a particular compound may refer to any form of that compound. In some embodiments, for example, a preparation of a single stereoisomer of a compound may be considered a form of that compound that is different from a racemic mixture of that compound, a particular salt of a compound may be considered a form that is different from another salt form of that compound, a preparation containing a structural isomer of a double bond ((Z) or (E)) may be considered a form that is different from one containing the other structural isomer of that double bond ((E) or (Z)), and a preparation in which one or more atoms are isotopically different from those present in a reference preparation may be considered a different form.

[0071] As used herein, the term "administration" refers to the administration of a composition (e.g., a compound or a preparation comprising a compound described herein) to a subject or system. Administration also includes administering to a subject a prodrug derivative or analog of a compound or a pharmaceutically acceptable salt of a compound or composition, which can form an equivalent amount of the active compound in the subject's body. Administration to an animal subject (e.g., a human) can be by any suitable route. For example, in some embodiments, administration can be bronchial (including bronchial infusion), buccal, enteral, interdermal, intraarterial, intradermal, intragastric, intramedullary, intramuscular, nasal, intraperitoneal, intrathecal, intravenous, intraventricular, mucosal, intranasal, oral, rectal, subcutaneous, sublingual, topical, intratracheal (including intratracheal infusion), transdermal, intravaginal, or intravitreal.

[0072] As used herein, the term "acetyl" refers to the group -C(O)CH3.

[0073] As used herein, the term "alkoxy" refers to an -O-C20 It means that the alkyl and alkoxy groups are attached to the remainder of the compound through an oxygen atom.

[0074] As used herein, the term "alkyl" refers to a saturated, straight- or branched-chain monovalent hydrocarbon group containing 1 to 20 (e.g., 1 to 10, or 1 to 6) carbons. In some embodiments, alkyl groups are unbranched (i.e., straight-chain), and in some embodiments, alkyl groups are branched. Alkyl groups are exemplified by, but not limited to, methyl, ethyl, n- and isopropyl, n-, sec-, iso- and tert-butyl, and neopentyl.

[0075] As used herein, the term "alkylene" refers to a saturated divalent hydrocarbon group derived from a straight or branched chain saturated hydrocarbon by removing two hydrogen atoms, and is exemplified by methylene, ethylene, isopropylene, and the like. x -C y "Alkylene" refers to an alkylene group having x to y carbons. Exemplary values ​​of x are 1, 2, 3, 4, 5, and 6, and exemplary values ​​of y are 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, or 20 (e.g., C1-C6, C1-C6). 10 , C2-C 20 , C2-C6, C2-C 10 , or C2-C 20 In some embodiments, the alkylene can be further substituted with 1, 2, 3, or 4 substituents, as defined herein.

[0076] As used herein, the term "alkenyl," unless otherwise specified, represents a monovalent straight- or branched-chain group of 2 to 20 carbons (e.g., 2 to 6, or 2 to 10 carbons) containing one or more carbon-carbon double bonds, and is exemplified by ethenyl, 1-propenyl, 2-propenyl, 2-methyl-1-propenyl, 1-butenyl, and 2-butenyl. Alkenyl includes both cis and trans isomers. As used herein, the term "alkenylene," unless otherwise specified, represents a divalent straight- or branched-chain group of 2 to 20 carbons (e.g., 2 to 6, or 2 to 10 carbons) containing one or more carbon-carbon double bonds.

[0077] As used herein, the term "alkynyl" refers to a monovalent straight or branched chain group of 2 to 20 carbon atoms (e.g., 2 to 4, 2 to 6, or 2 to 10 carbons) containing a carbon-carbon triple bond and is exemplified by ethynyl and 1-propynyl.

[0078] As used herein, the term "alkynyl sulfone" refers to a group having the structure [ka] wherein R is any suitable chemical substituent as described herein.

[0079] As used herein, the term "amino" refers to -N(R † )2, for example, -NH2 and -N(CH3)2.

[0080] As used herein, the term "aminoalkyl" refers to an alkyl moiety substituted on one or more carbon atoms with one or more amino moieties.

[0081] As used herein, the term "amino acid" refers to a molecule having a side chain, an amino group, and an acidic group (e.g., -COH or -SOH), where the amino acid is attached to the parent molecular group by the side chain, the amino group, or the acidic group (e.g., the side chain). As used herein, the term "amino acid" in its broadest sense refers to any compound or substance that can be incorporated into a polypeptide chain, for example, by the formation of one or more peptide bonds. In some embodiments, an amino acid has the general structure HN-C(H)(R)-COOH. In some embodiments, an amino acid is a naturally occurring amino acid. In some embodiments, an amino acid is a synthetic amino acid, in some embodiments, an amino acid is a D-amino acid, and in some embodiments, an amino acid is an L-amino acid. A "standard amino acid" refers to any of the 20 standard L-amino acids commonly found in naturally occurring peptides. Exemplary amino acids include alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, optionally substituted hydroxylnorvaline, isoleucine, leucine, lysine, methionine, norvaline, ornithine, phenylalanine, proline, pyrrolysine, selenocysteine, serine, taurine, threonine, tryptophan, tyrosine, and valine.

[0082] As used herein, "amino acid substitution" refers to the replacement of a wild-type amino acid in a protein with a non-wild-type amino acid. Amino acid substitutions can occur through genetic mutation and can alter one or more properties of a protein (e.g., can confer altered binding affinity or specificity, altered enzymatic activity, altered structure, or altered function).

[0083] As used herein, the term "aryl" refers to a monovalent monocyclic, bicyclic, or polycyclic ring system formed by carbon atoms, wherein the ring attached to the pendant group is aromatic. Examples of aryl groups are phenyl, naphthyl, phenanthrenyl, and anthracenyl. An aryl ring can be attached to its pendant group at any heteroatom or carbon ring atom that results in a stable structure, and unless otherwise specified, any of the ring atoms can be optionally substituted.

[0084] As used herein, the term "C" represents a bond. For example, part of the term -N(C(O)-(C-C alkylene-H)- includes -N(C(O)-(C alkylene-H)-, which is also represented by -N(C(O)-H)-.

[0085] As used herein, the terms "carbocyclic" and "carbocyclyl" refer to a monovalent, optionally substituted C- 12 "Carbocyclic" refers to a monocyclic, bicyclic, or tricyclic ring structure, which may be optionally bridged, fused, or spirocyclic, in which all rings are formed by carbon atoms and at least one ring is non-aromatic. Carbocyclic structures include cycloalkyl, cycloalkenyl, and cycloalkynyl groups. Examples of carbocyclyl groups are cyclohexyl, cyclohexenyl, cyclooctynyl, 1,2-dihydronaphthyl, 1,2,3,4-tetrahydronaphthyl, fluorenyl, indenyl, indanyl, decalinyl, and the like. A carbocyclic ring can be attached to its pendant group at any ring atom that results in a stable structure, and unless otherwise specified, any of the ring atoms can be optionally substituted.

[0086] As used herein, the term "carbonyl" refers to a C(O) group, which can also be represented as C=O.

[0087] As used herein, the term "carboxyl" refers to -CO2H, (C=O)(OH), COOH, or C(O)OH, or the unprotonated corresponding groups.

[0088] The term "combination therapy" refers to a method of treatment that includes administering to a subject at least two therapeutic agents, optionally in one or more pharmaceutical compositions, as part of a treatment regimen. For example, combination therapy can include administration of a single pharmaceutical composition containing at least two therapeutic agents and one or more pharmaceutically acceptable carriers, excipients, diluents, or surfactants. Combination therapy can include administration of two or more pharmaceutical compositions, each containing one or more therapeutic agents and one or more pharmaceutically acceptable carriers, excipients, diluents, or surfactants. The two or more agents can optionally be administered simultaneously (as a single or separate composition) or sequentially (as separate compositions). The therapeutic agents can be administered in effective amounts. The therapeutic agents can be administered in therapeutically effective amounts. In some embodiments, the effective amount of one or more therapeutic agents can be less when used in combination therapy than the therapeutic amount of the same therapeutic agent when used as a monotherapy, for example, due to an additive or synergistic effect of combining two or more therapeutic agents.

[0089] As used herein, the term "cyano" refers to a -CN group.

[0090] As used herein, the term "cycloalkyl" refers to a monovalent saturated cyclic hydrocarbon group, which, unless otherwise specified, may be bridged, fused, or spirocyclic having 3 to 8 carbons, and is exemplified by cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cycloheptyl.

[0091] As used herein, the term "cycloalkenyl" refers to a monovalent non-aromatic saturated cyclic hydrocarbon group, which, unless otherwise specified, may be bridged, fused, or spirocyclic having 3 to 8 carbons and containing one or more carbon-carbon double bonds.

[0092] As used herein, the term "diastereomers" means stereoisomers that are not mirror images of each other and are not superimposable with respect to one another.

[0093] As used herein, the term "dosage form" refers to a physically discrete unit of a compound (e.g., a compound of the present disclosure) for administration to a subject. Each unit contains a predetermined amount of the compound. In some embodiments, such amount is a unit dose (or a whole fraction thereof) appropriate for administration according to a dosing regimen (i.e., using a therapeutic dosing regimen) that has been determined to correlate with a desired or beneficial outcome when administered to a relevant population. Those skilled in the art will understand that the total amount of a therapeutic composition or compound administered to a particular subject will be determined by one or more attending physicians and may involve the administration of multiple dosage forms.

[0094] As used herein, the term "dosing regimen" refers to a collection of unit doses (usually two or more) administered individually to a subject, usually separated by a period of time. In some embodiments, a given therapeutic compound (e.g., a compound of the present disclosure) has a recommended dosing regimen, which may have one or more doses. In some embodiments, a dosing regimen includes multiple doses, each separated from the other by the same length of time period; in some embodiments, a dosing regimen includes multiple doses and at least two different time periods separating the individual doses. In some embodiments, all doses within a dosing regimen are the same unit dose. In some embodiments, different doses within a dosing regimen are different amounts. In some embodiments, a dosing regimen includes a first dose at a first dosage amount, followed by one or more additional doses at a second dosage amount that is different from the first dosage amount. In some embodiments, a dosing regimen includes a first dose at a first dosage amount, followed by one or more additional doses at a second dosage amount that is the same as the first dosage amount. In some embodiments, the dosing regimen correlates with a desired or beneficial outcome when administered across relevant populations (ie, is a therapeutic dosing regimen).

[0095] The term "disorder" is used in this disclosure to mean, and is used interchangeably with, the terms disease, condition, or illness, unless otherwise indicated.

[0096] As used herein, "enantiomer" means each individual optically active form of a compound of the invention having an optical purity or enantiomeric excess (as measured by standard methods in the art) of at least 80% (i.e., at least 90% of one enantiomer and up to 10% of the other enantiomer), preferably at least 90%, and more preferably at least 98%.

[0097] The term "guanidyl" means a compound having the structure [ka] wherein each R is independently any chemically suitable substituent described herein.

[0098] As used herein, the term "guanidinoalkylalkyl" refers to an alkyl moiety substituted on one or more carbon atoms with one or more guanidyl moieties.

[0099] As used herein, the term "haloacetyl" means an acetyl group in which at least one hydrogen has been replaced by a halogen.

[0100] As used herein, the term "haloalkyl" refers to an alkyl moiety substituted on one or more carbon atoms with one or more of the same or different halogen moieties.

[0101] As used herein, the term "halogen" refers to a halogen selected from bromine, chlorine, iodine, or fluorine.

[0102] As used herein, the term "heteroalkyl" refers to an "alkyl" group (as defined herein) in which at least one carbon atom has been replaced with a heteroatom (e.g., an O, N, or S atom). The heteroatom can occur at the center or end of the radical.

[0103] As used herein, the term "heteroaryl" refers to a monovalent monocyclic or polycyclic ring system containing at least one complete aromatic ring. That is, they contain 4n+2 pi electrons in the monocyclic or polycyclic ring system and at least one ring heteroatom selected from N, O, or S in the aromatic ring. Exemplary unsubstituted heteroaryl groups are those of 1 to 12 (e.g., 1 to 11, 1 to 10, 1 to 9, 2 to 12, 2 to 11, 2 to 10, or 2 to 9) carbons. The term "heteroaryl" includes bicyclic, tricyclic, and tetracyclic groups in which any of the above heteroaromatic rings are fused to one or more aryl or carbocyclic rings, such as phenyl or cyclohexane rings. Examples of heteroaryl groups include, but are not limited to, pyridyl, pyrazolyl, benzoxazolyl, benzimidazolyl, benzothiazolyl, imidazolyl, thiazolyl, quinolinyl, tetrahydroquinolinyl, and 4-azaindolyl. A heteroaryl ring can be attached to its pendant group at any ring atom that results in a stable structure, and unless otherwise specified, any ring atom can be optionally substituted. In some embodiments, a heteroaryl is substituted with 1, 2, 3, or 4 substituents.

[0104] As used herein, the term "heterocycloalkyl" refers to a monovalent monocyclic, bicyclic, or polycyclic ring system, which may be bridged, fused, or spirocyclic, in which at least one ring is non-aromatic and the non-aromatic ring contains 1, 2, 3, or 4 heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur. Five-membered rings have 0 to 2 double bonds, and six- and seven-membered rings have 0 to 3 double bonds. Exemplary unsubstituted heterocycloalkyl groups are those of 1 to 12 (e.g., 1 to 11, 1 to 10, 1 to 9, 2 to 12, 2 to 11, 2 to 10, or 2 to 9) carbons. The term "heterocycloalkyl" also refers to heterocyclic compounds having bridged polycyclic structures in which one or more carbons or heteroatoms bridge two non-adjacent members of a monocyclic ring, such as a quinuclidinyl group. The term "heterocycloalkyl" includes bicyclic, tricyclic, and tetracyclic groups in which any of the above heterocyclic rings is fused to one or more aromatic, carbocyclic, heteroaromatic, or heterocyclic rings, such as an aryl ring, a cyclohexane ring, a cyclohexene ring, a cyclopentane ring, a cyclopentene ring, a pyridine ring, or a pyrrolidine ring. Examples of heterocycloalkyl groups are pyrrolidinyl, piperidinyl, 1,2,3,4-tetrahydroquinolinyl, decahydroquinolinyl, dihydropyrrolopyridine, and decahydronaphthyridinyl. A heterocycloalkyl ring can be attached to its pendant group at any ring atom that results in a stable structure, and unless otherwise specified, any of the ring atoms can be optionally substituted.

[0105] As used herein, the term "hydroxy" refers to an --OH group.

[0106] As used herein, the term "hydroxyalkyl" refers to an alkyl moiety substituted on one or more carbon atoms with one or more --OH moieties.

[0107] As used herein, "isomer" refers to any tautomer, stereoisomer, atropisomer, enantiomer, or diastereomer of any compound of the invention. It is recognized that the compounds of the invention can have one or more chiral centers or double bonds and therefore exist as stereoisomers, e.g., double bond isomers (i.e., geometric E / Z isomers), or diastereomers (e.g., enantiomers (i.e., (+) or (-), or cis / trans isomers)). In accordance with the present invention, the chemical structures depicted herein, and hence the compounds of the invention, may be present in all their corresponding stereoisomers, i.e., stereomerically pure forms (e.g., geometrically pure, enantiomerically pure, or diastereomeric pure), as well as mirror image isomers. This includes both isomeric and stereoisomeric mixtures (e.g., racemates). Enantiomeric and stereoisomeric mixtures of the compounds of the invention can usually be resolved into their component enantiomers or stereoisomers by well-known methods, such as chiral-phase gas chromatography, chiral-phase high-performance liquid chromatography, crystallization of the compound as a chiral salt complex, or crystallization of the compound in a chiral solvent. Enantiomers and stereoisomers can also be obtained from stereomerically or enantiomerically pure intermediates, reagents, and catalysts by well-known asymmetric synthetic methods.

[0108] As used interchangeably herein, the terms "immunorefractory," "immune escape," or "cold tumor" refer to tumors, cancers, or patients with tumors or cancers for which prior immunotherapy, such as immune checkpoint inhibitors, has proven ineffective or intolerable. For example, patients with immunorefractory cancer include patients who have previously been treated with immunotherapy, such as immune checkpoint inhibitors, but the immunotherapy has proven ineffective or not sufficiently effective in slowing or stopping disease progression or alleviating symptoms associated with disease progression. Immunorefractory cancers include cancers that have become resistant to or have reduced sensitivity to immunotherapy treatment (e.g., the effectiveness of immunotherapy, such as immune checkpoint inhibitors, previously administered to the patient has diminished over time). Immunorefractory cancers can be identified by methods known to those skilled in the art or described herein. For example, immunorefractory cancers can be characterized by low immune cell infiltration in one or more tumors. Low immune cell infiltration may include a reduction or absence of lymphocytes, a reduction or absence of tumor-infiltrating lymphocytes (TILs), a reduction or absence of dendritic cells, a reduction or absence of myeloid cells, a reduction or absence of natural killer (NK) cells, a reduction or absence of macrophages, a reduction or absence of T cells, a reduction or absence of CD8+ T cells, a reduction or absence of CD4+ T cells, or a reduction or absence of CD4+ / CD8+ T cells. See, e.g., Chen and Mellman, Nature, 541:321 (2017). In contrast, in some embodiments, a "hot tumor" refers to a tumor, cancer, or a patient with a tumor or cancer that is not immune refractory. A cancer or tumor with a low number of cytotoxic T cells can be characterized as "immune desert." In some embodiments, a cancer or tumor with a cytotoxic T cell count of less than 1% of viable cells is considered "immune desert." In some embodiments, cancers or tumors in which the number of cytotoxic T cells is less than 0.5% of viable cells are considered "immune desert."In some embodiments, cancers or tumors in which the number of cytotoxic T cells is less than 0.25% of viable cells are considered "immune desert."

[0109] As used herein, the term "inhibitor" refers to a compound that prevents a biomolecule (e.g., a protein, a nucleic acid) from completing or initiating a reaction. An inhibitor can inhibit a reaction, for example, by competitive, uncompetitive, or noncompetitive means. With respect to its binding mechanism, an inhibitor can be an irreversible inhibitor or a reversible inhibitor. Exemplary inhibitors include, but are not limited to, nucleic acids; DNA; RNA; shRNA; siRNA; proteins; protein mimetics; peptides; peptidomimetics; antibodies; small molecules; chemicals; and analogs that mimic the binding site of an enzyme, receptor, or other protein. In some embodiments, the inhibitor is a small molecule, e.g., a low molecular weight organic compound, e.g., an organic compound having a molecular weight (MW) of less than 1200 daltons (Da). In some embodiments, the MW is less than 1100 Da. In some embodiments, the MW is less than 1000 Da. In some embodiments, the MW is less than 900 Da. In some embodiments, the MW of the small molecule ranges from 800 Da to 1200 Da. Small molecule inhibitors include cyclic and acyclic compounds. Small molecule inhibitors include natural products and their derivatives and analogs. Small molecule inhibitors can include, for example, covalent cross-linking groups that can form covalent cross-links with amino acid side chains of target proteins.

[0110] As used herein, the term "linker" refers to a divalent organic moiety that connects a first moiety (e.g., a macrocyclic moiety) to a second moiety (e.g., a bridging group). In some embodiments, the linker results in a compound that is capable of achieving an IC50 of 2 uM or less in the Ras-RAF disruption assay protocol provided herein.

[0111] The purpose of this biochemical assay is to measure the ability of a test compound to facilitate ternary complex formation between a nucleotide-loaded Ras isoform and cyclophilin A, and the resulting ternary complex is a BRAF RBD disrupting binding to the construct and inhibiting Ras signaling through RAF effectors.

[0112] In an assay buffer containing 25 mM HEPES (pH 7.3), 0.002% Tween 20, 0.1% BSA, 100 mM NaCl, and 5 mM MgCl, untagged cyclophilin A, His6-K-RasGMPPNP (or other Ras variants), and GST-BRAF RBD are combined in a 384-well assay plate at final concentrations of 25 μM, 12.5 nM, and 50 nM, respectively. Compounds are present in the plate wells as a 10-point, 3-fold dilution series starting at a final concentration of 30 μM. After 3 hours of incubation at 25°C, a mixture of anti-His Eu-W1024 and anti-GST allophycocyanin was added to the assay sample wells at final concentrations of 10 nM and 50 nM, respectively, and the reactions were incubated for an additional 1.5 hours. TR-FRET signals were read in a microplate reader (excitation 320 nm, emission 665 / 615 nm). Compounds that promote disruption of the Ras:RAF complex are identified as those that induce a decrease in the TR-FRET ratio relative to DMSO control wells.

[0113] In some embodiments, the linker comprises 20 or fewer linear chain atoms. In some embodiments, the linker comprises 15 or fewer linear chain atoms. In some embodiments, the linker comprises 10 or fewer linear chain atoms. In some embodiments, the linker has a molecular weight of less than 500 g / mol. In some embodiments, the linker has a molecular weight of less than 400 g / mol. In some embodiments, the linker has a molecular weight of less than 300 g / mol. In some embodiments, the linker has a molecular weight of less than 200 g / mol. In some embodiments, the linker has a molecular weight of less than 100 g / mol. In some embodiments, the linker has a molecular weight of less than 50 g / mol.

[0114] As used herein, the term "mutation" refers to any modification of a nucleic acid or polypeptide that results in a change in the nucleic acid or polypeptide. The term "mutation" can include, for example, point mutations, deletions, or insertions of single or multiple residues in a polynucleotide, and includes changes that occur within the protein-coding region of a gene, as well as changes in regions outside the protein-coding region, such as, but not limited to, regulatory or promoter sequences, as well as amplification, or chromosomal disruption or translocation. In certain embodiments, the mutation results in an amino acid substitution in the encoded protein.

[0115] A "patient" or "subject" is a mammal, e.g., a human, mouse, rat, guinea pig, dog, cat, horse, cow, pig, or non-human primate, e.g., a monkey, chimpanzee, baboon, or rhesus monkey.

[0116] The term "prevent" or "prevention" in relation to a subject refers to preventing the subject from suffering from a disease or disorder. Prevention includes prophylactic treatment. For example, prevention can include administering a compound disclosed herein to a subject before the subject suffers from a disease, which administration prevents the subject from suffering from the disease.

[0117] As used herein, the term "pharmaceutical composition" means a compound, such as a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, formulated together with a pharmaceutically acceptable excipient.

[0118] As used herein, "pharmaceutically acceptable excipient" refers to any inactive ingredient (e.g., a vehicle capable of suspending or dissolving an active compound) that is non-toxic and non-inflammatory in a subject. Typical excipients include, for example, anti-adhesive agents, antioxidants, binders, coating agents, compression aids, disintegrants, dyes (coloring agents), emollients, emulsifiers, fillers (diluents), film-forming or coating agents, flavorings, fragrances, glidants (flow improvers), lubricants, preservatives, printing inks, adsorbents, suspending or dispersing agents, sweeteners, or hydration water. Excipients include, but are not limited to, optionally substituted butylated hydroxyl toluene (BHT), calcium carbonate, calcium phosphate (dibasic), calcium stearate, croscarmellose, cross-linked polyvinylpyrrolidone, citric acid, crospovidone, cysteine, ethylcellulose, gelatin, optionally substituted hydroxyl propyl cellulose, optionally substituted hydroxyl propyl methylcellulose, lactose, magnesium stearate, maltitol, mannitol, methionine, methylcellulose, methylparaben, microcrystalline cellulose, polyethylene glycol, polyvinylpyrrolidone, povidone, pregelatinized starch, propylparaben, retinyl palmitate, shellac, silicon dioxide, sodium carboxymethylcellulose, sodium citrate, sodium starch glycolate, sorbitol, starch (corn), stearic acid, stearic acid, sucrose, talc, titanium dioxide, vitamin A, vitamin E, vitamin C, and xylitol. Those skilled in the art are familiar with the variety of agents and materials useful as excipients.See, e.g., Ansel, et al., Ansel's Pharmaceutical Dosage Forms and Drug Delivery Systems. Philadelphia: Lippincott, Williams & Wilkins, 2004; Gennaro, et al., Remington: The Science and Practice of Pharmacy. Philadelphia: Lippincott, Williams & Wilkins, 2000; and Rowe, Handbook of Pharmaceutical Excipients. Chicago, Pharmaceutical Press, 2005. In some embodiments, the composition comprises at least two different pharmaceutically acceptable excipients.

[0119] As used herein, the term "pharmaceutically acceptable salt" refers to those salts of the compounds described herein that are suitable for use in contact with the tissues of humans and other animals without undue toxicity, irritation, allergic response, etc., within the normal scope of sound medical judgment, and commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, pharmaceutically acceptable salts are described in Berge et al., J. Pharmaceutical Sciences 66:1-19, 1977 and Pharmaceutical Salts: Properties, Selection, and Use, (Eds. P.H. Stahl and C.G. Wermuth), Wiley-VCH, 2008. Salts can be prepared in situ during the final isolation and purification of the compounds described herein, or separately by reacting the free base group with a suitable organic acid.

[0120] The terms "RAS inhibitor" and "inhibitor of RAS" are used interchangeably and refer to any inhibitor that targets, i.e., selectively binds to or selectively inhibits, the RAS protein.

[0121] As used herein, the term "RAS(ON) inhibitor" refers to an inhibitor that targets, i.e., selectively binds to, or inhibits, the GTP-bound, activated state of RAS (e.g., more selectively than the GDP-bound, inactive state of RAS). Inhibition of the GTP-bound, activated state of RAS includes, for example, inhibition of oncogenic signaling from the GTP-bound, activated state of RAS. In some embodiments, a RAS(ON) inhibitor is an inhibitor that selectively binds to and inhibits the GTP-bound, activated state of RAS. In certain embodiments, a RAS(ON) inhibitor can also bind to or inhibit the GDP-bound, inactive state of RAS (e.g., with a lower affinity or inhibition constant than the GTP-bound, activated state of RAS). The RAS(ON) inhibitor may be a tri-complex RAS(ON) inhibitor having a mechanism of action that requires the formation of a high-affinity ternary complex between a synthetic ligand (the RAS(ON) inhibitor) and two intracellular proteins that do not interact under normal physiological conditions: the intended target protein RAS, and cyclophilin A, a cytosolic chaperone protein that is ubiquitously expressed in cells. See, e.g., WO2021091982. The RAS inhibitors of Formula 0 and Formula I and their sub-formulas herein are tri-complex RAS(ON) inhibitors.

[0122] As used herein, the term "RAS(OFF) inhibitor" refers to an inhibitor that targets, i.e., selectively binds to or inhibits, the GDP-bound inactive state of RAS (e.g., more selectively than the GTP-bound active state of RAS). RAS(OFF) inhibitors are known in the art. Non-limiting examples of RAS(OFF) inhibitors include ARS-853, ARS-1620, ERAS-3490, JAB-21822, IBI351 / GFH-925, JDQ443, D-1553, GDC-6036, AMG510, and MRTX849.

[0123] The terms "RAS pathway" and "RAS / MAPK pathway" are used interchangeably herein to refer to a signal transduction cascade downstream of various cell surface growth factor receptors, in which activation of RAS (and its various isoforms and allotypes) is a central event driving various cellular effector events that determine cell proliferation, activation, differentiation, mobilization, and other functional properties. SHP2 delivers positive signals from growth factor receptors to the RAS activation / deactivation cycle, which is regulated by guanine nucleotide exchange factors (GEFs, such as SOS1) that load GTP onto RAS to produce functionally active, GTP-bound RAS, as well as GTP-activating proteins (GAPs, such as NF1) that promote signal termination by converting GTP to GDP. The GTP-bound RAS produced by this cycle delivers essential positive signals to a series of serine / threonine kinases, including RAFs and MAP kinases, from which further signals propagate to various cellular effector functions.

[0124] As used herein, the term "resistant to therapy" refers to the treatment of a disorder with a therapeutic agent when the therapeutic agent is ineffective or when the therapeutic agent was previously effective but becomes less effective over time. Resistance to therapy includes acquired resistance to therapy, which refers to a decrease in the effectiveness of a treatment over a period of time when the therapeutic agent is administered to a subject. Acquired resistance to therapy may result from the acquisition of mutations in the target protein that render the treatment ineffective or less effective. Thus, resistance to therapy may persist even after the administration of a therapeutic agent has ceased. In particular, cancers may become resistant to immune checkpoint inhibitor treatment after treatment with an immune checkpoint inhibitor. Such cancers are also referred to herein as "immune refractory." Measuring the decrease in therapeutic effectiveness depends on the disorder being treated, and such methods are known to those skilled in the art. For example, the effectiveness of cancer treatment can be measured by disease progression. Effective treatment can slow or halt disease progression. Cancers that are resistant to treatment with therapeutic agents, such as immune checkpoint inhibitors, may fail to slow or halt the progression of the disease.

[0125] As used herein, the term "stereoisomer" refers to all possible different isomeric and conformational forms that a compound (e.g., a compound of any formula described herein) may possess, in particular all possible stereochemical and conformational isomeric forms, all diastereomers, enantiomers, or conformational isomers of a basic molecular structure, e.g., atropisomers. Some compounds of the present invention can exist in different tautomeric forms, all of the latter being included within the scope of the present invention.

[0126] As used herein, the term "sulfonyl" refers to the group -S(O)2-.

[0127] A "therapeutic agent" is any substance, e.g., a compound or composition, capable of treating a disease or disorder. In some embodiments, therapeutic agents useful in combination with the present disclosure include RAS inhibitors and cancer chemotherapy. Many such therapeutic agents are known in the art and are disclosed herein.

[0128] The term "therapeutically effective amount" refers to an amount sufficient to treat a disease, disorder, or condition when administered to a population afflicted with or suspected of having the disease, disorder, or condition in accordance with a therapeutic dosing regimen. In some embodiments, a therapeutically effective amount is an amount that reduces the occurrence or severity of one or more symptoms of the disease, disorder, or condition, or delays the onset of one or more symptoms of the disease, disorder, or condition. Those skilled in the art will appreciate that the term "therapeutically effective amount" does not, in fact, require that successful treatment be achieved in any particular individual. Rather, a therapeutically effective amount can be an amount that, when administered to subjects in need of such treatment, results in a specific, desired pharmacological response in a significant number of subjects. It is specifically understood that certain subjects may, in fact, be "refractory" to a "therapeutically effective amount." In some embodiments, reference to a therapeutically effective amount can be a reference to the amount measured in one or more specific tissues (e.g., tissues affected by a disease, disorder, or condition) or fluids (e.g., blood, saliva, serum, sweat, tears, urine). Those skilled in the art will appreciate that in some embodiments, a therapeutically effective amount can be formulated or administered in a single dose, hi some embodiments, a therapeutically effective amount can be formulated or administered in multiple doses, for example, as part of a dosing regimen.

