Preclinical studies testing TCR T cells for AML
| Reference . | Target epitope . | HLA . | Donor pool . | Pool type . | Specific stimulation . | Isolation . | Sequencing . | TCR modification . | Additional elements . | Transduction . | Recipient cells . | Testing methods . | Testing against . |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Overexpressed antigens | |||||||||||||
| Ruggerio et al36 | WT137-45 (VLDFAPPGA) | A∗02:01 | Healthy | Not specified | Direct peptide loading | CD137 | RACE PCR | - | CRISPR/Cas9 knockout of endogenous TCR | LV/AAV | Healthy donors Patient T cell | Flow cytometry cytokine release and cytotoxicity assays Live-cell imaging Luciferase assay | Cell lines, modified HLA expression (K562, 697, OCI-AML3) EBV-LCLs Peptide-loaded T2 Primary AML Healthy cells PDX NOD mouse model |
| Van Amerongen et al37 | WT1 (VLDFAPPGA) (ALLPAVPSL) (VLDFAPPGASAY) (TPYSSDNLY) | A∗02:01 A∗02:01 A∗01:01 B∗35:01 | Healthy | Allo | - | Multimer single-cell clone | PCR | Murine constant Cysteine modified | - | RV | Healthy donors | ELISA Chromium 51 | Cell lines, modified HLA/antigen expression, peptide loaded (Raji) EBV-LCLs Healthy cells |
| Lahman et al38 | WT137-45 (VLDFAPPGA) | A∗02:01 | Healthy | Not specified | Peptide-loaded DCs | Tetramer | RACE PCR | - | CD8 coreceptor | LV | Healthy HLA-A∗02:01 donors | Flow cytometry intracellular cytokine release assay | Primary AML Cell lines, modified HLA/antigen expression (K562) PHA blasts, peptide loaded EBV-LCLs CDX NOD and NSG mouse model |
| Xue et al39 | WT1126-134 (RMFPNAPYL) | A∗02:01 | Healthy | Allo | Peptide-loaded T2 cells | Vβ2+ and CD8+ (magnetic bead and FACS) | PCR | - | - | RV | HLA-A2+ Healthy Donor T cells Patient T cells | Chromium 51 IFNγ release | T2 peptide–loaded cells Cell lines (697, BV173, Lama81, Kyo-1) CDX NOD/SCID mouse model |
| Arber et al40 | Survivin96-10497M (LMLGEFLKL) | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | Single-cell clone testing | 5′ RACE PCR | Murine constant | - | RV | Healthy Donor CD8+ cells | Chromium 51 ELISPOT CFU assay | Cell lines (BV173, U266, K562, HL-60) Primary AML CDX NSG mouse model |
| Sandri et al41,42 | TERT865–873 | A∗02:01 | Patients (B-CLL) Mouse CTLs | Auto | Peptide-loaded DCs | Single-cell clone testing | PCR | - | - | RV | Healthy donor PBMCs | ELISA flow cytometry cytotoxicity activity assay | Cell lines (THP1) Peptide-loaded PBMCs CDX NOG mouse model |
| Depreter et al43 | TARP(P5L)4–13 | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | Single-cell clone testing | 5′ RACE PCR | Murine constant | - | LV/RV | Healthy donor CD8+ cells | Flow cytometry cytokine release and cytotoxicity assays Chromium 51 Luciferase assay | Cell lines, modified HLA/antigen expression (THP1, LNCaP, KG1a, Molm13, OCI-AML3, HL60) Primary samples |
| Cancer testes antigens | |||||||||||||
| Amir et al44 | PRAME425-433 (SLLQHLIGL) | A∗02:01 | Patient donors (after HSCT) | Auto- and allo-HSCT | - | Tetramer single-cell sorted | RT-PCR | Cysteine modified | - | RV | EBV-specific T cells | ELISA Chromium 51 | Peptide-loaded T2 cell lines, modified HLA/antigen expression (K562, COS) EBV-LCLs Primary samples Healthy cell subsets |
| Kang et al45 Jager et al46 | NY-ESO-1157-165 | A∗02:01 | Healthy | Auto | Primary tumor cell line | Not specified | Not specified | Cysteine-modified CDR3a dual aa substitution | - | LV | Healthy donor PBMCs | ELISA, ELISPOT LDH cytotoxicity assay | Cell lines (U937, HL60, Kasumi-1) CDX NCG mouse model |
| Nagai et al47 | AURKA207-215 (YLILEYAPL) | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | Not specified | 5′ RACE PCR | - | - | RV | Healthy donor T cells | ELISA, ELISPOT Chromium 51 CSFE proliferation assay Luciferase reporter TCR signaling assay | Cell lines, modified HLA expression (GANMO-1, MEG01, KAZZ, OUN-1) Primary AML samples CDX NOG mouse model |
