Immune landscape signatures for ocular inflammation

Abstract

The present disclosure relates to gene signatures of AAV-induced inflammatory response useful in gene therapy methods for the identification and treatment of individuals who are candidates to receive, or who have received an rAAV gene therapy treatment, and who have an increased likelihood of AAV-induced inflammation.

Description

Abstract This paper discloses genetic signatures involved in AAV-induced inflammatory responses that can be used in gene therapy to identify and treat individuals who may have received or have received rAAV gene therapy and who are at increased risk of AAV-induced inflammation.

Claims

CLAIMSWhat is claimed is:

1. A method of screening a patient prior to intravitreal rAAV therapy to identify a patient at risk of AAV- induced inflammation, the method comprising: (a) obtaining a biological sample from the patient; (b) detecting expression levels of each of a set of genes in the biological sample; (c) determining whether the detected expression levels indicate an increased likelihood of AAV-induced inflammation by comparison to a predetermined threshold expression levels for the set of genes; and (d) identifying the patient as in need of antiinflammatory treatment if an increased likelihood of AAV-induced inflammation is determined.

2. A method of monitoring a patient who has received or is receiving an rAAV treatment and who also is receiving an anti-inflammatory treatment for AAV-induced inflammation, the method comprising: (a) obtaining a biological sample from the patient; (b) detecting expression levels of each of a set of genes in the biological sample; (c) determining whether the detected expression levels indicate a reduced likelihood of AAV-induced inflammation by comparison to a predetermined threshold expression levels for the set of genes; and (d) identifying the patient as a candidate for reduced anti-inflammatory treatment if a reduced likelihood of AAV-induced inflammation is determined.

3. A method of identifying a patient in need of an anti-inflammatory treatment from a population of patients who have received intravitreal rAAV therapy, the method comprising: (a) obtaining a biological sample from the patient who has received an rAAV treatment; (b) detecting expression levels of each of a set of genes in the biological sample; (c) determining whether the detected expression levels indicate an increased likelihood of AAV- induced inflammation by comparison to a predetermined threshold expression levels for the set of genes; and (d) identifying the patient as in need of anti-inflammatory treatment if an increased likelihood of AAV-induced inflammation is determined.

4. A method of selecting and treating a patient who has an increased likelihood of AAV- induced inflammation from a population of patients who have received an rAAV treatment, the method comprising: (a) obtaining a biological sample from the patient who has received an rAAV treatment; (b) detecting expression levels of each of a set of genes in the biological sample; (c) determining whether the detected expression levels indicate an increased likelihood of AAV-induced inflammation by comparison to a predetermined threshold expression levels for the set of genes; and (d) administering an anti-inflammatory agent to the patient if an increased likelihood of AAV-induced inflammation is determined.

5. A method of providing immunosuppressive treatment in a patient who has received intravitreal rAAV therapy, the method comprising: (a) obtaining a biological sample from the patient who has received an rAAV treatment; (b) detecting expression levels of each of a set of genes in the biological sample; (c) determining whether the detected expression levels indicate an increased likelihood of AAV-induced inflammation by comparison to predetermined threshold levels for the genes; and (d) administering an immunosuppressive treatment to the patient if an increased likelihood of AAV-induced inflammation is determined.