[0129] As used herein, the term "thiocarbonyl" refers to a -C(S)- group.

[0130] The term "treatment" (plus "treat" or "treating"), in its broadest sense, refers to any administration of a substance (e.g., a compound of the present disclosure) that partially or completely alleviates, improves, reduces, or inhibits a particular disease, disorder, or condition; partially or completely delays the onset of a particular disease, disorder, or condition; partially or completely reduces the severity of a particular disease, disorder, or condition; or partially or completely reduces the occurrence of one or more symptoms, characteristics, or causes of a particular disease, disorder, or condition. In some embodiments, such treatment can be administered to a subject who does not exhibit signs of the associated disease, disorder, or condition, or who exhibits only early signs of a disease, disorder, or condition. Alternatively, or in addition, in some embodiments, treatment can be administered to a subject who exhibits established signs of one or more of the associated diseases, disorders, or conditions. In some embodiments, treatment can be in a subject who has been diagnosed as suffering from the associated disease, disorder, or condition. In some embodiments, treatment may be in a subject known to have one or more susceptibility factors that are statistically correlated with an increased risk of developing the relevant disease, disorder or condition.

[0131] As used herein, the term "vinyl ketone" refers to a group containing a carbonyl group attached directly to a carbon-carbon double bond.

[0132] As used herein, the term "vinyl sulfone" refers to a group containing a sulfonyl group attached directly to a carbon-carbon double bond.

[0133] The term "wild-type" refers to an entity having a structure or activity that is found in a "normal" (as opposed to a mutant, diseased, altered, etc.) state or context in nature. Those skilled in the art will understand that wild-type genes and polypeptides often exist in multiple alternative forms (e.g., alleles).

[0134] As used herein, the term "ynone" refers to a compound having the structure [ka] where R is any suitable chemical substituent as described herein.

[0135] RAS inhibitors Provided herein are compounds that inhibit RAS, and uses thereof. Also provided are pharmaceutical compositions comprising one or more RAS inhibitor compounds, or pharmaceutically acceptable salts thereof, and a pharmaceutically acceptable excipient. RAS inhibitor compounds can be used in the methods of inhibiting RAS (e.g., in a subject or cell) and treating cancer described herein. In some embodiments, the compounds of the present disclosure are or act as prodrugs, e.g., for administration to a cell or a subject in need thereof.

[0136] RAS proteins (KRAS, HRAS, and NRAS) play essential roles in various human cancers and are therefore suitable targets for anticancer therapeutics. Indeed, mutations in RAS proteins account for approximately 30% of all human cancers in the United States, many of which are fatal. Dysregulation of RAS proteins due to activating mutations, overexpression, or upstream activating genes is common in human tumors, and activating mutations in RAS are frequently found in human cancers. RAS switches between a GDP-bound "off" state and a GTP-bound "on" state. This state switch is facilitated by the interaction of guanine nucleotide exchange factor (GEF) proteins (e.g., SOS1), which load GTP onto RAS, with GTPase-activating protein (GAP) proteins (e.g., NF1), which hydrolyze GTP, resulting in RAS inactivation. Additionally, SH2 domain-containing protein tyrosine phosphatase 2 (SHP2) associates with the receptor signaling machinery and becomes active upon RTK activation, promoting RAS activation. Mutations in RAS proteins can lock the protein in the "on" state, resulting in a constitutively active signaling pathway that leads to uncontrolled cell proliferation. For example, activating mutations at codon 12 in RAS proteins function by inhibiting both the GAP-dependent and intrinsic hydrolysis rates of GTP, significantly skewing the population of RAS mutant proteins toward the "on" (GTP-bound) state (RAS(ON)), resulting in oncogenic MAPK signaling. Notably, RAS exhibits picomolar affinity for GTP, allowing RAS to be activated even in the presence of low concentrations of this nucleotide. Mutations at codon 13 (e.g., G13D) and codon 61 (e.g., Q61K) of RAS also contribute to oncogenic activity in some cancers.

[0137] Oncogenic pathways such as KRAS and the presence of immunosuppressive cell populations such as tumor-associated macrophages (TAMs) and myeloid-derived suppressor cells (MDSCs) have emerged as biological mechanisms of T cell elimination (Liu et al., Theranostics 2021).

[0138] KRAS mutations are prevalent in lung, pancreatic, and colon cancers, determining the immunosuppressive tumor microenvironment (TME) in these tumors (Gu et al., Cancers 2021). Oncogenic KRAS mutations mediate autocrine effects and crosstalk with the TME by inducing several inflammatory cytokines, chemokines, and signaling pathways that promote carcinogenesis and resistance to immunotherapy (Hamarsheh et al., Nat. Commun. 2020).

[0139] The RAS inhibitors described herein can enhance the sensitivity of immunorefractory lung cancer to immunotherapy. As described herein, treatment using the RAS inhibitors described herein can alter tumor immune infiltrates consisting of T cells, B cells, APCs, monocytes, MDSCs, TAMs, neutrophils, other monocyte-derived cells, tumor-associated stroma, cancer stem cells, or mesenchymal stem cells, resulting in enhanced anti-tumor therapeutic effects. In some embodiments, the RAS inhibitors described herein can enhance the sensitivity of subjects to immunotherapy, such as checkpoint inhibitor therapy.

[0140] The RAS inhibitors disclosed herein can form high-affinity ternary complexes or conjugates between a synthetic ligand and two intracellular proteins that do not interact under normal physiological conditions: the protein of interest (e.g., RAS) and a cytosolic chaperone (presenter protein) (e.g., cyclophilin A) that is widely expressed in cells. More specifically, in some embodiments, the RAS inhibitors described herein induce a novel binding pocket in RAS by driving the formation of a high-affinity tri-complex or conjugate between the RAS protein and the widely expressed cytosolic chaperone cyclophilin A (CYPA). Without being bound by theory, the inventors believe that one way in which the compounds of the present invention and the complexes or conjugates they form exert their inhibitory effect on RAS is by sterically blocking the interaction site between RAS and downstream effector molecules, such as RAF, that are necessary to propagate oncogenic signals. See, e.g., WO2021 / 091982, incorporated herein by reference in its entirety.

[0141] Accordingly, provided herein are compounds having the structure of Formula 0, or a pharmaceutically acceptable salt thereof: [ka] wherein the dotted lines represent 0, 1, 2, 3, or 4 non-adjacent double bonds; A is —N(H or CH3)C(O)—(CH2)—, where the amino nitrogen is —CH(R 10 )-, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene; B is absent or -CH(R 9 )-,>C=CR 9 R 9’ , or >CR 9 R 9’ wherein carbon is —N(R11 )C(O)—; optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or 5- to 6-membered heteroarylene; G is optionally substituted C1-C4 alkylene, optionally substituted C1-C4 alkenylene, optionally substituted C1-C4 heteroalkylene; [ka] [In the formula, [ka] -C(R 7 R 8 )-bonded to ], [ka] [In the formula, [ka] -C(R 7 R 8 )-; optionally substituted C1-C4 heteroalkylene, or 3- to 8-membered heteroarylene; L is absent or a linker; W is a bridging group comprising a vinyl ketone, vinyl sulfone, ynone, haloacetyl, or alkynyl sulfone; X 1 is an optionally substituted C1-C2 alkylene, NR, O, or S(O) n and X 2 is O or NH, X 3 is N or CH, n is 0, 1, or 2; R is hydrogen, cyano, optionally substituted C1-C4 alkyl, optionally substituted C2-C4 alkenyl, optionally substituted C2-C4 alkynyl, C(O)R', C(O)OR', C(O)N(R')2, S(O)R', S(O)2R', or S(O)2N(R')2; Each R ’ are independently H or optionally substituted C1-C4 alkyl; Y 1 is C, CH, or N, Y 2 , Y 3 , Y 4 , and Y 7 are independently C or N, Y 5 is CH, CH2, or N, Y 6 is C(O), CH, CH2, or N, R 1 is cyano, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 6-membered cycloalkenyl, optionally substituted 3- to 6-membered heterocycloalkyl, optionally substituted 6- to 10-membered aryl, or optionally substituted 5- to 10-membered heteroaryl; R 1 and R 2 combine with the atom to which they are attached to form an optionally substituted 3- to 14-membered heterocycloalkyl; R 2 is absent, hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 7-membered heterocycloalkyl, optionally substituted 6-membered aryl, optionally substituted 5- or 6-membered heteroaryl; R 3 does not exist, or R 2 and R 3combine with the atom to which they are attached to form an optionally substituted 3- to 8-membered cycloalkyl or an optionally substituted 3- to 14-membered heterocycloalkyl; R 4 is absent, hydrogen, halogen, cyano, or methyl optionally substituted with 1 to 3 halogens; R 5 is hydrogen; C1-C4 alkyl optionally substituted with halogen, cyano, hydroxy, or C1-C4 alkoxy; cyclopropyl, or cyclobutyl; R 6 is hydrogen or methyl, and R 7 is hydrogen, halogen, or optionally substituted C1-C3 alkyl, or R 6 and R 7 combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R 8 is hydrogen, halogen, hydroxy, cyano, optionally substituted C1-C3 alkoxy, optionally substituted C1-C3 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 8-membered cycloalkyl, optionally substituted 3- to 14-membered heterocycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 6- to 10-membered aryl, or R 7 and R 8 combine with the carbon atom to which they are attached, forming C=CR 7’ R 8’ forming C=N(OH), C=N(O-C1-C3 alkyl), C=O, C=S, C=NH, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted 3- to 7-membered heterocycloalkyl; R 7a and R 8a are independently hydrogen, halo, optionally substituted C1-C3 alkyl, or combined with the carbon to which they are attached to form a carbonyl; R7’ is hydrogen, halogen, or optionally substituted C1-C3 alkyl, and R 8’ is hydrogen, halogen, hydroxy, cyano, optionally substituted C1-C3 alkoxy, optionally substituted C1-C3 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 8-membered cycloalkyl, optionally substituted 3- to 14-membered heterocycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 6- to 10-membered aryl, or R 7’ and R 8’ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R 9 is H, F, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 7-membered heterocycloalkyl; or R 9 and L, combined with the atom to which they are attached, form an optionally substituted 3- to 14-membered heterocycloalkyl; R 9’ is hydrogen or optionally substituted C1-C6 alkyl, or R 9 and R 9’ combine with the atom to which they are attached to form a 3- to 6-membered cycloalkyl or a 3- to 6-membered heterocycloalkyl; R 10 is hydrogen, halo, hydroxy, C1-C3 alkoxy, or C1-C3 alkyl; R 10a is hydrogen or halo, R 11 is hydrogen or C1-C3 alkyl, and R 21is hydrogen or C1-C3 alkyl (e.g., methyl). In some embodiments, the compound of the present invention is selected from Table 1, or a pharmaceutically acceptable salt or stereoisomer thereof. In some embodiments, the compound of the present invention is selected from Table 1, or a pharmaceutically acceptable salt or atropisomer thereof.

[0142] In some embodiments, the present disclosure provides a method of treating immunorefractory lung cancer in a subject, comprising administering to the subject a RAS inhibitor of Formula 0, or a pharmaceutically acceptable salt thereof. In some embodiments, the compound of Formula 0 is a compound of Table 1 or Table 2, or a pharmaceutically acceptable salt thereof.

[0143] Thus, provided herein are compounds having the structure of Formula I, or a pharmaceutically acceptable salt thereof: [ka] wherein the dotted lines represent 0, 1, 2, 3, or 4 non-adjacent double bonds; A is —N(H or CH3)C(O)—(CH2)—, where the amino nitrogen is —CH(R 10 )-, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene; B is absent or -CH(R 9 )-,>C=CR 9 R 9’ , or >CR 9 R 9’ wherein carbon is —N(R 11 )C(O)—; optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or 5- to 6-membered heteroarylene; G is optionally substituted C1-C4 alkylene, optionally substituted C1-C4 alkenylene, optionally substituted C1-C4 heteroalkylene; [ka] [In the formula, [ka] -C(R 7 R 8 )-bonded to ], [ka] [In the formula, [ka] -C(R 7 R 8 )-; optionally substituted C1-C4 heteroalkylene, or 3- to 8-membered heteroarylene; L is a linker, said linker being acyclic or comprising a monocyclic, fused bicyclic, fused polycyclic, bridged bicyclic, or bridged polycyclic group; W is a bridging group comprising a vinyl ketone, vinyl sulfone, ynone, haloacetyl, or alkynyl sulfone; X 1 is an optionally substituted C1-C2 alkylene, NR, O, or S(O) n and X 2 is O or NH, X 3 is N or CH, n is 0, 1, or 2; R is hydrogen, cyano, optionally substituted C1-C4 alkyl, optionally substituted C2-C4 alkenyl, optionally substituted C2-C4 alkynyl, C(O)R', C(O)OR', C(O)N(R')2, S(O)R', S(O)2R', or S(O)2N(R')2; Each R’ are independently H or optionally substituted C1-C4 alkyl; Y 1 is C, CH, or N, Y 2 , Y 3 , Y 4 , and Y 7 are independently C or N, Y 5 is CH, CH2, or N, Y 6 is C(O), CH, CH2, or N, R 1 is cyano, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 6-membered cycloalkenyl, optionally substituted 3- to 6-membered heterocycloalkyl, optionally substituted 6- to 10-membered aryl, or optionally substituted 5- to 10-membered heteroaryl; R 1 and R 2 combine with the atom to which they are attached to form an optionally substituted 3- to 14-membered heterocycloalkyl; R 2 is absent, hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 7-membered heterocycloalkyl, optionally substituted 6-membered aryl, optionally substituted 5- or 6-membered heteroaryl; R 3 does not exist, or R 2 and R 3 combine with the atom to which they are attached to form an optionally substituted 3- to 8-membered cycloalkyl or an optionally substituted 3- to 14-membered heterocycloalkyl; R 4 is absent, hydrogen, halogen, cyano, or methyl optionally substituted with 1 to 3 halogens; R 5is hydrogen; C1-C4 alkyl optionally substituted with halogen, cyano, hydroxy, or C1-C4 alkoxy; cyclopropyl, or cyclobutyl; R 6 is hydrogen or methyl, and R 7 is hydrogen, halogen, or optionally substituted C1-C3 alkyl, or R 6 and R 7 combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R 8 is hydrogen, halogen, hydroxy, cyano, optionally substituted C1-C3 alkoxy, optionally substituted C1-C3 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 8-membered cycloalkyl, optionally substituted 3- to 14-membered heterocycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 6- to 10-membered aryl, or R 7 and R 8 combine with the carbon atom to which they are attached, forming C=CR 7’ R 8’ forming C=N(OH), C=N(O-C1-C3 alkyl), C=O, C=S, C=NH, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted 3- to 7-membered heterocycloalkyl; R 7a and R 8a are independently hydrogen, halo, optionally substituted C1-C3 alkyl, or combined with the carbon to which they are attached to form a carbonyl; R 7’ is hydrogen, halogen, or optionally substituted C1-C3 alkyl, and R 8’is hydrogen, halogen, hydroxy, cyano, optionally substituted C1-C3 alkoxy, optionally substituted C1-C3 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted 3- to 8-membered cycloalkyl, optionally substituted 3- to 14-membered heterocycloalkyl, optionally substituted 5- to 10-membered heteroaryl, or optionally substituted 6- to 10-membered aryl, or R 7’ and R 8’ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R 9 is H, F, optionally substituted C1-C6 alkyl, optionally substituted C1-C6 heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 7-membered heterocycloalkyl; or R 9 and L, combined with the atom to which they are attached, form an optionally substituted 3- to 14-membered heterocycloalkyl; R 9’ is hydrogen or optionally substituted C1-C6 alkyl, or R 9 and R 9’ combine with the atom to which they are attached to form a 3- to 6-membered cycloalkyl or a 3- to 6-membered heterocycloalkyl; R 10 is hydrogen, halo, hydroxy, C1-C3 alkoxy, or C1-C3 alkyl; R 10a is hydrogen or halo, R 11 is hydrogen or C1-C3 alkyl, and R 21 is H or C1-C3 alkyl.

[0144] In some embodiments, the present disclosure provides a method of treating immunorefractory lung cancer in a subject, comprising administering to the subject a RAS inhibitor of Formula I, or a pharmaceutically acceptable salt thereof. In some embodiments, the compound of Formula 0 is a compound of Table 1, or a pharmaceutically acceptable salt thereof.

[0145] In some embodiments, the compound of the present invention is selected from Table 1, or a pharmaceutically acceptable salt or stereoisomer thereof. In some embodiments, the compound of the present invention is selected from Table 1, or a pharmaceutically acceptable salt or atropisomer thereof.

[0146] [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6] [Table 1-7] [Table 1-8] [Table 1-9] [Table 1-10] Table 1-11 Table 1-12 Table 1-13 Table 1-14 Table 1-15 Table 1-16 Table 1-17 Table 1-18 Table 1-19 Table 1-20 Table 1-21 Table 1-22 Table 1-23 Table 1-24 Table 1-25 Table 1-26 Table 1-27 Table 1-28 Table 1-29 Table 1-30 Table 1-31 Table 1-32 Table 1-33 Table 1-34 Table 1-35 Table 1-36 Table 1-37 Table 1-38 Table 1-39 Table 1-40 Table 1-41 Table 1-42 Table 1-43 Table 1-44 Table 1-45 Table 1-46 Table 1-47 Table 1-48 Table 1-49 Table 1-50 Table 1-51 Table 1-52 Table 1-53 Table 1-54 Table 1-55 Table 1-56 Table 1-57 Table 1-58 Table 1-59 Table 1-60 Table 1-61 Table 1-62 Table 1-63 Table 1-64 Table 1-65 Table 1-66 Table 1-67 Table 1-68

Table 1-69

Table 1-90

[0147] In some embodiments, the compound of the present invention is selected from Table 2, or a pharmaceutically acceptable salt or stereoisomer thereof. In some embodiments, the compound of the present invention is selected from Table 2, or a pharmaceutically acceptable salt or atropisomer thereof.

[0148] [Table 2-1] [Table 2-2] [Table 2-3] [Table 2-4] [Table 2-5] [Table 2-6] [Table 2-7] [Table 2-8] [Table 2-9] [Table 2-10] [Table 2-11] Table 2-12 Table 2-13 Table 2-14 Table 2-15 Table 2-16 Table 2-17 Table 2-18

[0149] In some embodiments, the RAS inhibitor is selective for a RAS comprising a G12C amino acid substitution compared to wild-type RAS or other RAS mutants. In some embodiments, the RAS inhibitor is a KRAS inhibitor that is selective for a KRAS comprising a G12C amino acid substitution compared to wild-type KRAS or other KRAS mutants. In some embodiments, the RAS inhibitor is an NRAS inhibitor that is selective for an NRAS comprising a G12C amino acid substitution compared to wild-type NRAS or other NRAS mutants. In some embodiments, the RAS inhibitor is an HRAS inhibitor that is selective for an HRAS comprising a G12C amino acid substitution compared to wild-type NRAS or other NRAS mutants. In some embodiments, the RAS inhibitor that is selective for a RAS comprising a G12C amino acid substitution compared to wild-type NRAS or other NRAS mutants is a RAS(ON) inhibitor. In some embodiments, the RAS inhibitor that is selective for a RAS comprising a G12C compared to wild-type RAS or other RAS mutants is not a RAS(OFF) inhibitor.

[0150] Immune checkpoint inhibitors The compositions and methods described herein may include an immune checkpoint inhibitor (ICI). The immune checkpoint inhibitor may be administered or formulated in combination with a RAS inhibitor described herein. The immune checkpoint inhibitor may be administered or formulated in combination with a RAS inhibitor and an SHP2 inhibitor described herein.

[0151] Immune checkpoints refer to numerous inhibitory pathways built into the immune system that, under normal physiological conditions, are important in regulating the duration and amplitude of physiological immune responses in peripheral tissues to maintain self-tolerance and minimize collateral tissue damage in response to pathogenic infections. However, the expression of immune checkpoint proteins is often dysregulated by tumors as an important immune resistance and evasion mechanism.

[0152] Because many immune checkpoints are initiated by ligand-receptor interactions, they can be easily blocked by antibodies or modulated by recombinant forms of the ligand or receptor. Therefore, inhibition of these pathways has been used to activate therapeutic antitumor immunity. For example, cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) antibodies were the first in this class of immunotherapeutic agents to achieve U.S. Food and Drug Administration (FDA) approval. Preliminary clinical findings with inhibitors of additional immune checkpoint proteins, such as programmed cell death protein 1 (PD-1), suggest broad and diverse opportunities for enhancing antitumor immunity that may result in durable clinical responses.

[0153] Activation of T cells through immune checkpoint blockade has become a major focus of efforts to therapeutically manipulate endogenous antitumor immunity due to their ability to selectively recognize peptides derived from proteins within all cellular compartments, their ability to directly recognize and kill antigen-expressing cells (CD8+ effector T cells, also known as cytotoxic T lymphocytes (CTLs)), and their ability to orchestrate diverse immune responses (mediated by CD4+ helper T cells) that integrate adaptive and innate effector mechanisms. Thus, agonists of costimulatory receptors or antagonists of inhibitory signals, both of which result in the amplification of antigen-specific T cell responses, are currently agents of interest in clinical testing.

[0154] [Table 3]

[0155] Approved or investigational ICIs include YERVOY® (ipilimumab), OPDIVO® (nivolumab), KEYTRUDA® (pembrolizumab), tremelimumab, galiximab, MDX-1106, BMS-936558, MEDI4736, MPDL3280A, MEDI6469, BMS-986016, BMS-663513, and PF-05082. 566, IPH2101, KW-0761, CDX-1127, CP-870, CP-893, GSK2831781, MSB0010718C, MK3475, CT-011, AMP-224, MDX-1105, IMP321, and MGA271, as well as numerous other antibodies or fusion proteins against immune checkpoint proteins listed in Table 3. Common immune checkpoint proteins that can be targeted by ICI include, but are not limited to, B7.1, B7-H3, LAG3, CD137, KIR, CCR4, CD27, OX40, GITR, CD40, CTLA4, PD-1, and PD-L1.

[0156] In some embodiments, the ICI therapy is selected from one or more of anti-PD-1, anti-PD-L1, anti-CTLA-4, anti-LAG3, anti-B7.1, anti-B7H3, anti-B7H4, anti-TIM3, anti-VISTA, anti-CD137, anti-OX40, anti-CD40, anti-CD27, anti-CCR4, anti-GITR, anti-NKG2D, and anti-KIR. In some embodiments, the ICI therapy is an antibody (e.g., a monoclonal antibody selective for any of the targets in Table 3). In some embodiments, the ICI is an anti-PD-1 antibody. The antibody can be, for example, humanized or fully human.

[0157] In some embodiments, the checkpoint inhibitor is a fusion protein, e.g., an Fc receptor fusion protein. In some embodiments, the checkpoint inhibitor is an agent, such as an antibody, that interacts with a checkpoint protein. In some embodiments, the checkpoint inhibitor is an agent, such as an antibody, that interacts with a ligand of a checkpoint protein. In some embodiments, the checkpoint inhibitor is an inhibitor (e.g., an inhibitory antibody or small molecule inhibitor) of CTLA-4 (e.g., an anti-CTLA-4 antibody or fusion protein). In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist (e.g., an inhibitory antibody or small molecule inhibitor) of PD-1. In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist (e.g., an inhibitory antibody or small molecule inhibitor) of PD-L1. In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist (e.g., an inhibitory antibody or Fc fusion or small molecule inhibitor) of PD-L2 (e.g., a PD-L2 / Ig fusion protein). In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist (e.g., an inhibitory antibody or small molecule inhibitor) of B7-H3, B7-H4, BTLA, HVEM, TIM3, GAL9, LAG3, VISTA, KIR, 2B4, CD160, CGEN-15049, CHK1, CHK2, A2aR, a B-7 family ligand, or a combination thereof.In some embodiments, the checkpoint inhibitor is pembrolizumab, nivolumab, PDR001 (NVS), REGN2810 (Sanofi / Regeneron), a PD-L1 antibody such as avelumab, durvalumab, atezolizumab, pidilizumab, JNJ-63723283 (JNJ), BGB-A317 (also known as tislelizumab; BeiGene & Celgene), or Preusser, M. et al. al. (2015) Nat. Rev. Neurol., including, but not limited to, ipilimumab, tremelimumab, nivolumab, pembrolizumab, AMP224, AMP514 / MEDI0680, BMS936559, MED14736, MPDL3280A, MSB0010718C, BMS986016, IMP321, lirilumab, IPH2101, 1-7F9, and KW-6002.

[0158] SHP2 inhibitors The compositions and methods described herein can include an SHP2 inhibitor. The SHP2 inhibitor can be administered or formulated in combination with a RAS inhibitor described herein. The SHP2 inhibitor can be administered or formulated in combination with a RAS inhibitor and an immune checkpoint inhibitor.

[0159] SHP2, a non-receptor protein tyrosine phosphatase encoded by the PTPN11 gene, contributes to multiple cellular functions, including proliferation, differentiation, cell cycle maintenance, and migration. SHP2 contains two N-terminal Src homology 2 domains (N-SH2 and C-SH2), a catalytic domain (PTP), and a C-terminal tail. The two SH2 domains regulate the subcellular localization and functional regulation of SHP2. This molecule exists in an inactive, autoinhibited conformation stabilized by a network of bonds involving residues from both the N-SH2 and PTP domains. Stimulation by cytokines or growth factors acting through receptor tyrosine kinases (RTKs), for example, leads to exposure of the catalytic site, resulting in enzymatic activation of SHP2.

[0160] SHP2 is involved in signal transduction via the RAS mitogen-activated protein kinase (MAPK), JAK-STAT, or phosphoinositol 3-kinase-AKT pathways. Mutations in the PTPN11 gene and subsequent SHP2 mutations have been identified in several human developmental disorders, such as Noonan syndrome and Leopard syndrome, as well as human cancers such as juvenile myelomonocytic leukemia, neuroblastoma, melanoma, acute myeloid leukemia, and breast, lung, and colon cancer. Some of these mutations destabilize the autoinhibitory conformation of SHP2, promoting its self-activation or enhanced growth factor-driven activation. Therefore, SHP2 is a highly attractive target for the development of novel therapies for the treatment of various diseases, including cancer. SHP2 inhibitors (e.g., RMC-4550 or SHP099) in combination with RAS pathway inhibitors (e.g., MEK inhibitors) have been shown to inhibit the growth of multiple cancer cell lines (e.g., pancreatic, lung, ovarian, and breast cancer) in vitro.

[0161] Non-limiting examples of such SHP2 inhibitors are known in the art and include those described in: Chen et al. Mol Pharmacol. 2006, 70, 562; Sarver et al., J. Med. Chem. 2017, 62, 1793; Xie et al., J. Med. Chem. 2017, 60, 113734; and Igbe et al. al., Oncotarget, 2017, 8, 113734; and patent applications WO2023282702, WO2023280283, WO2023280237, WO2023018155, WO2023011513, WO2022271966, WO2022271964, WO2022271911, WO2022259157, WO2022242767, WO202224197, each of which is incorporated herein by reference. 5, WO2022237676, WO2022237367, WO2022237178, WO2022235822, WO2022234409, WO2022208408, WO2022207924, WO202216 7682, WO2022166844, WO2022161222, WO2022156765, WO2022135568, WO2022089406, WO2022089389, WO2022063190, WO2022 043865, WO2022042331, WO2022033430, WO2022017444, WO2022007869, WO2021259077, WO2021249449, WO2021249057, WO20212 44659, WO2021218755, WO2021281752, WO2021149817, WO2021148010, WO2021147879, WO2021143823, WO2021143701, WO2021143 680, WO2021121397, WO2021119525, WO2021115286, WO2021110796, WO2021088945, WO2021073439, WO2021061706, WO202106151 5, WO2021043077, WO2021033153, WO2021028362, WO2021033153, WO2021028362, WO2021018287, WO2020259679, WO2020249079,WO2020210384、WO2020201991、WO2020181283、WO2020177653、WO2020165734、WO2020165733、WO2020165732、WO2020156243、WO2020156242、WO2020108590、WO2020104635、WO2020094104、WO2020094018、WO2020081848、WO2020073949、WO2020073945、WO2020072656、WO2020065453、WO2020065452、WO2020063760、WO2020061103、WO2020061101、WO2020033828、WO2020033286、WO2020022323、WO2019233810、WO2019213318、WO2019183367、WO2019183364、WO2019182960、WO2019167000、WO2019165073、WO2019158019、WO2019152454、WO2019051469、WO2019051084、WO2018218133、WO2018172984、WO2018160731、WO2018136265、WO2018136264、WO2018130928、WO2018129402、WO2018081091、WO2018057884、WO2018013597、WO2017216706、WO2017211303、WO2017210134、WO2017156397、WO2017100279、WO2017079723、WO2017078499、WO2016203406、WO2016203405、WO2016203404、WO2016196591、WO2016191328、WO2015107495、WO2015107494、WO2015107493、WO2014176488、WO2014113584、CN115677661、CN115677660、CN115611869、CN115521305、CN115490697、CN115466273、CN115394612、CN115304613、CN115304612、CN115300513、CN115197225、CN114957162、CN114920759、CN114716448、CN114671879、CN114539223, CN114524772, CN114213417, CN114195799, CN114163457, CN113896710, CN113248521, CN113248449, CN1131 35924, CN113024508, CN112920131, CN112823796, CN112409334, CN112402385, CN112174935, 111848599, CN111704611, CN 111393459, CN111265529, CN110143949, CN108113848, US11179397, US11044675, US11034705, US11033547, US11001561, US10988466, US10954243, US10934302, US10858359, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof.