| Spranger et al48 | HMMR (MSFPKAPL) | A∗02:01 | Healthy | Allo | RNA-loaded DCs | CD137 single-cell clones | PCR | Murine constant | - | RV | Healthy donor T cells | Chromium 51 CFU assay | Cell line, modified HLA expression (K562, THP1) Primary AML CDX NSG mouse model HSCs |
| MiHA | |||||||||||||
| Van Loenen et al49 | HA-1 (VLHDDLLEA) | A∗02:01 | Healthy | Not specified | Not specified | Not specified | Not specified | Cysteine modifications | - | RV | Allo-restricted healthy donor T cells | ELISA Cytotoxicity assays | EBV-LCLs Primary AML |
| Dossa et al50 | HA-1 (VLHDDLLEA) | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | Single-cell clones | RACE PCR | Cysteine modifications | CD8 coreceptor iCasp9 safety switch | LV | Healthy donor T cells | Chromium 51 Intracellular cytokine release and CD107 degranulation flow cytometry | T2 peptide–loaded cells Primary AML |
| Pilunov et al51 | HA-1 | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | CD137/multimter | RACE PCR | Murine constant Cysteine modifications | CRISPR-Cas9 knock down of endogenous TCR | LV | Healthy donor CD8+ cells | Fluorescent reporter J76 cell line assay ELISA Flow cytometry cytotoxicity assay | Primary AML cell lines, modified HLA expressed, peptide loaded (K562) Healthy and patient PBMCs |
| Neoantigens | |||||||||||||
| Van der Lee et al52 | mutNPM1 (CLAVEEVSL) | A∗02:01 | Healthy (unsuccessful with AML patients) | Auto | - | Tetramer single-cell clones | PCR | Murine constant Cysteine modification | - | RV | Healthy PBMCs, later CD4/8 purified | ELISA Chromium 51 | Peptide-loaded T2 cells Primary AML CDX NSG mouse model |
| Van der Lee et al53 | mutNPM1 (AVEEVSLRK) | A∗11:01 | Healthy | Auto | - | Tetramer single-cell clones | PCR | Murine constant | - | RV | Healthy PBMCs, later CD4/8 purified | ELISA Chromium 51 | Peptide-loaded T2 cells EBV-LCLs Cell lines, modified HLA expression (K562, OCI-AML2, OCI-AML3) Primary AML CDX NSG mouse model |
| Biernacki et al54 | CBFB-MYH11 fusion (REEMEVHEL) | B∗40:01 | Healthy | Auto | Peptide-loaded DCs | Single-cell clones | RACE PCR | Cysteine modifications | - | LV | Healthy donor T cells | Chromium 51 Flow cytometry–based cytotoxicity and CD107 degranulation assays | Peptide-loaded LCLs Cell lines, modified HLA/antigen expression (NB4, ME-1) PDX MISTRG mouse model |
| Biernacki et al55 | U2AF1Q157R (DFREACCRR) | A∗33:01/03 | Healthy | Auto | Peptide-loaded DCs | Single-cell clones | RACE PCR | Murine constant Cysteine modifications | CRISPR-Cas9 knock down of endogenous TCR | LV | Healthy donor T cells | Chromium 51 Flow cytometry–based cytotoxicity and CD107 degranulation assays | Peptide-loaded LCLs Cell lines, modified antigen expression (TF-1) Primary AML Healthy PBSCs/MCs CDX NSG mouse model |
| Giannakopoulou et al56 | mutFLT3 - D835Y (YIMSDSNYV) | A2 | Healthy | Auto | RNA-loaded DCs | Multimer single-cell clones | RT-PCR | Murine constant Cysteine modifications | - | RV | Healthy donor PBMCs | ELISA Flow cytometry–based activity and cytotoxicity assays | Cell lines, modified HLA expression, peptide-loaded (K562) EBV-LCLs Primary AML Healthy A2+ cells CDX and PDX NSG mouse model |
| Reference . | Target epitope . | HLA . | Donor pool . | Pool type . | Specific stimulation . | Isolation . | Sequencing . | TCR modification . | Additional elements . | Transduction . | Recipient cells . | Testing methods . | Testing against . |
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Overexpressed antigens | |||||||||||||