6. The method of any one of claims 1 -5, wherein the set of genes comprises at least two genes selected from: A2M, B2M, BLNK, BTK, C1QA, C1 QB, C1QC, C1 R, C1S, C2, C3, C4B, CARD11 , CASP8, CCL3, CCL4L2, CCL5, CD14, CD19, CD22, CD247, CD28, CD3D, CD3E, CD3G, CD4, CD40, CD40LG, CD72, CD74, CD79A, CD79B, CD86, CD8A, CD8B, CFHR4, CHUK, CIITA, CPB2, CR1 , CR2, CTLA4, CTSS, CXCL10, CXCL11, CXCL8, CXCL9, CXCR3, DAPP1 , F12, F13A1 , F2RL3, FCGR1A, FCGR2A, FCGR3A, FYN, GRAP2, HCK, HLA-DMA, HLA-DMB, HLA-DOA, HLA-DOB, HLA-DPA1 , HLA-DPB1 , HLA- DQA1 , HLA-DRA, HLA-DRB1 , HLA-E, HLA-F, HLA-G, HSP90AA1 , ICOS, IFI30, IFITIvll, IFITM1 , IFNG, IKBKB, IKBKE, IKBKG, IL10, IL12A, IL12B, IL1 B, IL6, INPP5D, IRF7, ITGAM, ITGAX, ITGB2, ITK, IY96, KLRC2, KLRD1 , KPK3CD, LAT, LCK, LCP2, LGMN, LILRA1 , LILRA4, LILRB2, LILRB4, LR2, LY97, MAP3K7, MAPK13, MP3K7, MYD88, NCF1 , NFATC2, PAK5, PDCD1 , PIK3AP1 , PIK3CD, PLA2G4B, PLA2G4C, PLAU, PLCG2, PLD1 , PRKCD, PROC, PSME1 , PTPRC, RAC2, RASGRP3, SCIN, SERPINA1 , SERPINA5, SERPINE1 , STAT1 , SYK, TAP1 , TAP2, TLR1 , TLR4, TLR7, TLR8, TLR10, TNF, VASP, VAV1 , VAV2, VSIG4, WAS, WAV1 , and ZAP70.

7. The method of any one of claims 1-6, wherein the set of genes is selected from:(a) CXCL9, CXCL10, CXCL11 , CCL5 and CCL3;(b) CXCL9, CXCL11 , CXCL10, IL6, TNF, CCL5, CXCL8, CCL3, TLR8, CCL4L2, IY96, IL12A, IL12B, STAT1 , IKBKE, IL1 B, TLR1 , CASP8, IKBKG, TLR4, IRF7, CD14, MYD88, CD86, IKBKB, KPK3CD, MP3K7, CHUK, and CD40;(c) CXCL10, CXCL9, CCL5, CCL4L2, and MYD88;(d) CXCL10, CXCL9, CCL5, CXCL11 , IL1B, TLR8, CCL4L2, CD86, TLR2, STAT1 , TNF, LY97, TLR4, IRF7, TLR7 and IKBKE;(e) PLA2G4C, PRKCG, RAC2, PTPRC, FCGR3A, FCGR2A, SYK, and FCGR1A;(f) PLA2G4C, PTPRC, RAC2, LAT, NCF1 , FCGR3A, WAV1 , SYK, PLA2G4B, WAS, PLCG2, HCK, FCGR2A, INPP5D, FCGR2A, INPP5D, FCGR1A, PIK3CD, PRKCD, and VASP;(g) RAC2, SYK, VAV2, FCGR3A and PTPRC;(h) RAC2, FCGR2A, PTPRC, FCGR3A, SYK, VAV1 , NCF1 , PLA2G4C, WAS, SCIN, FCGR1A, INPP5D, HCK and PLD1;(i) C1R, ITGAM, C4B, F13A1, ITGB2 and C1QC;(j) SERPINA5, CR1, CR2, C4B, ITGAX, PROC, ITGAM, C3, F12, F13A1, ITGB2, SERPINE1, PLAU, C1S, C1QC, CFHR4, CIQAand C1R;(k) ITGAX, FGG and CPB2;(l) F13A1, ITGAX, C1QB, ITGAM, C1QC, C1QA, C2, ITGB2, C3, C4B, C1S, SERPINA1, VSIG4, SERPINA5, CR2, A2M, and F2RL3;(m)HLA-DOB, KLRC2, CD8B, TAP1, HLA-DRA, CTSS, HLA-DMB, IFNG, CD8Aand LGMN;(n) HLA-DOB, KLRD1, TNF, IFNG, TAP1, HLA-G, CIITA, KLRC2, CD8B, TAP2, HLA-E, CD8A, CTSS, HLA-DPB1, HLA-DPA1, HLA-DRB1 , HLA-DMB, HLA-DRA, HLA-DOA, HLA-F, CD74, HLA-DMA, IFI30, PSME1, HSP90AA1, and LGMN;(o) HLA-DRB1, HLA-DQA1, HLA-DPA1, and KLRD1;(p) HLA-DRA, HLA-G, B2M, HLA-DQA1, HLA-DRB1 , HLA-DPB1 , CIITA, CD74, HLA-DOA, HLA-DMB, HLA-DMA, HLA-E, CD8B, HLA-DPA1 , CD4, TAP1, IFI30, KLRD1, HLA- DOB, TNF, CD8A, TAP2, and PSME1;(q) CD79A, DAPP1, CD79B, CARD11, RAC2, BLNK, BTK, IFITIvll, and SYK;(r) CD79A, CD19, CARD11, CD79B, DAPP1, CR2, LILRA4, CD22, RAC2, BTK, LILRA1, BLNK, IFITM1, LILRB2, LILRB4, VAV1, SYK, PIK3AP1, PLCG2, INPP5D, CD72, IKBKG, IKBKB, PIK3CD, and CHUK;(s) DAPP1, RAC2, SYK, VAV2, and NFATC2;(t) RAC2, CD79A, CD79B, DAPP1 , LILRA4, SYK, VAV1, BTK, BLNK, LILRB2, INPP5D, NFATC2, LILRB4, CD22, CARD11, CR2, LILRA1, CD72 and RASGRP3;(u) LCK, CD40LG, CD3D, CD28, CD3E, CARD11, PAK5, CD247, CD8B, ITK, PTPRC, LCP2, IFNG, CD8A, and FYN;(v) LCK, CD3D, CARD11, CD28, TNF, ZAP70, CD40LG, CD3E, CD3G, PTPRC, ITK, IFNG, ICOS, PDCD1, LAT, LCP2, GRAP2, IL10, CD8B, CD8A, CD247, VAV1, PAK5, IKBKG, IKBKB, PIK3CD, MAPK13, MAP3K7, and CHUK;(w) VAV2, CD40LG, CD28, PTPRC, and NFATC2; and(x) CD3D, PTPRC, CD28, LCK, CD3G, VAV1, ICOS, CD8B, CD4, CD40LG, ITK, CD3E, TNF, CD8A, NFATC2, LCP2, CARD11, and CTLA4.