[0162] In some embodiments, the SHP2 inhibitor binds to the active site. In some embodiments, the SHP2 inhibitor is a mixed irreversible inhibitor. In some embodiments, the SHP2 inhibitor binds to an allosteric site, e.g., a non-covalent allosteric inhibitor. In some embodiments, the SHP2 inhibitor is a covalent SHP2 inhibitor, such as an inhibitor that targets a cysteine ​​residue (C333) outside the active site of the phosphatase. In some embodiments, the SHP2 inhibitor is a reversible inhibitor. In some embodiments, the SHP2 inhibitor is an irreversible inhibitor. In some embodiments, the SHP2 inhibitor is SHP099. In some embodiments, the SHP2 inhibitor is TNO155, which has the following structure: [ka] or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is RMC-4550, which has the structure: [ka] or a pharmaceutically acceptable salt, solvate, isomer (eg, stereoisomer), prodrug, or tautomer thereof.

[0163] In some embodiments, the SHP2 inhibitor is RMC-4630, which has the structure: [ka] or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is JAB-3068, which has the structure: [ka] or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is JAB-3312. In some embodiments, the SHP2 inhibitor is RLY-1971, which has the following structure: [ka] or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is ERAS-601, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is BBP-398, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof.

[0164] The present disclosure also provides pharmaceutical compositions. The pharmaceutical compositions comprise a RAS inhibitor, an SHP2 inhibitor, an immune checkpoint inhibitor, or a combination thereof as an active agent, and at least one pharmaceutically acceptable excipient. The pharmaceutically acceptable excipient may be a diluent, binder, filler, buffer, pH adjuster, disintegrant, dispersant, preservative, lubricant, taste masking agent, flavoring agent, or coloring agent. The amount and type of excipient used to form the pharmaceutical composition can be selected according to known pharmaceutical principles.

[0165] The compositions can be formulated into a variety of dosage forms and can be administered by several different means that deliver a therapeutically effective amount of the active agent(s). Such compositions can be administered orally (e.g., by inhalation) or parenterally in dosage unit formulations that optionally contain conventional non-toxic pharmaceutically acceptable carriers, adjuvants, and vehicles.

[0166] How to use In some embodiments, the present disclosure provides methods of treating a subject with immunorefractory lung cancer, the method generally comprising administering to the subject a compound or combination of compounds described herein. In some embodiments, the subject is administered a RAS inhibitor described herein (i.e., a RAS G12C inhibitor). In some embodiments, the subject is administered a RAS inhibitor described herein in combination with an immune checkpoint inhibitor, an SHP2 inhibitor, or a combination thereof. Suitable examples of RAS inhibitors, SHP2 inhibitors, and immune checkpoint inhibitors are described above and incorporated by reference into this section.

[0167] In some embodiments, the present disclosure provides a method of sensitizing an immunorefractory cancer to immunotherapy in a subject in need thereof, the method generally comprising administering to the subject a compound or combination of compounds described herein. In some embodiments, the subject is administered a RAS inhibitor described herein (i.e., a RAS G12C inhibitor). In some embodiments, the subject is administered a RAS inhibitor described herein in combination with an immune checkpoint inhibitor, an SHP2 inhibitor, or a combination thereof.

[0168] In some embodiments, the present disclosure provides a method for promoting the conversion of an immunologically cold tumor to an immunologically hot tumor in a subject in need thereof, the method generally comprising administering to the subject a compound or combination of compounds described herein, thereby enabling treatment of a previously immunologically cold tumor that has been converted to an immunologically hot tumor. In some embodiments, the subject is administered a RAS inhibitor described herein (i.e., a RAS G12C inhibitor). In some embodiments, the subject is administered a RAS inhibitor described herein in combination with an immune checkpoint inhibitor, an SHP2 inhibitor, or a combination thereof.

[0169] In some embodiments, the present disclosure provides a method for enhancing the effectiveness of other cancer treatments when administered in combination to a subject. In some embodiments, the subject is administered a RAS inhibitor (i.e., a RAS G12C inhibitor) described herein. In some embodiments, the subject is administered a RAS inhibitor described herein in combination with an immune checkpoint inhibitor, an SHP2 inhibitor, or a combination thereof.

[0170] In each of the above embodiments, the disclosed methods can alter the tumor immune infiltrate, which consists of T cells, B cells, APCs, monocytes, MDSCs, TAMs, neutrophils, other monocyte-derived cells, tumor-associated stroma, cancer stem cells, and mesenchymal stem cells, resulting in enhanced anti-tumor therapeutic effects.

[0171] Tumor type identification In various embodiments, the disclosure provides methods of treating lung cancer in a subject comprising administering to the subject a RAS inhibitor or combination of compounds described herein, wherein the subject has one or more tumors characterized as immunorefractory, immunoevasive, immunologically protected, immunologically "cold," microsatellite stable, microsatellite instability-low, containing a reduced immune infiltrate, containing a low tumor mutational burden, or exhibiting heterogeneity.

[0172] In various embodiments, the present disclosure provides methods for treating a tumor (e.g., lung cancer) in a subject characterized as being immunoevasive, immunologically protected, immunologically "cold," microsatellite stable, low microsatellite instability, containing a low immune infiltrate, containing a low tumor mutational burden, or exhibiting heterogeneity, comprising: (i) diagnosing the subject as having an immunorefractory, immunoevasive, immunologically protected, immunologically "cold," microsatellite stable, low microsatellite instability, containing a low immune infiltrate, containing a low tumor mutational burden, or exhibiting heterogeneity; and (ii) administering to the subject a RAS inhibitor or combination of compounds described herein. In various embodiments, diagnosing comprises assaying biomarkers / characteristics associated with tumors characterized as immunorefractory, immunoevasive, immunologically protected, immunologically "cold," microsatellite stable, low microsatellite instability, containing a low immune infiltrate, containing a low tumor mutational burden, or exhibiting heterogeneity. In various embodiments, the method further comprises (iii) determining whether the subject's tumor becomes immunoresponsive, and then (iv) administering immunotherapy, optionally in combination with a RAS inhibitor or a combination of compounds described herein. In some embodiments, the subject has previously been diagnosed with a tumor characterized as immunoevasive, immunologically protected, immunologically "cold," microsatellite stable, low microsatellite instability, containing a low immune infiltrate, containing a low tumor mutational burden, or exhibiting heterogeneity.

[0173] Also provided herein are methods for determining whether a subject is responsive or likely to respond to treatment with a RAS inhibitor or combination of compounds described herein, and treating the subject accordingly. In various embodiments, a patient diagnosed with cancer is tested to identify the tumor as a cold tumor, for example, using the methods described herein and other methods described in the art. The present disclosure provides methods for treating a subject with cancer, e.g., an immunorefractory cancer, with a RAS inhibitor or combination of compounds described herein, comprising obtaining a tumor sample from the subject, performing an assay to determine whether the tumor is a cold tumor, and, if the tumor is identified as a cold tumor, treating the subject with a RAS inhibitor or combination of compounds described herein. Assays for determining whether a tumor is a cold tumor include tumor mutation burden analysis, microsatellite instability (MSI) testing, and the introduction of immune cells (e.g., CD4 + T cells, CD8 + T cells, NK1.1 + The degree of infiltration of immune cells (NK cells, APCs, monocytes, and neutrophils), immune cell phenotype (e.g., PD-1 + , PD-L1 + , and PD-L2 + ), immune cell function (e.g., expression of IFN-γ, IL-12, IL-15, and MHCII), and the ratio of pro- to anti-inflammatory mediators in the tumor microenvironment (TME).

[0174] Several diagnostic tools designed to characterize tumors at the cellular and molecular level are FDA approved and commercially available. Examples of approved diagnostics include FOUNDATIONONE® CDX, FOUNDATIONONE® LIQUID, FOUNDATIONONE® HEME, BRACAnalysis CDx, therascreen EGFR RGQ PCR Kit, cobase EGFR Mutation Test v2, PD-L1 IHC 22C3 pharmDx, Abbott RealTime IDH1, MRDx BCR-ABL test, VENTANA ALK(D5F3)CDx Assay, Abbott RealTime IDH2, Praxis Extended RAS Panel, Oncomine Dx Target Test, LeukoStrat CDx FLT3 Mutation Assay, FoundationFocus CDx BRCA Assay, Vysis CLL FISH Probe Kit, KIT D816V Mutation Detection, PDGFRB FISH, cobas KRAS Mutation Test, therascreen KRAS RGQ PCR Kit, FerriScan, Dako c-KIT pharmDx, and INFORM Her-2 / neu, PathVysion HER-2 DNA Probe Kit, SPOT-LIGHT HER2 CISH Kit, Bond Oracle HER2 IHC System, HER2 CISH pharmDx Kit, INFORM HER2 DUAL ISH DNA Probe Cocktail, HercepTest, HER2 FISH pharmDx Kit, THXID BRAF Kit, Vysis ALK Break Apart FISH Probe Kit, cobas 4800 BRAF V600 Mutation Test, VENTANA PD-L1 (SP142) Assay, therascreen FGFR RGQ RT-PCR Kit, and therascreen PIK3CA RGQ PCR Kit.

[0175] In various embodiments, subjects are screened for eligibility for treatment with one or more immunotherapies described herein. In various embodiments, subjects who are not eligible for such immunotherapeutic treatment (e.g., who are non-responsive to one or more immunotherapies or have a cancer characterized as non-responsive to one or more immunotherapies) may first be treated with a RAS inhibitor or combination of compounds described herein according to the methods described herein. Non-limiting examples of immunotherapies include pembrolizumab (KEYTRUDA®, Merck Sharp & Dohme Corp), nivolumab (OPDIVO®, Bristol-Myers Squibb), atezolizumab (TECENTRIQ®), avelumab (BAVENCIO®), and durvalumab (IMFINZI®). Eligibility criteria for these immunotherapies are known in the art. For example, without limitation, pembrolizumab (KEYTRUDA®), nivolumab (OPDIVO®), and atezolizumab (TECENTRIQ®) have eligibility criteria based on PD-L1 expression levels. PD-L1 expression criteria and methods for measuring it can be found at keytrudahcp.com / biomarker-testing / pd-11-expression-testing / (pembrolizumab; KEYTRUDA®), or in the FDA-approved prescribing information for pembrolizumab (KEYTRUDA®, revised January 2020), atezolizumab (e.g., TECENTRIQ®, revised May 2020), and nivolumab (e.g., OPDIVO®, revised June 2020). Each of these publications is incorporated herein by reference in its entirety for all purposes. As described herein, treating such patients with a RAS inhibitor or combination of compounds described herein may promote the conversion of tumors that are ineligible for treatment with immunotherapy into immunogenic tumors, thereby making such tumors treatable with immunotherapy.In various embodiments, a subject's tumor that is ineligible for immunotherapy can be monitored throughout the course of treatment with a RAS inhibitor or combination of compounds described herein to determine when the tumor becomes eligible for immunotherapy treatment. Once the tumor becomes eligible for immunotherapy treatment, the subject can be administered immunotherapy, alone or in combination with a RAS inhibitor or SHP2 inhibitor, or a combination thereof.

[0176] In various embodiments, the present disclosure provides a method of treating lung cancer in a subject, comprising administering to the subject a RAS inhibitor or a combination of compounds described herein, wherein the subject has one or more tumors with a low immune infiltrate. In various embodiments, administration to a subject with one or more lung tumors with a low immune infiltrate alters the tumor immune infiltrate. In various embodiments, the tumor immune infiltrate comprises antigen-presenting cells, myeloid cells, and lymphoid cells. In various embodiments, antigen-presenting cells in the tumor immune infiltrate comprise macrophages or dendritic cells. In various embodiments, myeloid cells in the tumor immune infiltrate comprise monocytes, neutrophils, myeloid-derived suppressor cells (MDSCs), and tumor-associated macrophages (TAMs). In various embodiments, TAMs in the tumor immune infiltrate comprise M1 macrophages, M2 macrophages, and MARCO macrophages. + In various embodiments, the immune infiltrate includes lymphoid cells, T cells, B cells, NK T cells, and NK cells.

[0177] Qualitative and quantitative methods have been described for characterizing tumor immune infiltrates, including, but not limited to, microscopic analysis, histological assays, cytological assays, flow cytometry, polymerase chain reaction (PCR), quantitative polymerase chain reaction (qPCR), RNA sequencing (RNA-seq), single-cell RNA sequencing (scRNA-seq), next-generation sequencing, whole-exome sequencing, epigenetic sequencing, ATAC-seq, microarray analysis, and mass cytometry or CyTOF. Biomarkers can be used alone or in combination to assess immune cells and include cell surface markers and secreted proteins. Exemplary biomarkers for characterization of tumor immune infiltrates include CD45, CD3, CD4, CD8, CD25, CD44, CD134, CD252, CD137, CD79, CD39, FOXP3, PD-1, LAG-3, TIM-1, IFN-γ, granzyme, perforin, CD11b, CD11c, Ly6C, Ly6G, CD14, CD16, CD80, MARCO, CD68, CD115, CD206, CD163, CD103c, F4 / 80, PD-L1, PD-L2, arginase, iNOS, ROS, TN These include, but are not limited to, F-α, TGF-β, MHC-I, MHC-II, NK1.1, NKG2D, CD244, Ki67, CD19, CD20, CCR2, CXCR3, CCR4, CCR5, CCR6, CCR7, CCR10, CCL2, CCL5, Cx3CR1, CCL10, ICOS, CD40, CD40L, IL1α, IL1β, IL2, IL4, IL5, IL6, IL8, IL12, IL15, IL17, IL21, IL22, TCRγ / δ, TCRα / β, STAT3, ROR1c, and RORγt.

[0178] Cancer stem cells (CSCs) have been described as a subset of cells found in solid and hematological tumors that are tumorigenic, self-renewing, and capable of differentiation. Several reports have documented the importance of CSCs in the pathogenesis of various tumors, tumor recurrence after treatment, and the development of therapeutic resistance. Several cell surface markers can be used to distinguish CSCs within solid and hematological tumors. CSC markers include, but are not limited to, CD19, CD20, CD24, CD34, CD38, CD44, CD90, CD133, aldehyde dehydrogenase 1, CEACAM-6 / CD66c, BML-1, connexin 43 / GJA1, DLL4, EpCAM / TROP1, GL1-1, GL1-2, integrin, PON1, PTEN, ALCAM / CD166, DPPIV / CD26, Lgr5, Musashi-1, A20, ABCG2, CD15, fractalkine, HIF-2α, L1CAM, c-MAF, nestin, podoplanin, SOX2, CD96, CD117, FLT3, AFP, CD13, CD90, NF2 / Merlin, ABCB5, NGFR, syndecan-1, endoglin, STRO-1, and PON1.

[0179] In various embodiments, the present disclosure provides a method of treating cancer in a subject comprising administering to the subject a RAS inhibitor or a combination of compounds described herein, wherein the subject has one or more immunorefractory tumors. In various embodiments, the subject has one or more immunologically protected tumors. In various embodiments, the subject has one or more microsatellite-stable tumors. In various embodiments, the subject has one or more microsatellite-poor tumors. In various embodiments, the subject has one or more tumors with moderate microsatellite instability. In various embodiments, the subject has one or more tumors with low tumor mutational burden. In various embodiments, the subject has one or more tumors with moderate tumor mutational burden. In various embodiments, the subject has one or more tumors that are resistant to treatment. In various embodiments, the subject has one or more immunologically heterogeneous tumors. In various embodiments, the subject has a genetically heterogeneous tumor. In various embodiments, the subject has one or more refractory tumors. In one or more embodiments, the subject has a tumor that acquires resistance during the course of treatment.

[0180] In various embodiments, tumor characteristics are determined from one or more biological samples from a subject suffering from cancer. In various embodiments, tumor characteristics are determined by comparing one or more biological samples from a subject suffering from cancer with one or more biological samples from one or more healthy subjects. In various embodiments, tumor characteristics are determined from one or more biological samples selected from the group consisting of blood, cerebrospinal fluid, urine, feces, buccal swabs, nasal swabs, lavage fluid, tissue biopsy, bone marrow biopsy, and tumor biopsy. In various embodiments, tumor characteristics are determined from analysis of cells, proteins, or nucleic acids in one or more biological samples from a subject suffering from cancer. In various embodiments, tumor characteristics are determined by comparing analysis of cells, proteins, or nucleic acids in one or more biological samples from a subject suffering from cancer with analysis of one or more biological samples from one or more healthy subjects. In various embodiments, tumor characteristics are determined by comparing analysis of cells, proteins, or nucleic acids in one or more biological samples from a subject suffering from cancer with analysis of one or more biological samples from one or more subjects suffering from cancer and who are responsive to treatment. In various embodiments, the cells are selected from the group consisting of leukocytes, epithelial cells, mesenchymal cells, mesenchymal stem cells, stromal cells, endothelial cells, fibroblasts, cancer-associated fibroblasts (CAFs), pericytes, adipocytes, cancer stem cells, circulating tumor cells (CTCs), hematopoietic stem cells, and hematopoietic progenitor cells. In various embodiments, the proteins are selected from the group consisting of cytokines, chemokines, growth factors, signaling proteins, enzymes, proteases, and nucleases. In various embodiments, the nucleic acids are selected from the group consisting of DNA, ssDNA, circulating tumor DNA (ctDNA), RNA, mRNA, dsRNA, siRNA, miRNA, and lncRNA. In various embodiments, the nucleic acid analysis is performed by PCR, RT-PCR, qRT-PCR, next-generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single-cell sequencing.

[0181] In various embodiments, tumor characteristics of a subject suffering from cancer are determined from analysis of one or more blood samples taken from the subject. In various embodiments, tumor characteristics of a subject suffering from cancer are determined from analysis of cells, proteins, or nucleic acids in one or more blood samples taken from the subject. In various embodiments, tumor characteristics of a subject suffering from cancer are determined by comparing analysis of cells, proteins, or nucleic acids in one or more blood samples from a subject suffering from cancer with analysis of one or more blood samples from one or more healthy subjects. In various embodiments, the cells analyzed in the one or more blood samples are leukocytes, epithelial cells, mesenchymal cells, mesenchymal stem cells, stromal cells, endothelial cells, fibroblasts, cancer-associated fibroblasts (CAFs), pericytes, adipocytes, cancer stem cells, circulating tumor cells (CTCs), hematopoietic stem cells, and hematopoietic progenitor cells. In various embodiments, the leukocytes are myeloid cells and lymphoid cells. In various embodiments, the myeloid cells are monocytes, macrophages, neutrophils, granulocytes, dendritic cells, mast cells, eosinophils, and basophils. In various embodiments, the lymphoid cells are T cells, B cells, NK cells, NK-T cells, or iNK cells.

[0182] In various embodiments, analysis of cells from one or more blood samples taken from a subject suffering from cancer shows elevated levels of immunosuppressive cells compared to analysis of cells from one or more healthy subjects, or one or more subjects suffering from cancer and responsive to treatment (e.g., responsive to immunotherapy, such as an immune checkpoint inhibitor). In various embodiments, the immunosuppressive cells include myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), neutrophils, T cells, and the like. reg cells, and B regIn various embodiments, the MDSCs are monocytic MDSCs (M-MDSCs) and polymorphonuclear MDSCs (PMN-MDSCs). In various embodiments, the TAMs are M2 TAMs. In various embodiments, the immunosuppressive cells are CAFs. In various embodiments, the level of immunosuppressive cells in one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is about 5-100% (e.g., about 5%, about 10%, about 15%, about 20%) lower than one or more blood samples taken from one or more healthy subjects, or one or more subjects with cancer who are responsive to treatment. , about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50% increase. In various embodiments, the level of immunosuppressive cells in one or more blood samples from subjects with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by about 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more blood samples taken from one or more healthy subjects, or one or more subjects with cancer who are responsive to treatment. In various embodiments, the immunosuppressive cells are identified by assaying cell surface protein expression.In various embodiments, analysis of cells from one or more blood samples taken from a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows a reduced level or absence of activated pro-inflammatory immune cells (e.g., a reduction of about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges between these values) compared to a healthy subject, or a subject having cancer and who is responsive to treatment).

[0183] In various embodiments, analysis of cells from one or more blood samples taken from a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows a reduced level or absence of activated pro-inflammatory immune cells (e.g., about a 2-, 5-, 10-, 15-, 20-, 25-, 30-, 35-, 40-, 45-, 50-, 55-, 60-, 65-, 70-, 75-, 80-, 85-, 90-, 95-, or 100-fold reduction (including all values ​​and ranges therebetween) compared to a healthy subject or a subject having cancer and who is responsive to treatment). In various embodiments, the activated pro-inflammatory cells are dendritic cells (DCs), macrophages, M1 macrophages, T cells, B cells, NK cells, NK-T cells, and iNK cells. In various embodiments, the frequency of pro-inflammatory immune cells is 10% or less (e.g., about 9%, about 8%, about 7%, about 6%, about 5%, about 4%, about 3%, about 2%, or about 1%) of total white blood cells analyzed from one or more blood samples taken from the subject. In various embodiments, activated pro-inflammatory immune cells are identified by assaying cell surface protein expression.

[0184] In various embodiments, the analysis of cells in one or more blood samples from a subject suffering from cancer is performed by assaying for cell surface proteins, such as receptor tyrosine kinase (RTK), CD1c, CD2, CD3, CD4, CD5, CD8, CD9, CD10, CD11b, CD11c, CD14, CD15, CD16, CD18, CD19, CD20, CD21, CD22, CD23, CD24, TACI, CD25, CD27, CD28, CD30, CD30L, CD31, CD32, CD32b, CD34, CD33, CD38, CD39, CD40, CD40-L, CD41b, CD42a, CD42b, CD43, CD44, CD48 , CD47, CD45RA, CD45RO, CD48, CD52, CD55, CD56, CD58, CD61, CD66b, CD70, CD72, CD79, CD68, CD84, CD86, CD93, CD94, CD95, CRACC, BLAME, BCMA, CD103 , CD107, CD112, CD120a, CD120b, CD123, CD125, CD134, CD135, CD140a, CD141, CD154, CD155, CD160, CD163, CD172a, XCR1, CD203c, CD204, CD206, CD207CD226、CD244、CD267、CD268、CD269、CD355、CD358、NKG2A、NKG2B、NKG2 C、NKG2D、NKG2E、NKG2F、NKG2H、KIR2DL1、KIR2DL2、KIR2DL3、KIR2DL5A、 KIR2DL5B、KIR3DL1、KIR3DL2、KIR3DL3、KIR3DL4、KIR2DS1、KIR2DS2、KI R2DS3、KIR2DS4、KIR2DS5、DAP12、KIR3DS、NKp44、NKp46、TCR、BCR、インテグリン、FcβεRI、MHC-I、MHC-II、IL-1R、IL-2Rα、IL-2Rβ、IL-2Rγ、IL-3Rα、CSF2RB、IL-4R、IL-5Rα、IL-6Rα、gp130、IL-7Rα、IL-9R、IL-12Rβ1、IL-12R β2, IL-13Rα1, IL-13Rα2, IL-15Rα, IL-21R, IL-23R, IL-27Rα, IL-31Rα, OSMR, CSF-1R, IL-15, IL-10Rα, IL-10Rβ, IL-20Rα, IL-20Rβ, IL-22 Rα1、IL-22Rα2、IL-22Rβ、IL-28RA、PD-1、PD-1H、BTLA、CTLA-4、PD-L1、P D-L2、2B4、B7-1、B7-2、B7-H1、B7-H4、B7-DC、DR3、LIGHT、LAIR、LTα1β2、 LTβR、TIM-1、TIM-3、TIM-4、TIGIT、LAG-3、ICOS、ICOS-L、SLAM、SLAMF2、 OX-40、OX-40L、GITR、GITRL、TL1A、HVEM、41-BB、41BB-L、TL-1A、TRAF1、 TRAF2、TRAF3、TRAF5、BAFF、BAFF-R、APRIL、TRAIL、RANK、AITR、TRAMP、C CR1、CCR2、CCR3、CCR4、CCR5、CCR6、CCR7、CCR8、CCR9、CCR10、CCR11、CXC R1、CXCR2、CXCR3、CXCR4、CXCR5、CXCR6、CXCR7、CLECL9a、DC-SIGN、IGSF 4A、SIGLEC、EGFR、PDGFR、VEGFR、FAP、α-SMA、ビメンチン、ラミニン、FAS、FAS-L、F c, ICAM-1, ICAM-2, ICAM-3, ICAM-4, ICAM-5, PECAM-1, MICA, MICB, UL16, ULBP1, ULBP2, ILBP3, ULBP4, ULBP5, ULBP6, MULT1, RAE1 α, β, γ, δ, and ε, A1R, A 2A R.A. 2B In various embodiments, the integrin is selected from the group consisting of αR, αR, A3R, H60a, H60b, and H60c. In various embodiments, the integrin is selected from the group consisting of α1, α2, αIIb, α3, α4, α5, α6, α7, α8, α9, α10, α11, αD, αE, αL, αM, αV, αX, β1, β2, β3, β4, β5, β6, β7, β8, or a combination thereof. In various embodiments, the TCR is selected from the group consisting of α, β, γ, δ, ε, and ζ TCRs. Several methods for assaying cell surface protein expression have been described in the literature, including flow cytometry and mass cytometry (CyTOF). The presence or abundance of one or more of these cell surface proteins indicates that the patient is suitable for treatment with the methods disclosed herein.

[0185] In various embodiments, analysis of cells from one or more blood samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" exhibits a high neutrophil-to-lymphocyte ratio (NLR). In various embodiments, analysis of cells from one or more blood samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" exhibits an NLR of 2 or greater. In various embodiments, analysis of cells from one or more blood samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" exhibits an NLR of 2-10 (e.g., an NLR of 2, 3, 4, 5, 6, 7, 8, 9, and 10, including all values ​​and ranges therebetween). In various embodiments, the NLR is used to determine the prognosis of a subject afflicted with cancer and having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold." In various embodiments, an NLR of 2 or greater is considered to be a poor prognosis.

[0186] In various embodiments, the cells analyzed from one or more blood samples taken from a subject suffering from cancer are circulating tumor cells (CTCs). In various embodiments, assays of one or more blood samples taken from a subject suffering from cancer show an increased frequency of CTCs compared to analyses of one or more blood samples taken from one or more healthy subjects, or one or more subjects suffering from cancer and responsive to treatment. In various embodiments, the frequency of circulating tumor cells in one or more blood samples from a subject with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is greater than or equal to 3 CTCs or less than or equal to 5 CTCs per 7.5 ml of blood.

[0187] In various embodiments, the tumor signature of a subject suffering from cancer is determined from an analysis of proteins in one or more blood samples from the subject. In various embodiments, the tumor signature of a subject suffering from cancer is determined by comparing the analysis of proteins in one or more blood samples from the subject suffering from cancer with an analysis of one or more blood samples from one or more healthy subjects or one or more subjects suffering from cancer and who are responsive to treatment. In various embodiments, the protein is an intracellular protein or a secreted protein. In various embodiments, the protein is selected from the group consisting of a cytokine, a chemokine, a growth factor, an enzyme, a protease, and a nuclease. In various embodiments, the cytokines and chemokines include IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-20, IL-21, IL-22 , IL-23, IL-24, IL-25, IL-26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33, IL-35, I L-36, CCL1, CCL2, CCL3, CCL4, CCL5, CCL6, CCL7, CCL8, CCL9, CCL10, CCL11, CCL12, CCL14, C CL15, CCL16, CCL17, CCL18, CCL19, CCL20, CCL21, CCL22, CCL23, CCL24, CCL25, CCL26, CCL27, CCL28, CXCL1, CXCL2 (MCP-1), CXCL3 (MIP-1α), CXCL4 (MIP-1β), CXCL5 (RANTES), CXCL6, CXCL7, CXCL8, CXCL9, CXCL10, CXCL11, CXCL12, CXCL13, CXCL14, CXCL15, CXCL16, CXCL17, IFN-α, IFN-β, IFN-γ, granzyme B, perforin, TNF-α, TGF-β1, TGF-β2, and TGF-β3.In various embodiments, the growth factor is selected from the group consisting of EGF, FGF, NGF, PDGF, VEGF, IGF, GMCSF, GCSF, TGF, erythropoietin, TPO, BMP, HGF, GDF, neurotrophins, MSF, SGF, GDF, G-CSF, and GM-CSF. In various embodiments, the protein is a protease selected from the group consisting of an aspartic acid protease, a cysteine ​​protease, a metalloprotease, a serine protease, or a threonine protease. In some embodiments, the protein is a protease selected from the group consisting of ADAM1, ADAM2, ADAM7, ADAM8, ADAM9, ADAM10, ADAM11, ADAM12, ADAM15, ADAM17, ADAM18, ADAM19, ADAM20, ADAM21, ADAM22, ADAM23, ADAM28, ADAM29, ADAM30, ADAM33, MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11, MMP12, MMP13, MMP14, MMP15, MMP16, MMP17, MMP18, MMP19, MMP20, MMP21, MMP23A, MMP23B, MMP24, MMP25, MMP26, MMP27, and MMP28. In various embodiments, the protein is an enzyme selected from the group consisting of arginase, asparaginase, kynureninase, indoleamine 2,3 dioxygenase (IDO1 and IDO2), tryptophan 2,3 dioxygenase (TDO), and IL4I1. In various embodiments, the protein is associated with apoptosis. In various embodiments, the protein associated with apoptosis is selected from the group consisting of P53, caspase 1, caspase 2, caspase 3, caspase 4, caspase 5, caspase 6, caspase 7, caspase 8, caspase 9, caspase 10, caspase 11, caspase 12, caspase 13, caspase 14, BCL-2, BCL-XL, MCL-1, CED-9, A1, BFL1, BAX, BAK, DIVA, BCL-XS, BIK, BIM, BAD, BID, and EGL-1. Several methods for assaying proteins from blood samples have been described in the literature, including Western blot and ELISA.

