Biology subjects

Lehner, M.

Publications and source records attributed to Lehner, M..

8 recordsLinked to original sources

bioRxiv2026

Aurora kinase A is a synthetic lethal target in FANCA-deficient cancers

Loss-of-function genomic alterations in FANCA occur across multiple cancer types, yet no molecularly tailored therapies have successfully exploited this potential vulnerability. Using complementary unbiased approaches, including a genome-wide CRISPR/Cas9 loss-of-function screen and a high-throughput drug screen in isogenic cancer cell-based models, we identified Aurora kinase A (AURKA) as a reproducible synthetic lethal target of FANCA-deficient cancers. Inhibition of AURKA induced chromosomal instability, micronucleation, and early G2/M arrest selectively in FANCA-deficient cells, consistent with an increased reliance on mitotic checkpoint control. Mechanistically, FANCA deficiency is associated with an elevated AURKA expression at both the transcriptomic and protein levels, and with an upregulation of mitotic spindle and G2/M checkpoint gene signatures. Analysis of large-scale cancer genomics datasets, including over 650,000 clinically sequenced tumors, confirms that FANCA is the most frequently altered Fanconi anemia pathway gene across cancers, and that Fanconi anemia-defective tumors exhibit an increased tumor mutational burden and genomic instability. Collectively, our findings point to AURKA inhibition as a promising precision treatment strategy in FANCA-deficient cancers and provide a rationale to further explore this strategy in the clinic.

cancer biology↗

bioRxiv2025

Blocking Minor Intron Splicing Disrupts DNA Repair and Overcomes Therapy Resistance in Prostate and Breast Cancer

The minor spliceosome (MiS) is a specialized RNA-processing machinery upregulated in cancer, promoting oncogene expression. We uncovered an adaptive resistance mechanism driven by secretion of extracellular vesicles enriched in U6atac snRNA, which amplifies MiS activity and promotes therapy resistance. Here, we show that U6atac snRNA, a crucial MiS component, reverses this process when depleted, revealing it as a druggable vulnerability in therapy-resistant prostate and breast cancers. U6atac knockdown triggers R-loop-mediated DNA damage while impairing repair by downregulating key DNA repair factors, disabling both homologous recombination and non-homologous end joining. This dual effect sensitizes prostate and breast tumors to PARP inhibitors, cisplatin, and radiation, independent of BRCA status. Across multiple in vitro and in vivo models, MiS targeting demonstrates tumor-selective activity with minimal toxicity. These findings position U6atac as a central regulator of genome stability and establish MiS targeting as a promising approach to potentiate genotoxic therapy and overcome resistance. Statement of significanceU6atac, a minor spliceosome component, is a crucial regulator of genome stability in cancer. Its knockdown triggers R-loop-driven DNA damage, downregulates DNA repair genes, and sensitizes tumors to DNA-damaging therapies while simultaneously blocking resistance mechanisms. Thus, minor spliceosome knockdown is a tumour-selective and broadly applicable therapeutic strategy.

cancer biology↗

bioRxiv2025

Engineering human protein switches for functional control of CARs and transcription factors via oral drug administration

While CAR T cells have revolutionized the treatment of certain hematologic malignancies, they can cause severe toxicities, which are expected to be exacerbated with next-generation CAR Ts engineered for improved proliferation, persistence, and efficacy. Therefore, regulatory systems are urgently needed to be able to control these living drugs directly in patients. Here, we engineered a molecular switch, in which the interaction of two human proteins is efficiently induced with the orally available and non-toxic drug A1120. We demonstrate the versatility of this switch by regulating CAR signaling and transcriptional activity in human T cells in vitro and in vivo. Both systems were tightly controlled in the absence of the drug but strongly activated upon administration of the small molecule. Since this switch enables the regulation of diverse systems including CARs and transcription factors, we anticipate that it represents an important step towards next-generation cellular therapies with improved safety and efficacy.

synthetic biology↗

bioRxiv2025

MiniCARbids: Minimalistic human binding domains specifically tailored to CAR T applications

Traditionally, chimeric antigen receptor (CAR) T cells employ single-chain variable fragments (scFvs) as binding entities. While scFvs represent a convenient option due to their broad availability, they also come with drawbacks, in particular their tendency to cluster and their relatively large size. Moreover, most scFvs used in the CAR field are of non-human origin, potentially causing immunogenicity. Therefore, we established an engineering platform for minimalistic CAR binding domains (miniCARbids), which combine several critical advantages: (i) human origin, (ii) small size, (iii) efficient expression in T cells and (iv) single-domain architecture, among others. We demonstrate that miniCARbids can be engineered to recognize various antigens with antibody-like affinities, while being stable and aggregation-resistant. When miniCARbids are incorporated into CARs, they induce high anti-tumor potency in both adapter and conventional CAR formats. Remarkably, CD22-directed miniCARbid-based CARs showed similar or even more efficient tumor clearance in leukemia-bearing mice when compared with a CAR comprising the clinically tested m971-1xG4S scFv. Together, we introduce the miniCARbid engineering platform, enabling the generation of small, human antigen-binding domains with high potency in CAR T cells against virtually any target antigen.