8. The method of any one of claims 1-7, wherein the biological sample comprises a first biological sample obtained at a first time point and a second biological sample obtained at a second time point.

9. The method of any one of claims 1-8, wherein detecting the expression levels comprises sequencing RNA in the biological sample.

10. The method of any one of claims 1-9, wherein the threshold expression level is a Iog2 foldchange in expression > |0.6|.

11. The method of any one of claims 1-10, wherein the AAV-induced inflammation is ocular inflammation.

12. The method of any one of claims 1-11 , wherein the biological sample comprises ocular cells.

13. The method of any one of claims 1-12, wherein the biological sample comprises blood, serum, tears, or a combination thereof.

14. The method of any one of claims 1-13, wherein the rAAV treatment was administered to an eye of the subject.

15. The method of claim 14, wherein the rAAV treatment was administered to the contralateral eye of the patient.

16. The method of any one of claims 4-13, wherein the anti-inflammatory agent is administered to the eye.

17. The method of any one of claims 4-15, wherein the anti-inflammatory agent is a steroid; optionally wherein the steroid is selected from a corticosteroid, prednisone, and difluprednate.

18. The method of any one of claims 16-17, wherein the steroid is administered systemically.

19. The method of any one of claims 16-17, wherein the steroid is administered orally.

20. The method of any one of claims 16-17, wherein the steroid is administered topically.21 . The method of any one of claims 1-20, wherein the AAV-induced inflammation is in the ciliary body and / or in the iris.