[0188] In various embodiments, analysis of proteins from one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reveals elevated levels of tumor-promoting, anti-inflammatory, or immunosuppressive proteins. In various embodiments, the tumor-promoting, anti-inflammatory, or immunosuppressive proteins are cell surface proteins, intracellular proteins, or secreted proteins. In various embodiments, the tumor-promoting, anti-inflammatory, or immunosuppressive proteins include CD39, CD79, MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11, MMP12, MMP13, MMP14, MMP15, MMP16, MMP17, MMP18, MMP19, MMP20, MMP21, MMP23A, MMP23B, MMP24, MMP25, MMP26, MMP27, MMP28, MMP29, MMP30, MMP31, MMP32, MMP33, MMP34, MMP35, MMP36, MMP37, MMP38, MMP39, MMP40, MMP41, MMP42, MMP43, MMP44, MMP45, MMP46, MMP47, MMP48, MMP49, MMP50, MMP51, MMP52, MMP53, MMP54, MMP55, MMP56, MMP57, MMP58, MMP59, MMP60, MMP61, MMP62, MMP63, MMP64, MMP65, MMP66, MMP67, MMP68, MMP69, MMP70, MMP71, MMP72, MMP73, MMP74, MMP75, MMP76, MMP77, MMP78, MMP79, MMP80, MMP 27, MMP28, CXCL12, GM-CSF, G-CSF, TGF-β1, TGF-β2, TGF-β3, arginase, asparaginase, kynureninase, indoleamine 2,3 dioxygenase (IDO1 and IDO2), tryptophan 2,3 dioxygenase (TDO), myeloperoxidase (MPO), neutrophil elastase (NE), and IL4I1. In various embodiments, the level of a tumor-promoting, anti-inflammatory, or immunosuppressive protein in one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 5-100% (e.g., about 5%, about 10%, about 15%, An increase of about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100%.In various embodiments, the level of a tumor-promoting, anti-inflammatory, or immunosuppressive protein in one or more blood samples from subjects with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 2 to 100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more blood samples taken from one or more healthy subjects, or one or more subjects with cancer who are responsive to treatment. In various embodiments, analysis of proteins from one or more blood samples from subjects with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows reduced, low, or absent levels of tumor-inhibitory, anti-tumor, or pro-inflammatory proteins. In various embodiments, the tumor inhibitory, anti-tumor, or pro-inflammatory protein is IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL -10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-20, IL-21, IL-22, IL-23, IL-24, IL-25, I L-26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33, IL-35, IL-36, cell surface IL-15, CXCL2(MCP-1), CXCL3( MIP-1α), CXCL4 (MIP-1β), CXCL5 (RANTES), IFN-α, IFN-β, IFN-γ, granzyme B, perforin, and TNF-α.In various embodiments, the level of a tumor-inhibiting, anti-tumor, or pro-inflammatory protein in one or more blood samples from a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 5-100% (e.g., increased by 5-100% compared to one or more blood samples taken from one or more healthy subjects, or one or more subjects having cancer and who are responsive to treatment) A reduction of about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to the original amount. In various embodiments, the level of a tumor-inhibitory, anti-tumor, or pro-inflammatory protein in one or more blood samples from a subject with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is reduced by 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more blood samples taken from one or more healthy subjects, or subjects with cancer who are responsive to treatment. Several methods for assaying proteins from blood samples have been described in the literature, including Western blot and ELISA.

[0189] In various embodiments, analysis of one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows elevated levels of neutrophil extracellular traps (NETs). In various embodiments, analysis of one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows elevated levels of neutrophil extracellular traps (NETs) compared to analysis of one or more blood samples from one or more healthy subjects, or one or more subjects with cancer and responsive to treatment. In various embodiments, the level of NETs in one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows elevated levels of neutrophil extracellular traps (NETs) compared to one or more blood samples taken from one or more healthy subjects, or one or more subjects with cancer and responsive to treatment. , about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges between these values), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% increase. In various embodiments, the level of NETs in one or more blood samples from subjects with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more blood samples taken from one or more healthy subjects or one or more subjects with cancer who are responsive to treatment. Several methods for assaying NETs from blood samples have been described in the literature, including Western blot, ELISA, and flow cytometry.

[0190] In various embodiments, a tumor signature of a subject suffering from cancer is determined from analysis of nucleic acids in one or more blood samples from the subject. In various embodiments, a tumor signature of a subject suffering from cancer is determined by comparing analysis of nucleic acids in one or more blood samples from the subject suffering from cancer with analysis of one or more blood samples from one or more healthy subjects or one or more subjects suffering from cancer and responsive to treatment. In various embodiments, the nucleic acid is selected from the group including DNA, ssDNA, circulating tumor DNA (ctDNA), RNA, mRNA, dsRNA, siRNA, miRNA, and lncRNA. In various embodiments, analysis of ctDNA from one or more blood samples from a subject suffering from cancer with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates low levels or absence of one or more tumor mutations, tumor antigens, or neoantigens. In various embodiments, analysis of ctDNA from one or more blood samples from a subject suffering from cancer indicates low or absent tumor mutation burden. In various embodiments, analysis of ctDNA from one or more blood samples from subjects with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reveals a tumor mutational burden of between 5 and 0.001 somatic mutations per megabase pair (e.g., about 5, about 4, about 3, about 2, about 1, about 0.9, about 0.8, about 0.7, about 0.6, about 0.5, about 0.4, about 0.3, about 0.2, about 0.1, about 0.09, about 0.08, about 0.07, about 0.06, about 0.05, about 0.04, about 0.03, about 0.02, about 0.01, about 0.009, about 0.008, about 0.007, about 0.006, about 0.005, about 0.004, about 0.003, about 0.002, or 0.001 (including all values ​​and ranges therebetween). In various embodiments, the nucleic acid analysis is performed by PCR, RT-PCR, qRT-PCR, next generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single cell sequencing.

[0191] In various embodiments, the subject's tumor characteristics are determined from gene expression analysis of nucleic acids in one or more blood samples from subjects afflicted with cancer. In various embodiments, gene expression analysis of nucleic acids in one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows increased expression of tumor-promoting, tumor-permissive, or immunosuppressive genes compared to analysis of one or more blood samples from one or more healthy subjects, or one or more subjects afflicted with cancer and responsive to treatment. In various embodiments, expression of tumor-promoting, tumor-permissive, or immunosuppressive genes in one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows an increase in expression of tumor-promoting, tumor-permissive, or immunosuppressive genes compared to one or more blood samples taken from one or more healthy subjects, or one or more subjects afflicted with cancer and responsive to treatment. %, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges between these values), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50% increase. In various embodiments, expression of tumor-promoting, tumor-permissive, or immunosuppressive genes in one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 2 to 100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges between these values)) compared to one or more blood samples taken from one or more healthy subjects, or one or more subjects afflicted with cancer and responsive to treatment.In various embodiments, analysis of nucleic acids in one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows decreased expression of tumor-inhibitory, anti-tumor, or pro-inflammatory genes compared to analysis of one or more blood samples from one or more healthy subjects, or one or more subjects with cancer and responsive to treatment. In various embodiments, analysis of nucleic acids in one or more blood samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows reduced or no expression of tumor-inhibitory, anti-tumor, or anti-inflammatory genes compared to analysis of one or more blood samples from one or more healthy subjects, or one or more subjects with cancer and responsive to treatment. In various embodiments, the expression of tumor-inhibiting, anti-tumor, or pro-inflammatory genes in one or more blood samples of a subject having one or more tumors that are characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 100%, about 110%, about 120%, about 130%, about 140%, about 150%, about 160%, about 170%, about 180%, about 210%, about 220%, about 230%, about 240%, about 250%, about 260%, about 270%, about 280%, about 300%, about 310%, about 320%, about 330%, about 340%, about 350%, about 360%, about 370%, about 380%, about 390%, about 400%, about 410%, about 420%, about 430%, about 440%, about 450%, about 460%, about 470%, about 480%, about 490%, about 500%, about 510%, about 520%, about 530%, about 540%, about 550%, about 560%, about 570%, about 580%, about 590%, about 610%, about 620%, about 630%, about 640%, about 650%, about 660%, about 670%, about 680%, about 690%, about 700%, about 710%, about 720%, about 730%, about 740%, about 750%, about 76 A decrease of 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50%. In various embodiments, expression of tumor-inhibiting, anti-tumor, or pro-inflammatory genes in one or more blood samples of subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is reduced by 2 to 100-fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100-fold (including all values ​​and ranges therebetween) compared to a healthy subject or a subject having cancer and who is responsive to treatment).In various embodiments, gene expression analysis is performed by PCR, RT-PCR, qRT-PCR, next generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single cell sequencing.

[0192] In various embodiments, tumor characteristics of a subject suffering from cancer are determined from analysis of one or more tumor samples obtained from the subject. In various embodiments, the tumor samples are biopsies. In various embodiments, tumor characteristics of a subject suffering from cancer are determined from analysis of cells, proteins, or nucleic acids in one or more tumor samples obtained from the subject. In various embodiments, tumor characteristics of a subject suffering from cancer are determined by comparing analysis of cells, proteins, or nucleic acids in one or more tumor samples from a subject suffering from cancer with analysis of tissue samples from one or more healthy subjects or one or more subjects suffering from cancer and responsive to treatment. In various embodiments, the cells analyzed in the one or more tumor samples are leukocytes, epithelial cells, mesenchymal cells, mesenchymal stem cells, stromal cells, endothelial cells, fibroblasts, pericytes, adipocytes, and cancer stem cells. In various embodiments, the leukocytes are myeloid cells and lymphoid cells. In various embodiments, the myeloid cells are monocytes, macrophages, neutrophils, granulocytes, dendritic cells, mast cells, eosinophils, and basophils. In various embodiments, the lymphoid cells are T cells, B cells, NK cells, NK-T cells, or iNK cells.

[0193] In various embodiments, analysis of cells from one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates the presence of immunosuppressive cells. In various embodiments, analysis of one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates the presence of immunosuppressive cells in the tumor core. In various embodiments, analysis of cells from one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates elevated levels of immunosuppressive cells. In various embodiments, analysis of one or more tumor samples indicates elevated levels of immunosuppressive cells in the tumor core. In various embodiments, immunosuppressive cells include myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), neutrophils, T reg cells, and B regIn various embodiments, the MDSCs are monocytic MDSCs (M-MDSCs) and polymorphonuclear MDSCs (PMN-MDSCs). In various embodiments, the TAMs are M2 TAMs. In various embodiments, the immunosuppressive cells are CAFs. In various embodiments, the level of immunosuppressive cells in one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, An increase of about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges between these values), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50%. In various embodiments, the level of immunosuppressive cells in one or more tumor samples from a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 2 to 100-fold compared to one or more tissue samples from one or more healthy subjects, or one or more subjects with cancer and responsive to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100-fold (including all values ​​and ranges between these values) compared to healthy subjects, or subjects with cancer and responsive to treatment).

[0194] In various embodiments, analysis of cells from one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows an absence of leukocytes. In various embodiments, analysis of cells from one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows a reduction or low level of leukocytes. In various embodiments, the frequency of leukocytes is 50% or less, 40% or less, 30% or less, 20% or less, 10% or less, or 5% or less of total cells analyzed (including all values ​​and ranges therebetween).

[0195] In various embodiments, analysis of cells from one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates the absence of activated pro-inflammatory immune cells. In various embodiments, analysis of cells from one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates the absence of activated pro-inflammatory immune cells from the tumor core. In various embodiments, analysis of cells from one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates low or reduced levels of activated pro-inflammatory immune cells. In various embodiments, analysis of cells from one or more tumor samples taken from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates low or reduced levels of activated pro-inflammatory immune cells in the tumor core. In various embodiments, the activated pro-inflammatory cells are dendritic cells (DCs), macrophages, M1 macrophages, T cells, B cells, NK cells, NK-T cells, and iNK cells. In various embodiments, the frequency of pro-inflammatory immune cells is 50% or less, 40% or less, 30% or less, 20% or less, 10% or less, or 5% or less of the total cells analyzed (including all values ​​and ranges therebetween).

[0196] In various embodiments, tumor characteristics of a subject suffering from cancer are determined from an analysis of the location of immune cells in one or more tumor samples from the subject. In various embodiments, immune cells in one or more tumor samples from subjects with one or more of immunorefractory, immunologically protected, or immunologically "cold" conditions are located in the tumor periphery. In various embodiments, immune cells in one or more tumor samples from subjects with one or more of immunorefractory, immunologically protected, or immunologically "cold" conditions are absent from the tumor core. In various embodiments, immune cells in one or more tumor samples from subjects with one or more of immunorefractory, immunologically protected, or immunologically "cold" conditions are reduced in the tumor core. In various embodiments, immune cells in the tumor core are reduced by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50% compared to one or more samples from one or more healthy subjects, or one or more subjects afflicted with cancer and responsive to treatment.

[0197] In various embodiments, tumor characteristics of a subject suffering from cancer are determined from an analysis of the location of stromal cells in one or more tumor samples from the subject. In various embodiments, the stromal cells are CAFs, pericytes, adipocytes, and endothelial cells. In various embodiments, CAFs in one or more tumor samples from a subject having one or more immunorefractory, immunologically protected, or immunologically "cold" tumors are increased in the tumor periphery. In various embodiments, CAFs in one or more tumor samples from a subject having one or more immunorefractory, immunologically protected, or immunologically "cold" tumors are increased in the tumor core. In various embodiments, the frequency of CAFs in the tumor periphery is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tissue samples from one or more healthy subjects or subjects afflicted with cancer and responsive to treatment. In various embodiments, the frequency of CAFs in the tumor periphery is increased by 2 to 100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 fold (including all values ​​and ranges therebetween)) compared to one or more tissue samples from one or more healthy subjects or one or more subjects afflicted with cancer and responsive to treatment.In various embodiments, the frequency of CAFs in the tumor core is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tissue samples from one or more healthy subjects, or one or more subjects afflicted with cancer and responsive to the treatment. In various embodiments, the frequency of CAFs in the tumor core is increased by 2 to 100-fold compared to one or more healthy tissue samples (e.g., by about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95-fold (including all values ​​and ranges between these values) compared to a healthy subject or a subject afflicted with cancer and responsive to treatment).

[0198] In various embodiments, the analysis of cells in one or more tumor samples from a subject suffering from cancer is performed by assaying for cell surface proteins, such as receptor tyrosine kinase (RTK), CD1c, CD2, CD3, CD4, CD5, CD8, CD9, CD10, CD11b, CD11c, CD14, CD15, CD16, CD18, CD19, CD20, CD21, CD22, CD23, CD24, TACI, CD25, CD27, CD28, CD30, CD30L, CD31, CD32, CD32b, CD34, CD33, CD38, CD39, CD40, CD40-L, CD41b, CD42a, CD42b, CD43, CD44, and CD45. , CD47, CD45RA, CD45RO, CD48, CD52, CD55, CD56, CD58, CD61, CD66b, CD70, CD72, CD79, CD68, CD84, CD86, CD93, CD94, CD95, CRACC, BLAME, BCMA, CD103 , CD107, CD112, CD120a, CD120b, CD123, CD125, CD134, CD135, CD140a, CD141, CD154, CD155, CD160, CD163, CD172a, XCR1, CD203c, CD204, CD206, CD207CD226, CD244, CD267, CD268, CD269, CD355, CD358, NKG2A, NKG2B, NKG2C, NKG2D, NKG2E, NKG2F, NKG2H, KIR2DL1, KIR2DL2, KIR2DL3, KIR2DL5A, KIR2DL5B, KIR3DL1, KIR3DL2, KIR3DL3, KIR3DL4, KIR2DS1, KIR2DS2, KIR2DS3, KIR2DS4, KIR2DS5, DAP12, KIR3DS, NKp44, NKp46, TCR, BCR, Integrin, FcβcRI, MHC-I, MHC-II, IL-1R, IL-2Rα, IL-2Rβ, IL-2Rγ, IL-3Rα, CSF2RB, IL-4R, IL-5Rα, IL-6Rα, gp130, IL-7Rα, IL-9R, IL-12Rβ1, IL-12Rβ2, IL-13Rα1, IL-13Rα2, IL-15Rα, IL-21R, IL-23R, IL-27Rα, IL-31Rα, OSMR, CSF-1R, Cell surface IL-15, IL-10Rα, IL-10Rβ, IL-20Rα, IL-20Rβ, IL-22Rα1, IL-22Rα2, IL-22Rβ, IL-28RA, PD-1, PD-1H, BTLA, CTLA-4, PD-L1, PD-L2, 2B4, B7-1, B7-2, B7-H1, B7-H4, B7-DC, DR3, LIGHT, LAIR, LTa1β2, LTβR, TIM-1, TIM-3, TIM-4, TIGIT, LAG-3, ICOS, ICOS-L, SLAM, SLAMF2, OX-40, OX-40L, GITR, GITRL, TL1A, HVEM, 41-BB, 41BB-L, TL-1A, TRAF1, TRAF2, TRAF3, TRAF5, BAFF, BAFF-R, APRIL, TRAIL, RANK, AITR, TRAMP, CCR1, CCR2, CCR3, CCR4, CCR5, CCR6, CCR7, CCR8, CCR9, CCR10, CCR11, CXCR1, CXCR2, CXCR3, CXCR4, CXCR5, CXCR6, CXCR7, CLECL9a, DC-SIGN, IGSF4A, SIGLEC, EGFR, PDGFR, VEGFR, FAP, α-SMA, Vimentin, Laminin, FAS, FAS-L, F c, ICAM-1, ICAM-2, ICAM-3, ICAM-4, ICAM-5, PECAM-1, MICA, MICB, UL16, ULBP1, ULBP2, ILBP3, ULBP4, ULBP5, ULBP6, MULT1, RAE1α, β, γ, δ, and ε, A1R, A 2A R.A. 2B In various embodiments, the integrin is selected from the group consisting of αR, αR, A3R, H60a, H60b, and H60c. In various embodiments, the integrin is selected from the group consisting of α1, α2, αIIb, α3, α4, α5, α6, α7, α8, α9, α10, α11, αD, αE, αL, αM, αV, αX, β1, β2, β3, β4, β5, β6, β7, β8, or a combination thereof. In various embodiments, the TCR is selected from the group consisting of α, β, γ, δ, ε, and ζ TCRs. Several methods for assaying cell surface protein expression from tumor samples have been described in the literature, including immunohistochemistry, immunofluorescence, Western blot, flow cytometry, and mass cytometry (CyTOF).

[0199] The tumor core is generally described as the dense, central, mass-forming, differentiated region of the tumor. In contrast, the tumor periphery is generally described as the infiltrative periphery of the tumor that interacts with the surrounding stroma and parenchyma.

[0200] In various embodiments, a tumor characteristic of a subject suffering from cancer is determined from an analysis of proteins in one or more tumor samples from the subject. In various embodiments, a tumor characteristic of a subject suffering from cancer is determined by comparing an analysis of proteins in one or more tumor samples from the subject suffering from cancer with an analysis of one or more tissues from one or more healthy subjects or one or more subjects suffering from cancer and responsive to treatment. In various embodiments, the protein is intracellular or extracellular. In various embodiments, the protein is selected from the group consisting of cytokines, chemokines, growth factors, enzymes, proteases, and nucleases. In various embodiments, the cytokines and chemokines include IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-20, IL-21, IL-22 , IL-23, IL-24, IL-25, IL-26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33, IL-35, I L-36, CCL1, CCL2, CCL3, CCL4, CCL5, CCL6, CCL7, CCL8, CCL9, CCL10, CCL11, CCL12, CCL14, C CL15, CCL16, CCL17, CCL18, CCL19, CCL20, CCL21, CCL22, CCL23, CCL24, CCL25, CCL26, CCL27, CCL28, CXCL1, CXCL2 (MCP-1), CXCL3 (MIP-1α), CXCL4 (MIP-1β), CXCL5 (RANTES), CXCL6, CXCL7, CXCL8, CXCL9, CXCL10, CXCL11, CXCL12, CXCL13, CXCL14, CXCL15, CXCL16, CXCL17, IFN-α, IFN-β, IFN-γ, granzyme B, perforin, TNF-α, TGF-β1, TGF-β2, and TGF-β3.In various embodiments, the growth factor is selected from the group consisting of EGF, FGF, NGF, PDGF, VEGF, IGF, GMCSF, GCSF, TGF, erythropoietin, TPO, BMP, HGF, GDF, neurotrophins, MSF, SGF, GDF, G-CSF, and GM-CSF. In various embodiments, the protein is a protease selected from the group consisting of an aspartic acid protease, a cysteine ​​protease, a metalloprotease, a serine protease, or a threonine protease. In some embodiments, the protein is a protease selected from the group consisting of ADAM1, ADAM2, ADAM7, ADAM8, ADAM9, ADAM10, ADAM11, ADAM12, ADAM15, ADAM17, ADAM18, ADAM19, ADAM20, ADAM21, ADAM22, ADAM23, ADAM28, ADAM29, ADAM30, ADAM33, MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11, MMP12, MMP13, MMP14, MMP15, MMP16, MMP17, MMP18, MMP19, MMP20, MMP21, MMP23A, MMP23B, MMP24, MMP25, MMP26, MMP27, and MMP28. In various embodiments, the protein is an enzyme selected from the group consisting of arginase, asparaginase, kynureninase, indoleamine 2,3 dioxygenase (IDO1 and IDO2), tryptophan 2,3 dioxygenase (TDO), and IL4I1. In various embodiments, the protein is associated with apoptosis. In various embodiments, the protein associated with apoptosis is selected from the group consisting of P53, caspase 1, caspase 2, caspase 3, caspase 4, caspase 5, caspase 6, caspase 7, caspase 8, caspase 9, caspase 10, caspase 11, caspase 12, caspase 13, caspase 14, BCL-2, BCL-XL, MCL-1, CED-9, A1, BFL1, BAX, BAK, DIVA, BCL-XS, BIK, BIM, BAD, BID, and EGL-1. Several methods for assaying proteins from tumor samples have been described in the literature, including immunohistochemistry, immunofluorescence, Western blot, and ELISA.

[0201] In various embodiments, analysis of proteins from one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reveals elevated levels of proteins associated with tumor progression, anti-inflammatory activity, or immunosuppression. In various embodiments, the proteins associated with tumor progression, anti-inflammatory activity, or immunosuppression are cell surface proteins, intracellular proteins, or secreted proteins. In various embodiments, the proteins associated with tumor progression, anti-inflammatory activity, or immunosuppression are CD39, CD47, CD79, CD140a, CD163, CD206, FOXP3, FAP, PD-1, PD-L1, PD-L2, CSF-1R, A1R, A2R, A3R, A4R, A5R, A6R, A7R, A8R, A9R, A1 ... 2A R.A. 2BR, A3R, TIM-1, TIM-3, TIM-4, TIGIT, CSFR, SIGLEC, MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11, MMP12, M MP13, MMP14, MMP15, MMP16, MMP17, MMP18, MMP19, MMP20, MMP21, MMP23A, MMP23B, MMP24, MMP25, MMP26, MMP27, In various embodiments, the level of a protein associated with tumor progression, anti-inflammatory activity, or immunosuppression in one or more tumor samples from a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 5 to 100% (e.g., about 5%, about 10%, about 15%) compared to one or more tissue samples from one or more healthy subjects, or one or more subjects afflicted with cancer and responsive to treatment. %, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges between these values), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% increase.In various embodiments, the level of a protein associated with tumor progression, anti-inflammatory activity, or immunosuppression in one or more tumor samples from a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" is increased by 2 to 100 fold compared to one or more tissue samples from one or more healthy subjects, or subjects with cancer and responsive to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 fold (including all values ​​and ranges between these values) compared to healthy subjects, or subjects with cancer and responsive to treatment).

[0202] In various embodiments, analysis of proteins from one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows reduced levels, low levels, or absence of proteins associated with tumor growth inhibition, anti-tumor activity, or pro-inflammatory activity. In various embodiments, the protein associated with tumor growth inhibition, anti-tumor activity, or pro-inflammatory activity is CD44, CD56, CD103c, CD69, KG2A, NKG2B, NKG2C, NKG2D, NKG2E, NKG2F, NKG2H, ICOS, ICOS-L, SLAM, SLAMF2, OX-40, OX-40L, GITR, GITRL, TL-1A, HVEM, 41-BB, 41BB-L, TRAF1, TRAF2, TRAF3, TRAF5, BAFF, BAFF-R, APRIL, TRAIL, RANK, AITR, TRAMP, IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-19, IL-20, IL-21, IL-19, IL-19, IL-18, IL-19, IL-22, IL-19, IL-19, IL-19, IL-23, IL-19 ... L-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-20, IL-21, IL-22, IL-23, IL-24, IL-25, IL-26, IL-27, IL-28, IL-29, IL-30 , IL-31, IL-32, IL-33, IL-35, IL-36, CXCL2 (MCP-1), CXCL3 (MIP-1α), CXCL4 (MIP-1β), CXCL5 (RANTES), IFN-α, IFN-β, IFN-γ, granzyme B, perforin, and TNF-α.In various embodiments, the level of a protein associated with tumor growth inhibition, anti-tumor activity, or pro-inflammatory activity is reduced by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more samples taken from one or more healthy tissues or one or more tumor samples taken from a subject afflicted with cancer and responsive to the treatment. Several methods have been described for assaying proteins from tumor samples, including immunohistochemistry, immunofluorescence, Western blot, intracellular flow cytometry, and ELISA.

[0203] In various embodiments, tumor characteristics of a subject afflicted with cancer are determined from the tumor proportion score (TPS) of PD-L1 expression in one or more tumor samples from the subject. In various embodiments, a subject with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" has a TPS of 1 to 50 (e.g., 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, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50 (including all ranges between these values)). In various embodiments, subjects with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" have a TPS of less than or equal to 1. The TPS for PD-L1 expression is defined as the percentage of viable tumor cells that show partial or complete membrane staining by immunohistochemical analysis.

[0204] In various embodiments, tumor characteristics of a subject afflicted with cancer are determined from the combined positive score (CPS) of PD-L1 expression in one or more tumor samples from the subject. In various embodiments, a subject with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" has a CPS of 10 or less (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, including all ranges between these values). In various embodiments, the CPS is 1 or less. The CPS of PD-L1 expression is determined by immunohistochemically measuring the number of viable PD-L1 positive tumor cells, lymphocytes, and macrophages as a percentage of total viable tumor cells.

[0205] In various embodiments, a tumor characteristic of a subject suffering from cancer is determined from microsatellite instability testing of one or more tumor samples from the subject. In various embodiments, a tumor characteristic of a subject suffering from cancer is determined by comparing microsatellite instability testing of one or more tumor samples with microsatellite stability testing of one or more healthy tissues from the subject. In various embodiments, the microsatellite instability testing is an assay for a microsatellite marker. In various embodiments, the microsatellite instability testing is an assay for a mismatch repair marker. In various embodiments, the microsatellite marker is selected from the group consisting of BAT25, BAT26, D2S123, D5S346, and D17S250. In various embodiments, the mismatch repair marker is selected from the group consisting of MLH1, MSH2, MLH6, and PMS2. In various embodiments, the subject has one or more immunorefractory, immunologically protected, or immunologically "cold" tumors determined to have low microsatellite instability. In various embodiments, the subject has one or more immunorefractory, immunoprotected, or immunologically "cold" tumors that are determined to be microsatellite stable. In various embodiments, the subject has one or more immunorefractory, immunoprotected, or immunologically "cold" tumors that are mismatch repair competent.

[0206] In various embodiments, analysis of one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows elevated levels of neutrophil extracellular traps (NETs). In various embodiments, analysis of one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows elevated levels of neutrophil extracellular traps (NETs) compared to analysis of one or more tumor samples from one or more healthy subjects. In various embodiments, the level of NETs in one or more tumor samples from a subject afflicted with cancer is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tissue samples from one or more healthy subjects, or subjects afflicted with cancer and responsive to treatment. In various embodiments, the level of NETs in one or more tumor samples from a subject suffering from cancer is increased 2-100 fold compared to one or more tissue samples from one or more healthy subjects, or subjects suffering from cancer and responsive to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 fold (including all values ​​and ranges therebetween) compared to healthy subjects, or subjects suffering from cancer and responsive to treatment). Several methods for assaying NETs have been described in the literature, including Western blot, ELISA, and flow cytometry.

[0207] In various embodiments, tumor characteristics of a subject suffering from cancer are determined from analysis of nucleic acids in one or more tumor samples from the subject. In various embodiments, tumor characteristics of a subject suffering from cancer are determined by comparing analysis of nucleic acids in one or more tumor samples from the subject suffering from cancer with analysis of one or more tissue samples from one or more healthy subjects or subjects suffering from cancer and responsive to treatment. In various embodiments, the nucleic acid is selected from the group including DNA, ssDNA, RNA, mRNA, dsRNA, siRNA, miRNA, and lncRNA. In various embodiments, nucleic acid analysis is performed by PCR, RT-PCR, qRT-PCR, next-generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single-cell sequencing.

[0208] In various embodiments, analysis of nucleic acids from one or more tumor samples from subjects afflicted with cancer is used to determine tumor mutation burden. In various embodiments, analysis of nucleic acids from one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates a low tumor mutation burden. In various embodiments, analysis of nucleic acids from one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" indicates a tumor mutation burden of between 5 and 0.001 somatic mutations per megabase pair (e.g., about 5, about 4, about 3, about 2, about 1, about 0.9, about 0.8, about 0.7, about 0.6, about 0.5, about 0.6, about 0.7, about 0.8, about 0.9 ... 0.4, about 0.3, about 0.2, about 0.1, about 0.09, about 0.08, about 0.07, about 0.06, about 0.05, about 0.04, about 0.03, about 0.02, about 0.01, about 0.009, about 0.008, about 0.007, about 0.006, about 0.005, about 0.004, about 0.003, about 0.002, or 0.001 (including all values ​​and ranges therebetween). In various embodiments, the nucleic acid analysis is performed by PCR, RT-PCR, qRT-PCR, next generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single cell sequencing.

