Compositions and methods for assessing immune response

Abstract

The present invention provides methods and compositions that are useful for assessing gene expression for tumor immune response profile of a sample. In particular, a target-specific primer panel is provided that allows for selective amplification of immune response target sequences in a sample. In one aspect, the invention relates to target-specific primers useful for selective amplification of one or more target sequences associated with immune response. In some aspects, amplified target sequences obtained using the disclosed methods, and compositions can be used in various processes including nucleic acid sequencing and used to detect the presence of genetic variants and/or expression levels of one or more targeted sequences associated with immune response.

1 - 20 . (canceled) 
     
     
         21 . A method for determining immune response activity in a biological sample, comprising:
 multiplex amplification of a plurality of target sequences from the biological sample, wherein the amplification comprises contacting at least a portion of the sample with a plurality of sets of primer pair reagents and a polymerase under amplification conditions, wherein the plurality of target sequences are selected from the group consisting of the following functions: checkpoint pathways, T cell related signaling pathways, markers of tumor infiltrating lymphocytes (TILs), and tumor markers, and housekeeping genes, and wherein the plurality of primer pair reagents are selected from the group consisting of SEQ ID NOs: 399-1194, thereby producing amplified immune response target sequences; and   detecting a level of expression of each of the plurality of immune response target sequences, wherein a change in the level of expression of one or of the immune response target sequences as compared with a control sample determines a change in immune response activity in the sample.   
     
     
         22 . The method of  claim 21 , wherein the immune response target sequences are from target genes selected from the group consisting of immune checkpoint pathways and targets; T and B cell signaling genes, markers of lymphocyte subsets, interferon signaling genes, cytokine signaling genes; tumor markers, tumor antigens, proliferation markers; and housekeeping genes. 
     
     