22. The method of any one of claims 1 -20, wherein the AAV-induced inflammation is in the retina.

23. The method of any one of claims 1-20, wherein the AAV-induced inflammation is in the choroid.

24. The method of claim 11 , wherein the expression levels of the set of genes in the biological sample relative to the predetermined threshold levels indicates a likelihood of AAV-induced inflammation in the ciliary body and / or the iris, and wherein the set of genes is selected from:(a) CXCL9, CXCL10, CXCL11 , CCL5 and CCL3;(b) CXCL9, CXCL11 , CXCL10, IL6, TNF, CCL5, CXCL8, CCL3, TLR8, CCL4L2, IY96, IL12A, IL12B, STAT1 , IKBKE, IL1 B, TLR1 , CASP8, IKBKG, TLR4, IRF7, CD14, MYD88, CD86, IKBKB, KPK3CD, MP3K7, CHUK, and CD40;(c) PLA2G4C, PRKCG, RAC2, PTPRC, FCGR3A, FCGR2A, SYK, and FCGR1A;(d) PLA2G4C, PTPRC, RAC2, LAT, NCF1 , FCGR3A, WAV1 , SYK, PLA2G4B, WAS, PLCG2, HCK, FCGR2A, INPP5D, FCGR2A, INPP5D, FCGR1A, PIK3CD, PRKCD, and VASP;(e) C1 R, ITGAM, C4B, F13A1 , ITGB2 and C1QC;(f) SERPINA5, CR1 , CR2, C4B, ITGAX, PROC, ITGAM, C3, F12, F13A1 , ITGB2, SERPINE1 , PLAU, C1S, C1QC, CFHR4, CIQA and C1R;(g) HLA-DOB, KLRC2, CD8B, TAP1 , HLA-DRA, CTSS, HLA-DMB, IFNG, CD8A and LGMN;(h) HLA-DOB, KLRD1 , TNF, IFNG, TAP1 , HLA-G, CIITA, KLRC2, CD8B, TAP2, HLA-E, CD8A, CTSS, HLA-DPB1 , HLA-DPA1 , HLA-DRB1 , HLA-DMB, HLA-DRA, HLA-DOA, HLA-F, CD74, HLA-DMA, IFI30, PSME1 , HSP90AA1 , and LGMN;(i) CD79A, DAPP1 , CD79B, CARD11 , RAC2, BLNK, BTK, IFITIvl I, and SYK; and(j) CD79A, CD19, CARD11 , CD79B, DAPP1 , CR2, LILRA4, CD22, RAC2, BTK, LILRA1 , BLNK, IFITM1 , LILRB2, LILRB4, VAV1 , SYK, PIK3AP1 , PLCG2, INPP5D, CD72, IKBKG, IKBKB, PIK3CD, and CHUK;(k) LCK, CD40LG, CD3D, CD28, CD3E, CARD11 , PAK5, CD247, CD8B, ITK, PTPRC, LCP2, IFNG, CD8A, and FYN; and(l) LCK, CD3D, CARD11 , CD28, TNF, ZAP70, CD40LG, CD3E, CD3G, PTPRC, ITK, IFNG, ICOS, PDCD1 , LAT, LCP2, GRAP2, IL10, CD8B, CD8A, CD247, VAV1 , PAK5, IKBKG, IKBKB, PIK3CD, MAPK13, MAP3K7, and CHUK.

25. The method of claim 11 , wherein the expression levels of the set of genes in the biological sample relative to the predetermined threshold levels indicates a likelihood AAV-induced inflammation in the retina, and wherein the set of genes is selected from:(m) CXCL10, CXCL9, CCL5, CCL4L2, and MYD88;(n) CXCL10, CXCL9, CCL5, CXCL11 , IL1B, TLR8, CCL4L2, CD86, TLR2, STAT1 , TNF, LY97, TLR4, IRF7, TLR7 and IKBKE;(o) RAC2, SYK, VAV2, FCGR3A and PTPRC;(p) RAC2, FCGR2A, PTPRC, FCGR3A, SYK, VAV1 , NCF1 , PLA2G4C, WAS, SCIN, FCGR1A, INPP5D, HCK and PLD1 ;(q) ITGAX, FGG and CPB2;(r) F13A1 , ITGAX, C1QB, ITGAM, C1QC, C1QA, C2, ITGB2, C3, C4B, C1S, SERPINA1 , VSIG4, SERPINA5, CR2, A2M, and F2RL3;(s) HLA-DRB1 , HLA-DQA1 , HLA-DPA1 , and KLRD1 ;(t) HLA-DRA, HLA-G, B2M, HLA-DQA1 , HLA-DRB1 , HLA-DPB1 , CIITA, CD74, HLA-DOA, HLA-DMB, HLA-DMA, HLA-E, CD8B, HLA-DPA1 , CD4, TAP1 , IFI30, KLRD1 , HLA- DOB, TNF, CD8A, TAP2, and PSME1 ;(u) DAPP1 , RAC2, SYK, VAV2, and NFATC2;(v) RAC2, CD79A, CD79B, DAPP1 , LILRA4, SYK, VAV1 , BTK, BLNK, LILRB2, INPP5D, NFATC2, LILRB4, CD22, CARD11 , CR2, LILRA1, CD72 and RASGRP3(w) VAV2, CD40LG, CD28, PTPRC, and NFATC2; and(x) CD3D, PTPRC, CD28, LCK, CD3G, VAV1 , ICOS, CD8B, CD4, CD40LG, ITK, CD3E, TNF, CD8A, NFATC2, LCP2, CARD11 , and CTLA4.