[0209] In various embodiments, analysis of nucleic acids in one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows increased expression of genes associated with tumor-promoting, tumor-permissive, anti-inflammatory, or immunosuppressive activity compared to analysis of one or more tissue samples from one or more healthy subjects, or subjects afflicted with cancer and responsive to treatment. In various embodiments, genes associated with tumor-promoting, tumor-permissive, anti-inflammatory, or immunosuppressive activity include CD39, CD47, CD79, CD140a, CD163, CD206, FOXP3, FAP, PD-1, PD-L1, PD-L2, CSF-1R, A1R, A2R, A3R, A4R, A5R, A6R, A7R, A8R, A9R, A1 ... 2A R.A. 2BR, A3R, TIM-1, TIM-3, TIM-4, TIGIT, CSFR, SIGLEC, MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11, MMP12, M MP13, MMP14, MMP15, MMP16, MMP17, MMP18, MMP19, MMP20, MMP21, MMP23A, MMP23B, MMP24, MMP25, MMP26, MMP27, Selected from the group consisting of MMP28, CXCL12, GM-CSF, G-CSF, FAP, TGF-β1, TGF-β2, TGF-β3, arginase, asparaginase, kynureninase, indoleamine 2,3 dioxygenase (IDO1 and IDO2), tryptophan 2,3 dioxygenase (TDO), myeloperoxidase (MPO), neutrophil elastase (NE), and IL4I1. In various embodiments, expression of genes associated with tumor-promoting, tumor-permissive, anti-inflammatory, or immunosuppressive activity is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tissue samples from one or more healthy subjects, or from subjects afflicted with cancer and responsive to treatment. In various embodiments, gene expression analysis is performed by PCR, RT-PCR, qRT-PCR, next generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single cell sequencing.

[0210] In various embodiments, analysis of nucleic acids in one or more tumor samples from subjects having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" shows low or decreased expression of genes associated with tumor inhibitory, anti-tumor, or pro-inflammatory activity. In various embodiments, analysis of nucleic acids in one or more tumor samples from subjects afflicted with cancer shows no expression of genes associated with tumor inhibitory, anti-tumor, or pro-inflammatory activity. In various embodiments, the gene associated with tumor inhibitory, anti-tumor, or pro-inflammatory activity is CD44, CD56, CD103c, CD69, KG2A, NKG2B, NKG2C, NKG2D, NKG2E, NKG2F, NKG2H, ICOS, ICOS-L, SLAM, SLAMF2, OX-40, OX-40L, GITR, GITRL, TL-1A, HVEM, 41-BB, 41BB-L, TRAF1, TRAF2, TRAF3, TRAF5, BAFF, BAFF-R, APRIL, TRAIL, RANK, AITR, TRAMP, cell surface IL-15, IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-15β, IL-16, IL-17, IL-18, IL-19, IL-20, IL-21, IL-19, IL-16, IL-19, IL-18, IL-19, IL-22, IL-19, IL-19, IL-19, IL-19, IL-23, IL-19 ... L-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL- 18, IL-20, IL-21, IL-22, IL-23, IL-24, IL-25, IL-26, IL-27, IL-28, IL-29, IL-30, I selected from the group consisting of L-31, IL-32, IL-33, IL-35, IL-36, CXCL2 (MCP-1), CXCL3 (MIP-1α), CXCL4 (MIP-1β), CXCL5 (RANTES), IFN-α, IFN-β, IFN-γ, granzyme B, perforin, TNF-α, and p53.In various embodiments, expression of genes associated with tumor inhibitory, anti-tumor, or pro-inflammatory activity is reduced by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50% compared to a healthy subject or a subject afflicted with cancer and responsive to treatment. In various embodiments, gene expression analysis is performed by PCR, RT-PCR, qRT-PCR, next generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single cell sequencing.

[0211] In some embodiments, the methods of the present disclosure include analyzing a first and a second biological sample (e.g., a blood sample or a tumor sample) obtained from a subject, where "first" and "second" refer to the order in which the samples were collected. In some embodiments, a first, second, third, fourth, or fifth biological sample can be obtained and analyzed. The biological samples can be collected days, weeks, or months apart. Two or more biological samples can be analyzed as described herein above. For example, a first biological sample can be obtained from the subject before administration of a RAS inhibitor described herein, and a second biological sample can be obtained from the subject after administration of a RAS inhibitor described herein.

[0212] In various embodiments, the present disclosure provides a method of treating cancer in a subject, comprising administering to the subject a RAS inhibitor or a combination of compounds described herein, wherein the subject has one or more tumors that are resistant or refractory to treatment. In various embodiments, the subject has one or more tumors that are resistant or refractory to one or more treatments selected from the group consisting of surgery, radiation, chemotherapy, biologic agents, small molecules, cell-based therapies, hormonal therapies, and immunotherapy. In various embodiments, the treatment is standard of care, first-line therapy, second-line therapy, or third-line therapy. In various embodiments, the subject has one or more tumors that have progressed during one or more treatments that are standard of care, first-line therapy, second-line therapy, or third-line therapy.

[0213] First-line therapy is defined as a treatment administered to a subject with cancer who has not received any prior treatment. Second-line therapy is defined as a treatment administered to a subject with cancer who has previously received first-line therapy but experienced disease progression during the first-line therapy. Third-line therapy is defined as a treatment administered to a subject with cancer who has previously received first- and second-line therapy but experienced disease progression during the second-line therapy. Each specific type of cancer has first-line, second-line, and third-line therapies. First-, second-, and third-line therapies for each type of cancer are known in the art. Furthermore, FDA-approved drug labels indicate whether a particular drug is approved as a first-, second-, or third-line therapy.

[0214] In order to determine the effect of one or more treatments on the tumor of a subject suffering from cancer, several criteria and definitions published in the literature can be used. Based on these criteria, a tumor is defined as "responsive", "stable" or "progressive" if it improves, remains the same or worsens during treatment, respectively.

[0215] Examples of commonly used criteria published in the literature include Response Evaluation Criteria in Solid Tumors (RECIST), modified Response Evaluation Criteria in Solid Tumors (mRECIST), PET Response Criteria in Solid Tumors (PERCIST), Choi criteria, Lugano response criteria, European Association for the Study of the Liver (EASL) criteria, Response Evaluation Criteria in Liver Cancer (RECICL), and WHO criteria for tumor response.

[0216] In various embodiments, the present disclosure provides a method of treating cancer in a subject comprising administering to the subject a RAS inhibitor or a combination of compounds described herein, wherein the subject is intolerant to standard, first-line, second-line, or third-line therapy. In various embodiments, the present disclosure provides a method of treating cancer in a subject comprising administering to the subject a RAS inhibitor or a combination of compounds described herein, wherein the subject has experienced tumor recurrence after surgical resection of the primary tumor. In various embodiments, the present disclosure provides a method of treating cancer in a subject comprising administering to the subject a RAS inhibitor or a combination of compounds described herein, wherein the subject has a tumor that cannot be surgically removed. In various embodiments, the present disclosure provides a method of treating cancer in a subject comprising administering to the subject a RAS inhibitor or a combination of compounds described herein, wherein the subject has no available treatment options.

[0217] Some therapies used to treat cancer (e.g., chemotherapy) are cytotoxic and associated with significant side effects and toxicities associated with poor outcomes and poor response to treatment. Prior to administering such treatments, clinicians rely on several assessment tools to help determine a cancer patient's risk of experiencing treatment-related toxicities and adverse events. Based on the results of these assessments, a cancer patient is considered intolerant to treatment if they are determined to be at high risk of experiencing treatment-related toxicities and adverse events that result in poor outcomes. Examples of commonly used assessment tools used to determine treatment intolerance include the Karnofsky Performance Status (KPS), Eastern Cooperative Oncology Group Performance Status (ECOG PS), Timed Get Up and Go (TUG), Short Physical Performance Battery (SPPB), Comprehensive Geriatric Assessment (CGA), Cancer Aging Research Group (CARG) score, and the Chemotherapy Risk Assessment Scale for Elderly Patients (CRASH).

[0218] Treatment outcomes and clinical endpoints In various embodiments, the present disclosure provides a method for treating lung cancer in a subject, the method comprising administering to the subject a RAS inhibitor or a combination of compounds described herein, the subject having one or more immunorefractory tumors. In various embodiments, the administration alters the tumor immune infiltrate. In various embodiments, the administration alters the anti-tumor immune response. In various embodiments, the administration alters the tumor microenvironment, including tumor cells, immune cells, cancer stem cells, and stroma. In various embodiments, the administration converts an immunologically cool tumor to an immunologically hot tumor. In various embodiments, the administration reduces tumor size or inhibits tumor growth. In various embodiments, the administration induces tumor cell death, apoptosis, or necrosis via direct particle uptake by tumor cells.

[0219] In various embodiments, the present disclosure provides methods of treating cancer in a subject, the method comprising administering to the subject a RAS inhibitor or combination of compounds described herein, the subject having one or more tumors characterized as immunologically protected or immunorefractory. In various embodiments, the administration alters tumor-associated stroma, including fibroblasts, cancer-associated fibroblasts, adipocytes, pericytes, endothelium, vasculature, lymphatic vessels, tumor-associated vasculature, mesenchymal stromal cells, mesenchymal stem cells, and extracellular matrix.

[0220] It is contemplated that the methods herein reduce tumor size or tumor burden in a subject, or reduce metastasis in a subject. In various embodiments, the methods reduce tumor size by 10%, 20%, 30%, or more. In various embodiments, the methods reduce tumor size by 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% (or all values ​​and ranges therebetween).

[0221] When tumors become immunorefractory, the abundance of certain biomarkers may decrease.It is contemplated herein that after treatment with a RAS inhibitor or a combination of compounds described herein, the level of one or more of the biomarkers increases by an amount ranging from about 1.1 times to about 10 times, for example, about 1.1, about 1.5, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 5.5, about 6, about 6.5, about 7, about 7.5, about 8, about 8.5, about 9, about 9.5, or about 10 times.Similarly, when tumors become immunorefractory, the abundance of certain biomarkers increases. Following treatment with a RAS inhibitor or combination of compounds described herein, the levels of one or more of such biomarkers are reduced by an amount ranging from about 1.1-fold to about 10-fold, e.g., about 1.1, about 1.5, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 5.5, about 6, about 6.5, about 7, about 7.5, about 8, about 8.5, about 9, about 9.5, or about 10-fold. In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the levels of immunosuppressive cells in the blood. In various embodiments, the suppressive cells include myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), neutrophils, T cells, and the like. reg cells, and B regIn various embodiments, the MDSCs are monocytic MDSCs (M-MDSCs) and polymorphonuclear MDSCs (PMN-MDSCs). In various embodiments, the TAMs are M2 TAMs. In various embodiments, the immunosuppressive cells are CAFs. In various embodiments, the level of immunosuppressive cells is reduced by about 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50% compared to one or more blood samples taken from the subject prior to treatment. In various embodiments, the level of immunosuppressive cells is reduced by about 2 to 100-fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100-fold (including all values ​​and ranges between these values)) compared to one or more blood samples taken from the subject prior to treatment. In various embodiments, the immunosuppressive cells are identified by assaying cell surface protein expression. In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the level of activated pro-inflammatory immune cells by 5 to 100% (e.g., about 5%, about 10%, about 15%, about 20%) compared to one or more blood samples taken from the subject prior to treatment. , about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges between these values), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50% increase.In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" increases the level of activated pro-inflammatory immune cells by 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more blood samples taken from the subject prior to treatment. In various embodiments, the activated pro-inflammatory cells are dendritic cells (DCs), macrophages, M1 macrophages, T cells, B cells, NK cells, NK-T cells, and iNK cells. In various embodiments, the frequency of pro-inflammatory immune cells is increased to 10-50% (e.g., about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% (including all values ​​and ranges between these values)) of total white blood cells analyzed from one or more blood samples taken from the subject. In various embodiments, activated pro-inflammatory immune cells are identified by assay of cell surface protein expression.

[0222] In various embodiments, the analysis of cells in one or more blood samples from a subject suffering from cancer is performed by assaying for cell surface proteins, such as receptor tyrosine kinase (RTK), CD1c, CD2, CD3, CD4, CD5, CD8, CD9, CD10, CD11b, CD11c, CD14, CD15, CD16, CD18, CD19, CD20, CD21, CD22, CD23, CD24, TACI, CD25, CD27, CD28, CD30, CD30L, CD31, CD32, CD32b, CD34, CD33, CD38, CD39, CD40, CD40-L, CD41b, CD42a, CD42b, CD43, CD44, CD48 , CD47, CD45RA, CD45RO, CD48, CD52, CD55, CD56, CD58, CD61, CD66b, CD70, CD72, CD79, CD68, CD84, CD86, CD93, CD94, CD95, CRACC, BLAME, BCMA, CD103 , CD107, CD112, CD120a, CD120b, CD123, CD125, CD134, CD135, CD140a, CD141, CD154, CD155, CD160, CD163, CD172a, XCR1, CD203c, CD204, CD206, CD207CD226、CD244、CD267、CD268、CD269、CD355、CD358、NKG2A、NKG2B、NKG2 C、NKG2D、NKG2E、NKG2F、NKG2H、KIR2DL1、KIR2DL2、KIR2DL3、KIR2DL5A、 KIR2DL5B、KIR3DL1、KIR3DL2、KIR3DL3、KIR3DL4、KIR2DS1、KIR2DS2、KI R2DS3、KIR2DS4、KIR2DS5、DAP12、KIR3DS、NKp44、NKp46、TCR、BCR、インテグリン、FcβεRI、MHC-I、MHC-II、IL-1R、IL-2Rα、IL-2Rβ、IL-2Rγ、IL-3Rα、CSF2RB、IL-4R、IL-5Rα、IL-6Rα、gp130、IL-7Rα、IL-9R、IL-12Rβ1、IL-12R β2, IL-13Rα1, IL-13Rα2, IL-15Rα, IL-21R, IL-23R, IL-27Rα, IL-31Rα, OSMR, CSF-1R, IL-15, IL-10Rα, IL-10Rβ, IL-20Rα, IL-20Rβ, IL-22 Rα1、IL-22Rα2、IL-22Rβ、IL-28RA、PD-1、PD-1H、BTLA、CTLA-4、PD-L1、P D-L2、2B4、B7-1、B7-2、B7-H1、B7-H4、B7-DC、DR3、LIGHT、LAIR、LTα1β2、 LTβR、TIM-1、TIM-3、TIM-4、TIGIT、LAG-3、ICOS、ICOS-L、SLAM、SLAMF2、 OX-40、OX-40L、GITR、GITRL、TL1A、HVEM、41-BB、41BB-L、TL-1A、TRAF1、 TRAF2、TRAF3、TRAF5、BAFF、BAFF-R、APRIL、TRAIL、RANK、AITR、TRAMP、C CR1、CCR2、CCR3、CCR4、CCR5、CCR6、CCR7、CCR8、CCR9、CCR10、CCR11、CXC R1、CXCR2、CXCR3、CXCR4、CXCR5、CXCR6、CXCR7、CLECL9a、DC-SIGN、IGSF 4A、SIGLEC、EGFR、PDGFR、VEGFR、FAP、α-SMA、ビメンチン、ラミニン、FAS、FAS-L、F c, ICAM-1, ICAM-2, ICAM-3, ICAM-4, ICAM-5, PECAM-1, MICA, MICB, UL16, ULBP1, ULBP2, ILBP3, ULBP4, ULBP5, ULBP6, MULT1, RAE1 α, β, γ, δ, and ε, A1R, A 2A R.A. 2B In various embodiments, the integrin is selected from the group consisting of αR, αR, A3R, H60a, H60b, and H60c. In various embodiments, the integrin is selected from the group consisting of α1, α2, αIIb, α3, α4, α5, α6, α7, α8, α9, α10, α11, αD, αE, αL, αM, αV, αX, β1, β2, β3, β4, β5, β6, β7, β8, or a combination thereof. In various embodiments, the TCR is selected from the group consisting of α, β, γ, δ, ε, and ζ TCRs. Several methods for assaying cell surface protein expression have been described in the literature, including flow cytometry and mass cytometry (CyTOF). The presence or abundance of one or more of these cell surface proteins indicates that the patient will be responsive to treatment with the methods disclosed herein.

[0223] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the neutrophil-to-lymphocyte (NLR) in one or more blood samples from high to moderate, or from high to low. In various embodiments, analysis of cells from one or more blood samples taken from a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the NLR to 1 to 2 (e.g., 1, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, 1.9, and 2, including all values ​​and ranges therebetween). In various embodiments, after administration of a RAS inhibitor or combination of compounds described herein, the NLR is reduced. In various embodiments, after administration of a RAS inhibitor or combination of compounds described herein, the NLR is less than 2.

[0224] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the level of CTCs in one or more blood samples, hi various embodiments, the level of blood CTCs is reduced to 5, 4, 3, 2, 1, or 0 (including all values ​​and ranges between these values) per 7.5 ml of blood.

[0225] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the level of tumor-promoting, anti-inflammatory, or immunosuppressive proteins in one or more blood samples from the subject. In various embodiments, the tumor-promoting, anti-inflammatory, or immunosuppressive proteins include CD39, CD79, MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11, MMP12, MMP13, MMP14, MMP15, MMP16, MMP17, MMP18, MMP19, MMP20, MMP21, MMP23A, MMP23B, MMP24, MMP25, MMP26, MMP27, or the like. and MMP28, CXCL12, GM-CSF, G-CSF, TGF-β1, TGF-β2, and TGF-β3, arginase, asparaginase, kynureninase, indoleamine 2,3 dioxygenase (IDO1 and IDO2), tryptophan 2,3 dioxygenase (TDO), myeloperoxidase (MPO), neutrophil elastase (NE), and IL4I1. In various embodiments, the level of a tumor-promoting, anti-inflammatory, or immunosuppressive protein in one or more blood samples from the subject is reduced by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more blood samples taken before treatment. In various embodiments, the level of a tumor-promoting, anti-inflammatory, or immunosuppressive protein in one or more blood samples from the subject is reduced by 2 to 100-fold compared to one or more blood samples taken from the subject prior to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100-fold (including all values ​​and ranges therebetween) compared to one or more samples taken prior to treatment).

[0226] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" increases the level of a tumor-inhibiting, anti-tumor, or pro-inflammatory protein in one or more blood samples taken from the subject. In various embodiments, the tumor-inhibiting, anti-tumor, or pro-inflammatory protein is selected from the group consisting of IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-20, IL-21, IL-22, IL-23, IL-24, IL-25, IL-26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33, IL-34, IL-35, IL-36, IL-37, IL-38, IL-39, IL-40, IL-41, IL-42, IL-43, IL-44, IL-45, IL-46, IL-47, IL-48, IL-49, IL-50, IL-51, IL-52, IL-53, IL-54, IL-55, IL-56, IL-57, IL-58, IL-59, IL-60, IL-61, IL-62, IL-63, IL-64, IL-65, IL-66, IL-67, IL-68, IL-69, L-26, IL-27, IL-28, IL-29, IL-30, IL-31, IL-32, IL-33, IL-35, IL-36, cell surface IL-15, CXCL2(MCP-1), CXCL3( MIP-1α), CXCL4 (MIP-1β), CXCL5 (RANTES), IFN-α, IFN-β, IFN-γ, granzyme B, perforin, and TNF-α. In various embodiments, the level of an anti-tumor or pro-inflammatory protein is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more blood samples taken before treatment. In various embodiments, the level of the anti-tumor or pro-inflammatory protein is increased by 2-100 fold compared to one or more blood samples taken from the subject prior to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween) compared to one or more samples taken prior to treatment).Several methods for assaying proteins from blood samples have been described in the literature, including Western blot and ELISA.

[0227] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the level of neutrophil extracellular traps (NETs) in one or more blood samples taken from the subject. In various embodiments, the level of NETs in one or more blood samples is reduced by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more blood samples taken before treatment. In various embodiments, the level of NETs in one or more blood samples is reduced by 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more blood samples taken from the subject prior to treatment. Several methods for assaying NETs from blood samples have been described in the literature, including Western blot, ELISA, and flow cytometry.

[0228] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the expression of tumor-promoting, tumor-permissive, or immunosuppressive genes in one or more blood samples from the subject. In one or more embodiments, expression of a tumor-promoting, tumor-permissive, or immunosuppressive gene is reduced by 5-100% compared to one or more blood samples taken before treatment (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween) compared to levels in one or more blood samples taken before treatment). In one or more embodiments, expression of a tumor-promoting, tumor-permissive, or immunosuppressive gene is reduced by 2 to 100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more blood samples taken from the subject prior to treatment.

[0229] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" increases the expression of tumor-inhibitory, anti-tumor, or pro-inflammatory genes in one or more samples taken from the subject. In one or more embodiments, expression of a tumor-inhibiting, anti-tumor, or pro-inflammatory gene is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more blood samples taken before treatment. In various embodiments, expression of the tumor-inhibiting, anti-tumor, or pro-inflammatory gene is increased by 2-100 fold compared to one or more blood samples taken from the subject prior to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween) compared to one or more samples taken prior to treatment). In various embodiments, gene expression analysis is performed by PCR, RT-PCR, qRT-PCR, next-generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single-cell sequencing.

[0230] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" increases the level of leukocytes in the tumor, hi various embodiments, the level of leukocytes is increased in the tumor core or the tumor periphery. In various embodiments, white blood cells are increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50% compared to one or more tumor samples taken from the subject prior to treatment. In various embodiments, the level of leukocytes is increased by 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold, including all values ​​and ranges therebetween) compared to one or more tumor samples taken from the subject prior to treatment. In various embodiments, the frequency of leukocytes in the tumor core or peritumor area is 5% or more, 10% or more, 15% or more, 20% or more, 25% or more, 30% or more, 35% or more, 40% or more, 45% or more, or 50% or more of the total cells analyzed, including all values ​​and ranges therebetween.

[0231] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the level of immunosuppressive cells in the tumor. In various embodiments, the level of immunosuppressive cells is reduced in the tumor core or the tumor periphery. In various embodiments, the suppressive cells include myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), neutrophils, T cells, and the like. reg cells, and B regIn various embodiments, the MDSCs are monocytic MDSCs (M-MDSCs) and polymorphonuclear MDSCs (PMN-MDSCs). In various embodiments, the TAMs are M2 TAMs. In various embodiments, the immunosuppressive cells are CAFs. In various embodiments, the level of immunosuppressive cells is reduced by about 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50% compared to one or more tumor samples taken from the subject prior to treatment. In various embodiments, the level of immunosuppressive cells is reduced by about 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more tumor samples taken from the subject prior to treatment. In various embodiments, the immunosuppressive cells are identified by assaying cell surface protein expression.

[0232] The level of leukocytes in tumor samples can be assessed by several methods, including flow cytometry and immunohistochemistry. In various embodiments, administering a RAS inhibitor or a combination of compounds described herein to a subject with one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" increases the level of activated pro-inflammatory immune cells in the tumor. In various embodiments, the level of activated pro-inflammatory cells is increased in the tumor core or the tumor periphery.

[0233] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the level of activated pro-inflammatory immune cells in the tumor by 5 to 100% (e.g., about 5%, about 10%, about 15%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 100%, about 110%, about 120%, about 130%, about 140%, about 150%, about 160%, about 170%, about 180%, about 190%, about 210%, about 220%, about 230%, about 240%, about 250%, about 260%, about 270%, about 280%, about 290%, about 300%, about 310%, about 320%, about 330%, about 340%, about 350%, about 360%, about 370%, about 380%, about 390%, about 400%, about 410%, about 420%, about 430%, about 440%, about 450%, about 460%, about 470%, about 480%, about 490%, about 500%, about 510%, about 520%, about 530%, about 540%, about 550%, about 560%, about 570%, about 580%, about 590%, about 610%, about 620%, about 630%, about 640%, about 650%, about 660%, about 670%, about 680%, about 690%, about 700%, about 710%, about 720%, about 730%, about 740%, about 750%, An increase of 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges between these values), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, or 50%. In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" increases the level of activated pro-inflammatory immune cells by 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more tumor samples taken from the subject prior to treatment. In various embodiments, the activated pro-inflammatory cells are dendritic cells (DCs), macrophages, M1 macrophages, T cells, B cells, NK cells, NK-T cells, and NK cells. In various embodiments, the frequency of pro-inflammatory immune cells is about 10-50% (e.g., about 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, or 50% (including all values ​​and ranges between these values)) of total leukocytes analyzed from one or more tumor samples collected from the subject. In various embodiments, activated pro-inflammatory immune cells are identified by assaying cell surface protein expression.

[0234] In various embodiments, the analysis of cells in one or more tumor samples from a subject suffering from cancer is performed by assaying for cell surface proteins, such as receptor tyrosine kinase (RTK), CD1c, CD2, CD3, CD4, CD5, CD8, CD9, CD10, CD11b, CD11c, CD14, CD15, CD16, CD18, CD19, CD20, CD21, CD22, CD23, CD24, TACI, CD25, CD27, CD28, CD30, CD30L, CD31, CD32, CD32b, CD34, CD33, CD38, CD39, CD40, CD40-L, CD41b, CD42a, CD42b, CD43, CD44, CD48 , CD47, CD45RA, CD45RO, CD48, CD52, CD55, CD56, CD58, CD61, CD66b, CD70, CD72, CD79, CD68, CD84, CD86, CD93, CD94, CD95, CRACC, BLAME, BCMA, CD103 , CD107, CD112, CD120a, CD120b, CD123, CD125, CD134, CD135, CD140a, CD141, CD154, CD155, CD160, CD163, CD172a, XCR1, CD203c, CD204, CD206, CD207CD226、CD244、CD267、CD268、CD269、CD355、CD358、NKG2A、NKG2B、NKG2 C、NKG2D、NKG2E、NKG2F、NKG2H、KIR2DL1、KIR2DL2、KIR2DL3、KIR2DL5A、 KIR2DL5B、KIR3DL1、KIR3DL2、KIR3DL3、KIR3DL4、KIR2DS1、KIR2DS2、KI R2DS3、KIR2DS4、KIR2DS5、DAP12、KIR3DS、NKp44、NKp46、TCR、BCR、インテグリン、FcβεRI、MHC-I、MHC-II、IL-1R、IL-2Rα、IL-2Rβ、IL-2Rγ、IL-3Rα、CSF2RB、IL-4R、IL-5Rα、IL-6Rα、gp130、IL-7Rα、IL-9R、IL-12Rβ1、IL-12R β2, IL-13Rα1, IL-13Rα2, IL-15Rα, IL-21R, IL-23R, IL-27Rα, IL-31Rα, OSMR, CSF-1R, IL-15, IL-10Rα, IL-10Rβ, IL-20Rα, IL-20Rβ, IL-22 Rα1、IL-22Rα2、IL-22Rβ、IL-28RA、PD-1、PD-1H、BTLA、CTLA-4、PD-L1、P D-L2、2B4、B7-1、B7-2、B7-H1、B7-H4、B7-DC、DR3、LIGHT、LAIR、LTα1β2、 LTβR、TIM-1、TIM-3、TIM-4、TIGIT、LAG-3、ICOS、ICOS-L、SLAM、SLAMF2、 OX-40、OX-40L、GITR、GITRL、TL1A、HVEM、41-BB、41BB-L、TL-1A、TRAF1、 TRAF2、TRAF3、TRAF5、BAFF、BAFF-R、APRIL、TRAIL、RANK、AITR、TRAMP、C CR1、CCR2、CCR3、CCR4、CCR5、CCR6、CCR7、CCR8、CCR9、CCR10、CCR11、CXC R1、CXCR2、CXCR3、CXCR4、CXCR5、CXCR6、CXCR7、CLECL9a、DC-SIGN、IGSF 4A、SIGLEC、EGFR、PDGFR、VEGFR、FAP、α-SMA、ビメンチン、ラミニン、FAS、FAS-L、F c, ICAM-1, ICAM-2, ICAM-3, ICAM-4, ICAM-5, PECAM-1, MICA, MICB, UL16, ULBP1, ULBP2, ILBP3, ULBP4, ULBP5, ULBP6, MULT1, RAE1 α, β, γ, δ, and ε, A1R, A 2A R.A. 2B In various embodiments, the integrin is selected from the group consisting of αR, αR, A3R, H60a, H60b, and H60c. In various embodiments, the integrin is selected from the group consisting of α1, α2, αIIb, α3, α4, α5, α6, α7, α8, α9, α10, α11, αD, αE, αL, αM, αV, αX, β1, β2, β3, β4, β5, β6, β7, β8, or a combination thereof. In various embodiments, the TCR is selected from the group consisting of α, β, γ, δ, ε, and ζ TCRs. Several methods for assaying cell surface protein expression have been described in the literature, including flow cytometry and mass cytometry (CyTOF). The presence or abundance of one or more of these cell surface proteins indicates that the patient will be responsive to treatment with the methods disclosed herein.

[0235] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the level of a tumor-promoting, anti-inflammatory, or immunosuppressive protein in one or more tumor samples from the subject. In various embodiments, the tumor-promoting, anti-inflammatory, or immunosuppressive protein is selected from the group consisting of CD39, CD79, MMP1, MMP2, MMP3, MMP7, MMP8, MMP9, MMP10, MMP11, MMP12, MMP13, MMP14, MMP15, MMP16, MMP17, MMP18, MMP19, MMP20, MMP21, MMP23A, MMP23B, MMP24, MMP25, MMP26, MMP27, MMP28, MMP29, MMP30, MMP31, MMP32, MMP33, MMP34, MMP35, MMP36, MMP37, MMP38, MMP39, MMP40, MMP41, MMP42, MMP43, MMP44, MMP45, MMP46, MMP47, MMP48, MMP49, MMP50, MMP51, MMP52, MMP53, MMP54, MMP55, MMP56, MMP57, MMP58, MMP59, MMP60, MMP61, MMP62, MMP63, MMP64, MMP65, MMP66, MMP67, MMP68, MMP69, MMP70, MMP71, MMP72, MMP73, MMP74, MMP75, MMP76, MMP77, MMP78, MMP79, MMP70, MMP71, MMP72, MMP73, MMP7 27, MMP28, CXCL12, GM-CSF, G-CSF, TGF-β1, TGF-β2, TGF-β3, arginase, asparaginase, kynureninase, indoleamine 2,3 dioxygenase (IDO1 and IDO2), tryptophan 2,3 dioxygenase (TDO), myeloperoxidase (MPO), neutrophil elastase (NE), and IL4I1. In various embodiments, the level of a tumor-promoting, anti-inflammatory, or immunosuppressive protein in one or more tumor samples from the subject is reduced by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tumor samples taken before treatment. In various embodiments, the level of a tumor-promoting, anti-inflammatory, or immunosuppressive protein in one or more tumor samples from the subject is reduced by 2 to 100-fold compared to one or more tumor samples taken from the subject prior to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100-fold (including all values ​​and ranges therebetween) compared to one or more samples taken prior to treatment).