| Ruggerio et al36 | WT137-45 (VLDFAPPGA) | A∗02:01 | Healthy | Not specified | Direct peptide loading | CD137 | RACE PCR | - | CRISPR/Cas9 knockout of endogenous TCR | LV/AAV | Healthy donors Patient T cell | Flow cytometry cytokine release and cytotoxicity assays Live-cell imaging Luciferase assay | Cell lines, modified HLA expression (K562, 697, OCI-AML3) EBV-LCLs Peptide-loaded T2 Primary AML Healthy cells PDX NOD mouse model |
| Van Amerongen et al37 | WT1 (VLDFAPPGA) (ALLPAVPSL) (VLDFAPPGASAY) (TPYSSDNLY) | A∗02:01 A∗02:01 A∗01:01 B∗35:01 | Healthy | Allo | - | Multimer single-cell clone | PCR | Murine constant Cysteine modified | - | RV | Healthy donors | ELISA Chromium 51 | Cell lines, modified HLA/antigen expression, peptide loaded (Raji) EBV-LCLs Healthy cells |
| Lahman et al38 | WT137-45 (VLDFAPPGA) | A∗02:01 | Healthy | Not specified | Peptide-loaded DCs | Tetramer | RACE PCR | - | CD8 coreceptor | LV | Healthy HLA-A∗02:01 donors | Flow cytometry intracellular cytokine release assay | Primary AML Cell lines, modified HLA/antigen expression (K562) PHA blasts, peptide loaded EBV-LCLs CDX NOD and NSG mouse model |
| Xue et al39 | WT1126-134 (RMFPNAPYL) | A∗02:01 | Healthy | Allo | Peptide-loaded T2 cells | Vβ2+ and CD8+ (magnetic bead and FACS) | PCR | - | - | RV | HLA-A2+ Healthy Donor T cells Patient T cells | Chromium 51 IFNγ release | T2 peptide–loaded cells Cell lines (697, BV173, Lama81, Kyo-1) CDX NOD/SCID mouse model |
| Arber et al40 | Survivin96-10497M (LMLGEFLKL) | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | Single-cell clone testing | 5′ RACE PCR | Murine constant | - | RV | Healthy Donor CD8+ cells | Chromium 51 ELISPOT CFU assay | Cell lines (BV173, U266, K562, HL-60) Primary AML CDX NSG mouse model |
| Sandri et al41,42 | TERT865–873 | A∗02:01 | Patients (B-CLL) Mouse CTLs | Auto | Peptide-loaded DCs | Single-cell clone testing | PCR | - | - | RV | Healthy donor PBMCs | ELISA flow cytometry cytotoxicity activity assay | Cell lines (THP1) Peptide-loaded PBMCs CDX NOG mouse model |
| Depreter et al43 | TARP(P5L)4–13 | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | Single-cell clone testing | 5′ RACE PCR | Murine constant | - | LV/RV | Healthy donor CD8+ cells | Flow cytometry cytokine release and cytotoxicity assays Chromium 51 Luciferase assay | Cell lines, modified HLA/antigen expression (THP1, LNCaP, KG1a, Molm13, OCI-AML3, HL60) Primary samples |
| Cancer testes antigens | |||||||||||||
| Amir et al44 | PRAME425-433 (SLLQHLIGL) | A∗02:01 | Patient donors (after HSCT) | Auto- and allo-HSCT | - | Tetramer single-cell sorted | RT-PCR | Cysteine modified | - | RV | EBV-specific T cells | ELISA Chromium 51 | Peptide-loaded T2 cell lines, modified HLA/antigen expression (K562, COS) EBV-LCLs Primary samples Healthy cell subsets |
| Kang et al45 Jager et al46 | NY-ESO-1157-165 | A∗02:01 | Healthy | Auto | Primary tumor cell line | Not specified | Not specified | Cysteine-modified CDR3a dual aa substitution | - | LV | Healthy donor PBMCs | ELISA, ELISPOT LDH cytotoxicity assay | Cell lines (U937, HL60, Kasumi-1) CDX NCG mouse model |
| Nagai et al47 | AURKA207-215 (YLILEYAPL) | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | Not specified | 5′ RACE PCR | - | - | RV | Healthy donor T cells | ELISA, ELISPOT Chromium 51 CSFE proliferation assay Luciferase reporter TCR signaling assay | Cell lines, modified HLA expression (GANMO-1, MEG01, KAZZ, OUN-1) Primary AML samples CDX NOG mouse model |
| Spranger et al48 | HMMR (MSFPKAPL) | A∗02:01 | Healthy | Allo | RNA-loaded DCs | CD137 single-cell clones | PCR | Murine constant | - | RV | Healthy donor T cells | Chromium 51 CFU assay | Cell line, modified HLA expression (K562, THP1) Primary AML CDX NSG mouse model HSCs |
