Recent reports suggest that induced neurons (iNs), but not induced pluripotent stem cell (iPSC)-derived neurons, largely preserve age-associated traits. Here, we report on the extent of preserved epigenetic and transcriptional aging signatures in directly converted induced neural stem cells (iNSCs). Employing restricted and integration-free expression of SOX2 and c-MYC, we generated a fully functional, bona fide NSC population from adult blood cells that remains highly responsive to regional patterning cues. Upon conversion, low passage iNSCs display a profound loss of age-related DNA methylation signatures, which further erode across extended passaging, thereby approximating the DNA methylation age of isogenic iPSC-derived neural precursors. This epigenetic rejuvenation is accompanied by a lack of age-associated transcriptional signatures and absence of cellular aging hallmarks. We find iNSCs to be competent for modeling pathological protein aggregation and for neurotransplantation, depicting blood-to-NSC conversion as a rapid alternative route for both disease modeling and neuroregeneration.

Head and neck squamous cell carcinomas (HNSCC) caused by infections with high-risk human papillomaviruses (HPV) are responsible for an increasing number of head and neck cancers, particularly in the oropharynx. Despite the significant biological differences between HPV-driven and HPV-negative HNSCC, treatment strategies are similar and not HPV targeted. HPV-driven HNSCC are known to be more sensitive to treatment, particularly to radiotherapy, which is at least partially due to HPV-induced immunogenicity. The development of novel therapeutic strategies that are specific for HPV-driven cancers requires tumor models that reflect as closely as possible the characteristics and complexity of human tumors and their response to treatment. Current HPV-positive cancer models lack one or more hallmarks of their human counterpart. This study presents the development of a new HPV16 oncoprotein-dependent tumor model in MHC-humanized mice, modeling the major biologic features of HPV-driven tumors and presenting HLA-A2-restricted HPV16 epitopes. Furthermore, this model was developed to be orthotopic (base of tongue). Thus, it also reflects the correct tumor microenvironment of HPV-driven HNSCC. The cancer cells are implanted in a manner that allows the exact control of the anatomical location of the developing tumor, thereby homogenizing tumor growth. In conclusion, the new model is suited to study HPV16-specific therapeutic vaccinations and other immunotherapies, as well as tumor-targeted interventions, such as surgery or radiotherapy, or a combination of all these modalities.

Current cancer therapies focus on targeting driver alterations responsible for tumorigenesis - either oncogenic point mutations or oncogenic driver events within large somatic copy number alterations (SCNAs). However, these alterations are often not actionable or are only present in a small subset of patients. Thus, new therapeutic targets are urgently needed. Apart from oncogenic drivers, SCNAs usually also comprise neutral bystander genes without an active role in neoplastic transformation. We hypothesized that passenger events, specifically in amplified regions, could be therapeutically exploited by providing actionable molecules on the cell surface. Using publicly available multi-omics data, we screened cell surface protein-coding genes for their genomic status and expression levels in liver cancer. This screen identified MPZL1 (Myelin protein zero-like 1), a gene amplified in 75% of hepatocellular carcinomas, accompanied by high mRNA expression in tumors compared to normal livers. MPZL1 is located on chromosome 1q, one of the most commonly amplified regions across several solid cancer types, and codes for a glycosylated cell surface receptor. We further validated MPZL1 protein expression in a wide range of human cancer entities (n=2038 samples) and normal tissues (n=163 samples) by immunohistochemistry and assigned expression scores analogous to the clinically implemented HER2-score. A high percentage of tumors showed scores 2 or 3 (e.g. 48% of HCCs or 89% of TNBCs), whereas healthy tissues were mostly negative or just faintly positive (scores 0 or 1). Moreover, in order to target MPZL1-expressing cells, we generated a highly specific monoclonal antibody directed to the extracellular domain of human MPZL1 protein and thoroughly characterized its binding capacity. Next, the scFv sequence of this antibody was used to generate a CAR (chimeric antigen receptor) construct targeting MPZL1. Luciferase-based cytotoxic assays showed that T cells transduced with this construct can kill various MPZL1-high human cancer cell lines in vitro (liver, breast and lung malignant cell lines, among others), whereas they fail to kill respective isogenic cell lines harboring knockout of MPZL1. Upon antigen-specific exposure, MPZL1-28ζ CAR-T cells underwent antigen-dependent proliferation and showed increased cytokine production (IFNγ, TNFα, IL-2, GZMB), further confirming their specificity. In summary, our findings reveal MPZL1 as a new target for treatment of 1q-amplified cancers and implement a novel immunotherapeutic strategy based on MPZL1-28ζ CAR-T cells. Furthermore, our work provides a proof of concept that passenger events within large chromosomal amplifications can serve as potent therapeutic targets and thus opens a new avenue for innovative approaches in anti-cancer drug development. Citation Format: Sonia Jiménez Vázquez, Christos Patsis, Luise Butthof, Joan Crous Masó, Hendrik Wiethoff, Matthias Gaida, Thomas Longerich, Rosella Pellegrino, Ilse Hofmann, Judith Feucht, Darjus Felix Tschaharganeh. Repurposing passenger amplifications for specific therapeutic targeting of solid cancers. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5003.

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