[0236] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" increases the levels of proteins associated with tumor growth inhibition, anti-tumor activity, or pro-inflammatory activity. In various embodiments, the protein associated with tumor growth inhibition, anti-tumor activity, or pro-inflammatory activity is CD44, CD56, CD103c, CD69, KG2A, NKG2B, NKG2C, NKG2D, NKG2E, NKG2F, NKG2H, ICOS, ICOS-L, SLAM, SLAMF2, OX-40, OX-40L, GITR, GITRL, TL-1A, HVEM, 41-BB, 41BB-L, TRAF1, TRAF2, TRAF3, TRAF5, BAFF, BAFF-R, APRIL, TRAIL, RANK, AITR, TRAMP, IL-1α, IL-1β, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-19, IL-20, IL-21, IL-19, IL-19, IL-18, IL-19, IL-22, IL-19, IL-19, IL-19, IL-23, IL-19 ... L-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-20, IL-21, IL-22, IL-23, IL-24, IL-25, IL-26, IL-27, IL-28, IL-29, IL-30 , IL-31, IL-32, IL-33, IL-35, IL-36, CXCL2 (MCP-1), CXCL3 (MIP-1α), CXCL4 (MIP-1β), CXCL5 (RANTES), IFN-α, IFN-β, IFN-γ, granzyme B, perforin, and TNF-α. In various embodiments, the level of a protein associated with tumor growth inhibition, anti-tumor activity, or pro-inflammatory activity is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tumor samples taken prior to treatment.In various embodiments, the level of a protein associated with tumor growth inhibition, anti-tumor activity, or pro-inflammatory activity is increased 2-100 fold compared to one or more tumor samples taken from the subject prior to treatment (e.g., decreased by about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween) compared to one or more samples taken prior to treatment). Several methods for assaying proteins from tumor samples have been described in the literature, including Western blot and ELISA.

[0237] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the level of neutrophil extracellular traps (NETs) in one or more tumor samples taken from the subject. In various embodiments, the level of NETs in one or more tumor samples is reduced by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tumor samples taken prior to treatment. In various embodiments, the level of NETs in one or more tumor samples is reduced by 2-100 fold compared to one or more tumor samples taken from the subject prior to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween) compared to one or more samples taken prior to treatment). Several methods for assaying NETs from tumor samples have been described in the literature, including Western blot, ELISA, and flow cytometry.

[0238] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" reduces the expression of tumor-promoting, tumor-permissive, or immunosuppressive genes in one or more tumor samples from the subject. In one or more embodiments, expression of tumor-promoting, tumor-permissive, or immunosuppressive genes is reduced by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tumor samples taken before treatment. In one or more embodiments, expression of the tumor-promoting, tumor-permissive, or immunosuppressive gene is reduced by 2 to 100 fold compared to one or more tumor samples taken from the subject prior to treatment (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween) compared to one or more samples taken prior to treatment).

[0239] In various embodiments, administering a RAS inhibitor or combination of compounds described herein to a subject having one or more tumors characterized as immunorefractory, immunologically protected, or immunologically "cold" increases the expression of tumor-inhibitory, anti-tumor, or pro-inflammatory genes in one or more samples taken from the subject. In one or more embodiments, expression of the tumor-inhibiting, anti-tumor, or pro-inflammatory gene is increased by 5-100% (e.g., about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, about 90%, about 95%, or about 100% (including all values ​​and ranges therebetween)), 10-95%, 15-90%, 20-85%, 25-75%, 30-70%, 35-65%, 40-60%, 45-55%, 50%, or 100% compared to one or more tumor samples taken before treatment. In various embodiments, expression of the tumor-inhibiting, anti-tumor, or pro-inflammatory gene is increased by 2-100 fold (e.g., about 2, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 fold (including all values ​​and ranges therebetween)) compared to one or more tumor samples taken from the subject prior to treatment. In various embodiments, gene expression analysis is performed by PCR, RT-PCR, qRT-PCR, next-generation sequencing (NGS), RNA-seq, ATAC-seq, exome sequencing, Southern blot, microarray analysis, or single-cell sequencing.

[0240] In various embodiments, treating a subject with lung cancer with a RAS inhibitor or a combination of compounds described herein converts a cold tumor into a hot tumor. Such a conversion can be detected using methods described herein and known in the art. If a subject is diagnosed with a tumor that has converted from a cold tumor to a hot tumor, treatment may be continued by administering a RAS inhibitor or a combination of compounds described herein, which are useful for treating hot tumors, or tumors that are rich in immune cells or immunogenic. In other embodiments, when the tumor converts from a cold tumor to a hot tumor, the patient is stopped from treatment with a RAS inhibitor or a combination of compounds described herein, and the patient is started on a cancer treatment that is useful for treating hot tumors, or tumors that are rich in immune cells or immunogenic. Such cancer therapeutic agents include chemotherapeutic agents, cytokines, anti-angiogenic agents, enzymes, immune checkpoint modulators and monoclonal antibodies, hormonal therapies, including one or more cell-based therapies, such as adoptive cell transfer, tumor-infiltrating leukocyte therapy, chimeric antigen receptor T-cell therapy (CAR-T), NK cell therapy and stem cell therapy, or oncolytic viruses or oncolytic bacteria.

[0241] In various embodiments, the immune checkpoint modulator targets programmed cell death protein 1 (PD-1), programmed cell death protein ligand-1 (PD-L1), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), T-cell immunoglobulin and mucin domain-containing 3 (TIM-3), lymphocyte-activation gene 3 (LAG-3), or TIGIT (T-cell immunoreceptor with Ig and ITIM domains). In various embodiments, the immune checkpoint modulator is an antibody selected from the group consisting of ipilimumab, tremelimumab, pembrolizumab, nivolumab, atezolizumab, avelumab, cemiplimab, and durvalumab.

[0242] In various embodiments, subjects diagnosed with a cold tumor and receiving treatment with a RAS inhibitor or a combination of compounds described herein are monitored periodically to determine whether the tumor has transformed into a hot tumor. Monitoring may be performed, for example, monthly, every two months, every three months, every six months, or annually, as deemed necessary by a physician.

[0243] In various embodiments, the subject has previously been treated with immunotherapy but has developed resistance to the immunotherapy or has converted from a hot tumor to a cold tumor. Also provided are methods of treating a subject with cancer that has developed resistance to immunotherapy or has developed a cold tumor, comprising administering to the subject a RAS inhibitor or a combination of compounds described herein.

[0244] Cancers with oncogenic RAS mutations In some embodiments, the present invention discloses a method for treating lung cancer in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound of the present invention, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition comprising such a compound or salt, wherein the cancer is immunorefractory lung cancer.

[0245] The method for detecting Ras mutation is known in the art.Such means include but are not limited to direct sequencing and use highly sensitive diagnostic assays (CE-IVD marked), such as TheraScreen PCR; AmoyDx; PNAClamp; RealQuality; EntroGen; LightMix; StripAssay; Hybcell plexA; Devyser; Surveyor; Cobas; and TheraScreen Pyro, as described in, for example, Domagala, et al., Pol J Pathol 3:145-164(2012) (its entirety is incorporated herein by reference).See also, for example, WO2020 / 106640.

[0246] In some embodiments, the cancer is non-small cell lung cancer or any of the lung cancers described herein, and the Ras mutation comprises a K-Ras G12C mutation, an H-Ras G12C mutation, or an N-Ras G12C mutation. In some embodiments, the cancer is non-small cell lung cancer or any of the lung cancers described herein, and the Ras mutation comprises a K-Ras G12C mutation.

[0247] Also provided is a method for inhibiting Ras protein in cells, comprising contacting the cells with an effective amount of a compound of the present invention or a pharmaceutically acceptable salt thereof. Also provided is a method for inhibiting RAF-Ras binding, comprising contacting the cells with an effective amount of a compound of the present invention or a pharmaceutically acceptable salt thereof. The cells can be cancer cells. The cancer cells can be any type of cancer described herein. The cells can be in vivo or in vitro.

[0248] lung cancer In some embodiments, the present invention discloses a method of treating lung cancer, hi some embodiments, the lung cancer is immunorefractory lung cancer.

[0249] Lung cancer can be classified using different systems. In some systems, lung cancer includes adenocarcinoma (mixed, acinar, papillary, solid, micropapillary, lepidic non-mucinous, and lepidic mucinous), squamous cell carcinoma, large cell carcinoma (e.g., non-small cell lung cancer (NSCLC) (e.g., advanced or non-advanced, large cell carcinoma with neuroendocrine morphology (LCNEM), NSCLC not otherwise specified (NOS) / adenosquamous carcinoma, sarcomatoid carcinoma, adenosquamous carcinoma, and large cell neuroendocrine carcinoma (LCNEC)); and small cell lung cancer (SCLC)).

[0250] Alternatively, in different systems, lung cancer can be classified into preinvasive lesions, microinvasive adenocarcinoma, and invasive adenocarcinoma (invasive mucinous adenocarcinoma, mucinous bronchioloalveolar carcinoma (BAC), colloid, embryonal type (low-grade and high-grade), and intestinal type). Non-small cell lung cancer includes adenocarcinoma, squamous cell carcinoma, large cell carcinoma, or large cell neuroendocrine tumor.

[0251] More frequently, lung cancer can be classified as either small cell lung cancer ("SCLC") or non-small cell lung cancer ("NSCLC"). NSCLC can be further classified as squamous or non-squamous. An example of non-squamous NSCLC is adenocarcinoma.

[0252] In some embodiments, the lung cancer is bronchogenic carcinoma (squamous cell, small undifferentiated cell, large undifferentiated cell, adenocarcinoma), alveolar (bronchiolar) carcinoma, bronchial adenoma, sarcoma, lymphoma, chondroitin hamartoma, or mesothelioma.

[0253] Lung cancer may be newly diagnosed and previously untreated, or may be recurrent, refractory, recurrent and refractory, locally advanced, or metastatic. In some cases, lung cancer includes recurrent or refractory lung cancer. In some cases, lung cancer includes metastatic lung cancer. In some cases, the subject has been diagnosed with recurrent or refractory lung cancer. In further cases, the subject has been diagnosed with metastatic lung cancer.

[0254] Combination therapy The methods of the present invention may include the use of a compound of the present invention alone or in combination with one or more additional therapies (e.g., non-drug treatments or therapeutic agents). The dosage of one or more additional therapies (e.g., non-drug treatments or therapeutic agents) may be reduced from the standard dosage when administered alone. For example, dosage may be empirically determined from the combination and permutation of drugs, or estimated by isobolographic analysis (e.g., Black et al., Neurology 65:S3-S6 (2005)).

[0255] The compound of the present invention can be administered before, after, or simultaneously with such one or more additional therapies. When combined, the dosage of the compound of the present invention and the dosage of one or more additional therapies (e.g., non-drug treatments or therapeutic agents) provide a therapeutic effect (e.g., a synergistic or additive therapeutic effect). The compound of the present invention and the additional therapies, such as anti-cancer agents, can be administered together, for example, in a single pharmaceutical composition, or separately, and when administered separately, can be administered simultaneously or sequentially. Such sequential administration can be closely spaced or spaced apart.

[0256] In some embodiments, the additional therapy is the administration of a side effect limiting agent (e.g., an agent intended to reduce the occurrence or severity of side effects of treatment). For example, in some embodiments, the compounds of the present invention can also be used in combination with a therapeutic agent to treat nausea. Examples of agents that can be used to treat nausea include dronabinol, granisetron, metoclopramide, ondansetron, and prochlorperazine, or pharmaceutically acceptable salts thereof.

[0257] In some embodiments, the one or more additional therapies include a non-drug treatment (e.g., surgery or radiation therapy). In some embodiments, the one or more additional therapies include a therapeutic agent (e.g., a compound or biologic that is an anti-angiogenic agent, a signal transduction inhibitor, an anti-proliferative agent, a glycolysis inhibitor, or an autophagy inhibitor). In some embodiments, the one or more additional therapies include a non-drug treatment (e.g., surgery or radiation therapy) and a therapeutic agent (e.g., a compound or biologic that is an anti-angiogenic agent, a signal transduction inhibitor, an anti-proliferative agent, a glycolysis inhibitor, or an autophagy inhibitor). In other embodiments, the one or more additional therapies include two therapeutic agents. In yet other embodiments, the one or more additional therapies include three therapeutic agents. In some embodiments, the one or more additional therapies include four or more therapeutic agents.

[0258] In this Combination Therapy section, all references to the agents mentioned, whether explicitly stated or not, are incorporated by reference.

[0259] Non-drug therapy Examples of non-drug treatments include, but are not limited to, radiation therapy, cryotherapy, hyperthermia, surgery (eg, surgical removal of tumor tissue), and T cell adoptive transfer (ACT) therapy.

[0260] In some embodiments, the compounds of the present invention may be used as an adjuvant therapy after surgery. In some embodiments, the compounds of the present invention may be used as a neoadjuvant therapy before surgery.

[0261] Radiation therapy can be used to inhibit abnormal cell growth or treat hyperproliferative disorders, such as cancer, in a subject (e.g., a mammal (e.g., a human)). Techniques for administering radiation therapy are known in the art. Radiation therapy can be administered via one of several methods, or a combination of methods, including, but not limited to, external beam therapy, internal radiation therapy, implanted radiation, stereotactic radiotherapy, systemic radiation therapy, radiotherapy, and permanent or temporary interstitial brachytherapy. As used herein, the term "brachytherapy" refers to radiation therapy delivered by spatially restricted radioactive materials inserted within the body at or near the site of a tumor or other proliferative tissue disease. The term is intended to include, but is not limited to, exposure to radioactive isotopes (e.g., At-211, I-131, I-125, Y-90, Re-186, Re-188, Sm-153, Bi-212, P-32, and radioactive isotopes of Lu). Suitable radioactive sources for use as cell conditioning agents of the present invention include both solid and liquid sources. By way of non-limiting example, the radioactive source may be a radionuclide such as I-125, I-131, Yb-169, Ir-192, I-125 as a solid source, or other radionuclide that emits photons, beta particles, gamma rays, or other therapeutic rays. The radioactive material may also be a fluid made from any solution of radionuclide(s), such as a solution of I-125 or I-131, or a radioactive fluid may be produced using a slurry of a suitable fluid containing small particles of a solid radionuclide, such as Au-198 or Y-90. Furthermore, the radionuclide(s) may be embodied in a gel or radioactive microspheres.

[0262] In some embodiments, the compounds of the present invention can make abnormal cells more sensitive to radiation therapy, which is intended to kill or inhibit the growth of such cells.Therefore, the present invention further relates to a method for sensitizing abnormal cells in a mammal to radiation therapy, which method comprises administering to the mammal an amount of a compound of the present invention effective to sensitize the abnormal cells to radiation therapy.The amount of the compound in this method can be determined according to the means for determining the effective amount of such compounds described herein.In some embodiments, the compounds of the present invention can be used as adjuvant therapy after radiation therapy or as neoadjuvant therapy before radiation therapy.

[0263] In some embodiments, the non-drug treatment is T cell adoptive transfer (ACT) therapy. In some embodiments, the T cells are activated T cells. The T cells can be modified to express a chimeric antigen receptor (CAR). CAR-modified T (CAR-T) cells can be generated by any method known in the art. For example, CAR-T cells can be generated by introducing a suitable expression vector encoding a CAR into T cells. Prior to T cell expansion and genetic modification, a T cell source is obtained from a subject. T cells can be obtained from several sources, including peripheral blood mononuclear cells, bone marrow, lymph node tissue, umbilical cord blood, thymus tissue, tissue from an infection site, ascites, pleural effusion, spleen tissue, and tumors. In certain embodiments of the present invention, any number of T cell lines available in the art can be used. In some embodiments, the T cells are autologous T cells. Whether before or after genetic modification of the T cells to express a desired protein (e.g., a CAR), the T cells can be engineered to express any of the T cells described in, e.g., U.S. Patent Nos. 6,352,694, 6,534,055, 6,905,680, 6,692,964, 5,858,358, 6,887,466, 6,905,681, 7, Activation and proliferation can be achieved using methods described in US Pat. Nos. 144,575, 7,067,318, 7,172,869, 7,232,566, 7,175,843, 7,572,631, 5,883,223, 6,905,874, 6,797,514, and 6,867,041.

[0264] therapeutic agent The therapeutic agent can be a compound used in the treatment of cancer or a condition related thereto.

[0265] For example, the therapeutic agent can be a steroid. Thus, in some embodiments, the one or more additional therapies include a steroid. Suitable steroids include 21-acetoxypregnenolone, alclometasone, algestone, amcinonide, beclomethasone, betamethasone, budesonide, chloroprednisone, clobetasol, clocortolone, cloprednol, corticosterone, cortisone, cortivazol, deflazacort, desonide, desoximetasone, dexamethasone, diflorasone, diflucortolone, difuprednate, enoxolone, fluazacort, flucloronide, flumethasone, flunisolide, fluocinolone acetonide, fluocinonide, flucortine butyl, flucortolone, fluorometholone, fluperolone acetate, fluprednidene acetate, fluprednisolone, and flurandrenolide. , fluticasone propionate, formocortal, halcinonide, halobetasol propionate, halometasone, hydrocortisone, loteprednol etabonate, mazipredone, medrysone, meprednisone, methylprednisolone, mometasone furoate, paramethasone, prednicarbate, prednisolone, prednisolone 25-diethylaminoacetate, prednisolone sodium phosphate, prednisone, prednival, prednylidene, rimexolone, tixocortol, triamcinolone, triamcinolone acetonide, triamcinolone benetonide, triamcinolone hexacetonide, and salts or derivatives thereof.

[0266] Further examples of therapeutic agents that can be used in combination therapy with the compounds of the invention include compounds described in the following patents: U.S. Patent Nos. 6,258,812, 6,630,500, 6,515,004, 6,713,485, 5,521,184, 5,770,599, 5,747,498, 5,990,141, 6,235,764, and 8,623,885, and International Patent Application No. WO00 / 064994. Patent Applications WO01 / 37820, WO01 / 32651, WO02 / 68406, WO02 / 66470, WO02 / 55501, WO04 / 05279, WO04 / 07481, WO04 / 07458, WO04 / 09784, WO02 / 59110, WO99 / 45009, WO00 / 59509, WO99 / 61422, WO00 / 12089, and WO00 / 02871.

[0267] The therapeutic agent may be a biologic (e.g., a cytokine (e.g., an interferon or an interleukin such as IL-2)) used to treat cancer or a related condition. In some embodiments, the biologic is an immunoglobulin-based biologic, such as a monoclonal antibody (e.g., a humanized antibody, a fully human antibody, an Fc fusion protein, or a functional fragment thereof), which agonizes a target to stimulate an anti-cancer response or antagonizes an antigen important to cancer. Antibody-drug conjugates are also included.

[0268] The therapeutic agent can be a T cell checkpoint inhibitor. In one embodiment, the checkpoint inhibitor is an inhibitory antibody (e.g., a monospecific antibody, e.g., a monoclonal antibody). The antibody can be, for example, humanized or fully human. In some embodiments, the checkpoint inhibitor is a fusion protein, e.g., an Fc receptor fusion protein. In some embodiments, the checkpoint inhibitor is an agent, such as an antibody, that interacts with a checkpoint protein. In some embodiments, the checkpoint inhibitor is an agent, such as an antibody, that interacts with a ligand of a checkpoint protein. In some embodiments, the checkpoint inhibitor is an inhibitor (e.g., an inhibitory antibody or small molecule inhibitor) of CTLA-4 (e.g., an anti-CTLA-4 antibody or fusion protein). In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist of PD-1 (e.g., an inhibitory antibody or small molecule inhibitor). In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist of PD-L1 (e.g., an inhibitory antibody or small molecule inhibitor). In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist (e.g., an inhibitory antibody or Fc fusion or small molecule inhibitor) of PD-L2 (e.g., a PD-L2 / Ig fusion protein). In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist (e.g., an inhibitory antibody or small molecule inhibitor) of B7-H3, B7-H4, BTLA, HVEM, TIM3, GAL9, LAG3, VISTA, KIR, 2B4, CD160, CGEN-15049, CHK1, CHK2, A2aR, a B-7 family ligand, or a combination thereof.In some embodiments, the checkpoint inhibitor is pembrolizumab, nivolumab, PDR001 (NVS), REGN2810 (Sanofi / Regeneron), a PD-L1 antibody such as avelumab, durvalumab, atezolizumab, pidilizumab, JNJ-63723283 (JNJ), BGB-A317 (BeiGene & Celgene), or Preusser, M. et al. Checkpoint inhibitors disclosed in

[0016] al. (2015) Nat. Rev. Neurol. include, but are not limited to, ipilimumab, tremelimumab, nivolumab, pembrolizumab, AMP224, AMP514 / MEDI0680, BMS936559, MED14736, MPDL3280A, MSB0010718C, BMS986016, IMP321, lirilumab, IPH2101, 1-7F9, and KW-6002. Other checkpoint inhibitors are described herein.

[0269] The therapeutic agent can be an anti-TIGIT antibody, such as MBSA43, BMS-986207, MK-7684, COM902, AB154, MTIG7192A, or OMP-313M32 (etigilimab).

[0270] The therapeutic agent can be an agent that treats cancer or a related condition (e.g., a cytotoxic agent, a non-peptide small molecule, or other compound useful in treating cancer or a related condition, collectively an "anti-cancer agent"). The anti-cancer agent can be, for example, a chemotherapeutic agent or a targeted therapy agent.

[0271] Anticancer drugs include mitotic inhibitors, intercalating antibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes, topoisomerase inhibitors, biological response modifiers, alkylating agents, antimetabolites, folic acid analogs, pyrimidine analogs, purine analogs and related inhibitors, vinca alkaloids, epipodopirotoxins, antibiotics, L-asparaginase, topoisomerase inhibitors, interferons, platinum coordination complexes, anthracenedione-substituted ureas, methylhydrazine derivatives, adrenocortical suppressants, corticosteroids, progestins, estrogens, antiestrogens, androgens, antiandrogens, and gonadotropin-releasing hormone analogs. Additional anticancer drugs include leucovorin (LV), irenotecan, oxaliplatin, capecitabine, paclitaxel, and doxetaxel. In some embodiments, the one or more additional therapies comprise two or more anticancer drugs. Two or more anti-cancer drugs can be used in a cocktail that is administered in combination or separately. Suitable dosing regimens for combined anti-cancer drugs are known in the art and are described, for example, in Saltz et al., Proc. Am. Soc. Clin. Oncol. 18:233a (1999) and Douillard et al., Lancet 355(9209):1041-1047 (2000).

[0272] Other non-limiting examples of anti-cancer drugs include Gleevec® (imatinib mesylate); Kyprolis® (carfilzomib); Velcade® (bortezomib); Casodex (bicalutamide); Iressa® (gefitinib); alkylating agents, such as thiotepa and cyclophosphamide; alkyl sulfonates, such as busulfan, improsulfan, and piposulfan; aziridines, such as , benzodopa, carboquone, meturedopa, and uredopa; ethyleneimines and methylameramines, such as altretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, and trimethylolomelamine; acetogenins (especially bullatacin and bullatacinone); camptothecins (e.g., the synthetic analog topotecan); bryostatin; kallistatin; CC-1065 (e.g., its adzelesin, carzelesin, and synthetic analogs of beizeresin); cryptophycins (specifically, cryptophycin 1 and cryptophycin 8); dolastatins; duocarmycins (e.g., synthetic analogs KW-2189 and CB1-TM1); eleutherobin; pancratistatin; sarcodictyin A; spongistatins; nitrogen mustards, e.g., chlorambucil, chlornaphazine, colofosfamide, estramustine, ifosfamide, mechlorethamine, meclometamine, thiamin oxide hydrochloride, melphalan, nobembrine, phenesterine, prednimustine, trophosfamide, uracil mustard; nitrosoureas, e.g., camrustine, chlorozotocin, fotemustine, lomustine, nimustine, and ranimustine; antibiotics, e.g., enediyne antibiotics (e.g., calicheamicins, e.g., calicheamicin gamma 11 and calicheamicin omega 11 (see, e.g., Agnew, Chem. Intl. Ed Engl. 33:183-186 (1994)); dynemicins, such as dynemicin A; bisphosphonates, such as clodronate; esperamicin;Neocarzinostatin chromophore and related chromoprotein enediyne antibiotic chromophores, aclacinomycin, actinomycin, autramycin, azaserine, bleomycin, cactinomycin, calicheamicin, carabicin, caminomycin, carminomycin, carzinophilin, chromomycin, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, adriamycin (doxorubicin), morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino - doxorubicin, deoxydoxorubicin, epirubicin, esorubicin, idarubicin, marcelomycin, mitomycins such as mitomycin C, mycophenolic acid, nogalamycin, olivomycin, peplomycin, potfilomycin, puromycin, chelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; antimetabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogues such as denopterin, pteropterin, trimetrexate; Purine analogues such as ludarabine, 6-mercaptopurine, thiamiprine, and thioguanine; pyrimidine analogues such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, and floxuridine; androgens such as calucelone, dromostanolone propionate, epithiostanol, mepitiostane, and testolactone; antiadrenal agents such as aminoglutethimide, mitotane, and trilostane; folic acid replacement fluids such as furoic acid; aceglatone; aldophosphamide glycosides; aminolevulinic acid Acid; Eniluracil; Amsacrine; Bestravcil; Bisantrene; Edatrexate; Defofamine; Demecolcine; Diaziquone; Elfomithine; Elliptinium acetate; Epothilones such as epothilone B; Etoglucide; Gallium nitrate; Hydroxyurea; Lentinan; Maytansinoids such as lonidamine, maytansine, and ansamitocin; Mitoguazone; Mitoxantrone; Mopidamol; Nitracrine; Pentostatin; Fenameth; Pirarubicin; Rosoxantrone; Podophyllic acid;2-Ethylhydrazide; procarbazine; PSK® polysaccharide complex (JHS Natural Products, Eugene, OR); razoxane; rhizoxin; schizophyllan; spirogermanium; tenuazonic acid; triazicon; 2,2',2''-trichlorotriethylamine; trichothecenes such as T-2 toxin, veraculin A, roridin A, and anguidine; urethane; vindesine; dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside ("Ara-C"); Cyclophosphamide; thiotepa; taxoids such as Taxol® (paclitaxel), Abraxane® (a chromophore-free, albumin-engineered nanoparticle formulation of paclitaxel), and Taxotere® (doxetaxel); chlorambucil; tamoxifen (Nolvadex™); raloxifene; aromatase-inhibited 4(5)-imidazoles; 4-hydroxytamoxifen; trihydroxyphene; ketoxifene; LY 117018; onapristone; toremifene (Fareston®); flutamide, nilutamide, bicalutamide, leuprolide, goserelin; chlorambucil; Gemzar® gemcitabine; 6-thioguanine; mercaptopurine; platinum coordination complexes such as cisplatin, oxaliplatin, and carboplatin; vinblastine; platinum; etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; Navelbine® (vinorelbine); novantrone; teniposide; edatrexate; daunomycin; aminopterin; ibandronate; irinotecan (e.g., CPT-11); topoisomerase inhibitors RFS 2000; difluoromethylornithine (DMFO); retinoids such as retinoic acid; esperamicin; capecitabine (e.g., Xeloda®); and pharmaceutically acceptable salts of any of the above.

[0273] Additional non-limiting examples of anti-cancer drugs include trastuzumab (Herceptin®), bevacizumab (Avastin®), cetuximab (Erbitux®), rituximab (Rituxan®), Taxol®, Arimidex®, ABVD, avicin, abagovomab, acridine carboxamide, adecatumumab, 17-N-allylamino-17-demethoxygeldanamycin, alfalan, gin, alvocidib, 3-aminopyridine-2-carboxaldehyde thiosemicarbazone, amonafide, anthracenedione, anti-CD22 immunotoxin, antineoplastic agents (e.g., non-cell cycle specific antineoplastic agents, and other antineoplastic agents described herein), antineoplastic herbs, apaziquone, atiprimod, azathioprine, belotecan, bendamustine, BIBW2992, biricodar, brostallicin, bryostatin, buthionine sulfoximine, CBV (chemotherapeutic ), calyculin, dichloroacetic acid, discodermolide, elsamitrucin, enocitabine, eribulin, exatecan, exisulind, ferruginol, forodesin, fosfestrol, ICE chemotherapy regimen, IT-101, imexon, imiquimod, indolocarbazole, irofulven, laniquidar, larotaxel, lenalidomide, lucanton, lurtotecan, mafosfamide, mitozolomide, nafoxidine, nedaplatin, olaparib, orta These include taxel, PAC-1, pawpaw, pixantrone, proteasome inhibitors, rebeccamycin, resiquimod, rubitecan, SN-38, salinosporamide A, sapacitabine, Stanford V, swainsonine, talaporfin, tariquidar, tegafur-uracil, temodar, tesetaxel, triplatin tetranitrate, tris(2-chloroethyl)amine, troxacitabine, uramustine, vadimezan, vinflunine, ZD6126, and zosuquidar.

