SIRPα-αCD123 fusion antibodies targeting CD123 in conjunction with CD47 blockade enhance the clearance of AML-initiating cells

BACKGROUND Acute myeloid leukaemia (AML) stem cells (LSCs) cause disease relapse. The CD47 \textquotedbldon't eat me signal\textquotedbl is upregulated on LSCs and contributes to immune evasion by inhibiting phagocytosis through interacting with myeloid-specific signal regulatory protein alpha (SIRPα). Activation of macrophages by blocking CD47 has been successful, but the ubiquitous expression of CD47 on healthy cells poses potential limitations for such therapies. In contrast, CD123 is a well-known LSC-specific surface marker utilized as a therapeutic target. Here, we report the development of SIRPα-αCD123 fusion antibodies that localize the disruption of CD47/SIRPα signalling to AML while specifically enhancing LSC clearance. METHODS SIRPα-αCD123 antibodies were generated by fusing the extracellular domain of SIRPα to an αCD123 antibody. The binding properties of the antibodies were analysed by flow cytometry and surface plasmon resonance. The functional characteristics of the fusion antibodies were determined by antibody-dependent cellular phagocytosis and antibody-dependent cellular cytotoxicity assays using primary AML patient cells. Finally, an in vivo engraftment assay was utilized to assess LSC targeting. RESULTS SIRPα-αCD123 fusion antibodies exhibited increased binding and preferential targeting of CD123+ CD47+ AML cells even in the presence of CD47+ healthy cells. Furthermore, SIRPα-αCD123 fusion antibodies confined disruption of the CD47-SIRPα axis locally to AML cells. In vitro experiments demonstrated that SIRPα-αCD123 antibodies greatly enhanced AML cell phagocytosis mediated by allogeneic and autologous macrophages. Moreover, SIRPα-αCD123 fusion antibodies efficiently targeted LSCs with in vivo engraftment potential. CONCLUSIONS SIRPα-αCD123 antibodies combine local CD47 blockade with specific LSC targeting in a single molecule, minimize the risk of targeting healthy cells and efficiently eliminate AML LSCs. These results validate SIRPα-αCD123 antibodies as promising therapeutic interventions for AML

Accumulation of mutations in antibody and CD8 T cell epitopes in a B cell depleted lymphoma patient with chronic SARS-CoV-2 infection

Antibodies against the spike protein of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) can drive adaptive evolution in immunocompromised patients with chronic infection. Here we longitudinally analyze SARS-CoV-2 sequences in a B cell-depleted, lymphoma patient with chronic, ultimately fatal infection, and identify three mutations in the spike protein that dampen convalescent plasma-mediated neutralization of SARS-CoV-2. Additionally, four mutations emerge in non-spike regions encoding three CD8 T cell epitopes, including one nucleoprotein epitope affected by two mutations. Recognition of each mutant peptide by CD8 T cells from convalescent donors is reduced compared to its ancestral peptide, with additive effects resulting from double mutations. Querying public SARS-CoV-2 sequences shows that these mutations have independently emerged as homoplasies in circulating lineages. Our data thus suggest that potential impacts of CD8 T cells on SARS-CoV-2 mutations, at least in those with humoral immunodeficiency, warrant further investigation to inform on vaccine design

