Biomarkers and methods for treating NSCLC

Abstract

Biomarkers and methods for the treatment of non-small cell lung cancer (NSCLC). Biomarkers for patient selection and prognosis, and diagnostic kits for analyzing the biomarkers.

Description

Technical Field

[0001] The present application relates to biomarkers and methods for treating non-small cell lung cancer (NSCLC). In particular, the present application provides biomarkers for patient selection and prognosis, and diagnostic kits for analyzing the biomarkers. Background Art

[0002] Non-small cell lung cancer (NSCLC) refers to any type of epithelial lung cancer other than small cell lung cancer (SCLC). It accounts for about 85% of all lung cancers. Compared with small cell carcinoma, NSCLC is relatively insensitive to chemotherapy. The survival rate of NSCLC from stage I to stage IV decreases significantly as the disease progresses. The five-year survival rate is 47% for stage I, 30% for stage II, 10% for stage III, and only 1% for stage IV.

[0003] Treatment for people with advanced NSCLC is primarily aimed at relieving pain and suffering through chemotherapy that indiscriminately targets all rapidly dividing cells, such as cisplatin, and through targeted drugs that are more tailored to the specific genetic aberrations found within the tumor. However, there is a need for improved treatments. Summary of the invention

[0004] This application can meet clinical needs.

[0005] In one aspect, the present application provides a method for treating a subject with NSCLC, the method comprising: (a) determining the level of CD73 in a sample from the subject, (b) assessing whether the subject is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level, and (c) administering the combination therapy to the subject if the subject is more likely to respond to or benefit from the combination therapy.

[0006] On the other hand, the present application provides a method for selecting a therapy for a subject with NSCLC, the method comprising: (a) determining the level of CD73 in a sample from the subject, and (b) assessing whether the subject is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level, and (c) if the subject is more likely to respond to or benefit from the combination therapy, selecting the combination therapy for the subject.

[0007] On the other hand, the present application provides a method for identifying a subject with NSCLC who is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample of the subject, (b) assessing whether the subject is more likely to respond to or benefit from the combination therapy based on the CD73 level, and (c) if the subject is more likely to respond to or benefit from the combination therapy, providing a recommendation of the combination therapy to the subject.

[0008] On the other hand, the present application provides a method for predicting the responsiveness of a subject with NSCLC to a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample of the subject, (b) assessing whether the subject is more likely to respond to or benefit from the combination therapy based on the CD73 level, and (c) if the subject is more likely to respond to or benefit from the combination therapy, providing a recommendation of the combination therapy to the subject.

[0009] In another aspect, the present application provides a method for identifying a subject with NSCLC who is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample from the subject, wherein an increase in CD73 in the sample relative to CD73 in a reference sample indicates that the subject is more likely to respond to or benefit from the combination therapy, and (b) providing a suggestion that the subject will be more likely to respond to or benefit from the combination therapy.

[0010] In another aspect, the present application provides a method for predicting the responsiveness of a subject with NSCLC to a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample from the subject, wherein an increase in CD73 in the sample relative to CD73 in a reference sample indicates that the subject is more likely to respond to or benefit from the combination therapy, and (b) providing a suggestion that the subject will have an increased likelihood of responding to or benefiting from the combination therapy.

[0011] In another aspect, the present application provides a method for treating a subject having non-small cell lung cancer (NSCLC), the method comprising: (a) identifying a NSCLC subject as likely to respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level in a sample of the subject, and (b) administering a CD73 antagonist and a PD1 / PD-L1 antagonist to the identified subject.

[0012] On the other hand, the present application provides a method for selecting a therapy for a subject with NSCLC, the method comprising: (a) determining the level of CD73 in a sample of the subject, and (b) assessing whether the subject is likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level, and (c) when it is determined that the subject is likely to respond to or benefit from the combination therapy, selecting the combination therapy for the subject.

[0013] In another aspect, the present application provides a method for treating a subject with NSCLC, the method comprising: administering a CD73 antagonist and a PD1 / PD-L1 antagonist to a subject with NSCLC whose CD73 level in a sample indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist.

[0014] On the other hand, the present application provides a method for predicting the responsiveness of a subject with NSCLC to a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the CD73 level in a sample of the subject, (b) assessing whether the subject is likely to respond to or benefit from the combination therapy based on the CD73 level, and (c) when it is determined that the subject is likely to respond to or benefit from the combination therapy, providing a recommendation of the combination therapy to the subject.

[0015] In another aspect, the present application provides a method for identifying a subject with NSCLC who is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist than a reference patient, the method comprising: (a) determining the level of CD73 in a sample from the subject, wherein an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample from a reference patient indicates that the subject is more likely to respond to or benefit from the combination therapy than the reference patient, and (b) providing a suggestion that the subject will be more likely to respond to or benefit from the combination therapy than the reference patient.

[0016] In another aspect, the present application provides a method for predicting responsiveness of a subject with NSCLC to a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample from the subject, wherein an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample from a reference patient indicates that the subject is more likely to respond to or benefit from the combination therapy than the reference patient; and (b) providing a suggestion that the subject will have an increased likelihood of responding to or benefiting from the combination therapy compared to the reference patient.

[0017] In another aspect, the present application provides the use of a CD73 antagonist and a PD1 / PD-L1 antagonist for the manufacture of a medicament for treating a subject with NSCLC, wherein the level of CD73 in a sample of the subject indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist.

[0018] In another aspect, the present application provides the use of a CD73 antagonist for the manufacture of a medicament for treating a subject having NSCLC, wherein the level of CD73 in a sample of the subject indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist, wherein the subject is also administered a PD1 / PD-L1 antagonist.

[0019] On the other hand, the present application provides the use of a PD1 / PD-L1 antagonist for the manufacture of a medicament for treating a subject having NSCLC, wherein the level of CD73 in a sample of the subject indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist, wherein the subject is also administered a CD73 antagonist.

[0020] In some embodiments of the methods of the present application, the CD73 antagonist of the combined therapy is a CD73 antibody. In some embodiments, the CD73 antibody is selected from the group consisting of polyclonal antibodies, monoclonal antibodies, Fab, scFv, diantibodies, triantibodies, miniantibodies, VHH and sdAb. In some embodiments, the CD73 antibody comprises: (a) HCDR1, HCDR2 and HCDR3 in the heavy chain variable region (VH) having the sequence as shown in SEQ ID No: 1, and (b) LCDR1, LCDR2 and LCDR3 in the light chain variable region (VL) having the sequence as shown in SEQ ID No: 2, wherein HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are according to the Kabat numbering scheme.

[0021] In some embodiments, the CD73 antibody comprises (1) HCDR1 comprising the amino acid sequence as shown in SEQ ID NO:3; (2) HCDR2 comprising the amino acid sequence as shown in SEQ ID NO:4; (3) HCDR3 comprising the amino acid sequence as shown in SEQ ID NO:5; (4) LCDR1 comprising the amino acid sequence as shown in SEQ ID NO:6; (5) LCDR2 comprising the amino acid sequence as shown in SEQ ID NO:7; and (6) LCDR3 comprising the amino acid sequence as shown in SEQ ID NO:8.

[0022] In some embodiments, the CD73 antibody is selected from the group consisting of uliledlimab (I-Mab Biopharma), oleclumab (AstraZeneca), CPI-006 (Corvus Pharma), BMS-986179 (Bristol-Myers Squibb), AB-680 (Arcus Biosciences), NZV-930 (SRF373, Surface Oncolohgy / Novartis), JAB-BX102 (Jacobio), AK119 (Akesobio), Sym024 (Symphogen), IBI 325 (Innovent), BR101 (Hisun BioRay), and LY3475070 (Eli Lilly).

[0023] In some embodiments, the PD-1 / PD-L1 antagonist of the combination therapy is a PD-1 antibody or a PD-L1 antibody. In some embodiments, the PD-1 antibody is selected from the group consisting of: Pembrolizumab, nivolumab, toripalimab, pidilizumab, cemiplimab, sintilimab, cetrelimab, spartalizumab, camrelizumab, tislelizumab, balstilimab, dostarlimab, ABBV-181, penpulimab, genolimzumab ), retifanlimab, sasanlimab, AMP-224, AB122, F-520, MEDI-3387, MEDI-5771, MEDI-0680, SG-001, BCD-100, BAT-1306, BI-754091, CBT-501, GLS-010, LZM-009, Sym-021, CS-1003, HLX-10, AK-103, AM-0001, ENUM-244C8, ENUM-388D4, JTX-4014, RXI-762, STI-A1110, HLX-20, SSI-361, APL-501, TJ0141H, and SNA-01. In some embodiments, the PD-L1 antibody is selected from the group consisting of atezolizumab, manelimab, avelumab, cosibelimab, durvalumab, envafolimab, socazolimab, BGB-A333, CK-301, CS-1001, FAZ-053, APL-502, MDX-1105, IMC-001, KD-005, Gensci-047, LY-3300054, SHR-1316, MSB-2311, AVA-004, CBT-502, JS-003, B12, and KY-1003.

[0024] In some embodiments, the combination therapy further comprises chemotherapy. In some embodiments, the chemotherapy is selected from the group consisting of platinum drugs (such as cisplatin, carboplatin), taxane drugs (such as paclitaxel, albumin-bound paclitaxel, docetaxel), vinorelbine, vinblastine, etoposide, pemetrexed and gemcitabine, and any combination thereof.

[0025] In some embodiments, the combination therapy comprises the simultaneous or separate administration of a CD73 antagonist and a PD-1 / PD-L1 antagonist.

[0026] In some embodiments, the NSCLC is stage III or stage IV NSCLC. In some embodiments, the NSCLC is advanced NSCLC or metastatic NSCLC. In some embodiments, the NSCLC is recurrent NSCLC. In some embodiments, the NSCLC has squamous histology or non-squamous histology.

[0027] In some embodiments, the subject is untreated. In some embodiments, the treatment is the first line of treatment. In some embodiments, the subject does not meet the first line of treatment or is rejected by the first line of treatment. In some embodiments, the first line of treatment is selected from the group consisting of chemotherapy, PD-1 antibody therapy, PD-L1 antibody therapy, and any combination thereof. In some embodiments, chemotherapy is selected from the group consisting of platinum drugs (such as cisplatin, carboplatin), taxane drugs (such as paclitaxel, albumin-bound paclitaxel, docetaxel), vinorelbine, vinblastine, etoposide, pemetrexed and gemcitabine, and any combination thereof. In some embodiments, the PD-1 antibody is selected from the group consisting of: pembrolizumab, nivolumab, toripalimab, pidilizumab, cemiplimab, sintilimab, cetrelimab, spartalizumab, camrelizumab, tislelizumab, balstilimab, dostarlimab, ABBV-181, penpulimab, genolimzumab ), retifanlimab, sasanlimab, AMP-224, AB122, F-520, MEDI-3387, MEDI-5771, MEDI-0680, SG-001, BCD-100, BAT-1306, BI-754091, CBT-501, GLS-010, LZM-009, Sym-021, CS-1003, HLX-10, AK-103, AM-0001, ENUM-244C8, ENUM-388D4, JTX-4014, RXI-762, STI-A1110, HLX-20, SSI-361, APL-501, TJ0141H, and SNA-01. In some embodiments, the PD-L1 antibody is selected from the group consisting of atezolizumab, manelimab, avelumab, cosibelimab, durvalumab, envafolimab, socazolimab, BGB-A333, CK-301, CS-1001, FAZ-053, APL-502, MDX-1105, IMC-001, KD-005, Gensci-047, LY-3300054, SHR-1316, MSB-2311, AVA-004, CBT-502, JS-003, B12, and KY-1003.

[0028] In some embodiments, the level of CD73 is the protein level of CD73. In some embodiments, the protein level is determined by immunohistochemistry (IHC).

[0029] In some embodiments, the sample includes tumor cells, immune cells (such as tumor infiltrating immune cells (TIL)), stromal cells, and any combination thereof. In some embodiments, the sample is obtained prior to the combination therapy.

[0030] In some embodiments, the CD73 level includes the ratio (such as percentage) of CD73-positive cells in all cells in the sample. In some embodiments, the CD73 level includes the ratio of CD73-positive tumor cells in all cells in the sample (TC), or the ratio of CD73-positive immune cells in all cells in the sample (IC), whichever is higher (TC / IC). In some embodiments, 30% or more CD73 levels (TC / IC) identify the subject as likely to respond to or benefit from the combined therapy. In some embodiments, 35%, 40%, 45% or 50% or more CD73 levels (TC / IC) identify the subject as likely to respond to or benefit from the combined therapy.