26. A method of screening an rAAV composition for reduced AAV-induced inflammation comprising: (a) administering a dose of a test rAAV composition to a non-human subject; (b) obtaining a biological sample from the subject; (b) detecting expression levels of each of a set of genes in the biological sample; (d) comparing the detected expression levels for the test rAAV composition to expression levels detected for a dose of a control rAAV composition.

27. The method of claim 26, wherein the set of genes comprises at least two genes selected from A2M, B2M, BLNK, BTK, C1QA, C1QB, C1QC, C1 R, C1S, C2, C3, C4B, CARD11 , CASP8, CCL3, CCL4L2, CCL5, CD14, CD19, CD22, CD247, CD28, CD3D, CD3E, CD3G, CD4, CD40, CD40LG, CD72, CD74, CD79A, CD79B, CD86, CD8A, CD8B, CFHR4, CHUK, CIITA, CPB2, CR1 , CR2, CTLA4, CTSS, CXCL10, CXCL11 , CXCL8, CXCL9, CXCR3, DAPP1 , F12, F13A1 , F2RL3, FCGR1A, FCGR2A, FCGR3A, FYN, GRAP2, HCK, HLA- DMA, HLA-DMB, HLA-DOA, HLA-DOB, HLA-DPA1 , HLA-DPB1 , HLA-DQA1 , HLA-DRA, HLA-DRB1 , HLA-E, HLA-F, HLA-G, HSP90AA1 , ICOS, IFI30, IFITIvll, IFITM1 , IFNG,IKBKB, IKBKE, IKBKG, IL10, IL12A, IL12B, IL1B, IL6, INPP5D, IRF7, ITGAM, ITGAX, ITGB2, ITK, IY96, KLRC2, KLRD1, KPK3CD, LAT, LCK, LCP2, LGMN, LILRA1 , LILRA4, LILRB2, LILRB4, LR2, LY97, MAP3K7, MAPK13, MP3K7, MYD88, NCF1, NFATC2, PAK5, PDCD1, PIK3AP1, PIK3CD, PLA2G4B, PLA2G4C, PLAU, PLCG2, PLD1, PRKCD, PROC, PSME1, PTPRC, RAC2, RASGRP3, SCIN, SERPINA1, SERPINA5, SERPINE1, STAT1, SYK, TAP1, TAP2, TLR1, TLR4, TLR7, TLR8, TLR10, TNF, VASP, VAV1, VAV2, VSIG4, WAS, WAV1 , and ZAP70.