         23 . The method of  claim 22 , wherein the target genes are selected from the genes of CD63, CD69, CXCL1, KLRD1, HLA-DOB, CXCR5, IL12B, PTK7, CEACAM1, CXCL9, IL13, NT5E, VEGFA, ABCF1, D38, JAML, S100A8, MYC, IRF1, CCL22, CXCR2, IFIT1, IFIT2, CD68, M6PR, SH2D1A, ISG20, GBP1, TBP, STAT6, ID3, CX3CL1, KLRB1, TNFSF4, CD52, IL10RA, HLA-DOA, IFNB1, CCR5, IKZF3, STAT1, CD6, BRCA1, CORO1A, TBX21, KLRK1, CXCR6, PTEN, PMEL, DMBT1, IFI44L, LAPTM5, CD226, TNFSF13B, ICOS, CD160, TRIM29, LST1, ZBTB46, VTCN1, KREMEN1, PDCD1LG2, TUBB, CLEC4C, CD86, HAVCR2, GZMH, NFATC1, CD8B, BCL2, GADD45GIP1, CBLB, ITGA1, CD8A, IL2RA, EIF2AK2, MADCAM1, PTPN6, LRG1, ADGRE5, SH2D1B, ITGB2, HLA-DPA1, DGAT2, IGF1R TAGAP, LMNA, NCAM1, TIGIT, IL17F, HLA-F-AS1, CD247, CD79B, IDO2, IL4, TYROBP, BTLA, AKT1, IL2RG, POLR2A, ITGAX, IL1B, CSF2RB, DDX58, KIAA0101, CD274, LAMPS, TNFAIP8, FOXP3, IL12A, SAMHD1, SIT1, CD3E, ICOSLG, HGF, MELK, IGSF6, GNLY, TDO2, KRT7, HLA-E, HLADMA, LAMP1, NTN3, CD28, TARP, EGFR, CCR4, MAGEA3, BATF, KLRG1, IRS1, CSF1R, CTLA4, TNFSF18, POU2AF1, GZMA, PIK3CA, ITK, IFI27, EOMES, LCN2, CD80, CD83, CXCL13, MTOR, FCER1G, TFRC, RORC, MMP9, BST2, PIK3CD, FCGR2B, TNFRSF14, OAS3, GRAP2, CCNB2, MLANA, MAGEA12, VCAM1, CDKN3, NCR1, FAS, GZMB, IRF9, IFITM2, TNFSF14, HLA-B, SDHA, NRP1, EBB, EFNA4, PVR BUB1, SKAP2, PRF1, CCL20, TNFRSF18, CTSS, NKG7, ISG15, PDCD1, SNAI1, CXCL11, CIITA, IFI35, TNFSF9, TNFSF10, MMP2, EGR3, MAGEA1, CD163, IL6, KLRF1, B3GAT1, C1QA, OAS1, IKZF2, TLR9, KLF2, GUSB, NFKBIA, IL23A, HERC6, SLAMF8, IL15, TLR7, OAS2, HLA-DRA, CRTAM, MAGEC2, ICAM1, CD4, MAPK14, C1QB, NOTCH3, NCR3, STAT3, TLR8, CYBB, IKZF4, IFIH1, LCK, BCL2L11, ITGAM, ITGB7, JCHAIN, CD209, SLAMF7, IL10, IL1A, FCGR3A, IFNA17, EGR2, TOP2A, C10orf54, FOXM1, AXL, MS4A1, IFI6, CD3D, GPR18, CD3G, ZAP70, HMBS, IL7, IFIT3, RBI, PTGS2, TGFB1, NCF1, TWIST1, CA4, SELL, LILRB1, CD14, ALOX15B, PECAM1, NOS2, FASLG, CD44, ENTPD1, CMKLR1, CD53, TNF, CXCL8, CD40LG, HLA-F, GATA3, LYZ, ARG1, IL2RB, NECTIN2, MPO, CCR2, BRCA2, ADORA2A, G6PD, TAP1, MXI, HLA-DQB2, CD27, CD276, STAT4, PTPN7, PTPRC, PSMB9, CD244, CXCR4, MAPK1, TP63, IRF4, CCL3, CCL18, IL7R, HLA-DRB1, CEACAM8, CXCL10, CCL2, SRGN, CD19, ITGB1, IFITM1, CCL21, MRC1, PGF, ITGAL, ID2, CD22, CCL17, ITGAE, IL3RA, CCR7, CD1C, MAD2L1, PYGL, CD40, LY9, HLA-G, TLR3, CD48, STAT5A, FCRLA, BCL6, ZEB1, CCL5, IDOL IL18, TNFRSF9, HIF1A, HLA-DPB1, FOXO1, CD33, S100A9, HLA-DMB, HLAA, SNAI2, TNFRSF17, LRP1, MAGEA4, HLA-DQA1, CD1D, RPS6, MKI67, GZMK, CD79A, CD37, FUT4, AIF1, CCR1, PRDM1, CD47, CD74, LAGS, TNFRSF4, CD2, CCL4, BAGE, LEXM, CCR6, CD70, CDK1, CTAG1B, CTAG2, CX3CR1, GAGE1, GAGE121, GAGE12F, GAGE12J, GAGE2C, GAGE2A, GAGE2E, GAGE10, GAGE13, IKZF1, IL17A, IL2, IL21, IL22, KIR2DL2, KIR2DL3, MAGEA10, MIF, PTPRCAP, SSX2, TCF7, XAGE1B, CXCR3, FCGR1A, FCGR3B, FYB, HLA-C, HLA-DQA2, IFNG, KIR2DL1, KRT5, and PTPN11. 
     