[0031] In some embodiments, the CD73 level includes the ratio of CD73-positive tumor cells among all cells in the sample (TC(+)), or the ratio of CD73-positive immune cells among all cells in the sample (IC(+)), whichever is higher, wherein the positivity is at least moderately positive (TC / IC(+)).

[0032] In some embodiments, a CD73 level (TC / IC(+)) of 10% or more identifies the subject as likely to respond to or benefit from the combination therapy. In some embodiments, a CD73 level (TC / IC(+)) of 15%, 20%, 25%, 30%, 35% or 40% or more identifies the subject as likely to respond to or benefit from the combination therapy.

[0033] In some embodiments, the reference sample comprises a reference tissue or a reference cell. In some embodiments, the reference sample is derived from a healthy subject or a non-disease sample of a subject.

[0034] In some embodiments, responsiveness comprises a relative increase in one or more of: overall survival (OS), progression-free survival (PFS), complete response (CR), partial response (PR), and combinations thereof.

[0035] In another aspect, the present application provides a kit or article of manufacture for use in the methods of the present application, comprising 1) one or more reagents for determining the level of CD73 in a sample from a subject with NSCLC; and (2) a package insert, wherein the package insert recommends treating the subject with a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level.

[0036] In another aspect, the present application provides a kit or article of manufacture for use in the methods of the present application, comprising: (1) one or more reagents for determining the level of CD73 in a sample from a subject with NSCLC; and (2) a package insert, wherein the package insert suggests that an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample indicates that the subject is more likely to respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist.

[0037] In some embodiments of the kit or article of manufacture, the agent is an anti-CD73 antibody. In some embodiments, the antibody is selected from the group consisting of: D7F9A (CST, Cat. No. 13160), 606117 (R&D), EPR6114 (Abcam, Cat. No. ab133582), 4G6E3 (Abcam, Cat. No. ab202122), NT5E / 2503 (Abcam, Cat. No. ab257309), NT5E / 2505 (Abcam, Cat. No. ab257310), NT5E / 2545 (Abcam, Cat. No. ab257311), cam, Cat.No.ab257311), NT5E / 2646 (Abcam, Cat.No.ab257312), 7G2 (Thermo Fisher, Cat.No.1D7), JM11-40 (Thermo Fisher, Cat.No.JM11-40), CD73 recombinant rabbit monoclonal antibody (1) (Invitrogen, Cat.No.MA5-29454), BLR054F (Thermo Fisher, Cat.No.BLR054F), sc-32299 (Santa Cruz, Cat.No.IE9) and AD2 (Biorad, Cat.No.AD2).

[0038] It should be understood that one, some or all of the various embodiments described herein can be combined to form other embodiments of the present disclosure. For those skilled in the art, these and other aspects of the present disclosure will be apparent. The following detailed description of the invention will further set forth these and other embodiments of the present disclosure. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1 Exemplary tumor cells or immune cells with CD73 staining intensities of 0-3 are shown.

[0040] Figure 2 Example sections of NSCLC tumor samples after CD73 staining are shown.

[0041] Figure 3A and 3B Shown are the mean CD73 levels of subjects who did or did not receive first-line therapy under different IHC scoring systems.

[0042] Figure 4A and 4B The correlation of CD73 levels with subjects' responses to combination therapy with CD73 antibody and PD-1 antibody under different IHC scoring systems is shown.

[0043] Figure 5A and 5B The correlation between CD73 levels and subjects' responses to combined treatment with CD73 antibodies and PD-1 antibodies under different IHC scoring systems was shown, and the Pearson Correlation Coefficient was used for analysis.

[0044] Fig. 6A and 6B Preliminary ROC analysis of CD73 levels under different IHC scoring systems is shown. DETAILED DESCRIPTION definition

[0045] Before describing the embodiments of the present invention in detail, it should be understood that the present application disclosure is not limited to specific compositions or biological systems, and of course, these compositions or biological systems can change. It should also be understood that the terminology used in the application is only for the purpose of describing specific embodiments and is not intended to be restrictive.

[0046] As used in this specification and the appended claims, the singular forms "a", "an", and "the" include plural references unless the content clearly dictates otherwise. Thus, for example, reference to "a molecule" may optionally include a combination of two or more such molecules, and so forth.

[0047] The term "about" used in this application refers to the usual error range of each numerical value known to those skilled in the art. "About" a value or parameter mentioned in this application includes (and describes) embodiments for the value or parameter itself.

[0048] It should be understood that various aspects and embodiments of the present disclosure include "comprising / including" various aspects and embodiments, "consisting of various aspects and embodiments", and "consisting essentially of various aspects and embodiments.

[0049] As used herein, the term "antibody" is used in the broadest sense and specifically covers intact antibodies (such as full-length antibodies), antibody fragments (including but not limited to Fab, F(ab')2, scFv, scFv-Fc, single domain antibodies, single heavy chain antibodies and single light chain antibodies), monoclonal antibodies and polyclonal antibodies, so long as they exhibit the desired biological activity (such as epitope binding).

[0050] As used herein, the term "isolated" antibody may refer to an antibody that is substantially free of other cellular material. In one embodiment, an isolated antibody is substantially free of other proteins from the same species. In another embodiment, an isolated antibody is expressed by cells from a different species and is substantially free of other proteins from a different species. In some embodiments, an "isolated" antibody refers to an antibody that has been identified and separated and / or recovered from components in its natural environment. Contaminating components in its natural environment refer to substances that interfere with the diagnostic or therapeutic use of the antibody and may include enzymes, hormones, and other proteinaceous or non-proteinaceous solutes. The antibody may be isolated using protein purification techniques known in the art to be substantially free of naturally associated components (or components associated with the cellular expression system used to produce the antibody). In some embodiments, the antibody will be purified: (1) greater than 75% by weight of the antibody as determined by the Lowry method, and most preferably greater than 80%, 90%, 95%, or 99% by weight; or (2) to homogeneity by SDS-PAGE under reducing or non-reducing conditions using Coomassie blue or, preferably, silver stain. Isolated antibody includes the antibody in situ within recombinant cells since at least one component of the antibody's natural environment will not be present. Ordinarily, however, isolated antibody will be prepared by at least one purification step.

[0051] As used in this application, the terms "natural antibodies and immunoglobulins" are usually heterotetrameric glycoproteins of about 150,000 daltons, composed of two identical light chains (L) and two identical heavy chains (H). Each light chain is connected to the heavy chain by one covalent disulfide bond (also called a "VH / VL pair"), while the number of disulfide bonds between the heavy chains of different immunoglobulin isotypes varies. Each heavy and light chain also has regularly spaced intrachain disulfide bridges. Each heavy chain has a variable domain (VH) at one end, followed by a number of constant domains. Each light chain has a variable domain (VL) at one end and a constant domain at its other end; the constant domain of the light chain is aligned with the first constant domain of the heavy chain, and the variable domain of the light chain is aligned with the variable domain of the heavy chain. It is believed that specific amino acid residues form an interface between the light chain and heavy chain variable domains. See, for example, Chothia et al., J. Mol. Biol., 186:651 (1985); Novotny and Haber, Proc. Natl. Acad. Sci. USA, 82:4592 (1985).

[0052] As used herein, the term "variable" refers to the fact that some parts of the variable domains differ greatly in sequence between antibodies and are used for the binding and specificity of each specific antibody to its specific antigen. However, variability is not evenly distributed throughout the variable domains of antibodies. It is concentrated in three segments called complementary determining regions (CDRs) or hypervariable regions in both the light chain and heavy chain variable domains. The more highly conserved parts in the variable domains are called frameworks (FRs). The variable domains of native heavy and light chains each contain four FR regions, most of which adopt a β sheet structure, connected by three CDRs, which form a loop connecting the β sheet structure, and in some cases constitute a part of the β sheet structure. The CDRs on each chain are kept together in close proximity by the FR region, and together with the CDRs on the other chain, contribute to the formation of the antigen binding site of the antibody. For example, see Kabat et al., Sequences of Proteins of Immunological Interest, Fifth Edition, National Institute of Health, Bethesda, Maryland, (1991). The constant domains are not directly involved in the binding of an antibody to an antigen, but exhibit various effector functions, such as participation in antibody-dependent cellular cytotoxicity. Variable region sequences of interest include humanized variable region sequences of CD47 antibodies described in detail elsewhere in this application.

[0053] The term "hypervariable region (HVR)" or "complementarity determining region (CDR)" may refer to subdomains of the VH and VL domains that are characterized by enhanced sequence variability and / or form defined loops. These include the three CDRs (H1, H2, and H3) in the VH domain and the three CDRs (L1, L2, and L3) in the VL domain. H3 is believed to be key to conferring good binding specificity, while L3 and H3 show the highest levels of diversity. See Johnson and Wu, in Methods in Molecular Biology 248: 1-25 (Lo, ed., Human Press, Totowa, NJ, 2003).

[0054] Many descriptions of CDR / HVR are known. Kabat complementarity determining regions (CDRs) are based on sequence variability and are the most commonly used (Kabat et al., Sequences of Proteins of Immunological Interest, 5th Edition, Public Health Service, National Institutes of Health, Bethesda, Maryland (1991)). Chothia refers to the position of structural loops (Chothia and Lesk J. Mol. Biol. 196: 901-917 (1987)). AbM HVR represents a compromise between Kabat HVR and Chothia structural loops and is adopted by Oxford Molecular's AbM antibody modeling software. "Contact" HVR is based on analysis of existing complex crystal structures. The residues of each of these HVR / CDRs are recorded as follows: "framework" or "FR" residues refer to those variable domain residues other than HVR / CDR residues.

[0055] Also known are "extended" HVRs: 24-36 or 24-34 (L1), 46-56 or 50-56 (L2), and 89-97 or 89-96 (L3) in VL, 26-35 (H1), 50-65 or 49-65 (H2), and 93-102, 94-102 or 95-102 (H3) in VH (Kabat numbering).

[0056] "Numbering according to Kabat" may refer to the numbering system used for the heavy chain variable domain or light chain variable domain compiled by Kabat et al. above. The actual linear amino acid sequence may contain fewer or additional amino acids, corresponding to the shortening of the FR or HVR of the variable domain or the insertion of the FR or HVR of the variable domain. The Kabat numbering of the residues of a given antibody can be determined by comparing the homologous region of the antibody sequence with the "standard" Kabat numbering sequence. Typically, when referring to the residues in the variable domain (approximately residues 1-107 of the light chain and residues 1-113 of the heavy chain), Kabat numbering is used, and when referring to the residues in the heavy chain constant region, the EU numbering system or index (such as the EU index in Kabat, according to the numbering of EU IgG1) is generally used.

[0057] As used herein, the term "antibody fragment" and all its grammatical variants are defined as a portion of an intact antibody that contains the antigen binding site or variable region of the intact antibody and, in some cases, does not contain the constant heavy chain domains (i.e., CH2, CH3 and / or CH4, depending on the antibody isotype) of the Fc region of the intact antibody. Examples of antibody fragments include Fab, Fab', Fab'-SH, F(ab') 2 and Fv fragments; diabodies; any antibody fragment of a polypeptide having a primary structure consisting of an uninterrupted sequence of continuous amino acid residues (referred to herein as a "single-chain antibody fragment" or "single-chain polypeptide"), including but not limited to (1) single-chain Fv (scFv) molecules, (2) single-chain polypeptides containing only one light chain variable domain or fragments thereof containing the three CDRs of the light chain variable domain without the associated heavy chain portion, and (3) single-chain polypeptides containing only one heavy chain variable region or fragments thereof containing the three CDRs of the heavy chain variable region without the associated light chain portion; and multispecific or multivalent structures formed by antibody fragments. In an antibody fragment comprising one or more heavy chains, the heavy chain may comprise any constant domain sequence found in the non-Fc region of an intact antibody (such as CH1 in the IgG isotype), and / or may comprise any hinge region sequence found in an intact antibody, and / or may comprise a leucine zipper sequence fused to or located within a hinge region sequence or a constant domain sequence of a heavy chain.

[0058] The Fab fragment also contains the constant domain of the light chain and the first constant domain of the heavy chain (CH 1 ) The difference between the Fab' fragment and the Fab fragment is that the heavy chain CH 1 A few residues are added to the carboxyl terminus of the domain, including one or more cysteines from the antibody hinge region. Fab'-SH is herein referred to as Fab' in which the cysteine ​​residues of the constant domains bear a free thiol group. F(ab') 2Antibody fragments originally were produced as pairs of Fab' fragments with hinge cysteines between them. Other chemical couplings of antibody fragments are also known.