[0274] Further non-limiting examples of anti-cancer agents include natural products, such as vinca alkaloids (e.g., vinblastine, vincristine, and vinorelbine), epidipodophyllotoxins (e.g., etoposide and teniposide), antibiotics (e.g., dactinomycin (actinomycin D), daunorubicin, and idarubicin), anthracyclines, mitoxantrone, bleomycin, plicamycin (mithramycin), mitomycin, enzymes (e.g., L-asparagine systemically), and the like. L-asparaginase, which metabolizes and depletes cells that do not have the ability to synthesize their own asparagine), antiplatelet agents, antiproliferative / antimitotic alkylating agents, such as nitrogen mustards (e.g., mechlorethamine, cyclophosphamide and analogs, melphalan, and chlorambucil), ethylenimines and methylmelamines (e.g., hexamethylmelamine and thiotepa), CDK inhibitors (e.g., CDK4 / 6 inhibitors, e.g., abemaciclib, ribociclib, palbociclib;seliciclib, UCN-01, P1446A-05, PD-0332991, dinaciclib, P27-00, AT-7519, RGB286638, and SCH727965), alkylsulfonates (e.g., busulfan), nitrosoureas (e.g., carmustine (BCNU) and analogs, and streptozocin), trazenes-dacarbazinine (DTIC), antiproliferative / antimitotic antimetabolites, e.g., folic acid analogs, pyrimidine analogs (e.g., fluorouracil, floxuridine, and cytarabine), purine analogs and related inhibitors (e.g., mercaptopurine, thioguanine, pentostatin, and 2-chlorodeoxyadenosine), aromatase inhibitors (e.g., anastrozole, exemestane, and letrozole), and platinum coordination complexes (e.g., cisplatin and carboplatin), procarbazine, hydroxyurea, mitotane, aminoglutethimide, histone deacetylase (HDAC) inhibitors (e.g., trichostatin, sodium butyrate, apicidan, suberoylanilide hydrochloride, anilide hydroamic acid), vorinostat, LBH589, romidepsin, ACY-1215, and panobinostat), mTOR inhibitors (e.g., bistusertib, temsirolimus, everolimus, ridaforolimus, and sirolimus), KSP (Eg5) inhibitors (e.g., Array520), DNA binders (e.g., Zalypsis®), PI3K inhibitors such as PI3K delta inhibitors (e.g., GS-1101 and TGR-1202), PI3K delta and gamma inhibitors (e.g., CAL-130), copanlisib, alpelisib, and idelalisib;Multikinase inhibitors (e.g., TG02 and sorafenib), hormones (e.g., estrogen), and hormone agonists, such as luteinizing hormone-releasing hormone (LHRH) agonists (e.g., goserelin, leuprolide, and triptorelin), BAFF neutralizing antibodies (e.g., LY2127399), IKK inhibitors, p38MAPK inhibitors, anti-IL-6 (e.g., CNT0328), telomerase inhibitors (e.g., GRN163L), Aurora kinase inhibitors (e.g., MLN8237), cell surface monoclonal antibodies (e.g., anti-CD38 (HUMAX-CD38)), anti-CS1 (e.g., elotuzumab), HSP90 inhibitors (e.g., 17AAG and KOS953), P13K / Akt inhibitors (e.g., perifosine), Akt inhibitors (e.g., GSK-2141795), PKC inhibitors (e.g., enzastaurin), FTIs (e.g., Zarnestra™), anti-CD138 (e.g., BT062), Torcl / 2-specific kinase inhibitors (e.g., INK128), ER / UPR targeting agents (e.g., MKC-3946), cFMS inhibitors (e.g., ARRY-382), JAK1 / 2 inhibitors (e.g., CYT387), PARP inhibitors (e.g., olaparib and veliparib (ABT-888)), and BCL-2 antagonists;

[0275] In some embodiments, the anti-cancer agent is selected from mechlorethamine, camptothecin, ifosfamide, tamoxifen, raloxifene, gemcitabine, Navelbine®, sorafenib, or any analog or derivative variant of the foregoing.

[0276] In some embodiments, the anti-cancer agent is a HER2 inhibitor. Non-limiting examples of HER2 inhibitors include monoclonal antibodies such as trastuzumab (Herceptin®) and pertuzumab (Perjeta®); small molecule tyrosine kinase inhibitors such as gefitinib (Iressa®), erlotinib (Tarceva®), piritinib, CP-654577, CP-724714, canertinib (CI 1033), HKI-272, lapatinib (GW-572016; Tykerb®), PKI-166, AEE788, BMS-599626, HKI-357, BIBW 2992, ARRY-334543, and JNJ-26483327.

[0277] In some embodiments, the anticancer agent is an ALK inhibitor. Non-limiting examples of ALK inhibitors include ceritinib, TAE-684 (NVP-TAE694), PF02341066 (crizotinib or 1066), alectinib, brigatinib, entrectinib, ensartinib (X-396), lorlatinib, ASP3026, CEP-37440, 4SC-203, TL-398, PLB1003, TSR-011, CT-707, TPX-0005, and AP26113. Additional examples of ALK kinase inhibitors are described in Examples 3-39 of WO05016894.

[0278] In some embodiments, the anti-cancer agent is an inhibitor of a downstream member of a receptor tyrosine kinase (RTK) / growth factor receptor (e.g., an SHP2 inhibitor (e.g., SHP099, TNO155, RMC-4550, RMC-4630, JAB-3068, JAB-3312, RLY-1971, ERAS-601, SH3809, PF-07284892, or BBP-398, or other SHP2 inhibitors described herein), an SOS1 inhibitor (e.g., BI-1701963, BI-3406, SDR5, RMC-5845, MRTX-0902, or BAY-293), a Raf inhibitor, a MEK inhibitor, an ERK inhibitor, a PI3K inhibitor, a PTEN inhibitor, an AKT inhibitor, or an mTOR inhibitor (e.g., an mTORC1 inhibitor or an mTORC2 inhibitor). In some embodiments, the anti-cancer agent is JAB-3312.

[0279] In some embodiments, the anti-cancer agent is an additional Ras inhibitor or a Ras vaccine, or another therapy designed to directly or indirectly reduce the oncogenic activity of Ras. In some embodiments, the anti-cancer agent is an additional Ras inhibitor. In some embodiments, the Ras inhibitor targets Ras in its active or GTP-bound state. In some embodiments, the Ras inhibitor targets Ras in its inactive or GDP-bound state. In some embodiments, the Ras inhibitor is an inhibitor of K-Ras G12C, such as AMG 510, MRTX1257, MRTX849, JNJ-74699157, LY3499446, ARS-1620, ARS-853, BPI-421286, LY3537982, JDQ443, JAB-21822, JAB-21000, IBI351, ERAS-3490, or GDC-6036. In some embodiments, the Ras inhibitor is a K-Ras G12D inhibitor, such as MRTX1133 or JAB-22000. In some embodiments, the Ras inhibitor is a K-Ras G12V inhibitor, such as JAB-23000. In some embodiments, the Ras inhibitor is RMC-6236. In some embodiments, the Ras inhibitor is selected from the Ras(ON) inhibitors (i.e., Ras in the GTP-bound state) disclosed in the following: WO2022235870, WO2022235864, WO2021091982, WO2021091967, WO2021091956, and WO2020132597 (each of which is incorporated by reference in its entirety), or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. Other examples of Ras inhibitors that can be combined with the Ras inhibitors of the present invention are set forth below, which are incorporated by reference in their entirety: WO2023287896, WO2023287730, WO2023284881, WO2023284730, WO2023284537, WO2023283933, WO2023283213, WO2023280960, WO2023280280, WO2023280136, WO2023280026, WO2023278600, WO2023274383,WO2023274324, WO2023020523, WO2023020521, WO2023020519, WO2023020518, WO2023018812, WO2023018810, WO2023018809, WO2023018699, WO202301 5559、WO2023014979、WO2023014006、WO2023010121、WO2023009716、WO202 3009572、WO2023004102、WO2023003417、WO2023001141、WO2023001123、WO2 022271923, WO2022271823, WO2022271810, WO2022271658, WO2022269508, WO2022266167, WO2022266069, WO2022266015, WO2022265974, WO202226115 4、WO2022261154、WO2022251576、WO2022251296、WO2022237815、WO2022232332、WO2022232331、WO2022232320、WO2022232318、WO2022223037、WO2022 221739、WO2022221528、WO2022221386、WO2022216762、WO2022192794、WO2 022192790、WO2022188729、WO2022187411、WO2022184178、WO2022173870、W O2022173678, WO2022135346, WO2022133731, WO2022133038, WO2022133345, WO2022132200, WO2022119748, WO2022109485, WO2022109487, WO2022066 805、WO2022002102、WO2022002018、WO2021259331、WO2021257828、WO2021 252339、WO2021248095、WO2021248090、WO2021248083、WO2021248082、WO20 21248079、WO2021248055、WO2021245051、WO2021244603、WO2021239058、WO 2021231526、WO2021228161、WO2021219090、WO2021219090、WO2021219072、WO2021218939、WO2021217019、WO2021216770、WO2021215545、WO2021215544、WO2021211864、WO2021190467、WO2021185233、WO2021180181、WO2021175199、2021173923、WO2021169990、WO2021169963、WO2021168193、WO2021158071、WO2021155716、WO2021152149、WO2021150613、WO2021147967、WO2021147965、WO2021143693、WO2021142252、WO2021141628、WO2021139748、WO2021139678、WO2021129824、WO2021129820、WO2021127404、WO2021126816、WO2021126799、WO2021124222、WO2021121371、WO2021121367、WO2021121330、WO2020050890、WO2020047192、WO2020035031、WO2020028706、WO2019241157、WO2019232419、WO2019217691、WO2019217307、WO2019215203、WO2019213526、WO2019213516、WO2019155399、WO2019150305、WO2019110751、WO2019099524、WO2019051291、WO2018218070、WO2018217651、WO2018218071、WO2018218069、WO2018206539、WO2018143315、WO2018140600、WO2018140599、WO2018140598、WO2018140514、WO2018140513、WO2018140512、WO2018119183、WO2018112420、WO2018068017、WO2018064510、WO2017201161、WO2017172979、WO2017100546、WO2017087528、WO2017058807、WO2017058805、WO2017058728、WO2017058902、WO2017058792、WO2017058768、WO2017058915、WO2017015562, WO2016168540, WO2016164675, WO2016049568, WO2016049524, WO2015054572, WO2014152588, WO2014143659 and WO2013155223.

[0280] In some embodiments, therapeutic agents that can be combined with compounds of the invention are inhibitors of the MAP kinase (MAPK) pathway (or "MAPK inhibitors"). MAPK inhibitors include, but are not limited to, one or more MAPK inhibitors described in Cancers (Basel) 2015 Sep;7(3):1758-1784. For example, MAPK inhibitors include trametinib, binimetinib, selumetinib, cobimetinib, LErafAON (NeoPharm), ISIS 5132; vemurafenib, pimasertib, TAK733, RO4987655 (CH4987655); CI-1040; PD-0325901; CH5126766; MAP855; AZD6244; refametinib (RDEA119 / BAY86-9766); GDC-0973 / XL581; AZD8330 (ARRY-424704 / ARRY-704); RO5126766 (Roche, described in PLoS One. 2014 Nov 25; 9(11)); and GSK1120212 (or JTP-74057, Clin Cancer Res. 2011 Mar 1;17(5):989-1000). The MAPK inhibitor can be PLX8394, LXH254, GDC-5573, or LY3009120.

[0281] In some embodiments, the anticancer drug is an inhibitor or disruptor of the RAS-RAF-ERK or PI3K-AKT-TOR or PI3K-AKT signaling pathway. The PI3K / AKT inhibitor may include, but is not limited to, one or more PI3K / AKT inhibitors described in Cancers (Basel) 2015 Sep;7(3):1758-1784. For example, the PI3K / AKT inhibitor may be selected from one or more of the following: NVP-BEZ235; BGT226; XL765 / SAR245409; SF1126; GDC-0980; PI-103; PF-04691502; PKI-587; GSK2126458.

[0282] In some embodiments, the anti-cancer agent is a PD-1 or PD-L1 antagonist.

[0283] In some embodiments, the additional therapeutic agent includes an ALK inhibitor, a HER2 inhibitor, an EGFR inhibitor, an IGF-1R inhibitor, a MEK inhibitor, a PI3K inhibitor, an AKT inhibitor, a TOR inhibitor, an MCL-1 inhibitor, a BCL-2 inhibitor, an SHP2 inhibitor, a proteasome inhibitor, and an immunotherapy, e.g., an immune checkpoint inhibitor. In some embodiments, the therapeutic agent can be a pan-RTK inhibitor, e.g., afatinib.

[0284] IGF-1R inhibitors include linsitinib, or a pharmaceutically acceptable salt thereof.

[0285] EGFR inhibitors include, but are not limited to, small molecule antagonists, antibody inhibitors, or specific antisense nucleotides or siRNA. Useful antibody inhibitors of EGFR include cetuximab (Erbitux®), panitumumab (Vectibix®), zalutumumab, nimotuzumab, and matuzumab. Additional antibody-based EGFR inhibitors include any anti-EGFR antibody or antibody fragment that can partially or completely block EGFR activation by its natural ligand. Non-limiting examples of antibody-based EGFR inhibitors include those described in Modjtahedi et al., Br. J. Cancer 1993, 67:247-253; Teramoto et al., Cancer 1996, 77:639-645; Goldstein et al., Clin. Cancer Res. 1995, 1:1311-1318; Huang et al., 1999, Cancer Res. 15:59(8):1935-40; and Yang et al., Cancer Res. 1999, 59:1236-1243. The EGFR inhibitor may be the monoclonal antibody Mab E7.6.3 (Yang, 1999 (supra)), or Mab C225 (ATCC Accession No. HB-8508), or an antibody or antibody fragment having the binding specificity thereof.

[0286] Small molecule antagonists of EGFR include gefitinib (Iressa®), erlotinib (Tarceva®), and lapatinib (TykerB®). See, e.g., Yan et al., Pharmacogenetics and Pharmacogenomics in Oncology Therapeutic Antibody Development, BioTechniques 2005, 39(4):565-8; and Paez et al., EGFR Mutations in Lung Cancer Correlation With Clinical Response To Gefitinib Therapy, Science 2004, 304(5676):1497-500. In some embodiments, the EGFR inhibitor is osimertinib (Tagrisso®). Further non-limiting examples of small molecule EGFR inhibitors include any of the EGFR inhibitors described in the following patent publications, and any pharmaceutically acceptable salts of such EGFR inhibitors: EP 0520722; EP 0566226; WO 96 / 33980; U.S. Patent No. 5,747,498; WO 96 / 30347; EP 0787772; WO 97 / 30034; WO 97 / 30044; WO 97 / 38994; WO 97 / 49688; EP 837063; WO 98 / 02434; WO 97 / 38983; WO 95 / 19774; WO 95 / 19970; WO 97 / 13771; WO 98 / 02437; WO 9 8 / 02438;WO97 / 32881;DE19629652;WO98 / 33798;WO97 / 32880;WO97 / 32880;EP68202 7;WO97 / 02266;WO97 / 27199;WO98 / 07726;WO97 / 34895;WO96 / 31510;WO98 / 14449;WO 98 / 14450; WO98 / 14451; WO95 / 09847; WO97 / 19065; WO98 / 17662; U.S. Patent Nos. 5,789,427; 5,650,415; 5,656,643; WO99 / 35146; WO99 / 35132; WO99 / 07701; and WO92 / 20642.Additional non-limiting examples of small molecule EGFR inhibitors include any of the EGFR inhibitors described in Traxler et al., Exp. Opin. Ther. Patents 1998, 8(12):1599-1625. In some embodiments, the EGFR inhibitor is an ERBB inhibitor. In humans, the ERBB family contains HER1 (EGFR, ERBB1), HER2 (NEU, ERBB2), HER3 (ERBB3), and HER (ERBB4).

[0287] MEK inhibitors include, but are not limited to, pimasertib, selumetinib, cobimetinib (Cotellic®), trametinib (Mekinist®), and binimetinib (Mektovi®). In some embodiments, the MEK inhibitor targets a MEK mutation that is a Class I MEK1 mutation selected from D67N; P124L; P124S; and L177V. In some embodiments, the MEK mutation is a Class II MEK1 mutation selected from ΔE51-Q58; ΔF53-Q58; E203K; L177M; C121S; F53L; K57E; Q56P; and K57N.

[0288] PI3K inhibitors include wortmannin; 17-hydroxywortmannin analogs described in WO 06 / 044453; 4-[2-(1H-indazol-4-yl)-6-[[4-(methylsulfonyl)piperazin-1-yl]methyl]thieno[3,2-d]pyrimidin-4-yl]morpholine (also known as pictilisib or GDC-0941, and described in WO 09 / 036082 and WO 09 / 055730); 2-methyl-2-[4-[3-methyl-2-oxo-8-(quinolin-3-yl)-2,3-dihydroimidazo[ 4,5-c]quinolin-1-yl]phenyl]propionitrile (also known as BEZ235 or NVP-BEZ235 and described in WO 06 / 122806); (S)-l-(4-((2-(2-aminopyrimidin-5-yl)-7-methyl-4-morpholinothieno[3,2-d]pyrimidin-6-yl)methyl)piperazin-1-yl)-2-hydroxypropan-1-one (described in WO 08 / 070740); LY294002 (2-(4-morpholinyl)-8-phenyl-4H-l-benzopyran-4-one (Axon Available from Medchem; PI103 hydrochloride (3-[4-(4-morpholinylpyrido-[3',2':4,5]furo[3,2-d]pyrimidin-2-yl]phenol hydrochloride (available from Axon Medchem); PIK75 (2-methyl-5-nitro-2-[(6-bromoimidazo[1,2-a]pyridin-3-yl)methylene]-1-methylhydrazide-benzenesulfonic acid, monohydrochloride) (available from Axon Medchem); PIK90 (N-(7,8-dimethoxy-2,3-dihydro-imidazo[l,2-c]quinazolin-5-yl)-nicotinamide (Axon available from Medchem); AS-252424 (5-[l-[5-(4-fluoro-2-hydroxy-phenyl)-furan-2-yl]-meth-(Z)-ylidene]-thiazolidine-2,4-dione (available from Axon Medchem); TGX-221 (7-methyl-2-(4-morpholinyl)-9-[1-(phenylamino)ethyl]-4H-pyrido-[1,2-a]pyrilnidin-4-one (available from Axon Medchem); XL-765;Other PI3K inhibitors include, but are not limited to, demethoxyviridine, perifosine, CAL101, PX-866, BEZ235, SF1126, INK1117, IPI-145, BKM120, XL147, XL765, Palomid529, GSK1059615, ZSTK474, PWT33597, IC87114, TGI00-115, CAL263, PI-103, GNE-477, CUDC-907, and AEZS-136;

[0289] AKT inhibitors include Akt-1-1 (inhibits Aktl) (Barnett et al., Biochem. J. 2005, 385(Pt. 2):399-408); Akt-1-1,2 (inhibits Akl and 2) (Barnett et al., Biochem. J. 2005, 385(Pt. 2):399-408); API-59CJ-Ome (e.g., Jin et al., Br. J. Cancer 2004, 91:1808-12); 1-H-imidazo[4,5-c]pyridinyl compounds (e.g., WO05 / 011700); indole-3-carbinol and its derivatives (e.g., U.S. Pat. No. 6,656,963; Sarkar and Li J Nutr. 2004, 134(12) Suppl):3493S-3498S); perifosine (e.g., interferes with Akt membrane localization; Dasmahapatra et al. Clin. Cancer Res. 2004, 10(15):5242-52); phosphatidylinositol ether lipid analogs (e.g., Gills and Dennis Expert. Opin. Investig. Drugs 2004, 13:787-97); and triciribine (TCN or API-2 or NCI identifier: NSC154020; Yang et al., Cancer Res. 2004, 64:4394-9).

[0290] mTOR inhibitors include ATP-competitive mTORC1 / mTORC2 inhibitors, e.g., PI-103, PP242, PP30; Torin1; FKBP12 enhancers; 4H-1-benzopyran-4-one derivatives; and rapamycin (also known as sirolimus) and its derivatives (temsirolimus (Torisel®); everolimus (Afinitor®; WO 94 / 09010); ridaforolimus (also known as deforolimus or AP23573); rapalogs disclosed, for example, in WO 98 / 02441 and WO 01 / 14387, e.g., AP23464 and AP23841; 40-(2-hydroxyethyl)rapamycin; 40-[3-hydroxy(hydroxymethyl)methylpropanoate]-rapamycin (also known as CC1779); 40-epi-(tetrazolito)-rapamycin 16-pentynyloxy-32(S)-dihydrorapamycin (also known as ABT578); 32-deoxorapamycin; 16-pentynyloxy-32(S)-dihydrorapamycin; derivatives disclosed in WO 05 / 005434; U.S. Patent Nos. 5,258,389, 5,118,677, 5,118,678, 5,100,883, 5,151,413, 5,120,842, and 5,256,790, and WO 94 / 005434. 090101, WO92 / 05179, WO93 / 111130, WO94 / 02136, WO94 / 02485, WO95 / 14023, WO94 / 02136, WO95 / 16691, WO96 / 41807, WO96 / 41807, and WO2018204416; and phosphorus-containing rapamycin derivatives (e.g., WO05 / 016252). In some embodiments, the mTOR inhibitor is a bisteric inhibitor (see, e.g., WO2018204416, WO2019212990, and WO2019212991), such as a compound having the structure: [ka] The RMC-5552 has the following.

[0291] BRAF inhibitors that can be used in combination with the compounds of the present invention include, for example, vemurafenib, dabrafenib, and encorafenib. BRAF can include class 3 BRAF mutations. In some embodiments, class 3 BRAF mutations are selected from one or more of the following amino acid substitutions in human BRAF: D287H; P367R; V459L; G466V; G466E; G466A; S467L; G469E; N581S; N581I; D594N; D594G; D594A; D594H; F595L; G596D; G596R and A762E.

[0292] MCL-1 inhibitors include, but are not limited to, AMG-176, MIK665, and S63845. Myeloid cell leukemia-1 (MCL-1) protein is one of the major anti-apoptotic members of the B-cell lymphoma-2 (BCL-2) protein family. Overexpression of MCL-1 is closely associated with tumor progression and resistance to not only conventional chemotherapy but also targeted therapeutics, including BCL-2 inhibitors such as ABT-263.

[0293] In some embodiments, the additional therapeutic agent is selected from the group consisting of a MEK inhibitor, a HER2 inhibitor, a SHP2 inhibitor, a CDK4 / 6 inhibitor, an mTOR inhibitor, a SOS1 inhibitor, and a PD-L1 inhibitor. In some embodiments, the additional therapeutic agent is selected from the group consisting of a MEK inhibitor, a SHP2 inhibitor, and a PD-L1 inhibitor. See, e.g., Hallin et al., Cancer Discovery, DOI: 10.1158 / 2159-8290 (October 28, 2019) and Canon et al., Nature, 575:217 (2019). In some embodiments, a Ras inhibitor of the present invention is used in combination with a MEK inhibitor and a SOS1 inhibitor. In some embodiments, a Ras inhibitor of the present invention is used in combination with a PD-L1 inhibitor and a SOS1 inhibitor. In some embodiments, a Ras inhibitor of the present invention is used in combination with a PD-L1 inhibitor and a SHP2 inhibitor. In some embodiments, a Ras inhibitor of the present invention is used in combination with a MEK inhibitor and a SHP2 inhibitor. In some embodiments, the cancer is colorectal cancer and the treatment comprises administering a Ras inhibitor of the present invention in combination with a second or third therapeutic agent.

[0294] Proteasome inhibitors include, but are not limited to, carfilzomib (Kyprolis®), bortezomib (Velcade®), and oprozomib.

[0295] Immunotherapies include, but are not limited to, monoclonal antibodies, immunomodulatory imides (IMiDs), GITR agonists, genetically engineered T cells (e.g., CAR-T cells), bispecific antibodies (e.g., BiTEs), and anti-PD-1, anti-PD-L1, anti-CTLA4, anti-LAG1, and anti-OX40 agents.

[0296] Immunomodulatory agents (IMiDs) are a class of immunomodulatory drugs (drugs that modify the immune response) that contain an imide group. The IMiD class includes thalidomide and its analogs (lenalidomide, pomalidomide, and apremilast).

[0297] Exemplary anti-PD-1 antibodies, and methods for their use, are described by Goldberg et al., Blood 2007, 110(1):186-192; Thompson et al., Clin. Cancer Res. 2007, 13(6):1757-1761; and WO06 / 121168 A1), and in addition, elsewhere herein.

[0298] GITR agonists include GITR fusion proteins and anti-GITR antibodies (e.g., bivalent anti-GITR antibodies), such as those described in U.S. Pat. Nos. 6,111,090, 8,586,023, WO2010 / 003118, and WO2011 / 090754, or those described in U.S. Pat. No. 7,025,962, EP1947183, U.S. Pat. Nos. 7,812,135, 8,388,967, and 8,591 ,886, 7,618,632, EP1866339, and the anti-GITR antibodies described in WO2011 / 028683, WO2013 / 039954, WO05 / 007190, WO07 / 133822, WO05 / 055808, WO99 / 40196, WO01 / 03720, WO99 / 20758, WO06 / 083289, WO05 / 115451, and WO2011 / 051726.

[0299] Another example of a therapeutic agent that can be used in combination with the compounds of the present invention is an anti-angiogenic agent.Anti-angiogenic agents include, but are not limited to, in vitro synthetically prepared chemical compositions, antibodies, antigen-binding regions, radionuclides, and combinations and conjugates thereof.Anti-angiogenic agents can be agonists, antagonists, allosteric modulators, toxins, or more generally, they can act to inhibit or stimulate their targets (e.g., activate or inhibit receptors or enzymes), thereby promoting cell death or stopping cell proliferation.In some embodiments, one or more additional therapies include an anti-angiogenic agent.

[0300] Antiangiogenic agents can be MMP-2 (matrix metalloproteinase 2) inhibitors, MMP-9 (matrix metalloproteinase 9) inhibitors, and COX-II (cyclooxygenase 11) inhibitors. Non-limiting examples of antiangiogenic agents include rapamycin, temsirolimus (CCI-779), everolimus (RAD001), sorafenib, sunitinib, and bevacizumab. Examples of useful COX-II inhibitors include alecoxib, valdecoxib, and rofecoxib. Examples of useful matrix metalloproteinase inhibitors are those disclosed in WO96 / 33172, WO96 / 27583, WO98 / 07697, WO98 / 03516, WO98 / 34918, WO98 / 34915, WO98 / 33768, WO98 / 30566, WO90 / 05719, WO99 / 52910, WO99 / 52 889, WO99 / 29667, WO99007675, EP0606046, EP0780386, EP1786785, EP1181017, EP0818442, EP1004578, and US20090012085, as well as U.S. Patent Nos. 5,863,949 and 5,861,510. Preferred MMP-2 and MMP-9 inhibitors are those that have little or no activity inhibiting MMP-1. More preferred are those that selectively inhibit MMP-2 or MMP-9 relative to other matrix metalloproteinases (i.e., MMP-1, MMP-3, MMP-4, MMP-5, MMP-6, MMP-7, MMP-8, MMP-10, MMP-11, MMP-12, and MMP-13). Some specific examples of MMP inhibitors are AG-3340, RO32-3555, and RS13-0830.

