IntroductionThe cell line MC38 is a commonly used murine model for colorectal carcinoma. It has a high mutational burden, is sensitive to immune checkpoint immunotherapy and endogenous CD8+ T cell... Show moreIntroductionThe cell line MC38 is a commonly used murine model for colorectal carcinoma. It has a high mutational burden, is sensitive to immune checkpoint immunotherapy and endogenous CD8+ T cell responses against neoantigens have been reported. MethodsHere, we re-sequenced exomes and transcriptomes of MC38 cells from two different sources, namely Kerafast (originating from NCI/NIH, MC38-K) and the Leiden University Medical Center cell line collection (MC38-L), comparing the cell lines on the genomic and transcriptomic level and analyzing their recognition by CD8+ T cells with known neo-epitope specificity. ResultsThe data reveals a distinct structural composition of MC38-K and MC38-L cell line genomes and different ploidies. Further, the MC38-L cell line harbored about 1.3-fold more single nucleotide variations and small insertions and deletions than the MC38-K cell line. In addition, the observed mutational signatures differed; only 35.3% of the non-synonymous variants and 5.4% of the fusion gene events were shared. Transcript expression values of both cell lines correlated strongly (p = 0.919), but we found different pathways enriched in the genes that were differentially upregulated in the MC38-L or MC38-K cells, respectively. Our data show that previously described neoantigens in the MC38 model such as Rpl18(mut) and Adpgk(mut) were absent in the MC38-K cell line resulting that such neoantigen-specific CD8+ T cells recognizing and killing MC38-L cells did not recognize or kill MC38-K cells. ConclusionThis strongly indicates that at least two sub-cell lines of MC38 exist in the field and underlines the importance of meticulous tracking of investigated cell lines to obtain reproducible results, and for correct interpretation of the immunological data without artifacts. We present our analyses as a reference for researchers to select the appropriate sub-cell line for their own studies. Show less
Simple Summary: Acute myeloid leukemia (AML) is an aggressive hematological malignancy with poor prognosis. For AML relapses after chemotherapy, new and effective therapies are needed. In 30-35% of... Show moreSimple Summary: Acute myeloid leukemia (AML) is an aggressive hematological malignancy with poor prognosis. For AML relapses after chemotherapy, new and effective therapies are needed. In 30-35% of AMLs, a frameshift mutation in the nucleophosmin 1 gene (dNPM1) creates potential neoantigens that are attractive targets for immunotherapy. We previously isolated a T-cell receptor (TCR) that targets an HLA-A*02:01-binding dNPM1 neoantigen on primary AML. Here, we investigated whether AVEEVSLRK is another dNPM1 neoantigen that can be targeted by TCR gene transfer. We isolated various T-cells, cloned the HLA-A*11:01-restricted TCR from one T-cell clone and, upon transfer to CD8 cells, demonstrated targeting of dNPM1 primary AMLs in vitro. However, the TCR failed to mediate an anti-tumor effect in immunodeficient mice engrafted with dNPM1 OCI-AML3 cells. Our results demonstrate that AVEEVSLRK is an HLA-A*11:01-binding neoantigen on dNPM1 AML. Whether the isolated TCR is of sufficient affinity to treat patients remains uncertain.Acute myeloid leukemia (AML) is a hematological malignancy caused by clonal expansion of myeloid progenitor cells. Most patients with AML respond to chemotherapy, but relapses often occur and infer a very poor prognosis. Thirty to thirty-five percent of AMLs carry a four base pair insertion in the nucleophosmin 1 gene (NPM1) with a C-terminal alternative reading frame of 11 amino acids. We previously identified various neopeptides from the alternative reading frame of mutant NPM1 (dNPM1) on primary AML and isolated an HLA-A*02:01-restricted T-cell receptor (TCR) that enables human T-cells to kill AML cells upon retroviral gene transfer. Here, we isolated T-cells recognizing the dNPM1 peptide AVEEVSLRK presented in HLA-A*11:01. The TCR cloned from a T-cell clone recognizing HLA-A*11:01+ primary AML cells conferred in vitro recognition and lysis of AML upon transfer to CD8 cells, but failed to induce an anti-tumor effect in immunodeficient NSG mice engrafted with dNPM1 OCI-AML3 cells. In conclusion, our data show that AVEEVSLRK is a dNPM1 neoantigen on HLA-A*11:01+ primary AMLs. CD8 cells transduced with an HLA-A*11:01-restricted TCR for dNPM1 were reactive against AML in vitro. The absence of reactivity in a preclinical mouse model requires further preclinical testing to predict the potential efficacy of this TCR in clinical development. Show less
Tondini, E.; Arakelian, T.; Oosterhuis, K.; Camps, M.; Duikeren, S. van; Han, W.D.; ... ; Ossendorp, F. 2019