     
         24 . The method of  claim 23 , wherein the target genes comprise the genes of CD63, CD69, CXCL1, KLRD1, HLA-DOB, CXCR5, IL12B, PTK7, CEACAM1, CXCL9, IL13, NT5E, VEGFA, ABCF1, D38, JAML, S100A8, MYC, IRF1, CCL22, CXCR2, IFIT1, IFIT2, CD68, M6PR, SH2D1A, ISG20, GBP1, TBP, STAT6, ID3, CX3CL1, KLRB1, TNFSF4, CD52, IL10RA, HLA-DOA, IFNB1, CCR5, IKZF3, STAT1, CD6, BRCA1, CORO1A, TBX21, KLRK1, CXCR6, PTEN, PMEL, DMBT1, IFI44L, LAPTM5, CD226, TNFSF13B, ICOS, CD160, TRIM29, LST1, ZBTB46, VTCN1, KREMEN1, PDCD1LG2, TUBB, CLEC4C, CD86, HAVCR2, GZMH, NFATC1, CD8B, BCL2, GADD45GIP1, CBLB, ITGA1, CD8A, IL2RA, EIF2AK2, MADCAM1, PTPN6, LRG1, ADGRE5, SH2D1B, ITGB2, HLA-DPA1, DGAT2, IGF1R TAGAP, LMNA, NCAM1, TIGIT, IL17F, HLA-F-AS1, CD247, CD79B, IDO2, IL4, TYROBP, BTLA, AKT1, IL2RG, POLR2A, ITGAX, IL1B, CSF2RB, DDX58, KIAA0101, CD274, LAMPS, TNFAIP8, FOXP3, IL12A, SAMHD1, SIT1, CD3E, ICOSLG, HGF, MELK, IGSF6, GNLY, TDO2, KRT7, HLA-E, HLADMA, LAMP1, NTN3, CD28, TARP, EGFR, CCR4, MAGEA3, BATF, KLRG1, IRS1, CSF1R, CTLA4, TNFSF18, POU2AF1, GZMA, PIK3CA, ITK, IFI27, EOMES, LCN2, CD80, CD83, CXCL13, MTOR, FCER1G, TFRC, RORC, MMP9, BST2, PIK3CD, FCGR2B, TNFRSF14, OAS3, GRAP2, CCNB2, MLANA, MAGEA12, VCAM1, CDKN3, NCR1, FAS, GZMB, IRF9, IFITM2, TNFSF14, HLA-B, SDHA, NRP1, EBB, EFNA4, PVR BUB1, SKAP2, PRF1, CCL20, TNFRSF18, CTSS, NKG7, ISG15, PDCD1, SNAI1, CXCL11, CIITA, IFI35, TNFSF9, TNFSF10, MMP2, EGR3, MAGEA1, CD163, IL6, KLRF1, B3GAT1, C1QA, OAS1, IKZF2, TLR9, KLF2, GUSB, NFKBIA, IL23A, HERC6, SLAMF8, IL15, TLR7, OAS2, HLA-DRA, CRTAM, MAGEC2, ICAM1, CD4, MAPK14, C1QB, NOTCH3, NCR3, STAT3, TLR8, CYBB, IKZF4, IFIH1, LCK, BCL2L11, ITGAM, ITGB7, JCHAIN, CD209, SLAMF7, IL10, IL1A, FCGR3A, IFNA17, EGR2, TOP2A, C10orf54, FOXM1, AXL, MS4A1, IFI6, CD3D, GPR18, CD3G, ZAP70, HMBS, IL7, IFIT3, RBI, PTGS2, TGFB1, NCF1, TWIST1, CA4, SELL, LILRB1, CD14, ALOX15B, PECAM1, NOS2, FASLG, CD44, ENTPD1, CMKLR1, CD53, TNF, CXCL8, CD40LG, HLA-F, GATA3, LYZ, ARG1, IL2RB, NECTIN2, MPO, CCR2, BRCA2, ADORA2A, G6PD, TAP1, MXI, HLA-DQB2, CD27, CD276, STAT4, PTPN7, PTPRC, PSMB9, CD244, CXCR4, MAPK1, TP63, IRF4, CCL3, CCL18, IL7R, HLA-DRB1, CEACAM8, CXCL10, CCL2, SRGN, CD19, ITGB1, IFITM1, CCL21, MRC1, PGF, ITGAL, ID2, CD22, CCL17, ITGAE, IL3RA, CCR7, CD1C, MAD2L1, PYGL, CD40, LY9, HLA-G, TLR3, CD48, STAT5A, FCRLA, BCL6, ZEB1, CCL5, IDOL IL18, TNFRSF9, HIF1A, HLA-DPB1, FOXO1, CD33, S100A9, HLA-DMB, HLAA, SNAI2, TNFRSF17, LRP1, MAGEA4, HLA-DQA1, CD1D, RPS6, MKI67, GZMK, CD79A, CD37, FUT4, AIF1, CCR1, PRDM1, CD47, CD74, LAGS, TNFRSF4, CD2, CCL4, BAGE, LEXM, CCR6, CD70, CDK1, CTAG1B, CTAG2, CX3CR1, GAGE1, GAGE121, GAGE12F, GAGE12J, GAGE2C, GAGE2A, GAGE2E, GAGE10, GAGE13, IKZF1, IL17A, IL2, IL21, IL22, KIR2DL2, KIR2DL3, MAGEA10, MIF, PTPRCAP, SSX2, TCF7, XAGE1B, CXCR3, FCGR1A, FCGR3B, FYB, HLA-C, HLA-DQA2, IFNG, KIR2DL1, KRT5, and PTPN11. 
     
     
         25 . The method of  claim 21 , wherein the plurality of immune response target sequences comprise a plurality of amplicon sequences selected from SEQ ID NOs: 1-398. 
     
     
         26 . The method of  claim 25 , wherein the plurality of target sequences comprises each of the amplicon sequences of SEQ ID NOs: 1-398. 
     
     
         27 . The method of  claim 26 , wherein the plurality of target sequences consists of each of the amplicon sequences of SEQ ID NOs: 1-398. 
     
     
         28 . The method of  claim 21 , wherein the plurality of primer pair reagents comprises each of the primer pairs of SEQ ID NOs: 399-1194. 
     
     
         29 . The method of  claim 28 , wherein the plurality of primer pair reagents consists of each of the primer pairs of SEQ ID NOs: 399-1194. 
     
     
         30 . The method of  claim 21 , wherein the one or more housekeeping genes are selected from ABCF1, G6PD, GUSB, HMBS, LMNA, LRP1, POLR2A, SDHA, TBP, TFRC, and TUBB. 
     
     
         31 . The method of  claim 21 , wherein the biological sample comprises a tumor, tissue surrounding a tumor, or both. 
     
     
         32 . The method of  claim 21 , wherein the biological sample and the control sample are from the same individual.