ISOFORM-SPECIFIC, CONTEXT-PERMISSIVE TGFB1 INHIBITORS AND USE THEREOF

1 . A method for treating a disease associated with TGFβ1 dysregulation in a subject, the method comprising a step of administering to the subject a therapeutically effective amount of a composition comprising an isoform-specific inhibitor of TGFβ1 and a pharmaceutically acceptable excipient,

wherein the inhibitor targets both ECM-associated TGFβ1 and immune cell-associated TGFβ1 but does not target TGF82 or TGFβ3 in vivo, and,

wherein the disease is characterized by dysregulation or impairment at least two of the following attributes:

a) regulatory T cells (Treg);

b) effector T cell (Teff) proliferation or function;

c) myeloid cell proliferation or differentiation;

d) monocyte recruitment or differentiation;

e) macrophage function;

f) epithelial-to-mesencymal transition (EMT) and/or endothelial-to-mesenchymal transition (EndMT);

g) gene expression in one or more of marker genes selected from the group consisting of: PAI-1, ACTA2, CCL2, Col1 al, Col3a1, FN-1, CTGF, and TGFβ1;

h) ECM components or function; and,

i) fibroblast activation/differentiation.

2 . The method of claim 1 , wherein the ECM-associated TGFβ1 is LTBP1-presented TGFβ1 and/or LTBP3-presented TGF81; and, wherein the immune cell-associated TGFβ1 is GARP-presented TGFβ1 and/or LRRC33-presented TGF81.

3 . The method of claim 1 , wherein the disease involves a proliferative component and/or a fibrotic component.

4 . The method of claim 3 , wherein the proliferative component of the disease comprises abnormal cell proliferation.

5 . The method of claim 4 , wherein the disease is cancer.

6 . The method of claim 5 , wherein the cancer is a metastatic cancer.

7 . The method of claim 5 , wherein the cancer comprises a solid tumor, wherein the solid tumor is TGFβ1-positive.

8 . The method of claim 7 , wherein TGFβ1 expression in a clinical sample of the subject is greater than TGF62 or TGF63 expression.

9 . The method of claim 7 , wherein the solid tumor is a desmoplastic tumor.

10 . The method of claim 5 , wherein the cancer is associated with an increased number of Tregs, TAMs, TANs, MDSCs, CAFs, or any combinations thereof.

11 . The method of claim 5 , wherein the subject is poorly responsive to a therapy selected from the group consisting of: radiation therapy, chemotherapy and checkpoint inhibitor therapy.

12 . The method of claim 11 , wherein the checkpoint inhibitor therapy comprises a PD-1 antagonist, PD-L1 antagonist or CTLA-4 antagonist.

13 . The method of claim 11 , wherein the subject has an immune checkpoint inhibitor-resistant cancer selected from the group consisting of:

myelofibrosis, melanoma, renal cell carcinoma, bladder cancer, colon cancer, hematologic malignancies, non-small cell carcinoma, non-small cell lung cancer (NSCLC), lymphoma (classical Hodgkin's and non-Hodgkin's), head and neck cancer, urothelial cancer, cancer with high microsatellite instability, cancer with mismatch repair deficiency, gastric cancer, renal cancer, and hepatocellular cancer.

14 . The method of claim 1 , wherein a clinical sample of the human subject is GARP-positive and/or LRRC33-positive in expression.

15 . The method of claim 5 , wherein the therapeutically effective amount is an amount effective to achieve one or more of the following clinical effects:

a) reduced tumor growth;

b) reduced metastasis;

c) reduced tumor invasion;

d) reduced angiogenesis and vascularization/vascularity;

e) reduced M2 TAM infiltration of the tumor;

f) increased ratios of M1 over M2 (TAM-like) macrophage populations at a tumor site;

g) reduced number of CAFs at a tumor site;

h) reduced immuno-suppression;

i) enhanced responsiveness to a cancer therapy;

j) prolonged survival;

k) prolonged refractory period;

I) increased rates of complete remission or complete responses;

m) decreased ratios of Treg/Teff cells at a tumor site;

n) increased number of Teff cells at a tumor site;

o) reduced number of Treg cells at a tumor site;

p) reduced number of MDSCs and/or TANs in the subject; and,

wherein the clinical effect(s) is/are achieved with an acceptable level of toxicities in the subject.

16 . The method of claim 5 , wherein the cancer is a myeloproliferative disorder.

17 . The method of claim 16 , wherein the myeloproliferative disorder is essential thrombocythemia (ET), polycythemia vera (PV) or primary myelofibrosis (PMF).

18 . The method of claim 16 , wherein the therapeutically effective amount is an amount effective to achieve at least two of the following clinical benefits:

a) reduced fibrosis in a bone marrow;

b) enhanced hematopoiesis of differentiated blood cells in a bone marrow;

c) reduced proliferation of abnormal stem cells in the bone marrow, wherein optionally the abnormal stem cells are CD133-positive;

d) reduced megakaryocytes in a bone marrow and/or spleen;

e) reduced occurrence and/or extent of extramedullary hematopoiesis in the subject, wherein optionally the extramedullary hematopoiesis is in spleen;

f) reduced need for bone marrow transplantation;

g) prolonged survival;

h) normalized levels of one or more of expression markers, wherein the expression marker optionally is selected from the group consisting of BMP1, BMP6, BMP7, and BMP-receptor 2, PLOD2, TGFβ, bFGF, platelet-derived growth factor (PDGF), Col1, metalloproteinases, FN1, CXCL12, VEGF, CXCR4, IL-2, IL-3, IL-9, CXCL1, IL-5, IL-12, TNFα, Bmp2, Bmp5, Acvr11, Tgfblil, Igf1, Cdkn1a, Ltbp1, Gdf2, Lefty1 and Nodal; and,

i) reduced chronic inflammation in the bone marrow.