synthetic biology↗

bioRxiv2025

Caffeine-regulated molecular switches for functional control of CAR T cells in vivo

The limited controllability of CAR T cells in patients represents a key challenge of this highly potent immunotherapy. A molecular ON-switch, which can be regulated with a non-toxic and readily available small molecule drug, would represent a major advance towards controllable CAR T therapeutics. For that purpose, we engineered caffeine-responsive heterodimeric ON-switches (CaffSwitches) and demonstrate their high caffeine-dependency and virtually absent leakiness. When incorporating these CaffSwitches into CARs, the resulting CaffCARs were efficiently activated by caffeine concentrations achieved in human plasma after drinking one cup of coffee. Moreover, CaffCAR T cells also showed efficient tumor clearance in an in vivo mouse model, which was completely abolished in the absence of caffeine. This tight control was even observed with c-Jun overexpressing CaffCAR T cells, despite their massive expansion. Together, we anticipate that these novel CaffSwitches will be valuable tools for the development of safe and efficient next generation CAR T cells.

synthetic biology↗

bioRxiv2024

Loss of PI5P4Kalpha slows the progression of a Pten mutant basal cell model of prostate cancer

While early prostate cancer (PCa) depends on the androgen receptor (AR) signaling pathway, which is predominant in luminal cells, there is much to be understood about the contribution of epithelial basal cells in cancer progression. Herein, we observe cell-type specific differences in the importance of the metabolic enzyme phosphatidylinositol 5-phosphate 4-kinase alpha (PI5P4K; gene name PIP4K2A) in the prostate epithelium. We report the development of a basal-cell-specific genetically engineered mouse model (GEMM) targeting Pip4k2a alone or in combination with the tumor suppressor phosphatase and tensin homolog (Pten). PI5P4K is enriched in basal cells, and no major histopathological changes were detectable following gene deletion. Notably, the combined loss of Pip4k2a slowed the development of Pten mutant mouse prostatic intraepithelial neoplasia (mPIN). Through the inclusion of a lineage tracing reporter, we utilize single-cell RNA sequencing to evaluate changes resulting from in vivo downregulation of Pip4k2a and characterize cell populations influenced in the established Probasin-Cre and Cytokeratin 5 (CK5)- Cre driven GEMMs. Transcriptomic pathway analysis points towards the disruption of lipid metabolism as a mechanism for reduced tumor progression. This was functionally supported by shifts of carnitine lipids in LNCaP PCa cells treated with siPIP4K2A. Overall, these data nominate PI5P4K as a target for PTEN mutant PCa. One Sentence SummaryLoss of PI5P4K slows cancer progression in prostate basal cells.

cancer biology↗

bioRxiv2023

NK cells shape the clonal evolution of B-ALL cells by IFN-γ production

The term cancer immunoediting describes the dual role by which the immune system can suppress and promote tumour growth and is divided into three phases: elimination, equilibrium and escape. The role of NK cells has mainly been attributed to the elimination phase. Here we show that NK cells play a role in all three phases of cancer immunoediting. Extended co-culturing of DNA barcoded mouse BCR/ABLp185+ B acute lymphoblastic leukaemia (B-ALL) cells with NK cells allowed for a quantitative measure of NK cell-mediated immunoediting. Whereas most tumour cell clones were efficiently eliminated by NK cells, a certain fraction of tumour cells harboured an intrinsic primary resistance. Furthermore, DNA barcoding revealed tumour cell clones with secondary resistance, which stochastically acquired resistance to NK cells. NK cell cytotoxicity put a selective pressure on B-ALL cells, which led to an outgrowth of primary and secondary resistant tumour cell clones, which were characterised by an IFN-{gamma} signature. Besides well-known regulators of immune evasion, our analysis of NK cell resistant tumour cells revealed the upregulation of genes, including Ly6a, which we found to promote NK cell resistance in leukaemic cells. Translation of our findings to the human system showed that high expression of LY6E on tumour cells impaired the physical interaction with NK cells and led to worse prognosis in leukaemia patients. Our results demonstrate that tumour cells are actively edited by NK cells during the equilibrium phase and use different avenues to escape NK cell-mediated eradication.

cancer biology↗

bioRxiv2023

TCR/CD3-based synthetic antigen receptors (TCC) convey superior antigen sensitivity combined with high fidelity of activation

Low antigen sensitivity and a gradual loss of effector functions limit the clinical applicability of chimeric antigen receptor (CAR)-modified T-cells and call for alternative antigen receptor designs for effective T-cell-based cancer immunotherapy. Here we applied advanced microscopy to demonstrate that TCR/CD3-based synthetic constructs (TCC) outperform second-generation CAR formats with regard to conveyed antigen sensitivities by up to a thousand-fold. TCC-based antigen recognition occurred without adverse non-specific signaling, which is typically observed in CAR-T-cells, and did not depend - unlike sensitized peptide/MHC detection by conventional T-cells - on CD4- or CD8- coreceptor engagement. TCC-endowed signaling properties may prove critical when targeting antigens in low abundance and aiming for a durable anti-cancer response.

immunology↗