[0059] As used herein, the term "monoclonal antibody" (mAb) refers to an antibody obtained from a population of substantially homogeneous antibodies, i.e., the individual antibodies constituting the population are identical except for a small amount of possible naturally occurring mutations. Monoclonal antibodies are highly specific and are directed only to a single antigenic site. Each mAb is directed to a single determinant on the antigen. In addition to its specificity, the advantage of monoclonal antibodies is that they can be synthesized by hybridoma culture without being contaminated by other immunoglobulins. The modifier "monoclonal" indicates that the characteristic of the antibody is obtained from a substantially homogeneous antibody population, and should not be construed as requiring the production of antibodies by any particular method. For example, the monoclonal antibodies used according to the disclosure of the present application can be made in immortalized B cells or their hybridomas, or can also be made by recombinant DNA methods.

[0060] The monoclonal antibodies in the present application include hybrid antibodies and recombinant antibodies produced by splicing the variable (including hypervariable) domains of CD47 antibodies with constant domains (e.g., "humanized" antibodies), or splicing light chains with heavy chains, or splicing chains from one species with chains from another species, or fusion with heterologous proteins, regardless of their source species or immunoglobulin class or subclass designations, as well as antibody fragments (e.g., Fab, F(ab') 2 and Fv), as long as they exhibit the desired biological activity.

[0061] The monoclonal antibodies in the present application specifically include chimeric antibodies (immunoglobulins) in which a portion of the heavy chain and / or light chain is identical or homologous to the corresponding sequence in an antibody derived from a specific species or belonging to a specific antibody class or subclass, while the remainder of the chain is identical or homologous to the corresponding sequence in an antibody derived from another species or belonging to another antibody class or subclass, as well as fragments of such antibodies, as long as they exhibit the desired biological activity.

[0062] As used in this application, the term "treatment" refers to a clinical intervention intended to change the natural course of the treated individual or cell in the clinical pathological process. The ideal therapeutic effect includes reducing the rate of disease progression, improving or alleviating the disease state, and alleviating or improving prognosis. For example, if one or more symptoms related to cancer are alleviated or eliminated, including but not limited to reducing the proliferation of cancer cells (or eliminating cancer cells), reducing the symptoms caused by the disease, improving the quality of life of individuals with the disease, reducing the dosage of other drugs required for treating the disease, and / or extending the survival period of the individual, then the individual is successfully "treated". In some embodiments, "treating" a disease (such as cancer) refers to delaying the development of the disease, that is, postponing, hindering, slowing down, delaying, stabilizing and / or delaying the development of a disease (such as cancer). This delay can be of varying lengths, depending on the disease and / or the medical history of the individual being treated. As is apparent to those skilled in the art, sufficient or significant delays can actually cover prevention, which is that the individual will not have disease development, for example, advanced cancer (such as metastasis) may be postponed.

[0063] "Effective amount" is at least the minimum amount required to produce a measurable improvement or preventive effect on a specific disease (e.g., cancer). The effective amount of the present application may vary according to factors such as the patient's disease state, age, sex, and weight, as well as the ability of the therapeutic agent (or combination of therapeutic agents) to cause the expected response in an individual. An effective amount also refers to an amount in which any toxic or harmful effects of the treatment are exceeded by the beneficial effects of the treatment. For therapeutic use, beneficial or expected results include clinical results, such as alleviating one or more symptoms caused by the disease, improving the quality of life of patients with the disease, reducing the dose of other drugs required to treat the disease, enhancing the effect of another drug (e.g., through targeting), delaying the progression of the disease and / or prolonging survival. In the case of cancer or tumors, an effective amount of the drug may have the following effects: reducing the number of cancer cells; reducing tumor volume; inhibiting (i.e., slowing down or ideally preventing) the infiltration of cancer cells into peripheral organs; inhibiting (i.e., slowing down and ideally preventing) tumor metastasis; inhibiting tumor growth to a certain extent; and / or alleviating one or more symptoms associated with the disease to a certain extent. The effective amount can be administered in one or more administrations. For the purposes of the present disclosure, an effective amount of a drug, compound, or pharmaceutical composition refers to an amount sufficient to directly or indirectly complete a therapeutic treatment. As understood in clinical semantics, an effective amount of a drug, compound, or pharmaceutical composition may or may not be achieved in combination with another drug, compound, or pharmaceutical composition. Thus, an "effective amount" may be considered in the context of administering one or more therapeutic agents, and a single drug may be considered to be administered in an effective amount if a desired effect can or is achieved in combination with one or more other drugs.

[0064] As used herein, the term "subject" for purposes of treatment refers to any animal classified as a mammal, including humans, livestock and farm animals, and zoo, sport or pet animals, such as dogs.

[0065] As used in this application, the term "PD-1 binding antagonist" refers to a molecule that can reduce, block, inhibit, abolish or interfere with the signal transduction generated by the interaction of PD-1 with one or more of its binding partners (such as PD-L1, PD-L2). In some embodiments, the PD-1 binding antagonist is a molecule that inhibits the binding of PD-1 to its binding partner. In a specific aspect, the PD-1 binding antagonist inhibits the binding of PD-1 to PD-L1 and / or PD-L2. For example, PD-1 binding antagonists include PD-1 antibodies, antigen-binding fragments thereof, immunoadhesins, fusion proteins, oligopeptides and other molecules that can reduce, block, inhibit, abolish or interfere with the signal transduction generated by the interaction of PD-1 with PD-L1 and / or PD-L2. In some embodiments, the PD-1 binding antagonist reduces negative signals mediated by or through cell surface proteins expressed on T lymphocytes and other cells or mediated by PD-1 or PD-L1 signaling, thereby making dysfunctional T cells less dysfunctional.

[0066] As used in this application, the term "PD-L1 binding antagonist" refers to a molecule that can reduce, block, inhibit, abolish or interfere with the signal transduction generated by the interaction of PD-L1 with one or more of its binding partners (such as PD-1, B7-1). In some embodiments, the PD-L1 binding antagonist is a molecule that inhibits the binding of PD-L1 to its binding partner. In some embodiments, the PD-L1 binding antagonist inhibits the binding of PD-L1 to PD-1 and / or B7-1. In some embodiments, the PD-L1 binding antagonist includes PD-L1 antibodies, antigen-binding fragments thereof, immunoadhesins, fusion proteins, oligopeptides and other molecules that can reduce, block, inhibit, abolish or interfere with the signal transduction generated by the interaction of PD-L1 with one or more of its binding partners (such as PD-1, B7-1). In some embodiments, a PD-L1 binding antagonist can reduce negative signals mediated by or through cell surface proteins expressed on T lymphocytes and other cells or through PD-L1 or PD-1 signaling, thereby making dysfunctional T cells less dysfunctional.

[0067] As used herein, the term "biomarker" refers to an indicator that can be detected in a sample, such as a predictive, diagnostic and / or prognostic indicator. A biomarker can be used as an indicator of a specific subtype of a disease or condition (such as cancer) characterized by specific molecular, pathological, histological and / or clinical features. Biomarkers include, but are not limited to, polynucleotides, polynucleotide copy number changes, polypeptides, post-translational modifications, carbohydrates and / or glycolipid-based molecules.

[0068] The "amount" or "level" of a biomarker associated with an increased clinical benefit to an individual refers to the level that can be detected in a biological sample. These can be measured by methods known to those skilled in the art (such as IHC). The assessed biomarker level or amount can be used to determine the response to treatment.

[0069] As used herein, the terms "level of expression" or "expression level" are often used interchangeably and generally refer to the amount of a biomarker in a biological sample. "Expression" generally refers to the process by which information (such as genetic code and / or epigenetic information) is converted into structures that exist and function in cells.

[0070] "Increased expression", "increased expression level" or "increased level" refers to increased expression or increased level of a biomarker in an individual relative to a reference, such as one or more individuals not suffering from a disease or condition (such as cancer) or an internal control (such as an internal reference biomarker). CD73

[0071] In the past few years, more studies have shown that the adenosine pathway plays a role in tumor progression and metastasis. In the tumor environment, increased extracellular adenosine can activate adenosine receptor subtypes A2A and A2B, thereby limiting the function of effector T cells, inducing immunosuppression and stimulating angiogenesis.

[0072] The production of extracellular adenosine is regulated by multiple enzymes. The classical pathway for extracellular adenosine production is based on sequential reactions mediated by ectonucleases. In particular, extracellular adenosine triphosphate is first hydrolyzed into adenosine monophosphate (AMP) by CD39 and then dephosphorylated to adenosine by CD73. Alternatively, CD38 and CD203a can convert NAD+ to ADP-ribose and ADP-ribose to AMP, respectively. In turn, AMP is dephosphorylated to adenosine by CD73. In both pathways, CD73 plays a key role in the production of extracellular adenosine.

[0073] CD73, cluster of differentiation 73, also known as 5'-nucleotidase (5'-NT) or ecto-5'-nucleotidase, is an enzyme used to convert AMP to adenosine. CD73 catalyzes the formation of extracellular adenosine, which contributes to the immunosuppressive tumor environment. CD73 exists in two forms: one form is fixed to the membrane of many cells or on extracellular vesicles through glycosylphosphatidylinositol; the second form is cleaved from the membrane by proteases or phospholipases, and it is present in biological fluids. CD73 antagonists

[0074] Preclinical evidence shows that CD73 inhibition prevents adenosine-mediated lymphocyte suppression, enhances the activity of CD8+ effector cells, and reduces both MDSCs and Tregs. Several anti-CD73 antibodies are being developed as potential anticancer drugs, but none has been approved for clinical use.

[0075] Any suitable CD73 antagonist can be used in the present application. In some embodiments, the CD73 antagonist is a CD73 antibody. Any CD73 antibody known in the art can be used in the present application. In some embodiments, the CD73 antibody is selected from the group consisting of full-length antibodies, Fab, Fab', F(ab')2, scFv and sdAb. In some embodiments, the CD73 antibody is a full-length antibody.

[0076] In some embodiments, the CD73 antibody comprises HCDR1, HCDR2, and HCDR3, which respectively comprise the amino acid sequences of CDR1, CDR2, and CDR3 in the heavy chain variable region (VH) as shown in SEQ ID NO: 1. In some embodiments, the CD73 antibody comprises HCDR1, HCDR2, and HCDR3, which respectively comprise the amino acid sequences of CDR1, CDR2, and CDR3 in the heavy chain variable region (VH) as shown in SEQ ID NO: 1, wherein the CDR1, CDR2, and CDR3 are according to the Kabat numbering system. In some embodiments, the CD73 antibody comprises (1) a HCDR1 comprising an amino acid sequence as shown in SEQ ID NO:3 or an amino acid sequence having one or more substitutions compared to SEQ ID NO:3, (2) a HCDR2 comprising an amino acid sequence as shown in SEQ ID NO:4 or an amino acid sequence having one or more substitutions compared to SEQ ID NO:4, and (3) a HCDR3 comprising an amino acid sequence as shown in SEQ ID NO:5 or an amino acid sequence having one or more substitutions compared to SEQ ID NO:5.

[0077] In some embodiments, the CD73 antibody comprises a heavy chain variable region (VH) comprising an amino acid sequence as shown in SEQ ID NO:1, or an amino acid sequence that is at least 80%, 85%, 87%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO:1.

[0078] In some embodiments, the CD73 antibody comprises LCDR1, LCR2 and LCDR3, which respectively comprise the amino acid sequences of CDR1, CDR2 and CDR3 in the light chain variable region (VL) shown in SEQ ID NO: 2. In some embodiments, the CD73 antibody comprises LCDR1, LCR2 and LCDR3, which respectively comprise the amino acid sequences of CDR1, CDR2 and CDR3 in the light chain variable region (VL) shown in SEQ ID NO: 2, wherein the CDR1, CDR2 and CDR3 are according to the Kabat numbering system. In some embodiments, the CD73 antibody comprises: (1) LCDR1 comprising the amino acid sequence as shown in SEQ ID NO:6 or an amino acid sequence having one or more substitutions compared to SEQ ID NO:6, (2) LCDR2 comprising the amino acid sequence as shown in SEQ ID NO:7 or an amino acid sequence having one or more substitutions compared to SEQ ID NO:7, and (3) LCDR3 comprising the amino acid sequence as shown in SEQ ID NO:8 or an amino acid sequence having one or more substitutions compared to SEQ ID NO:8.

[0079] In some embodiments, the CD73 antibody comprises a light chain variable region (VL) comprising an amino acid sequence as shown in SEQ ID NO:2, or an amino acid sequence that is at least 80%, 85%, 87%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO:2.