[0301] Further exemplary anti-angiogenic agents include KDR (kinase domain receptor) inhibitors (e.g., antibodies and antigen binding regions that specifically bind to kinase domain receptors), anti-VEGF agents (e.g., antibodies or antigen binding regions that specifically bind to VEGF (e.g., bevacizumab), or soluble VEGF receptors or ligand binding regions thereof), e.g., VEGF-TRAP™, and anti-VEGF receptor agents (e.g., antibodies or antigen binding regions that specifically bind thereto), EGFR inhibitors (e.g., antibodies or antigen binding regions that specifically bind thereto), e.g., Vectibix® (panitumumab), erlotinib (Tarceva®), anti-Ang1 and anti-Ang2 agents (e.g., antibodies or antigen binding regions that specifically bind thereto or their receptors, e.g., Tie2 / Tek), and anti-Tie2 kinase inhibitors (e.g., antibodies or antigen binding regions that specifically bind thereto). Other anti-angiogenic agents include Campath, IL-8, B-FGF, Tek antagonists (US2003 / 0162712; US6,413,932), anti-TWEAK agents (e.g., specifically binding antibodies or antigen-binding regions, or soluble TWEAK receptor antagonists; see US6,727,225), ADAM disintegrin domains to antagonize integrin binding to its ligands (US2002 / 0042368), specifically binding anti-eph receptor or anti-ephrin antibodies. or antigen-binding regions (U.S. Patent Nos. 5,981,245; 5,728,813; 5,969,110; 6,596,852; 6,232,447; 6,057,124 and patent family members thereof), and anti-PDGF-BB antagonists (e.g., antibodies or antigen-binding regions that specifically bind), as well as antibodies or antigen-binding regions that specifically bind to PDGF-BB ligands, and PDGFR kinase inhibitors (e.g., antibodies or antigen-binding regions that specifically bind thereto). Additional anti-angiogenic agents include SD-7784 (Pfizer, USA), cilengitide (Merck KGaA, Germany, EPO0770622), pegaptanib octasodium (Gilead Sciences,USA), alphastatin (BioActa, UK), M-PGA (Celgene, USA, US5712291), ilomastat (Arriva, USA, US5892112), emaxanib (Pfizer, USA, US5792783), vatalanib (Novartis, Switzerland), 2-methoxyestradiol (EntreMed, USA), TLC ELL-12 (Elan, Ireland), anecortave acetate (Alcon, USA), alpha-D148 Mab (Amgen, USA), CEP-7055 (Cephalon, USA), anti-Vn Mab (Crucell, Netherlands), DAC antiangiogenic agent (ConjuChem, Canada), angiocidin (InKine Pharmaceutical, USA), KM-2550 (Kyowa Hakko, Japan), SU-0879 (Pfizer, USA), CGP-79787 (Novartis, Switzerland, EP0970070), ARGENT technology (Ariad, USA), YIGSR-Stealth (Johnson & Johnson, USA), fibrinogen-E fragment (BioActa, UK), angiogenesis inhibitors (Trigen, UK), TBC-1635 (Encysive Pharmaceuticals, USA), SC-236 (Pfizer, USA), ABT-567 (Abbott, USA), metastatin (EntreMed, USA), maspin (Sosei, Japan), 2-methoxyestradiol (Oncology Sciences Corporation, USA), ER-68203-00 (IV AX, USA), BeneFin (Lane Labs, USA), Tz-93 (Tsumura, Japan), TAN-1120 (Takeda, Japan), FR-111142 (Fujisawa, Japan, JP02233610), platelet factor 4 (RepliGen, USA, EP407122), vascular endothelial growth factor antagonist (Borean, Denmark), bevacizumab (pINN) (Genentech, USA), angiogenesis inhibitor (SUGEN, USA), XL 784 (Exelixis, USA), XL 647 (Exelixis,(USA), MAb, alpha5beta3 integrin, second generation (Applied Molecular Evolution, USA and MediImmune, USA), enzastaurin hydrochloride (Lilly, USA), CEP 7055 (Cephalon, USA and Sanofi-Synthelabo, France), BC 1 (Genoa Institute of Cancer Research, Italy), rBPI 21 and BPI-derived antiangiogenic agents (XOMA, USA), PI 88 (Progen, Australia), cilengitide (Merck KGaA, Germany, Munich Technical University, Germany, Scripps Clinic and Research Foundation, USA), AVE 8062 (Ajinomoto, Japan), AS 1404 (Cancer Research Laboratory, New Zealand), SG 292 (Telios, USA), endostatin (Boston Children's Hospital, USA), ATN 161 (Attenuon, USA), 2-methoxyestradiol (Boston Children's Hospital, USA), ZD 6474 (AstraZeneca, UK), ZD 6126 (Angiogene Pharmaceuticals, UK), PPI 2458 (Praecis, USA), AZD 9935 (AstraZeneca, UK), AZD 2171 (AstraZeneca, UK), vatalanib (pINN) (Novartis, Switzerland and Schering AG, Germany), tissue factor pathway inhibitor (EntreMed, USA), pegaptanib (Pinn) (Gilead Sciences, USA), xanthorrhizol (Yonsei University, South Korea), vaccine, gene-based VEGF-2 (Scripps Clinic and Research Foundation, USA), SPV5.2 (Supratek, Canada), SDX 103 (University of of California at San Diego,USA), PX 478(ProlX,(USA), metastatin (EntreMed, USA), troponin I (Harvard University, USA), SU 6668 (SUGEN, USA), OXI 4503 (OXiGENE, USA), o-guanidine (Dimensional Pharmaceuticals, USA), motuporamine C (British Columbia University, Canada), CDP 791 (Celltech Group, UK), atiprimod (pINN) (GlaxoSmithKline, UK), E 7820 (Eisai, Japan), CYC 381 (Harvard University, USA), AE 941 (Aeterna, Canada), vaccine, angiogenic agent (EntreMed, USA), urokinase plasminogen activator inhibitor (Dendreon, USA), oglufanide (pINN) (Melmotte, USA), HIF-lalfa inhibitor (Xenova, UK), CEP 5214 (Cephalon, USA), BAY RES 2622 (Bayer, Germany), angiocidin (InKine, USA), A6 (Angstrom, USA), KR 31372 (Korea Research Institute of Chemical Technology, South Korea), GW 2286 (GlaxoSmithKline, UK), EHT 0101 (ExonHit, France), CP 868596 (Pfizer, USA), CP 564959 (OSI, USA), CP 547632 (Pfizer, USA), 786034 (GlaxoSmithKline, UK), KRN 633 (Kirin Brewery, Japan), drug delivery system, intraocular, 2-methoxyestradiol, Anginex (Maastricht University, Netherlands and University of Minnesota, USA), ABT 510 (Abbott, USA), AAL 993 (Novartis, Switzerland), VEGI (ProteomTech, USA), tumor necrosis factor-alpha inhibitor, SU 11248 (Pfizer, USA and SUGEN, USA), ABT 518 (Abbott,USA), YH16 (Yantai Rongchang, China), S-3APG (Boston Children's Hospital, USA and EntreMed, USA), MAb, KDR (ImClone Systems, USA), MAb, alpha5beta (Protein Design, USA), KDR kinase inhibitor (Celltech Group, UK and Johnson & Johnson, USA), GFB 116 (University of South Florida, USA and Yale University, USA), CS 706 (Sankyo, Japan), combretastatin A4 prodrug (Arizona State University, USA), chondroitinase AC (IBEX, Canada), BAY RES 2690 (Bayer, Germany), AGM 1470 (Harvard University, USA, Takeda, Japan, and TAP, USA), AG 13925 (Agouron, USA), tetrathiomolybdate (University of Michigan, USA), GCS 100 (Wayne State University, USA), CV 247 (Ivy Medical, UK), CKD 732 (Chong Kun Dang, South Korea), irsogladine (Nippon Shinyaku, Japan), RG 13577 (Aventis, France), WX 360 (Wilex, Germany), squalamine (Genaera, USA), RPI 4610 (Sirna, USA), heparanase inhibitor (InSight, Israel), KL 3106 (Kolon, South Korea), honokiol (Emory University, USA), ZK CDK (Schering AG, Germany), ZK Angio (Schering AG, Germany), ZK 229561 (Novartis, Switzerland and Schering AG, Germany), XMP 300 (XOMA, USA), VGA 1102 (Taisho, Japan), VE-cadherin-2 antagonist (ImClone Systems,(USA), vasostatin (National Institutes of Health, USA), Flk-1 (ImClone Systems, USA), TZ 93 (Tsumura, Japan), tumstatin (Beth Israel Hospital, USA), truncated soluble FLT 1 (vascular endothelial growth factor receptor 1) (Merck & Co, USA), Tie-2 ligand (Regeneron, USA), and and thrombospondin 1 inhibitor (Allegheny Health, Education and Research Foundation, USA).

[0302] Further examples of therapeutic agents that may be used in combination with the compounds of the invention include agents (e.g., antibodies, antigen-binding regions, or soluble receptors) that specifically bind and inhibit the activity of growth factors, such as antagonists of hepatocyte growth factor (HGF, also known as scatter factor), and antibodies or antigen-binding regions that specifically bind to its receptor, c-Met.

[0303] Another example of a therapeutic agent that can be used in combination with the compounds of the present invention is an autophagy inhibitor. Autophagy inhibitors include, but are not limited to, chloroquine, 3-methyladenine, hydroxychloroquine (Plaquenil™), bafilomycin A1, 5-amino-4-imidazolecarboxamide riboside (AICAR), okadaic acid, autophagy-inhibiting algal toxins that inhibit type 2A or type 1 protein phosphatases, cAMP analogs, and drugs that increase cAMP levels, such as adenosine, LY204002, N6-mercaptopurine riboside, and vinblastine. Antisense or siRNA inhibitors that inhibit the expression of proteins, including but not limited to ATG5 (involved in autophagy), can also be used. In some embodiments, one or more additional therapies include an autophagy inhibitor.

[0304] Another example of a therapeutic agent that can be used in combination with the compound of the present invention is an anti-cancer agent. In some embodiments, one or more additional therapies include an anti-cancer agent. Non-limiting examples of an anti-cancer agent include acemannan, aclarubicin, aldesleukin, alemtuzumab, alitretinoin, altretamine, amifostine, aminolevulinic acid, amrubicin, amsacrine, anagrelide, anastrozole, ancer, ancestim, aruglavin, arsenic trioxide, BAM-002 (Novelos), bexarotene, bicalutamide, broxuridine, capecitabine, celmoleukin, cetrorelix, cladribine, clotrimazole, cytarabine octophosphate, DA 3030 (Dong-A), daclizumab, denileukin diftitox, deslorelin, dexrazoxane, dilazep, docetaxel, docosanol, doxercalciferol, doxifluridine, doxorubicin, bromocriptine, carmustine, cytarabine, fluorouracil, HIT diclofenac, interferon alpha, daunorubicin, doxorubicin, tretinoin, edelfosine, edrecolomab, eflornithine, emiteflu, epirubicin, epoetin beta, etoposide phosphate, exemestane, exisulind, fadrozole, filgrastim, finasteride, fludarabine phosphate, formestane, fotemustine, gallium nitrate, gemcitabine, gemtuzumab zogamicin zogamicin), gimeracil / oteracil / tegafur combination, glycopin, goserelin, heptaplatin, human chorionic gonadotropin, human fetal alpha-fetoprotein, ibandronate, idarubicin (imiquimod, int...

Claims

1. A method of treating immunorefractory lung cancer in a subject, comprising administering to the subject a RAS Ｇ１２Ｃ (ON) inhibitor.

2. A method for immunologically transforming the tumor microenvironment of cold lung cancer in a subject in need thereof, comprising administering to the subject a RAS. Ｇ１２Ｃ (ON) inhibitor.

3. The RAS Ｇ１２Ｃ (ON) Inhibitors of the tricomplex RAS Ｇ１２Ｃ The method according to claim 1 or 2, wherein the compound is an (ON) inhibitor.

4. 1. A method of treating immunorefractory lung cancer in a subject, comprising administering to the subject a RAS inhibitor of formula I: 【Chemistry 1】 or a pharma- ceutically acceptable salt thereof wherein the dotted lines represent 0, 1, 2, 3, or 4 non-adjacent double bonds; A is -N(H or CH ３ )C(O)-(CH ２ )—[wherein the amino nitrogen is —CH(R １０ )-, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene; B is absent or -CH(R ９ )-, >C=CR ９ R ９’ , or >CR ９ R ９’ [wherein carbon is —N(R １１ )C(O)-- is bonded to the carbonyl carbon thereof; optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or 5- to 6-membered heteroarylene; G is an optionally substituted C １ -C ４ Alkylene, optionally substituted C １ -C ４ Alkenylene, optionally substituted C １ -C ４ Heteroalkylene, 【Chemistry 2】 [In the formula, 【Chemistry 3】 HA -C(R ７ R ８ )-bonded to 【Chemistry 4】 [In the formula, 【Chemistry 5】 HA -C(R ７ R ８ )-.], optionally substituted C １ -C ４ heteroalkylene or 3- to 8-membered heteroarylene; L is a linker, said linker being acyclic or comprising a monocyclic, fused bicyclic, fused polycyclic, bridged bicyclic, or bridged polycyclic group; W is a bridging group comprising a vinyl ketone, vinyl sulfone, ynone, haloacetyl, or alkynyl sulfone; X １ is an optionally substituted C １ -C ２ Alkylene, NR, O, or S(O) ｎ and X ２ is O or NH, X ３ is N or CH; n is 0, 1, or 2; R is hydrogen, cyano, optionally substituted C １ -C ４ Alkyl, optionally substituted C ２ -C ４ Alkenyl, optionally substituted C ２ -C ４ Alkynyl, C(O)R', C(O)OR', C(O)N(R') ２ , S(O)R', S(O) ２ R', or S(O) ２ N(R') ２ and Each R ’ are independently H or optionally substituted C １ -C ４ is alkyl, Y １ is C, CH, or N; Y ２ , Y ３ , Y ４ , and Y ７ are independently C or N; Y ５ , CH, CH ２ or N, Y ６ is C(O), CH, CH ２ or N, R １ is cyano, optionally substituted C １ -C ６ Alkyl, optionally substituted C １ -C ６ heteroalkyl, an optionally substituted 3- to 6-membered cycloalkyl, an optionally substituted 3- to 6-membered cycloalkenyl, an optionally substituted 3- to 6-membered heterocycloalkyl, an optionally substituted 6- to 10-membered aryl, or an optionally substituted 5- to 10-membered heteroaryl; or R １ and R ２ combine with the atom to which they are attached to form an optionally substituted 3- to 14-membered heterocycloalkyl; R ２ is absent, hydrogen, or optionally substituted C １ -C ６ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 7-membered heterocycloalkyl, optionally substituted 6-membered aryl, optionally substituted 5- or 6-membered heteroaryl; R ３ does not exist, or R ２ and R ３ combine with the atom to which they are attached to form an optionally substituted 3- to 8-membered cycloalkyl or an optionally substituted 3- to 14-membered heterocycloalkyl; R ４ is absent, hydrogen, halogen, cyano, or methyl optionally substituted with 1 to 3 halogens; R ５ is hydrogen; halogen, cyano, hydroxy, or C １ -C ４ C optionally substituted with alkoxy １ -C ４ alkyl; cyclopropyl, or cyclobutyl; R ６ is hydrogen or methyl; R ７ is hydrogen, halogen, or optionally substituted C １ -C ３ is alkyl, or R ６ and R ７ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R ８ is hydrogen, halogen, hydroxy, cyano, optionally substituted C １ -C ３ Alkoxy, optionally substituted C １ -C ３ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, an optionally substituted 3- to 8-membered cycloalkyl, an optionally substituted 3- to 14-membered heterocycloalkyl, an optionally substituted 5- to 10-membered heteroaryl, or an optionally substituted 6- to 10-membered aryl; or R ７ and R ８ combine with the carbon atom to which they are attached to form C=CR ７’ R ８’ ;C=N(OH), C=N(OC １ -C ３ alkyl), C=O, C=S, C=NH, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted 3- to 7-membered heterocycloalkyl; R ７ａ and R ８ａ are independently hydrogen, halo, optionally substituted C １ -C ３ alkyl or, when combined with the carbon to which they are attached, form a carbonyl; R ７’ is hydrogen, halogen, or optionally substituted C １ -C ３ alkyl, R ８’ is hydrogen, halogen, hydroxy, cyano, optionally substituted C １ -C ３ Alkoxy, optionally substituted C １ -C ３ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, an optionally substituted 3- to 8-membered cycloalkyl, an optionally substituted 3- to 14-membered heterocycloalkyl, an optionally substituted 5- to 10-membered heteroaryl, or an optionally substituted 6- to 10-membered aryl; or R ７’ and R ８’ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R ９ is H, F, optionally substituted C １ -C ６ Alkyl, optionally substituted C １ -C ６ heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 7-membered heterocycloalkyl; or R ９ and L, taken together with the atom to which they are attached, form an optionally substituted 3- to 14-membered heterocycloalkyl; R ９’ is hydrogen or an optionally substituted C １ -C ６ is alkyl, or R ９ and R ９’ combine with the atom to which they are attached to form a 3- to 6-membered cycloalkyl or a 3- to 6-membered heterocycloalkyl; R １０ is hydrogen, halo, hydroxy, C １ -C ３ Alkoxy, or C １ -C ３ is alkyl, R １０ａ is hydrogen or halo, R １１ is hydrogen or C １ -C ３ is alkyl, and R ２１ is H or C １ -C ３ wherein R is an alkyl group.

5. 1. A method of immunologically altering the tumor microenvironment of cold lung cancer in a subject in need thereof, comprising administering to the subject a RAS inhibitor of formula I: 【Chemistry 6】 or a pharma- ceutically acceptable salt thereof wherein the dotted lines represent 0, 1, 2, 3, or 4 non-adjacent double bonds; A is -N(H or CH ３ )C(O)-(CH ２ )—[wherein the amino nitrogen is —CH(R １０ )-, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene; B is absent or -CH(R ９ )-, >C=CR ９ R ９’ , or >CR ９ R ９’ [wherein carbon is —N(R １１ )C(O)-- is bonded to the carbonyl carbon thereof; optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or 5- to 6-membered heteroarylene; G is an optionally substituted C １ -C ４ Alkylene, optionally substituted C １ -C ４ Alkenylene, optionally substituted C １ -C ４ Heteroalkylene, 【Chemistry 7】 [In the formula, 【Chemistry 8】 HA -C(R ７ R ８ )-bonded to 【Chemistry 9】 [In the formula, 【Chemistry 10】 HA -C(R ７ R ８ )-.], optionally substituted C １ -C ４ heteroalkylene or 3- to 8-membered heteroarylene; L is a linker, said linker being acyclic or comprising a monocyclic, fused bicyclic, fused polycyclic, bridged bicyclic, or bridged polycyclic group; W is a bridging group comprising a vinyl ketone, vinyl sulfone, ynone, haloacetyl, or alkynyl sulfone; X １ is an optionally substituted C １ -C ２ Alkylene, NR, O, or S(O) ｎ and X ２ is O or NH, X ３ is N or CH; n is 0, 1, or 2; R is hydrogen, cyano, optionally substituted C １ -C ４ Alkyl, optionally substituted C ２ -C ４ Alkenyl, optionally substituted C ２ -C ４ Alkynyl, C(O)R', C(O)OR', C(O)N(R') ２ , S(O)R', S(O) ２ R', or S(O) ２ N(R') ２ and Each R ’ are independently H or optionally substituted C １ -C ４ is alkyl, Y １ is C, CH, or N; Y ２ , Y ３ , Y ４ , and Y ７ are independently C or N; Y ５ , CH, CH ２ or N, Y ６ is C(O), CH, CH ２ or N, R １ is cyano, optionally substituted C １ -C ６ Alkyl, optionally substituted C １ -C ６ heteroalkyl, an optionally substituted 3- to 6-membered cycloalkyl, an optionally substituted 3- to 6-membered cycloalkenyl, an optionally substituted 3- to 6-membered heterocycloalkyl, an optionally substituted 6- to 10-membered aryl, or an optionally substituted 5- to 10-membered heteroaryl; or R １ and R ２ combine with the atom to which they are attached to form an optionally substituted 3- to 14-membered heterocycloalkyl; R ２ is absent, hydrogen, or optionally substituted C １ -C ６ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 7-membered heterocycloalkyl, optionally substituted 6-membered aryl, optionally substituted 5- or 6-membered heteroaryl; R ３ does not exist, or R ２ and R ３ combine with the atom to which they are attached to form an optionally substituted 3- to 8-membered cycloalkyl or an optionally substituted 3- to 14-membered heterocycloalkyl; R ４ is absent, hydrogen, halogen, cyano, or methyl optionally substituted with 1 to 3 halogens; R ５ is hydrogen; halogen, cyano, hydroxy, or C １ -C ４ C optionally substituted with alkoxy １ -C ４ alkyl; cyclopropyl, or cyclobutyl; R ６ is hydrogen or methyl; R ７ is hydrogen, halogen, or optionally substituted C １ -C ３ is alkyl, or R ６ and R ７ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R ８ is hydrogen, halogen, hydroxy, cyano, optionally substituted C １ -C ３ Alkoxy, optionally substituted C １ -C ３ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, an optionally substituted 3- to 8-membered cycloalkyl, an optionally substituted 3- to 14-membered heterocycloalkyl, an optionally substituted 5- to 10-membered heteroaryl, or an optionally substituted 6- to 10-membered aryl; or R ７ and R ８ combine with the carbon atom to which they are attached to form C=CR ７’ R ８’ ;C=N(OH), C=N(OC １ -C ３ alkyl), C=O, C=S, C=NH, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted 3- to 7-membered heterocycloalkyl; R ７ａ and R ８ａ are independently hydrogen, halo, optionally substituted C １ -C ３ alkyl or, when combined with the carbon to which they are attached, form a carbonyl; R ７’ is hydrogen, halogen, or optionally substituted C １ -C ３ alkyl, R ８’ is hydrogen, halogen, hydroxy, cyano, optionally substituted C １ -C ３ Alkoxy, optionally substituted C １ -C ３ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, an optionally substituted 3- to 8-membered cycloalkyl, an optionally substituted 3- to 14-membered heterocycloalkyl, an optionally substituted 5- to 10-membered heteroaryl, or an optionally substituted 6- to 10-membered aryl; or R ７’ and R ８’ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R ９ is H, F, optionally substituted C １ -C ６ Alkyl, optionally substituted C １ -C ６ heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 7-membered heterocycloalkyl; or R ９ and L, taken together with the atom to which they are attached, form an optionally substituted 3- to 14-membered heterocycloalkyl; R ９’ is hydrogen or an optionally substituted C １ -C ６ is alkyl, or R ９ and R ９’ combine with the atom to which they are attached to form a 3- to 6-membered cycloalkyl or a 3- to 6-membered heterocycloalkyl; R １０ is hydrogen, halo, hydroxy, C １ -C ３ Alkoxy, or C １ -C ３ is alkyl, R １０ａ is hydrogen or halo, R １１ is hydrogen or C １ -C ３ is alkyl, and R ２１ is H or C １ -C ３ wherein R is an alkyl group.

6. 6. The method of claim 5, wherein the subject is resistant to an immune checkpoint inhibitor prior to the transformation of the tumor microenvironment.

7. 7. The method of claim 5 or 6, wherein administration of the RAS inhibitor alters the tumor microenvironment, thereby sensitizing the cancer to treatment with an immune checkpoint inhibitor.

8. The method of any one of claims 1 to 7, further comprising administering to the subject an SHP2 inhibitor.

9. A method of treating immunorefractory lung cancer in a subject, comprising administering to the subject a RAS Ｇ１２Ｃ The method comprises administering an (ON) inhibitor, an SHP2 inhibitor, and an immune checkpoint inhibitor.

10. A method for immunologically transforming the tumor microenvironment of cold lung cancer in a subject in need thereof, comprising administering to the subject a RAS. Ｇ１２Ｃ The method comprises administering an (ON) inhibitor, an SHP2 inhibitor, and an immune checkpoint inhibitor.

11. The RAS Ｇ１２Ｃ (ON) The inhibitor is a compound of formula I: 【Chemistry 11】 or a pharma- ceutically acceptable salt thereof wherein the dotted lines represent 0, 1, 2, 3, or 4 non-adjacent double bonds; A is -N(H or CH ３ )C(O)-(CH ２ )—[wherein the amino nitrogen is —CH(R １０ )-, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene; B is absent or -CH(R ９ )-, >C=CR ９ R ９’ , or >CR ９ R ９’ [wherein carbon is —N(R １１ )C(O)-- is bonded to the carbonyl carbon thereof; optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 6-membered arylene, or 5- to 6-membered heteroarylene; G is an optionally substituted C １ -C ４ Alkylene, optionally substituted C １ -C ４ Alkenylene, optionally substituted C １ -C ４ Heteroalkylene, 【Chemistry 12】 [In the formula, 【Chemistry 13】 HA -C(R ７ R ８ )-bonded to 【Chemistry 14】 [In the formula, 【Chemistry 15】 HA -C(R ７ R ８ )-.], optionally substituted C １ -C ４ heteroalkylene or 3- to 8-membered heteroarylene; L is a linker, said linker being acyclic or comprising a monocyclic, fused bicyclic, fused polycyclic, bridged bicyclic, or bridged polycyclic group; W is a bridging group comprising a vinyl ketone, vinyl sulfone, ynone, haloacetyl, or alkynyl sulfone; X １ is an optionally substituted C １ -C ２ Alkylene, NR, O, or S(O) ｎ and X ２ is O or NH, X ３ is N or CH; n is 0, 1, or 2; R is hydrogen, cyano, optionally substituted C １ -C ４ Alkyl, optionally substituted C ２ -C ４ Alkenyl, optionally substituted C ２ -C ４ Alkynyl, C(O)R', C(O)OR', C(O)N(R') ２ , S(O)R', S(O) ２ R', or S(O) ２ N(R') ２ and Each R ’ are independently H or optionally substituted C １ -C ４ is alkyl, Y １ is C, CH, or N; Y ２ , Y ３ , Y ４ , and Y ７ are independently C or N; Y ５ , CH, CH ２ or N, Y ６ is C(O), CH, CH ２ or N, R １ is cyano, optionally substituted C １ -C ６ Alkyl, optionally substituted C １ -C ６ heteroalkyl, an optionally substituted 3- to 6-membered cycloalkyl, an optionally substituted 3- to 6-membered cycloalkenyl, an optionally substituted 3- to 6-membered heterocycloalkyl, an optionally substituted 6- to 10-membered aryl, or an optionally substituted 5- to 10-membered heteroaryl; or R １ and R ２ combine with the atom to which they are attached to form an optionally substituted 3- to 14-membered heterocycloalkyl; R ２ is absent, hydrogen, or optionally substituted C １ -C ６ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, optionally substituted 3- to 6-membered cycloalkyl, optionally substituted 3- to 7-membered heterocycloalkyl, optionally substituted 6-membered aryl, optionally substituted 5- or 6-membered heteroaryl; R ３ does not exist, or R ２ and R ３ combine with the atom to which they are attached to form an optionally substituted 3- to 8-membered cycloalkyl or an optionally substituted 3- to 14-membered heterocycloalkyl; R ４ is absent, hydrogen, halogen, cyano, or methyl optionally substituted with 1 to 3 halogens; R ５ is hydrogen; halogen, cyano, hydroxy, or C １ -C ４ C optionally substituted with alkoxy １ -C ４ alkyl; cyclopropyl, or cyclobutyl; R ６ is hydrogen or methyl; R ７ is hydrogen, halogen, or optionally substituted C １ -C ３ is alkyl, or R ６ and R ７ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R ８ is hydrogen, halogen, hydroxy, cyano, optionally substituted C １ -C ３ Alkoxy, optionally substituted C １ -C ３ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, an optionally substituted 3- to 8-membered cycloalkyl, an optionally substituted 3- to 14-membered heterocycloalkyl, an optionally substituted 5- to 10-membered heteroaryl, or an optionally substituted 6- to 10-membered aryl; or R ７ and R ８ combine with the carbon atom to which they are attached to form C=CR ７’ R ８’ ;C=N(OH), C=N(OC １ -C ３ alkyl), C=O, C=S, C=NH, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted 3- to 7-membered heterocycloalkyl; R ７ａ and R ８ａ are independently hydrogen, halo, optionally substituted C １ -C ３ alkyl or, when combined with the carbon to which they are attached, form a carbonyl; R ７’ is hydrogen, halogen, or optionally substituted C １ -C ３ alkyl, R ８’ is hydrogen, halogen, hydroxy, cyano, optionally substituted C １ -C ３ Alkoxy, optionally substituted C １ -C ３ Alkyl, optionally substituted C ２ -C ６ Alkenyl, optionally substituted C ２ -C ６ alkynyl, an optionally substituted 3- to 8-membered cycloalkyl, an optionally substituted 3- to 14-membered heterocycloalkyl, an optionally substituted 5- to 10-membered heteroaryl, or an optionally substituted 6- to 10-membered aryl; or R ７’ and R ８’ combine with the carbon atom to which they are attached to form an optionally substituted 3- to 6-membered cycloalkyl or an optionally substituted 3- to 7-membered heterocycloalkyl; R ９ is H, F, optionally substituted C １ -C ６ Alkyl, optionally substituted C １ -C ６ heteroalkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted 3- to 7-membered heterocycloalkyl; or R ９ and L, taken together with the atom to which they are attached, form an optionally substituted 3- to 14-membered heterocycloalkyl; R ９’ is hydrogen or an optionally substituted C １ -C ６ is alkyl, or R ９ and R ９’ combine with the atom to which they are attached to form a 3- to 6-membered cycloalkyl or a 3- to 6-membered heterocycloalkyl; R １０ is hydrogen, halo, hydroxy, C １ -C ３ Alkoxy, or C １ -C ３ is alkyl, R １０ａ is hydrogen or halo, R １１ is hydrogen or C １ -C ３ is alkyl, and R ２１ is H or C １ -C ３ The method according to any one of claims 1 to 3 and 8 to 10, wherein

12. The method of any one of claims 1 to 11, wherein the RAS inhibitor is a compound of Table 1 or a pharma- ceutically acceptable salt thereof.

13. The RAS inhibitor is 【Chemistry 16】 or a pharma- ceutically acceptable salt thereof.

14. The method of any one of claims 1 to 8 and 11 to 13, further comprising administering to the subject an immune checkpoint inhibitor.

15. 1. A method of treating immunorefractory lung cancer in a subject, comprising administering to the subject a RAS inhibitor, a SHP2 inhibitor, and an immune checkpoint inhibitor, wherein the RAS inhibitor: 【Chemistry 17】 or a pharma- ceutically acceptable salt thereof; The SHP2 inhibitor is 【Chemistry 18】 or a pharma- ceutically acceptable salt thereof.

16. 16. The method of any one of claims 9, 10, 14, and 15, wherein the immune checkpoint inhibitor is a PD-1 inhibitor.

17. The method of any one of claims 1 to 16, wherein the subject has previously been administered an immune checkpoint inhibitor.

18. The method of any one of claims 1 to 17, wherein the subject is resistant to treatment with an immune checkpoint inhibitor.

19. 20. The method of claim 18, wherein the subject has acquired resistance to treatment with an immune checkpoint inhibitor.

20. The method of any one of claims 1 to 19, wherein administration of the RAS inhibitor sensitizes the cancer to treatment with an immune checkpoint inhibitor.

21. The method of any one of claims 8 to 20, wherein the inhibitors are administered simultaneously or sequentially.

22. The method of any one of claims 8 to 21, wherein the inhibitors are administered as a single formulation or as separate formulations.

23. The method of any one of claims 1 to 22, wherein the subject has one or more tumors with a low tumor mutation burden.

24. The method of any one of claims 1 to 23, wherein the subject has a tumor that is one or more microsatellite stable.

25. The method of any one of claims 1 to 24, wherein the subject has one or more tumors with low microsatellite instability.

26. The method of any one of claims 1 to 25, wherein the subject has one or more tumors with low tumor immune infiltrate.

27. The method of any one of claims 1 to 26, wherein administration of the RAS inhibitor alters the tumor immune infiltrate compared to the tumor immune infiltrate in the absence of the RAS inhibitor.

28. 28. The method of claim 26 or 27, wherein the tumor immune infiltrate comprises antigen-presenting cells, myeloid cells, or lymphoid cells.

29. The method of any one of claims 1 to 28, wherein administration of the RAS inhibitor alters an anti-tumor immune response.

30. 30. The method of any one of claims 1-29, wherein administration of the RAS inhibitor alters the tumor microenvironment compared to the tumor microenvironment in the absence of the RAS inhibitor.

31. 31. The method of any one of claims 1 to 30, wherein administration of the RAS inhibitor converts an immunologically cold tumor into an immunologically hot tumor.

32. The method of any one of claims 1 to 31, wherein the method reduces tumor size or inhibits tumor growth.

33. The method of any one of claims 1 to 32, wherein the immunorefractory lung cancer is non-small cell lung cancer or small cell lung cancer.

34. 34. The method of any one of claims 1 to 33, wherein the immunorefractory lung cancer comprises a K-Ras G12C, H-Ras G12C, or N-Ras G12C mutation.

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