Immunmonitoring unter Tyrosinkinaseinhibitor-Behandlung im Maus-Tumormodell

Hintergrund: Tyrosinkinaseinhibitoren (TKIs) stellen einen neuen Weg einer zielgerichteten Therapie in der Onkologie dar. Einige davon werden bereits erfolgreich in der Klinik eingesetzt, so zum Beispiel der TKI Imatinib. Imatinib blockiert zielgerichtet die Tyrosinkinase bcr-abl, die in vielen Fällen ursächlich für die CML ist. In einer Vorarbeit wurde deren Auswirkung und auch die des TKIs Nilotinib auf das Immunsystem untersucht. Ebenso wurde eine Steigerung des therapeutischen Erfolgs durch die Kombination mit dem Zytokin IL-2 erreicht. Ziel dieser Arbeit war es, denselben Effekt auf die Tumorreduktion mithilfe der TKIs Sorafenib und Dasatinib nachzuweisen, sowie auch deren Auswirkung auf bestimmte Zellen des Immunsystems zu analysieren. Hauptaugenmerk liegt hierbei auf den NK-Zellen und deren Subpopulationen. Methoden: Für die in vivo Versuche diente das B16F10 Melanom Modell, in welchem die Mäuse nach Tumorinfiltration jeweils für 10 bis 13 Tage mit einem TKI behandelt wurden. Einerseits wurde die Zahl der sichtbaren Lungenmetastasen ausgewertet und andererseits erfolgte eine durchflusszytometrische Analyse der Immunzellen. Ergebnisse: Letztendlich konnte durch diese Arbeit gezeigt werden, dass der TKI Sorafenib eine vergleichbare Tumorreduktion erzielt, wie das in den Vorarbeiten bereits für Nilotinib gezeigt werden konnte. Diesen Effekt konnte man wiederum mit IL-2 steigern. Der TKI Dasatinib hingegen erwies sich als kaum antitumoral in diesem Modell. Ferner wurde beobachtet, dass reife NK-Zellen unter Kombinationstherapie von Nilotinib und Sorafenib mit jeweils IL-2 zum Tumor migrieren. Die Bedeutung der NK-Zellen bei der antitumoralen Wirkung zeigte sich auch in verschiedenen Untersuchungen mit Knockout-Mäusen, insbesondere mit solchen, die kein IFN-γ bilden. Schlussfolgerung: Es konnte eine signifikante antitumorale Aktivität von Sorafenib, nicht jedoch von Dasatinib, in Kombination mit IL-2 gezeigt werden. Dieser Effekt scheint stark von IFN-γ abhängig zu sein, was zu einem wesentlichen Teil auf die Stimulation reifer NK-Zellen zurückzuführen ist.Background: Tyrosine kinase inhibitors (TKIs) build a new way of target-directed therapies in oncology. Some of them are used successfully in clinic, for example the TKI Imatinib. Imatinib blocks specifically the tyrosine kinase brc-abl, which is mostly the cause of CML. In previous work the impact of Imatinib and Nilotinib on the immune system has been shown. Furthermore, an increase of the therapeutical success could be achieved by the combination with the cytokine IL-2. Aim of the work was to investigate the effect on tumor reduction induced by the TKIs Sorafenib and Dasatinib, and to analyse their impact on specific immune cells. The main focus was the role of NK cells and their subpopulations. Methods: The in vivo experiments have been performed using the B16F10 melanoma model, in which mice were treated with a TKI for 10 to 13 days after injection of tumor cells. Not only the amount of apparent lung metastases, but also flow cytometric analysis of the immune cells have been performed. Results: Finally, our results demonstrate that Sorafenib had a similar effect on tumor reduction as Nilotinib. This effect was further increased by additional treatment with IL-2. In contrast, the TKI Dasatinib did not show any antitumoral effect. In addition, we observed that mature NK cells migrated to the tumor while treated with either Nilotinib or Sorafenib each plus IL-2. Additional experiments using different types of knockout mice, especially with a defect to build IFN-γ, showed the importance of NK cells for the described antitumoral effect. Conclusion: Our data demonstrate that Sorafenib, but not Dasatinib, plus IL-2 has significant antitumoral activity. The antitumoral effect seemed to be dependent on IFN-γ that is mainly produced by mature NK cells

T cells armed with C-X-C chemokine receptor type 6 enhance adoptive cell therapy for pancreatic tumours

The efficacy of adoptive cell therapy for solid tumours is hampered by the poor accumulation of the transferred T cells in tumour tissue. Here, we show that the forced expression of the C-X-C chemokine receptor type 6 (CXCR6, whose ligand is highly expressed by human and murine pancreatic cancer cells and by tumour-infiltrating immune cells) in antigen-specific T cells enhanced the recognition and lysis of pancreatic cancer cells and the efficacy of adoptive cell therapy for pancreatic cancer. In mice with subcutaneous pancreatic tumours treated with T cells with either a transgenic T-cell receptor or a murine chimeric antigen receptor targeting the tumour-associated antigen epithelial cell-adhesion molecule, and in mice with orthotopic pancreatic tumours or patient-derived xenografts treated with T cells expressing a chimeric antigen receptor targeting mesothelin, the T cells exhibited enhanced intratumoral accumulation, exerted sustained antitumoral activity and prolonged animal survival only when co-expressing CXCR6. Arming tumour-specific T cells with tumour-specific chemokine receptors may represent a promising strategy for the realization of adoptive cell therapy for solid tumours