[0080] In some embodiments, the CD73 antibody comprises: (1) HCDR1, HCDR2 and HCDR3, which respectively comprise the amino acid sequences of CDR1, CDR2 and CDR3 in the heavy chain variable region (VH) as shown in SEQ ID NO: 1, and (2) LCDR1, LCR2 and LCDR3, which respectively comprise the amino acid sequences of CDR1, CDR2 and CDR3 in the light chain variable region (VL) as shown in SEQ ID NO: 2. In some embodiments, the CD73 antibody comprises: (1) HCDR1, HCDR2 and HCDR3, which respectively comprise the amino acid sequences of CDR1, CDR2 and CDR3 in the heavy chain variable region (VH) as shown in SEQ ID NO: 1, and (2) LCDR1, LCR2 and LCDR3, which respectively comprise the amino acid sequences of CDR1, CDR2 and CDR3 in the light chain variable region (VL) as shown in SEQ ID NO: 2, wherein CDR1, CDR2 and CDR3 are according to the Kabat numbering system. In some embodiments, the CD73 antibody comprises: (1) HCDR1 comprising the amino acid sequence as shown in SEQ ID NO:3, or an amino acid sequence having one or more substitutions compared to SEQ ID NO:3, (2) HCDR2 comprising the amino acid sequence as shown in SEQ ID NO:4, or an amino acid sequence having one or more substitutions compared to SEQ ID NO:4, (3) HCDR3 comprising the amino acid sequence as shown in SEQ ID NO:5, or an amino acid sequence having one or more substitutions compared to SEQ ID NO:5, (4) LCDR1 comprising the amino acid sequence as shown in SEQ ID NO:6, or an amino acid sequence having one or more substitutions compared to SEQ ID NO:6, (5) LCDR2 comprising the amino acid sequence as shown in SEQ ID NO:7, or an amino acid sequence having one or more substitutions compared to SEQ ID NO:7, and (6) LCDR3 comprising the amino acid sequence as shown in SEQ ID NO:8, or an amino acid sequence having one or more substitutions compared to SEQ ID NO:8.

[0081] In some embodiments, the CD73 antibody comprises: (1) a heavy chain variable region (VH) comprising an amino acid sequence as set forth in SEQ ID NO: 1, or an amino acid sequence at least 80%, 85%, 87%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO: 1, and (2) a light chain variable region (VL) comprising an amino acid sequence as set forth in SEQ ID NO:2, or an amino acid sequence at least 80%, 85%, 87%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% identical to SEQ ID NO:2.

[0082] In some embodiments, the CD73 antibody is selected from the group consisting of uliledlimab (I-MabBiopharma), oleclumab (AstraZeneca), CPI-006 (Corvus Pharma), BMS-986179 (Bristol-Myers Squibb), AB-680 (Arcus Biosciences), NZV-930 (SRF373, Surface Oncolohgy / Novartis), JAB-BX102 (Jacobio), AK119 (Akesobio), Sym024 (Symphogen), IBI325 (Innovent), BR101 (HisunBioRay), and LY3475070 (Eli Lilly). In some embodiments, the CD73 antibody is uliledlimab (I-MabBiopharma) or oleclumab (AstraZeneca). PD-1 / PD-L1 antagonists

[0083] The methods of the present application may use any suitable PD-1 / PD-L1 antagonist known in the art. In some embodiments, the PD-1 / PD-L1 antagonist includes a small molecule inhibitor of PD-1 / PD-L1 signaling. In some embodiments, the PD-1 / PD-L1 antagonist includes an antibody or an antigen-binding fragment thereof, such as a PD-1 antibody, a PD-L1 antibody, or an antigen-binding fragment thereof.

[0084] In some embodiments, the PD-1 / PD-L1 signal inhibitor is a PD-1 antibody or an antigen-binding fragment thereof. Any suitable PD-1 antibody can be used in the present application. Exemplary PD-1 antibodies that can be used in the methods of the present application include, but are not limited to, polyclonal antibodies, monoclonal antibodies, Fab, scFv, diantibodies, triantibodies, miniantibodies, VHHs, and sdAbs. Exemplary PD-1 antibodies include, but are not limited to, pidilizumab, cemiplimab, sintilimab, cetrelimab, spartalizumab, camrelizumab, tislelizumab, balstilimab, toripalimab, dostarlimab, ABBV-181, penpulimab, Pembrolizumab, genolimzumab, retifanlimab ), sasanlimab, AMP-224, AB122, F-520, MEDI-3387, MEDI-5771, MEDI-0680, SG-001, nivolumab, BCD-100, BAT-1306, BI-754091, CBT-501, GLS-010, LZM-009, Sym-021, CS-1003, HLX-10, AK-103, AM-0001, ENUM-244C8, ENUM-388D4, JTX-4014, RXI-762, STI-A1110, HLX-20, SSI-361, APL-501, TJ0141H, and SNA-01. In some embodiments, the PD-1 antibody is toripalimab.

[0085] Any suitable PD-L1 antibody can be used in the present application. Exemplary PD-L1 antibodies that can be used in the present application method include, but are not limited to, polyclonal antibodies, monoclonal antibodies, Fab, scFv, diantibodies, triantibodies, miniantibodies, VHHs, and sdAbs. Exemplary PD-L1 antibodies include, but are not limited to, Manelimab, atezolizumab, avelumab, Cosibelimab, durvalumab, Envafolimab, Socazolimab, BGB-A333, CK-301, CS-1001, FAZ-053, APL-502, MDX-1105, IMC-001, KD-005, Gensci-047, LY-3300054, SHR-1316, MSB-2311, AVA-004, CBT-502, JS-003, B12, and KY-1003. Combination therapy

[0086] As used herein, the terms "combination treatment," "combo therapy," "combo treatment," or "combinational therapy" are used interchangeably and refer to the simultaneous or separate administration of a CD73 antagonist and a PD-1 / PD-L1 antagonist described herein to a subject.

[0087] In some embodiments, the combination therapy comprises a CD73 antagonist and a PD-1 / PD-L1 antagonist as described herein. In some embodiments of the combination therapy, the CD73 antagonist and the PD-1 / PD-L1 antagonist are administered intravenously to the subject.

[0088] In some embodiments of the combined treatment, the CD73 antagonist is a CD73 antibody described in the present application. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 2 to about 30 mg / kg body weight. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 2 to about 20 mg / kg body weight. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 5 to about 20 mg / kg body weight. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 10 to about 20 mg / kg body weight. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 15 to about 20 mg / kg body weight. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 2 to about 15 mg / kg body weight. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 5 to about 15 mg / kg body weight. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 10 to about 15 mg / kg body weight. In some embodiments, the CD73 antibody is administered to the subject at a dose of about 2 to about 10 mg / kg body weight. In some embodiments, the CD73 antibody is administered to a subject at a dose of about 5 to about 10 mg / kg body weight. In some embodiments, the CD73 antibody is administered to a subject at a dose of about 2 to about 5 mg / kg body weight. In some embodiments, the CD73 antibody is administered to a subject at a dose of about 20 to about 30 mg / kg body weight. In some embodiments, the CD73 antibody is administered to a subject at a dose of about 2, about 5, about 10, about 15, or about 20 mg / kg body weight. In some embodiments, the CD73 antibody is administered to a subject at a dose of about 20 mg / kg body weight. In some embodiments, the CD73 antibody is administered to a subject at a dose of about 30 mg / kg body weight.

[0089] In some embodiments of the combination therapy, the CD73 antibody is administered to the subject once a week (QW), once every two weeks (Q2W), once every three weeks (Q3W), once every four weeks (Q4W), or once a month. In some embodiments, the CD73 antibody is administered to the subject once every three weeks (Q3W). In some embodiments, the CD73 antibody is administered to the subject at a dose of 20 mg / kg, once every three weeks (Q3W). In some embodiments, the CD73 antibody is administered to the subject at a dose of 30 mg / kg, once every three weeks (Q3W).

[0090] In some embodiments of the combination therapy, CD73 antibodies include but are not limited to uliledlimab (I-MabBiopharma), oleclumab (AstraZeneca), CPI-006 (Corvus Pharma), BMS-986179 (Bristol-Myers Squibb), AB-680 (Arcus Biosciences), NZV-930 (SRF373, Surface Oncolohgy / Novartis), JAB-BX102 (Jacobio), AK119 (Akesobio), Sym024 (Symphogen), IBI 325 (Innovent), BR 101 (Hisun BioRay) and LY3475070 (Eli Lilly). In some embodiments, the CD73 antibody is uliledlimab (I-MabBiopharma) or oleclumab (AstraZeneca).

[0091] In some embodiments of combined therapy, the PD-1 / PD-L1 antagonist is a PD-1 antibody or a PD-L1 antibody described in the present application. In some embodiments of combined therapy, the PD-L1 antibody includes but is not limited to manelimab, atezolizumab, avelumab, Cosibelimab, durvalumab, Envafolimab, Socazolimab, BGB-A333, CK-301, CS-1001, FAZ-053, APL-502, MDX-1105, IMC-001, KD-005, Gensci-047, LY-3300054, SHR-1316, MSB-2311, AVA-004, CBT-502, JS-003, B12 and KY-1003. In some embodiments, the PD-L1 antibody is atezolizumab.

[0092] In some embodiments of the combination therapy, PD-1 antibodies include but are not limited to pidilizumab, cemiplimab, sintilimab, cetrelimab, spartalizumab, camrelizumab, tislelizumab, balstilimab, toripalimab, dostarlimab, ABBV-181, penpulimab, pembrolizumab, genolimzumab, retifan limab), sasanlimab, AMP-224, AB122, F-520, MEDI-3387, MEDI-5771, MEDI-0680, SG-001, nivolumab, BCD-100, BAT-1306, BI-754091, CBT-501, GLS-010, LZM-009, Sym-021, CS-1003, HLX-10, AK-103, AM-0001, ENUM-244C8, ENUM-388D4, JTX-4014, RXI-762, STI-A1110, HLX-20, SSI-361, APL-501, TJ0141H, and SNA-01. In some embodiments, the PD-1 antibody is pembrolizumab, nivolumab, or toripalimab.

[0093] In some embodiments, the PD-1 antibody or PD-L1 antibody is administered to the subject once a week (QW), once every two weeks (Q2W), once every three weeks (Q3W), once every four weeks (Q4W), or once a month. In some embodiments, the PD-1 antibody or PD-L1 antibody is administered to the subject once every three weeks (Q3W).

[0094] In some embodiments, the combination therapy comprises administering uliledlimab at a dose of 20 mg / kg intravenously to the subject once every three weeks (Q3W) and a fixed dose of 240 mg of toripalimab once every three weeks (Q3W). In some embodiments, the combination therapy comprises administering oleclumab at a dose of 20 mg / kg intravenously to the subject once every three weeks (Q3W) and a fixed dose of 240 mg of toripalimab once every three weeks (Q3W).

[0095] In some embodiments, the combination therapy comprises administering uliledlimab at a dose of 30 mg / kg intravenously to the subject once every three weeks (Q3W) and a fixed dose of 240 mg of toripalimab once every three weeks (Q3W). In some embodiments, the combination therapy comprises administering oleclumab at a dose of 30 mg / kg intravenously to the subject once every three weeks (Q3W) and a fixed dose of 240 mg of toripalimab once every three weeks (Q3W).

[0096] In some embodiments, the combination therapy comprises administering uliledlimab at a dose of 20 mg / kg intravenously to the subject once every three weeks (Q3W) and a fixed dose of 1200 mg of atezolizumab once every three weeks (Q3W). In some embodiments, the combination therapy comprises administering oleclumab at a dose of 20 mg / kg intravenously to the subject once every three weeks (Q3W) and a fixed dose of 1200 mg of atezolizumab once every three weeks (Q3W).

[0097] In some embodiments, the combination therapy comprises administering uliledlimab at a dose of 30 mg / kg intravenously to the subject once every three weeks (Q3W) and a fixed dose of 1200 mg of atezolizumab once every three weeks (Q3W). In some embodiments, the combination therapy comprises administering oleclumab at a dose of 30 mg / kg intravenously to the subject once every three weeks (Q3W) and a fixed dose of 1200 mg of atezolizumab once every three weeks (Q3W).