19 . The method of claim 3 , wherein the proliferative component of the disease comprises overexpression and deposition of an ECM component.

20 . The method of claim 19 , wherein the ECM component is Collagen I.

21 . The method of claim 19 , wherein the disease is a fibrotic disorder.

22 . The method of claim 21 , wherein the fibrotic disorder is an organ fibrosis.

23 . The method of claim 22 , wherein the organ fibrosis is liver fibrosis, lung fibrosis, kidney fibrosis, skin fibrosis and/or cardiac fibrosis.

24 . The method of claim 22 , wherein the subject is not a candidate for organ transplantation.

25 . The method of claim 21 , wherein the fibrotic disorder comprises chronic inflammation.

26 . The method of claim 25 , wherein the fibrotic disorder is a muscular dystrophy.

27 . The method of claim 26 , wherein the muscular dystrophy is DMD.

28 . The method of claim 1 , wherein the isoform-selective inhibitor inhibits three or more of the following TGFβ1 activities:

a) GARP-mediated TGFβ1 activity;

b) LRRC33-mediated TGFβ1 activity;

c) LTBP1-mediated TGFβ1 activity, and

d) LTBP3-mediated TGFβ1 activity.

29 . The method of claim 28 , wherein the inhibitor inhibits all of the TGFβ1 activities (a)-(d).

30 . The method of claim 1 , wherein the inhibitor inhibits TGFβ1 activation.

31 . The method of claim 1 , wherein the inhibitor is a monoclonal antibody or fragment thereof.

32 . The method of claim 31 , wherein the monoclonal antibody or fragment thereof binds a protein complex comprising a pro/latent TGFβ1.

33 . The method of claim 32 , wherein the protein complex comprises a presenting molecule selected from the group consisting of: LTBP1, LTBP3, GARP and LRRC33.

34 . The method of claim 32 , wherein the antibody or fragment thereof specifically binds an epitope within pro/latent TGFβ.

35 . The method of claim 32 , wherein the epitope is within a prodomain of the pro/latent TGFβ1.

36 . The method of claim 31 , wherein the antibody or fragment thereof specifically binds a combinatorial epitope.

37 . The method of claim 31 , wherein the antibody or fragment thereof specifically binds a conformational epitope.

38 . The method of claim 31 , wherein the antibody or fragment thereof inhibits release of mature TGFβ1 growth factor from a latent protein complex comprising pro/latent TGFβ1.

39 . The method of claim 31 , wherein the antibody or the fragment thereof is a fully human or humanized antibody.

40 . The method of claim 31 , wherein the antibody is a human IgG4 antibody.

41 . The method of claim 40 , wherein the human IgG4 antibody comprises a backbone substitution.

42 . The method of claim 31 , wherein the antibody or fragment thereof has the following CDR sequences, with three or fewer substitutions:

CDR-H1:

(SEQ ID NO: 85)

NYAMS;

CDR-H2:

(SEQ ID NO: 86)

SISGSGGATYYADSVKG;

CDR-H3:

(SEQ ID NO: 87)

ARVSSGHWDFDY;

CDR-L1:

(SEQ ID NO: 88)

RASQSISSYLN;

CDR-L2:

(SEQ ID NO: 89)

SSLQS;

and,

CDR-L3:

(SEQ ID NO: 90)

QQSYSAPFT.

43 . The method of claim 42 , wherein the antibody or fragment thereof has no substitutions.

44 . The method of claim 1 , further comprising the steps of

determining relative expression levels of TGFβ, TGFβ2 and TGFβ3 in a clinical sample of the human subject, and, if TGFβ1 is a dominant isoform expressed in the clinical sample, then,

selecting the human subject as a candidate for the isoform-specific inhibitor of TGFβ1 treatment.

45 . The method of claim 44 , further comprising:

determining relative expression levels of LTBP1/3, GARP and LRRC33 in a clinical sample of the human subject, and, if at least one type of ECM-associated presenting molecule and one type of cell-associated presenting molecule are co-expressed in the clinical sample, then,

selecting the human subject as a candidate for the isoform-specific inhibitor of TGFβ1 treatment.

46 . A pharmaceutical composition comprising an antibody comprising a heavy chain variable region polypeptide that is at least 90% identical to an amino acid sequence set forth in SEQ ID NO:95, and, a light chain variable region polypeptide that is at least 90% identical to an amino acid sequence set forth in SEQ ID NO:97.

47 . An isolated antibody having the following CDR sequences optionally with three or fewer substitutions:

CDR-H1:

(SEQ ID NO: 85)

NYAMS;

CDR-H2:

(SEQ ID NO: 86)

SISGSGGATYYADSVKG;

CDR-H3:

(SEQ ID NO: 87)

ARVSSGHWDFDY;

CDR-L1:

(SEQ ID NO: 88)

RASQSISSYLN;

CDR-L2:

(SEQ ID NO: 89)

SSLQS;

and,

CDR-L3:

(SEQ ID NO: 90)

QQSYSAPFT.

48 . The isolated antibody of claim 47 , wherein the antibody has the CDR sequences with no substitutions.