28. The method of any one of claims 26-27, wherein the set of genes is selected from:(a) CXCL9, CXCL10, CXCL11, CCL5 and CCL3;(b) CXCL9, CXCL11, CXCL10, IL6, TNF, CCL5, CXCL8, CCL3, TLR8, CCL4L2, IY96, IL12A, IL12B, STAT1, IKBKE, IL1B, TLR1, CASP8, IKBKG, TLR4, IRF7, CD14, MYD88, CD86, IKBKB, KPK3CD, MP3K7, CHUK, and CD40;(c) CXCL10, CXCL9, CCL5, CCL4L2, and MYD88;(d) CXCL10, CXCL9, CCL5, CXCL11, IL1B, TLR8, CCL4L2, CD86, TLR2, STAT1, TNF, LY97, TLR4, IRF7, TLR7 and IKBKE;(e) PLA2G4C, PRKCG, RAC2, PTPRC, FCGR3A, FCGR2A, SYK, and FCGR1A;(f) PLA2G4C, PTPRC, RAC2, LAT, NCF1, FCGR3A, WAV1, SYK, PLA2G4B, WAS, PLCG2, HCK, FCGR2A, INPP5D, FCGR2A, INPP5D, FCGR1A, PIK3CD, PRKCD, and VASP;(g) RAC2, SYK, VAV2, FCGR3A and PTPRC;(h) RAC2, FCGR2A, PTPRC, FCGR3A, SYK, VAV1 , NCF1 , PLA2G4C, WAS, SCIN, FCGR1A, INPP5D, HCK and PLD1;(i) C1R, ITGAM, C4B, F13A1, ITGB2 and C1QC;(j) SERPINA5, CR1, CR2, C4B, ITGAX, PROC, ITGAM, C3, F12, F13A1, ITGB2, SERPINE1, PLAU, C1S, C1QC, CFHR4, CIQAand C1R;(k) ITGAX, FGG and CPB2;(l) F13A1, ITGAX, C1QB, ITGAM, C1QC, C1QA, C2, ITGB2, C3, C4B, C1S, SERPINA1, VSIG4, SERPINA5, CR2, A2M, and F2RL3;(m)HLA-DOB, KLRC2, CD8B, TAP1, HLA-DRA, CTSS, HLA-DMB, IFNG, CD8Aand LGMN;(n) HLA-DOB, KLRD1, TNF, IFNG, TAP1, HLA-G, CIITA, KLRC2, CD8B, TAP2, HLA-E, CD8A, CTSS, HLA-DPB1, HLA-DPA1, HLA-DRB1 , HLA-DMB, HLA-DRA, HLA-DOA, HLA-F, CD74, HLA-DMA, IFI30, PSME1, HSP90AA1, and LGMN;(o) HLA-DRB1, HLA-DQA1, HLA-DPA1, and KLRD1;(p) HLA-DRA, HLA-G, B2M, HLA-DQA1, HLA-DRB1 , HLA-DPB1 , CIITA, CD74, HLA-DOA, HLA-DMB, HLA-DMA, HLA-E, CD8B, HLA-DPA1 , CD4, TAP1, IFI30, KLRD1, HLA- DOB, TNF, CD8A, TAP2, and PSME1;(q) CD79A, DAPP1 , CD79B, CARD11 , RAC2, BLNK, BTK, IFITIvll, and SYK;(r) CD79A, CD19, CARD11, CD79B, DAPP1 , CR2, LILRA4, CD22, RAC2, BTK, LILRA1 , BLNK, IFITM1 , LILRB2, LILRB4, VAV1 , SYK, PIK3AP1 , PLCG2, INPP5D, CD72, IKBKG, IKBKB, PIK3CD, and CHUK;(s) DAPP1 , RAC2, SYK, VAV2, and NFATC2; and(t) RAC2, CD79A, CD79B, DAPP1 , LILRA4, SYK, VAV1 , BTK, BLNK, LILRB2, INPP5D, NFATC2, LILRB4, CD22, CARD11 , CR2, LILRA1, CD72 and RASGRP3(u) LCK, CD40LG, CD3D, CD28, CD3E, CARD11 , PAK5, CD247, CD8B, ITK, PTPRC, LCP2, IFNG, CD8A, and FYN;(v) LCK, CD3D, CARD11 , CD28, TNF, ZAP70, CD40LG, CD3E, CD3G, PTPRC, ITK, IFNG, ICOS, PDCD1 , LAT, LCP2, GRAP2, IL10, CD8B, CD8A, CD247, VAV1 , PAK5, IKBKG, IKBKB, PIK3CD, MAPK13, MAP3K7, and CHUK;(w) VAV2, CD40LG, CD28, PTPRC, and NFATC2; and(x) CD3D, PTPRC, CD28, LCK, CD3G, VAV1 , ICOS, CD8B, CD4, CD40LG, ITK, CD3E, TNF, CD8A, NFATC2, LCP2, CARD11 , and CTLA4.

29. The method of any one of claims 26-28, wherein the test rAAV composition and control rAAV compositions are the same, and the dose of the test rAAV composition is different from the dose of the control rAAV composition.

30. The method of any one of claims 26-28, wherein the test rAAV composition and control rAAV compositions are different, and the dose of the test rAAV composition is the same as the dose of the control rAAV composition.

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