[0098] In some embodiments, the combination therapy also includes chemotherapy. Exemplary chemotherapy that can be used in the combination therapy of the present application includes, but is not limited to, 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 methylpropylamines, including altretamine, triethylenemelamine, trietylenephosphoramide, triethiylenethiophosphoramide and trimethylolomelamine; acetogenins (especially bullatacin and bullatacinone); camptothecins (including the synthetic analog topotecan); bryostatin; callystatin; CC-1065 (including its synthetic analogs adozelesin, carzelesin, and bizelesin); cryptophycins (particularly cryptophycin 1 and cryptophycin 8); dolastatin; duocarmycin (including synthetic analogs, KW-2189 and CB1-TM1); eleutherobin; pancratistatin; sarcodictyin; spongistatin; nitrogen mustards mustards, such as chlorambucil, chlornaphazine, cholophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, phenesterine, prednimustine, trofosfamide, uracil mustard, nitrosureas (e.g., carmustine), chlorozotocin, fotemustine, lomustine, nimustine and ranimnustine; antibiotics, such as enediyne antibiotics antibiotics; dynemicins, including dynemicin A; bisphosphonates, such as clodronate; esperamicin;and neocarzinostatin chromophores and related chromoprotein enediyne antiobiotic chromophores), aclacinomysins, actinomycin, authramycin, azaserine, bleomycins, cactinomycin, carabicin, carminomycin, carzinophilin, chromomycinis, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, Doxorubicin (including morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin), epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins (such as mitomycin C), mycophenolic acid (such as acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; antimetabolites, such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs, such as denopterin, rin), methotrexate, pteropterin, trimetrexate; purine analogs, for example, fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs, for example, ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine;androgens (e.g., calusterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone); anti-adrenals (e.g., aminoglutethimide, mitotane, trilostane); folic acid replenishers (e.g., frolinic acid); aceglatone; aldophosphamide glycoside; aminolevulinic acid acid; eniluracil; amsacrine; bestrabucil; bisantrene; edatraxate; defofamine; demecolcine; diaziquone; elfornithine; elliptinium acetate; epothilone; etoglucid; gallium nitrate nitrate; hydroxyurea; lentinan; lonidainine; maytansinoids, such as maytansine and ansamitocins; mitoguazone; mitoxantrone; mopidanmol; nitraerine; pentostatin; phenamet; pirarubicin; losoxantrone; podophyllinic acid; 2-ethylhydrazide; procarbazine; Polysaccharide complex ( polysaccharide complex (JHS Natural Products, Eugene, Oreg.); razoxane; rhizoxin; sizofiran; spirogermanium; tenuazonic acid; triaziquone; 2,2',2"-trichlorotriethylamine; trichothecenes (especially T-2 toxin, verracurin A, roridin A) A) and anguidine); urethan; vindesine; dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxoids, such as Paclitaxel (Bristol-Myers Squibb Oncology, Princeton, NJ), A hydrogenated castor oil-free, albumin-engineered paclitaxel nanoparticle formulation (American Pharmaceutical Partners, Schaumburg, Ill.), and Docetaxel ( doxetaxel) (Rhone-Poulenc Rorer, Antony, France); chloranbucil; Gemcitabine ( gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogs, such as cisplatin, oxaliplatin, and carboplatin; vinblastine, platinum, etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; Vinorelbine vinorelbine; novantrone; teniposide; edatrexate; daunomycin; aminopterin; xeloda; ibandronate; irinotecan (Camptosar, CPT-11) (including irinotecan with 5-FU and folinic acid); topoisomerase inhibitor RFS2000 topoisomerase inhibitor RFS2000; difluoromethlhylornithine (DMFO); retinoids, such as retinoic acid; capecitabine; combretastatin; leucovorin (LV); oxaliplatin, including the oxaliplatin regimen (FOLFOX); PKC-α inhibitors, Raf inhibitors, H-Ras inhibitors, EGFR inhibitors (such as erlotinib) that reduce cell proliferation ) and VEGF-A inhibitors, and pharmaceutically acceptable salts, acids or derivatives of any of the foregoing.

[0099] Other non-limiting exemplary chemotherapies include anti-hormonal agents that modulate or inhibit the effects of hormones on cancer, such as antiestrogens and selective estrogen receptor modulators (SERMs), including, for example, tamoxifen (including tamoxifen), raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, and Toremifene aromatase inhibitors that inhibit aromatase, which regulates estrogen production in the adrenal glands, such as 4(5)-imidazoles, aminoglutethimide, Megestrol acetate ( megestrol acetate), Exemestane exemestane), formestanie, fadrozole, Vorozole ( vorozole), Letrozole ( letrozole) and Anastrozole ( anastrozole; and antiandrogens, such as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin; and troxacitabine (a 1,3-dioxolane nucleoside cytosine analog); antisense oligonucleotides, particularly those that inhibit the expression of genes in signaling pathways involved in the proliferation of adherent cells (e.g., PKC-α, Ralf, and H-Ras); ribozymes, such as inhibitors of VEGF expression (e.g., ribozymes) and HER2 expression inhibitors; vaccines, such as gene therapy vaccines, e.g. vaccine, Vaccines and vaccine; rIL-2; Topoisomerase 1 inhibitors; rmRH; and pharmaceutically acceptable salts, acids or derivatives of any of the foregoing. method

[0100] In one aspect, the present application provides a method for treating a subject with NSCLC, the method comprising: (a) determining the level of CD73 in a sample from the subject, (b) assessing whether the subject is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level, and (c) administering the combination therapy to the subject if the subject is more likely to respond to or benefit from the combination therapy.

[0101] On the other hand, the present application provides a method for selecting a therapy for a subject with NSCLC, the method comprising: (a) determining the level of CD73 in a sample from the subject, and (b) assessing whether the subject is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level, and (c) if the subject is more likely to respond to or benefit from the combination therapy, selecting the combination therapy for the subject.

[0102] On the other hand, the present application provides a method for identifying a subject with NSCLC who is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample of the subject, (b) assessing whether the subject is more likely to respond to or benefit from the combination therapy based on the CD73 level, and (c) if the subject is more likely to respond to or benefit from the combination therapy, providing a recommendation of the combination therapy to the subject.

[0103] On the other hand, the present application provides a method for predicting the responsiveness of a subject with NSCLC to a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample of the subject, (b) assessing whether the subject is more likely to respond to or benefit from the combination therapy based on the CD73 level, and (c) if the subject is more likely to respond to or benefit from the combination therapy, providing a recommendation of the combination therapy to the subject.

[0104] In some embodiments of the methods of the present application, the NSCLC is stage III or stage IV NSCLC. In some embodiments, the NSCLC is stage IV NSCLC. In some embodiments, the NSCLC is advanced NSCLC. In some embodiments, the NSCLC is a metastatic NSCLC. In some embodiments, the NSCLC is a recurrent NSCLC. In some embodiments, the NSCLC is a refractory NSCLC. In some embodiments, the NSCLC has squamous histology or non-squamous histology.

[0105] In some embodiments of the methods of the present application, CD73 levels refer to CD73 protein levels. In the methods of the present application, any suitable method known in the art can be used to determine or measure protein levels. In some embodiments, the method determines the level of CD73 protein expressed on the cell membrane. In some embodiments, the method determines the level of CD73 protein expressed in the cytoplasm. In some embodiments, the method determines the level of CD73 protein expressed on the cell membrane and in the cytoplasm.

[0106] In some embodiments, protein levels are determined by immunohistochemistry (IHC), which measures the amount of CD73 protein in a cancer tissue sample (e.g., a NSCLC tissue sample). In some embodiments, the sample is a formalin-fixed and paraffin-embedded (FFPE) sample. In some embodiments, the sample is an archived sample. In some embodiments, the sample is a fresh sample. In some embodiments, the sample is a frozen sample. In some embodiments, the sample is obtained prior to the combined treatment described herein.

[0107] In some embodiments of the method of the present application, in IHC detection, according to the staining intensity, the content of CD73 protein in each cell (including cell membrane and cytosol) can be set to an intensity of 0, 1, 2 or 3 based on the standards shown in Table 1. Table 1. CD73 staining criteria Staining intensity standard 0 No staining 1 Faint or weak staining 2 Medium or moderate staining 3 Intense or strong staining

[0108] Intensity grading can be done manually by an experienced pathologist according to routinely determined criteria. Alternatively, a variety of automated / standardized intensity grading systems have been developed. For example, a quantitative microimmunohistochemistry detection method was published in Kashyap et al., "Quantitative microimmunohistochemistry for the grading of immunostains on tumour tissues," Nature Biomedical Engineering, 3, 478–490 (2019). This quantitative detection method is able to obtain dynamic information on the relative antigen density of quantitative tissue surfaces by measuring the evolution of immunohistochemical signals during tissue staining.

[0109] In some embodiments, the IHC assay determines the percentage of cells with staining intensities of 1, 2, and 3 relative to the total cells. In some embodiments, the IHC assay determines the percentage of cells with intensities of 2 and 3 relative to the total cells. In some embodiments, the IHC assay determines the percentage of cells with an intensity of 3 relative to the total cells.

[0110] In some embodiments, the cells include, but are not limited to, tumor cells, immune cells (e.g., tumor infiltrating immune cells (TILs)), stromal cells, vascular cells, and any combination thereof.

[0111] In some embodiments, the IHC assay determines the stained tumor relative to the total tumor cells. In some embodiments, the IHC assay determines the percentage of tumor cells with staining intensities of 1, 2, and 3 relative to the total tumor cells (TC). In some embodiments, the IHC assay determines the percentage of tumor cells with staining intensities of 2 and 3 relative to the total tumor cells (TC(+)). In some embodiments, the IHC assay determines the percentage of tumor cells with staining intensity of 3 relative to the total tumor cells (TC(++)).

[0112] The H score captures both the intensity and ratio of the biomarker from the IHC image and includes values ​​between 0 and 300, thereby providing a dynamic range for quantifying the abundance of the biomarker. In some embodiments of the methods of the present application, the IHC detection determines the H score (TCH) of the tumor cell. In some embodiments, the H score of the tumor cell refers to the sum of TC, TC(+) and TC(++).

[0113] In some embodiments, the IHC assay determines the stained immune cells relative to the total immune cells. In some embodiments, the IHC assay determines the percentage (IC) of immune cells with staining intensities of 1, 2, and 3 relative to the total immune cells. In some embodiments, the IHC assay determines the percentage (IC) of immune cells with staining intensities of 2 and 3 relative to the total immune cells. In some embodiments, the IHC assay determines the percentage (IC(+)) of immune cells with staining intensities of 3 relative to the total immune cells.

[0114] In some embodiments of the methods of the present application, IHC detection determines the H score (ICH) of the immune cell. In some embodiments, the H score of the immune cell refers to the sum of IC, IC(+) and IC(++).

[0115] In some embodiments, IHC detection determines the tumor cells and immune cells stained relative to total tumor cells and immune cells. In some embodiments, IHC detection determines the percentage of tumor cells with staining intensities of 1, 2, and 3 relative to total tumor cells and the sum of the percentages of immune cells with staining intensities of 1, 2, and 3 relative to total immune cells (TC+IC). In some embodiments, IHC detection determines the percentage of tumor cells with staining intensities of 2 and 3 relative to total tumor cells and the sum of the percentages of immune cells with staining intensities of 2 and 3 relative to total immune cells (TC+IC(+)). In some embodiments, IHC detection determines the percentage of tumor cells with staining intensities of 3 relative to total tumor cells and the sum of the percentages of immune cells with staining intensities of 3 relative to total immune cells (TC+IC(++)).

[0116] In some embodiments, IHC detection determines the tumor cells or immune cells stained relative to total tumor cells or immune cells. In some embodiments, IHC detection determines the percentage of tumor cells with staining intensities of 1, 2, and 3 relative to total tumor cells and the percentage of immune cells with staining intensities of 1, 2, and 3 relative to total immune cells (TC / IC). In some embodiments, IHC detection determines the percentage of tumor cells with staining intensities of 2 and 3 relative to total tumor cells and the percentage of immune cells with staining intensities of 2 and 3 relative to total immune cells (TC / IC(+)). In some embodiments, IHC detection determines the percentage of tumor cells with staining intensities of 3 relative to total tumor cells and the percentage of immune cells with staining intensities of 3 relative to total immune cells (TC / IC(++)).

[0117] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% TC in her / his sample as measured by an IHC assay described herein.

[0118] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% TC(+) in her / his sample as measured by an IHC assay described herein.

[0119] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% TC(++) in her / his sample as measured by an IHC assay described herein.

[0120] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has a Tumor Cell H-Score (TCH) of at least 1, at least 5, at least 10, at least 20, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 110, at least 120, at least 130, at least 140, at least 150, at least 160, at least 170, at least 180, at least 190, at least 200, at least 210, at least 220, at least 230, at least 240, at least 250, at least 260, at least 270, at least 280, at least 290, or 300.

[0121] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has an IC of at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% in her / his sample as measured by an IHC assay described herein.

[0122] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has an IC(+) in her / his sample of at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% as measured by an IHC assay described herein.

[0123] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has an IC(++) of at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% in her / his sample as measured by an IHC assay described herein.