| MiHA | |||||||||||||
| Van Loenen et al49 | HA-1 (VLHDDLLEA) | A∗02:01 | Healthy | Not specified | Not specified | Not specified | Not specified | Cysteine modifications | - | RV | Allo-restricted healthy donor T cells | ELISA Cytotoxicity assays | EBV-LCLs Primary AML |
| Dossa et al50 | HA-1 (VLHDDLLEA) | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | Single-cell clones | RACE PCR | Cysteine modifications | CD8 coreceptor iCasp9 safety switch | LV | Healthy donor T cells | Chromium 51 Intracellular cytokine release and CD107 degranulation flow cytometry | T2 peptide–loaded cells Primary AML |
| Pilunov et al51 | HA-1 | A∗02:01 | Healthy | Auto | Peptide-loaded DCs | CD137/multimter | RACE PCR | Murine constant Cysteine modifications | CRISPR-Cas9 knock down of endogenous TCR | LV | Healthy donor CD8+ cells | Fluorescent reporter J76 cell line assay ELISA Flow cytometry cytotoxicity assay | Primary AML cell lines, modified HLA expressed, peptide loaded (K562) Healthy and patient PBMCs |
| Neoantigens | |||||||||||||
| Van der Lee et al52 | mutNPM1 (CLAVEEVSL) | A∗02:01 | Healthy (unsuccessful with AML patients) | Auto | - | Tetramer single-cell clones | PCR | Murine constant Cysteine modification | - | RV | Healthy PBMCs, later CD4/8 purified | ELISA Chromium 51 | Peptide-loaded T2 cells Primary AML CDX NSG mouse model |
| Van der Lee et al53 | mutNPM1 (AVEEVSLRK) | A∗11:01 | Healthy | Auto | - | Tetramer single-cell clones | PCR | Murine constant | - | RV | Healthy PBMCs, later CD4/8 purified | ELISA Chromium 51 | Peptide-loaded T2 cells EBV-LCLs Cell lines, modified HLA expression (K562, OCI-AML2, OCI-AML3) Primary AML CDX NSG mouse model |
| Biernacki et al54 | CBFB-MYH11 fusion (REEMEVHEL) | B∗40:01 | Healthy | Auto | Peptide-loaded DCs | Single-cell clones | RACE PCR | Cysteine modifications | - | LV | Healthy donor T cells | Chromium 51 Flow cytometry–based cytotoxicity and CD107 degranulation assays | Peptide-loaded LCLs Cell lines, modified HLA/antigen expression (NB4, ME-1) PDX MISTRG mouse model |
| Biernacki et al55 | U2AF1Q157R (DFREACCRR) | A∗33:01/03 | Healthy | Auto | Peptide-loaded DCs | Single-cell clones | RACE PCR | Murine constant Cysteine modifications | CRISPR-Cas9 knock down of endogenous TCR | LV | Healthy donor T cells | Chromium 51 Flow cytometry–based cytotoxicity and CD107 degranulation assays | Peptide-loaded LCLs Cell lines, modified antigen expression (TF-1) Primary AML Healthy PBSCs/MCs CDX NSG mouse model |
| Giannakopoulou et al56 | mutFLT3 - D835Y (YIMSDSNYV) | A2 | Healthy | Auto | RNA-loaded DCs | Multimer single-cell clones | RT-PCR | Murine constant Cysteine modifications | - | RV | Healthy donor PBMCs | ELISA Flow cytometry–based activity and cytotoxicity assays | Cell lines, modified HLA expression, peptide-loaded (K562) EBV-LCLs Primary AML Healthy A2+ cells CDX and PDX NSG mouse model |
aa, amino acid; AAV, adeno-associated virus; B-CLL, B-cell chronic lymphocytic leukaemia; Cas9, CRISPR-associated protein 9; iCas9, inducible caspase 9; CDX, cell line-derived xenograft; CFU, colony-forming unit; CTLs, cytotoxic T lymphocytes; DCs, dendritic cells; EBV, Epstein-Barr virus; ELISA, enzyme-linked immunosorbent assay; ELISPOT, enzyme-linked immunospot; FACS, fluorescence-activated cell sorting; INFγ, interferon-γ; LCLs, lymphoblastoid cell lines; LDH, lactate dehydrogenase; LV, lentivirus; NSG, NOD scid gamma; NOD/SCID, nonobese diabetic/severe combined immunodeficiency; NOG, NOD/shi-scid IL2γr(null); PDX, patient-derived xenograft; PHA, phytohemagglutinin; RACE, rapid amplification of complementary DNA ends; RT, reverse transcription; RV, retrovirus; PCR, polymerase chain reaction.