[0124] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has an immune cell H score (ICH) of at least 1, at least 5, at least 10, at least 20, at least 30, at least 40, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 110, at least 120, at least 130, at least 140, at least 150, at least 160, at least 170, at least 180, at least 190, at least 200, at least 210, at least 220, at least 230, at least 240, at least 250, at least 260, at least 270, at least 280, at least 290, or 300.

[0125] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 100%, at least 101%, at least 102%, at least 103%, at least 104%, at least 105%, at least 106%, at least 107%, at least 108%, at least 109%, at least 110%, at least 111%, at least 112%, at least 113%, at least 114%, at least 115%, %, at least 180%, at least 185%, at least 190%, at least 195%, or 200% of a TC+IC as measured by an IHC assay described herein.

[0126] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 100%, at least 101%, at least 102%, at least 103%, at least 104%, at least 105%, at least 106%, at least 107%, at least 108%, at least 109%, at least 110%, at least 111%, at least 112%, %, at least 180%, at least 185%, at least 190%, at least 195%, or 200% TC+IC(+) as measured by an IHC assay described herein.

[0127] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 100%, at least 101%, at least 102%, at least 103%, at least 104%, at least 105%, at least 106%, at least 107%, at least 108%, at least 109%, at least 110%, at least 111%, at least 112%, %, at least 180%, at least 185%, at least 190%, at least 195%, or 200% TC + IC(++) as measured by an IHC assay described herein.

[0128] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has a TC / IC of at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% in her / his sample as measured by an IHC assay described herein.

[0129] like Figure 4A As shown in , non-responders have a median TC / IC of about 20%, while responders have a median TC / IC of about 50%. Therefore, a TC / IC score of 25% or higher indicates that the corresponding subject is likely to respond to or benefit from the combination therapy. Therefore, in some embodiments, a TC / IC score of 30%, 35%, 40%, 45%, 50% or 55% or higher indicates that the corresponding subject is likely to respond to or benefit from the combination therapy.

[0130] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has a TC / IC(+) in her / his sample of at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% as measured by an IHC assay described herein.

[0131] like Figure 4B As shown in , non-responders have a median TC / IC(+) of about 10%, while responders have a median TC / IC(+) of about 40%. Therefore, a TC / IC(+) score of 15% or more indicates that the corresponding subject is likely to respond to or benefit from the combination therapy. Therefore, in some embodiments, a TC / IC(+) score of 20%, 25%, 30%, 35%, 40% or 45% or more indicates that the corresponding subject is likely to respond to or benefit from the combination therapy.

[0132] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has a TC / IC(++) in her / his sample of at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% as measured by an IHC assay described herein.

[0133] In another aspect, the present application provides a method for identifying a subject with NSCLC who is more likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample from the subject, wherein an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample indicates that the subject is more likely to respond to or benefit from the combination therapy, and (b) providing a suggestion that the subject will be more likely to respond to or benefit from the combination therapy.

[0134] In another aspect, a method for predicting responsiveness of a subject having NSCLC to a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist is provided, the method comprising: (a) determining the level of CD73 in a sample from the subject, wherein an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample indicates that the subject is more likely to respond to or benefit from the combination therapy, and (b) providing a suggestion that the subject will have an increased likelihood of responding to or benefiting from the combination therapy.

[0135] In some embodiments, the reference sample comprises a reference tissue or a reference cell. In some embodiments, the reference sample is derived from a healthy subject, a subject who has undergone treatment for the first time, or a non-disease sample of a subject.

[0136] In another aspect, the present application provides a method for treating a subject having non-small cell lung cancer (NSCLC), the method comprising: (a) identifying a NSCLC subject who may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level in a sample from the subject, and (b) administering a CD73 antagonist and a PD1 / PD-L1 antagonist to the identified subject.

[0137] In another aspect, the present application provides a method for treating a subject with NSCLC, the method comprising administering a CD73 antagonist and a PD1 / PD-L1 antagonist to a subject with NSCLC, wherein the level of CD73 in a sample of the subject indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist.

[0138] In another aspect, the present application provides the use of a CD73 antagonist and a PD1 / PD-L1 antagonist for the manufacture of a medicament for treating a subject with NSCLC, wherein the level of CD73 in a sample of the subject indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist.

[0139] In another aspect, the present application provides the use of a CD73 antagonist for the manufacture of a medicament for treating a subject having NSCLC, wherein the level of CD73 in a sample of the subject indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist, wherein the subject is also administered a PD1 / PD-L1 antagonist.

[0140] In another aspect, the present application provides the use of a PD1 / PD-L1 antagonist for the manufacture of a medicament for treating a subject having NSCLC, wherein the level of CD73 in a sample of the subject indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist, wherein the subject is also administered a CD73 antagonist.

[0141] In some embodiments, the increase is determined by TC. In some embodiments, the increase is determined by TC(+). In some embodiments, the increase is determined by the H-score of tumor cells. In some embodiments, the increase is determined by TC(++). In some embodiments, the increase is determined by IC. In some embodiments, the increase is determined by IC(+). In some embodiments, the increase is determined by IC(++). In some embodiments, the increase is determined by the H-score of immune cells. In some embodiments, the increase is determined by TC / IC. In some embodiments, the increase is determined by TC / IC(+). In some embodiments, the increase is determined by TC / IC(++). In some embodiments, the increase is determined by TC+IC. In some embodiments, the increase is determined by TC+IC(+). In some embodiments, the increase is determined by TC+IC(++).

[0142] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% increase in CD73 compared to a reference sample.

[0143] In some embodiments of the methods of the present application, subjects who are more likely to respond to or benefit from the combination therapy as described herein have at least a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% increase in TC compared to a reference sample. In some embodiments of the methods of the present application, subjects who are more likely to respond to or benefit from the combination therapy as described herein have at least a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% increase in TC (+) compared to a reference sample. In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% increase in TC(++) compared to a reference sample.

[0144] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% increase in tumor cell H score compared to a reference sample.

[0145] In some embodiments of the present method, subjects who are more likely to respond to or benefit from the combined treatment as described herein have an IC increase of at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% compared to a reference sample. In some embodiments of the present method, subjects who are more likely to respond to or benefit from the combined treatment as described herein have an IC(+) increase of at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% compared to a reference sample. In some embodiments of the methods of the present application, subjects who are more likely to respond to or benefit from the combination therapy as described herein have an increase in IC(++) of at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% compared to a reference sample.

[0146] In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has an increase in immune cell H score of at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% compared to a reference sample.

[0147] In some embodiments of the methods of the present application, subjects who are more likely to respond to or benefit from the combination therapy as described herein have at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% TC / IC increase compared to a reference sample. In some embodiments of the methods of the present application, subjects who are more likely to respond to or benefit from the combination therapy as described herein have at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% TC / IC(+) increase compared to a reference sample. In some embodiments of the methods of the present application, a subject who is more likely to respond to or benefit from a combination therapy as described herein has at least a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 250%, 300%, 400%, 500% increase in TC / IC(++) compared to a reference sample.

[0148] In some embodiments of the present application methods, responsiveness includes a relative increase in one or more of the following: overall survival (OS), progression-free survival (PFS), complete response (CR), partial response (PR), stable disease (SD) and combinations thereof. In some embodiments, responsiveness includes eligibility for surgery (e.g., surgical operation).

[0149] In some embodiments of the present method, the subject is untreated. In some embodiments, the treatment is a first-line treatment. In some embodiments, the subject is ineligible for or refused first-line treatment. In some embodiments, the first-line treatment is selected from the group consisting of chemotherapy, PD-1 antibody therapy, PD-L1 antibody therapy, and any combination thereof.

[0150] In some embodiments, chemotherapy includes but is not limited to 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 methylpropylamines, including altretamine, triethylenemelamine, trietylenephosphoramide, triethiylenethiophosphoramide, and trimethylolomelamine; acetogenins (especially bullataci n) and bullatacinone); camptothecin (including the synthetic analogue topotecan); bryostatin; callystatin; CC-1065 (including its synthetic analogues adozelesin, carzelesin and bizelesin); cryptophycins (particularly cryptophycin 1 and cryptophycin 8); dolastatin; duocarmycin (including synthetic analogues, KW-2189 and CB1-TM1); eleutherobin; pancratistatin; sarcodictyin; spongistatin;Nitrogen mustards, for example, chlorambucil, chlornaphazine, cholophosphamide, estramustine, ifosfamide, mechlorethamine, mechlorethamine oxide hydrochloride, melphalan, novembichin, phenesterine, prednimustine, trofosfamide, uracil mustard mustard, nitrosureas (e.g. carmustine), chlorozotocin, fotemustine, lomustine, nimustine and ranimnustine; antibiotics, such as enediyne antibiotics, in particular calicheamicin gamma 1I and calicheamicin omega 1I; dynemicins, including dynemicin A; bisphosphonates, such as clodronate; esperamicin;and neocarzinostatin chromophores and related chromoprotein enediyne antibiotic chromophores), aclacinomysins, actinomycin, authramycin, azaserine, bleomycins, cactinomycin, carabicin, carminomycin, carzinophilin, chromomycinis, dactinomycin, daunorubicin, detorubicin, 6-diazo-5-oxo-L-norleucine, Doxorubicin (including morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino-doxorubicin and deoxydoxorubicin), epirubicin, esorubicin, idarubicin, marcellomycin, mitomycins (such as mitomycin C), mycophenolic acid (such as acid, nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin, quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin, ubenimex, zinostatin, zorubicin; antimetabolites, such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs, such as denopterin, rin), methotrexate, pteropterin, trimetrexate; purine analogs, for example, fludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidine analogs, for example, ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine;androgens (e.g., calusterone, dromostanolone propionate, epitiostanol, mepitiostane, testolactone); anti-adrenals (e.g., aminoglutethimide, mitotane, trilostane); folic acid replenishers (e.g., frolinic acid); aceglatone; aldophosphamide glycoside; aminolevulinic acid acid; eniluracil; amsacrine; bestrabucil; bisantrene; edatraxate; defofamine; demecolcine; diaziquone; elfornithine; elliptinium acetate; epothilone; etoglucid; gallium nitrate nitrate; hydroxyurea; lentinan; lonidainine; maytansinoids, such as maytansine and ansamitocins; mitoguazone; mitoxantrone; mopidanmol; nitraerine; pentostatin; phenamet; pirarubicin; losoxantrone; podophyllinic acid; 2-ethylhydrazide; procarbazine; Polysaccharide complex ( polysaccharide complex (JHS Natural Products, Eugene, Oreg.); razoxane; rhizoxin; sizofiran; spirogermanium; tenuazonic acid; triaziquone; 2,2',2"-trichlorotriethylamine; trichothecenes (especially T-2 toxin, verracurin A, roridin A) A) and anguidine); urethan; vindesine; dacarbazine; mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine; arabinoside ("Ara-C"); cyclophosphamide; thiotepa; taxoids, such as Paclitaxel (Bristol-Myers Squibb Oncology, Princeton, NJ), A hydrogenated castor oil-free, albumin-engineered paclitaxel nanoparticle formulation (American Pharmaceutical Partners, Schaumburg, Ill.), and Docetaxel ( doxetaxel) (Rhone-Poulenc Rorer, Antony, France); chloranbucil; Gemcitabine ( gemcitabine; 6-thioguanine; mercaptopurine; methotrexate; platinum analogs, such as cisplatin, oxaliplatin, and carboplatin; vinblastine, platinum, etoposide (VP-16); ifosfamide; mitoxantrone; vincristine; Vinorelbine vinorelbine; novantrone; teniposide; edatrexate; daunomycin; aminopterin; xeloda; ibandronate; irinotecan (Camptosar, CPT-11) (including irinotecan with 5-FU and folinic acid); topoisomerase inhibitor RFS2000 topoisomerase inhibitor RFS2000; difluoromethlhylornithine (DMFO); retinoids, such as retinoic acid; capecitabine; combretastatin; leucovorin (LV); oxaliplatin, including the oxaliplatin regimen (FOLFOX); PKC-α inhibitors, Raf inhibitors, H-Ras inhibitors, EGFR inhibitors (such as erlotinib) that reduce cell proliferation ) and VEGF-A inhibitors, and pharmaceutically acceptable salts, acids or derivatives of any of the foregoing.

[0151] Other non-limiting exemplary chemotherapies include anti-hormonal agents that modulate or inhibit the effects of hormones on cancer, such as antiestrogens and selective estrogen receptor modulators (SERMs), including, for example, tamoxifen (including tamoxifen), raloxifene, droloxifene, 4-hydroxytamoxifen, trioxifene, keoxifene, LY117018, onapristone, and Toremifene aromatase inhibitors that inhibit aromatase, which regulates estrogen production in the adrenal glands, such as 4(5)-imidazoles, aminoglutethimide, Megestrol acetate ( megestrol acetate), Exemestane exemestane), formestanie, fadrozole, Vorozole ( vorozole), Letrozole ( letrozole) and Anastrozole ( anastrozole; and antiandrogens, such as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin; and troxacitabine (a 1,3-dioxolane nucleoside cytosine analog); antisense oligonucleotides, particularly those that inhibit the expression of genes in signaling pathways involved in the proliferation of adherent cells (e.g., PKC-α, Ralf, and H-Ras); ribozymes, such as inhibitors of VEGF expression (e.g., ribozymes) and HER2 expression inhibitors; vaccines, such as gene therapy vaccines, e.g. vaccine, Vaccines and vaccine; rIL-2; Topoisomerase 1 inhibitors; rmRH; and pharmaceutically acceptable salts, acids or derivatives of any of the foregoing.

[0152] In some embodiments, exemplary PD-1 antibodies for first-line treatment include, but are not limited to, pembrolizumab, nivolumab, toripalimab, pidilizumab, cemiplimab, sintilimab, cetrelimab, spartalizumab, camrelizumab, tislelizumab, balstilimab, dostarlimab, ABBV-181, penpulimab, genolimzumab, mab), retifanlimab, sasanlimab, AMP-224, AB122, F-520, MEDI-3387, MEDI-5771, MEDI-0680, SG-001, BCD-100, BAT-1306, BI-754091, CBT-501, GLS-010, LZM-009, Sym-021, CS-1003, HLX-10, AK-103, AM-0001, ENUM-244C8, ENUM-388D4, JTX-4014, RXI-762, STI-A1110, HLX-20, SSI-361, APL-501, TJ0141H, and SNA-01.

[0153] In some embodiments, exemplary PD-L1 antibodies for first-line treatment include, but are not limited to, atezolizumab, manelimab, avelumab, cosibelimab, durvalumab, envafolimab, socazolimab, BGB-A333, CK-301, CS-1001, FAZ-053, APL-502, MDX-1105, IMC-001, KD-005, Gensci-047, LY-3300054, SHR-1316, MSB-2311, AVA-004, CBT-502, JS-003, B12, and KY-1003.

[0154] In another aspect, a combination of a CD73 antagonist and a PD1 / PD-L1 antagonist is provided for use in treating NSCLC in a subject in need thereof, wherein the treatment comprises (a) determining the level of CD73 in a sample from the subject, wherein an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample indicates that the subject is more likely to respond to or benefit from the combination therapy, and (b) administering the combination therapy to the subject. Kit or product

[0155] In another aspect, the present application provides a kit or article of manufacture for selecting a therapy for a subject with NSCLC, comprising: (1) one or more reagents for determining the level of CD73 in a sample from a subject with NSCLC; and (2) a package insert, wherein the package insert recommends treating the subject with a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the level of CD73.

[0156] In another aspect, the present application provides a kit or article of manufacture for identifying a subject with NSCLC who is more likely to respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist, the kit or article of manufacture comprising: (1) one or more reagents for determining the level of CD73 in a sample from a subject with NSCLC; and (2) a package insert, wherein the package insert recommends treating the subject with a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist based on the CD73 level.

[0157] In another aspect, the present application provides a kit or article of manufacture for use in the methods of the present application, the kit or article of manufacture comprising (1) one or more reagents for determining the level of CD73 in a sample from a subject with NSCLC; and (2) a package insert, wherein the package insert recommends treating the subject with a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the level of CD73.

[0158] On the other hand, the present application provides a kit or article of manufacture for use in the methods of the present application, the kit or article of manufacture comprising: (1) one or more reagents for determining the level of CD73 in a sample from a subject with NSCLC; and (2) a package insert, wherein the package insert suggests that an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample indicates that the subject is more likely to respond to a combination treatment with a CD73 antagonist and a PD1 / PD-L1 antagonist.

[0159] In some embodiments, the reagent in the kit or article is a CD73 antibody. Exemplary CD73 antibodies that can be used in the kit or article include, but are not limited to, D7F9A (CST, Cat. No. 13160), 606117 (R&D), EPR6114 (Abcam, Cat. No. ab133582), 4G6E3 (Abcam, Cat. No. ab202122), NT5E / 2503 (Abcam, Cat. No. ab257309), NT5E / 2505 (Abcam, Cat. No. ab257310), NT5E / 2545 (Abcam, Cat. No. ab2573 11), NT5E / 2646 (Abcam, Cat. No. ab257312), 7G2 (Thermo Fisher, Cat. No. 1D7), JM11-40 (Thermo Fisher, Cat. No. JM11-40), CD73 recombinant rabbit monoclonal antibody (1) (Invitrogen, Cat. No. MA5-29454), BLR054F (Thermo Fisher, Cat. No. BLR054F), sc-32299 (Santa Cruz, Cat. No. IE9) and AD2 (Biorad, Cat. No. AD2).

[0160] In some embodiments, the kit or article comprises a container and a label or package insert on or associated with the container. Suitable containers include, for example, bottles, vials, syringes, etc. The container can be made of a variety of materials (e.g., glass or plastic).

[0161] In some embodiments, the kit or article of manufacture further includes a CD73 antagonist and a PD1 / PD-L1 antagonist for combination therapy described herein.

[0162] All publications and patent applications cited in this specification are incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference. Example

[0163] The following examples are presented to provide a person of ordinary skill in the art with a complete disclosure and description of how to make and use the disclosure of the present application, and are not intended to limit the scope of what the inventors regard as their disclosure, nor are they intended to represent that the following experiments are all or the only experiments performed. Efforts have been made to ensure the accuracy of the numbers used (e.g., quantities, temperatures, etc.), but some experimental errors and deviations should also be taken into account. Unless otherwise indicated, parts are by weight, molecular weights are weight average molecular weights, temperatures are in degrees Celsius, and pressures are at or near atmospheric pressure. Example 1. Staining of CD73 by IHC

[0164] Formalin-fixed paraffin-embedded (FFPE) tumor tissue sample sections from NSCLC subjects were analyzed by IHC assay using the Ventana Benchmark Ultra system. Briefly, FFPE sections were dewaxed before antigen retrieval, blocked and incubated with Abcam anti-CD73 antibody EPR6114 (Abcam, Cat. No. ab133582), followed by incubation with HRP-labeled secondary antibodies. Ultra View Universal DAB Detection Kit (Roche 760-500), an HRP substrate, was used to visualize positive areas of the tissue.

[0165] In tumor samples, under a 4X-20X microscope, the intensity of CD73 staining of cells has a dynamic range from weak light brown to strong dark brown. Based on this intensity, staining can be divided into no staining (0), faint / weak staining (1), medium / moderate staining (2) and strong / intense staining (3). The intensity of 0-3 is shown in the figure. Figure 1 shown.

[0166] like Figure 2 As shown, the tumor region in a tumor sample typically includes tumor cells as well as tumor-associated stroma, including immune cells such as tumor infiltrating immune cells (TILs) that infiltrate into and are adjacent to the main tumor mass. Example 2. Scoring of CD73 levels by IHC

[0167] To quantify CD73 in tumor samples, a new CD73 IHC scoring system was established to measure CD73 levels in tumor cells and / or immune cells, as shown in Table 2 below. Table 2. CD73 IHC scoring system Rating system Measurement instructions TC Percentage of tumor cells stained with any intensity (1, 2, and 3) TC(+) Percentage of tumor cells staining with moderate or strong intensity (2 and 3) TC(++) Percentage of tumor cells stained with strong intensity (3) TCH Tumor cell H score, the sum of TC, TC(+) and TC(++) IC Percentage of immune cell staining, with any intensity (1, 2, and 3) IC(+) Percentage of immune cells staining with moderate or strong intensity (2 and 3) IC(++) Percentage of immune cells staining with strong intensity (3) ICH Immune cell H score, the sum of IC, IC(+) and IC(++) TC / IC The higher of TC and IC TC / IC(+) The higher of TC(+) and IC(+) TC / IC(++) The higher of TC(++) and IC(++) TC+IC The sum of TC and IC TC+IC(+) The sum of TC(+) and IC(+) TC+IC(++) The sum of TC(++) and IC(++)

[0168] In an ongoing Phase I / II study of the CD73 antibody Uliledlimab developed by I-Mab Biopharma, the combination of Uliledlimab and the PD-1 antibody Toriparlimab was introduced into one of the arms during the dose extension phase of the trial. The subjects included in this group were divided into two subgroups, including:

[0169] (I) Subjects who have failed first-line treatment (including SoC chemotherapy and SoC post-PD-1 / PD-L1 antibody treatment), and

[0170] (II) Subjects who have not received first-line treatment, including subjects who are not eligible for SoC chemotherapy or have been rejected by SoC chemotherapy.

[0171] Before treatment, FFPE sections of the subject's tumor samples were prepared and IHC staining was performed as described above. The staining results are summarized in Table 3 below. Table 3. IHC staining of CD73 Example 3. Correlation between CD73 levels and subjects' response to combined therapy

[0172] Before the combined treatment, CD73 levels in samples from subjects in groups (I) and (II) were analyzed. Then, Uliledlimab (20 mg / kg, Q3W) and PD-1 antibody Teplizumab (240 mg, Q3W) were administered to subjects in groups (I) and (II) for approximately 42 ± 7 days. After treatment, the correlation between the subjects' response to the combined treatment and their CD73 levels before treatment was further analyzed, and PR was defined as tumor regression of at least 30% from baseline according to RECISTv1.1.

[0173] Figure 3A and 3B The results showed that before combined treatment, the mean CD73 levels of Group (I) and Group (II) were comparable under both the TC / IC and TC / IC(+) scoring systems. In the subjects of Group (I), no significant difference in CD73 levels was detected between responders and non-responders. Surprisingly, Figure 4A and 4B As shown, subjects with higher CD73 levels in group (II) showed significantly stronger responses to the combined treatment of CD73 antibody and PD-1 antibody, regardless of the TC / IC (p=0.091) or TC / IC(+) (p=0.0019) scoring system.

[0174] The correlation between CD73 levels and response to combination therapy (measured as the best tumor change from baseline) in groups (I) and (II) was further evaluated by Pearson correlation coefficient. Figure 5A and 5B As shown, under the TC / IC (rho: -0.38 vs -0.02) and TC / IC(+) (rho: -0.53 vs -0.023) scoring systems, the CD73 level in group (II) was more strongly correlated with the response to combination therapy than the correlation presented in group (I), supporting the use of CD73 levels as a suitable biomarker to select subjects who have not undergone first-line treatment for combination therapy with anti-CD73 antibodies and anti-PD-1 antibodies. Example 4. Preliminary ROC evaluation

[0175] Receiver operating characteristic (ROC) analysis was used to evaluate the cutoff value of CD73 levels for selecting subjects who had not undergone first-line treatment (Group II) for combination therapy. Fig. 6A and 6B The preliminary ROC curves under the TC / IC and TC / IC(+) scoring systems are shown respectively. Table 4 below also summarizes the preliminary results from the ROC curves for the TC / IC and TC / IC(+) scoring systems, indicating that both scoring systems are reliable methods for selecting subjects. Table 4. Preliminary summary of ROC curves Rating system Area under the ROC curve Youden Index TC / IC 0.8385 0.6462 TC / IC(+) 0.9769 0.9231 Youden index = sensitivity + specificity - 1

[0176] The disclosure of the present application has been described to include the preferred mode of the practice of the disclosure of the present application according to the specific embodiment found or proposed by the inventor. It will be appreciated by those skilled in the art that, according to the disclosure of the present application, without departing from the expected scope of the disclosure of the present application, a large number of modifications and changes can be made to the specific embodiment of the examples. For example, due to codon redundancy, potential DNA sequences can be changed without affecting the protein sequence. In addition, for the consideration of biological function equivalence, protein structure can be changed without affecting its biological action in kind or quantity. All these modifications are expected to be included in the scope of the appended claims. Sequence Listing

Claims

1. A method for treating a subject having non-small cell lung cancer (NSCLC), the method comprising: (a) identifying a NSCLC subject who is likely to respond to or benefit from a combination of a CD73 antagonist and a PD1 / PD-L1 antagonist based on the level of CD73 in a sample from the subject, and (b) administering a CD73 antagonist and a PD1 / PD-L1 antagonist to the identified subject.

2. A method for selecting a therapy for a subject with NSCLC, the method comprising: (a) determining the level of CD73 in a sample from the subject, and (b) based on the CD73 level, assessing whether the subject is likely to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, (c) selecting the combination therapy for the subject when it is determined that the subject is likely to respond to or benefit from the combination therapy.

3. A method for treating a subject with NSCLC, the method comprising administering a CD73 antagonist and a PD1 / PD-L1 antagonist to a subject with NSCLC whose CD73 level in a sample indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist.

4. A method for predicting responsiveness of a subject with NSCLC to a combination treatment with a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample from the subject, (b) based on the CD73 level, assessing whether the subject is likely to respond to or benefit from the combination therapy, and (c) when it is determined that the subject is likely to respond to or benefit from the combination therapy, providing the subject with a recommendation for the combination therapy.

5. A method for identifying a subject with NSCLC who is more likely than a reference patient to respond to or benefit from a combination therapy of a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample from the subject, wherein an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample from a reference patient indicates that the subject is more likely to respond to or benefit from the combination therapy than the reference patient, and (b) providing a suggestion that the subject will be more likely than a reference patient to respond to or benefit from the combination therapy.

6. A method for predicting responsiveness of a subject with NSCLC to a combination treatment with a CD73 antagonist and a PD1 / PD-L1 antagonist, the method comprising: (a) determining the level of CD73 in a sample from the subject, wherein an increase in CD73 in the sample relative to CD73 in a reference sample from a reference patient indicates that the subject is more likely to respond to or benefit from the combination therapy than the reference patient, and (b) providing a suggestion that the subject will have an increased likelihood, compared to the reference patient, of responding to or benefiting from the combination therapy.

7. The method according to any one of claims 1-6, wherein the CD73 antagonist of the combination therapy is a CD73 antibody.

8. The method of claim 7, wherein the CD73 antibody is selected from the group consisting of a polyclonal antibody, a monoclonal antibody, a Fab, a scFv, a diabody, a triabody, a minibody, a VHH, and a sdAb.

9. The method of claim 7 or 8, wherein the CD73 antibody comprises: (a) HCDR1, HCDR2 and HCDR3 in the heavy chain variable region (VH) having the sequence as shown in SEQ ID No: 1, and (b) LCDR1, LCDR2 and LCDR3 in the light chain variable region (VL) having the sequence as shown in SEQ ID No: 2, Among them, HCDR1, HCDR2, HCDR3, LCDR1, LCDR2 and LCDR3 are according to the Kabat numbering scheme.

10. The method of any one of claims 7-9, wherein the CD73 antibody comprises: (1) HCDR1 comprising the amino acid sequence shown in SEQ ID NO: 3, (2) HCDR2 comprising the amino acid sequence shown in SEQ ID NO: 4, (3) HCDR3 comprising the amino acid sequence shown in SEQ ID NO: 5, (4) LCDR1 comprising the amino acid sequence shown in SEQ ID NO: 6, (5) LCDR2 comprising the amino acid sequence shown in SEQ ID NO: 7, and (6) LCDR3 comprising the amino acid sequence shown in SEQ ID NO:

8.

11. The method of any one of claims 7-10, wherein the CD73 antibody is selected from the group consisting of uliledlimab (I-Mab Biopharma), oleclumab (AstraZeneca), CPI-006 (Corvus Pharma), BMS-986179 (Bristol-Myers Squibb), AB-680 (Arcus Biosciences), NZV-930 (SRF373, Surface Oncolohgy / Novartis), JAB-BX102 (Jacobio), AK119 (Akesobio), Sym024 (Symphogen), IBI 325 (Innovent), BR 101 (Hisun BioRay) and LY3475070 (Eli Lilly).

12. The method according to any one of claims 1-11, wherein the PD-1 / PD-L1 antagonist of the combination therapy is a PD-1 antibody or a PD-L1 antibody.

13. The method of claim 12, wherein the PD-1 antibody is selected from the group consisting of: pembrolizumab, nivolumab, toripalimab, pidilizumab, cemiplimab, sintilimab, cetrelimab, spartalizumab, camrelizumab, tislelizumab, balstilimab, dostarlimab, ABBV-181, penpulimab, genoli mzumab), retifanlimab, sasanlimab, AMP-224, AB122, F-520, MEDI-3387, MEDI-5771, MEDI-0680, SG-001, BCD-100, BAT-1306, BI-754091, CBT-501, GLS-010, LZM-009, Sym-021, CS-1003, HLX-10, AK-103, AM-0001, ENUM-244C8, ENUM-388D4, JTX-4014, RXI-762, STI-A1110, HLX-20, SSI-361, APL-501, TJ0141H, and SNA-01.

14. The method of claim 12, wherein the PD-L1 antibody is selected from the group consisting of atezolizumab, manelimab, avelumab, cosibelimab, durvalumab, envafolimab, socazolimab, BGB-A333, CK-301, CS-1001, FAZ-053, APL-502, MDX-1105, IMC-001, KD-005, Gensci-047, LY-3300054, SHR-1316, MSB-2311, AVA-004, CBT-502, JS-003, B12 and KY-1003.

15. The method according to any one of claims 1-14, wherein the combination therapy further comprises chemotherapy.

16. The method of claim 15, wherein the chemotherapy is selected from the group consisting of platinum drugs (such as cisplatin, carboplatin), taxane drugs (such as paclitaxel, albumin-bound paclitaxel, docetaxel), vinorelbine, vinblastine, etoposide, pemetrexed and gemcitabine, and any combination thereof.

17. The method according to any one of claims 1 to 16, wherein: The combination therapy comprises the simultaneous or separate administration of a CD73 antagonist and a PD-1 / PD-L1 antagonist.

18. The method of any one of claims 1-17, wherein the NSCLC is stage III or stage IV NSCLC.

19. The method of any one of claims 1-18, wherein the NSCLC is advanced NSCLC or metastatic NSCLC.

20. The method of any one of claims 1-19, wherein the NSCLC is recurrent NSCLC.

21. The method of any one of claims 1-20, wherein the NSCLC has squamous histology or non-squamous histology.

22. The method according to any one of claims 1-21, wherein the subject is treatment naive.

23. The method of claim 22, wherein the treatment is first line treatment.

24. The method of claim 23, wherein the subject is ineligible for or has been rejected by a first line of treatment.

25. The method of claim 23 or 24, wherein the first line of treatment is selected from the group consisting of chemotherapy, PD-1 antibody therapy, PD-L1 antibody therapy, and any combination thereof.

26. The method of claim 25, wherein the chemotherapy is selected from the group consisting of platinum drugs (such as cisplatin, carboplatin), taxane drugs (such as paclitaxel, albumin-bound paclitaxel, docetaxel), vinorelbine, vinblastine, etoposide, pemetrexed and gemcitabine, and any combination thereof.

27. The method of claim 25, wherein the PD-1 antibody is selected from the group consisting of: pembrolizumab, nivolumab, toripalimab, pidilizumab, cemiplimab, sintilimab, cetrelimab, spartalizumab, camrelizumab, tislelizumab, balstilimab, dostarlimab, ABBV-181, penpulimab, genoli mzumab), retifanlimab, sasanlimab, AMP-224, AB122, F-520, MEDI-3387, MEDI-5771, MEDI-0680, SG-001, BCD-100, BAT-1306, BI-754091, CBT-501, GLS-010, LZM-009, Sym-021, CS-1003, HLX-10, AK-103, AM-0001, ENUM-244C8, ENUM-388D4, JTX-4014, RXI-762, STI-A1110, HLX-20, SSI-361, APL-501, TJ0141H, and SNA-01.

28. The method of claim 25, wherein the PD-L1 antibody is selected from the group consisting of atezolizumab, manelimab, avelumab, cosibelimab, durvalumab, envafolimab, socazolimab, BGB-A333, CK-301, CS-1001, FAZ-053, APL-502, MDX-1105, IMC-001, KD-005, Gensci-047, LY-3300054, SHR-1316, MSB-2311, AVA-004, CBT-502, JS-003, B12 and KY-1003.

29. The method of any one of claims 1-28, wherein the level of CD73 is the protein level of CD73.

30. The method of claim 29, wherein protein levels are determined by immunohistochemistry (IHC).

31. The method of any one of claims 1-30, wherein the sample comprises cells selected from the group consisting of tumor cells, immune cells (such as tumor infiltrating immune cells), stromal cells, and any combination thereof.

32. The method of any one of claims 1-31, wherein the sample is a formalin-fixed and paraffin-embedded (FFPE) sample, an archived sample, a fresh sample, or a frozen sample.

33. The method according to any one of claims 1-32, wherein the sample is obtained prior to the combination therapy.

34. The method of any one of claims 1-33, wherein the reference sample comprises a reference tissue or a reference cell.

35. The method of claim 34, wherein the reference sample is derived from a healthy subject or a non-disease sample of a subject.

36. The method of any one of claims 1-35, wherein responsiveness comprises a relative increase in one or more of: overall survival (OS), progression-free survival (PFS), complete response (CR), partial response (PR), and combinations thereof.

37. The method of any one of claims 1-36, wherein the level of CD73 comprises the proportion of CD73-positive cells among all cells in the sample.

38. The method of claim 37, wherein the CD73 level comprises the proportion of CD73-positive tumor cells among all tumor cells in the sample, or the proportion of CD73-positive immune cells among all immune cells in the sample, whichever is higher.

39. The method of claim 38, wherein a CD73 level of 30% or greater identifies the subject as likely to respond to or benefit from the combination therapy.

40. The method of claim 38, wherein a CD73 level of 35%, 40%, 45% or 50% or more identifies the subject as likely to respond to or benefit from the combination therapy.

41. The method of claim 37, wherein the CD73 level comprises the proportion of CD73-positive tumor cells among all tumor cells in the sample, or the proportion of CD73-positive immune cells among all immune cells in the sample, whichever is higher, wherein positive is at least moderately positive.

42. The method of claim 41, wherein a CD73 level of 10% or greater identifies the subject as likely to respond to or benefit from the combination therapy.

43. The method of claim 41, wherein a CD73 level of 15%, 20%, 25%, 30%, 35% or 40% or more identifies the subject as likely to respond to or benefit from the combination therapy.

44. A kit or article of manufacture for use in the method of any one of claims 1-43, the kit or article of manufacture comprising: (1) one or more reagents for determining the level of CD73 in a sample from a subject with NSCLC, and (2) a package insert, wherein the package insert recommends treating the subject with a CD73 antagonist and a PD1 / PD-L1 antagonist based on CD73 levels.

45. A kit or article of manufacture for use in the method of any one of claims 1-43, the kit or article of manufacture comprising: (1) one or more reagents for determining the level of CD73 in a sample from a subject with NSCLC, and (2) a package insert, wherein the package insert advises that an increase in the level of CD73 in the sample relative to the level of CD73 in a reference sample indicates that the subject is more likely to respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist.

46. ​​The kit or article of manufacture of claim 44 or 45, wherein the agent is a CD73 antibody.

47. The kit or article of manufacture according to claim 46, wherein the CD73 antibody is selected from the group consisting of: D7F9A (CST, Cat. No. 13160), 606117 (R&D), EPR6114 (Abcam, Cat. No. ab133582), 4G6E3 (Abcam, Cat. No. ab202122), NT5E / 2503 (Abcam, Cat. No. ab257309), NT5E / 2505 (Abcam, Cat. No. ab257310), NT 5E / 2545 (Abcam, Cat. No. ab257311), NT5E / 2646 (Abcam, Cat. No. ab257312), 7G2 (Thermo Fisher, Cat. No. 1D7), JM11-40 (Thermo Fisher, Cat. No. JM11-40), CD73 recombinant rabbit monoclonal antibody (1) (Invitrogen, Cat. No. MA5-29454), BLR054F (Thermo Fisher, Cat. No. BLR054F), sc-32299 (Santa Cruz, Cat. No. IE9) and AD2 (Biorad, Cat. No. AD2).

48. Use of a CD73 antagonist and a PD1 / PD-L1 antagonist in the manufacture of a medicament for treating a subject with NSCLC, wherein the level of CD73 in a sample of the subject indicates that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist.

49. Use of a CD73 antagonist for the manufacture of a medicament for treating a subject having NSCLC, wherein a sample from the subject has a level of CD73 indicating that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist, wherein the subject is also administered a PD1 / PD-L1 antagonist.

50. Use of a PD1 / PD-L1 antagonist for the manufacture of a medicament for treating a subject having NSCLC, wherein a sample of the subject has a level of CD73 indicating that the subject may respond to or benefit from a combination therapy with a CD73 antagonist and a PD1 / PD-L1 antagonist, wherein the subject is also administered a CD73 antagonist.

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