Peter Sajjakulnukit

In Type 1 Diabetes (T1D), disease onset and immunotherapy responses vary widely, reflecting heterogeneous stage-specific immune dysregulation that remains undefined. To investigate this heterogeneity, we engineered microporous polycaprolactone scaffolds that form synthetic immunological niches (INs) subcutaneously and capture tissue-associated systemic immune changes. Building on our prior demonstration of IN-based longitudinal T1D monitoring, we use multimodal IN profiling to define the immune changes underlying divergent disease trajectories and immunotherapy responses. In non-obese diabetic (NOD) mice, longitudinal transcriptomic profiling of IN identified early myeloid associated changes, followed by increasing T cell-associated dysregulation that distinguished progressors from non-progressors. An early-stage IN-based T1D gene signature stratified NOD progressors from non-progressors by 6 weeks of age and was conserved across human T1D tissues, distinguishing T1D from non-diabetic individuals in spleen and pancreatic lymph node samples, but not in peripheral blood. Signature-based stratification revealed macrophage-associated TNFα pathway enrichment in pancreas of NOD progressors, which was validated in human T1D islets. Given the heterogeneous response to anti-TNFα therapy, IN profiling identified genesets associated with anti-TNFα efficacy and generated a response score that prospectively distinguished treatment-sensitive from resistant mice. These findings establish IN as a platform for detecting stage-wise immune dysregulation and stratifying immunotherapy response in T1D.

Rapid repair of genotoxic therapy induced DNA damage mediates treatment resistance in glioblastoma (GBM). The role of non-malignant cell types in the tumor microenvironment in accelerating DNA repair in neoplastic cells is poorly understood. Using spatial transcriptomics, immunofluorescence, metabolomics, patient tissues and preclinical models, we show that tumor associated macrophages (TAMs) in GBM promote DNA repair and treatment resistance in neoplastic cells through the secretion of acetylated amino acids. These acetylated amino acids are consumed by GBM cells, leading to enhanced acetyl Co-A levels, histone acetylation and nucleotide synthesis. Interrupting histone acetylation via inhibition of the acetyltransferase KAT5 breaks these metabolic links and reverses the protective capacity of microenvironment-derived acetylated amino acids and TAMs. Blocking the exchange of acetylated amino acids and their downstream effects is a potential strategy for the treatment of GBM.

Mutant isocitrate dehydrogenase 1 (mIDH1) catalyzes 2-hydroxyglutarate (2HG) production which leads to epigenetic reprogramming in astrocytomas with tumor protein p53 (TP53)/α-thalassemia/mental retardation, X-linked (ATRX) loss. RNA-sequencing, single-cell RNA-sequencing, and Chromatin Immunoprecipitation sequencing (ChIP-seq) followed by bioinformatics analysis shows that human and mouse mIDH1 gliomas exhibit downregulated gene ontologies (GOs) related to mitochondrial metabolism and upregulated autophagy-related GOs. Decreased mitochondrial metabolism is accompanied by decreased glycolysis, rendering autophagy as a source of energy in mIDH1 gliomas. Mutant IDH1 glioma cells exhibit increased expression of autophagy-related proteins and enhanced microtubule-associated protein 1 light chain 3 (LC3) I/II conversion, indicating augmented autophagy. Inhibiting autophagy in vivo by administration of synthetic protein nanoparticles (SPNPs) encapsulating autophagy related gene 7 (ATG7) silencing RNA sensitizes mIDH1 glioma cells to radiation, resulting in tumor regression, long-term survival, and immunological memory. This work uncovers autophagy as a critical pathway for survival in mIDH1 gliomas and its inhibition elicits radiosensitivity in vitro in human and mouse mIDH1 glioma cells, and in vivo in mIDH1 models.

Background: Dordaviprone (ONC201) is the first FDA-approved therapy for H3K27M-mutant diffuse midline glioma (DMG). However, therapeutic response varies among patients, and no imaging biomarkers currently predict long-term benefit. Diffusion-weighted MRI and apparent diffusion coefficient (ADC) obtained during standard clinical imaging reflect tissue cellularity and necrosis, offering potential as an early response indicator. Methods: Clinical and imaging data from patients treated with dordaviprone (n = 83) on completed studies underwent centralized neuroradiology review, ADC histogram analysis, and functional diffusion mapping (fDM), which were compared and correlated to CSF tumor DNA (tDNA) and metabolomics when available. Final analysis included 38 patients receiving standardized dordaviprone dosing and treatment schedule. Results: In the pre-progression cohort, early changes in volumetric or cross-sectional RAPNO criteria were not predictive of overall (OS) or progression-free survival (PFS). Importantly, in patients treated pre-progression with dordaviprone post-radiation, tumors with increased diffusion ADC (reduced cellularity) at cycle 3 demonstrated median OS of 732 versus 360 days for those with decreased ADC (increased cellularity, P = .0149); with corresponding PFS of 334 versus 110 days (P = .0132). fDM analysis of spatio-temporal changes of ADC did not correlate with outcomes in a sub-cohort analysis but demonstrated predictive patterns in individual patient time course series. CSF metabolomics stratified by ADC change revealed differences in reductive stress metabolites among dordaviprone responders, linking ADC elevation to dordaviprone-driven metabolic effects. Conclusion: Early increase in ADC correlates with improved response to dordaviprone in H3K27M-DMG and correlates with CSF metabolic changes, suggesting ADC warrants prospective validation as a biomarker.

Chromosome instability is highly prevalent in cancer and drives large-scale chromosomal imbalances, known as aneuploidies^1,2,3,4. How aneuploidy contributes to tumorigenesis remains difficult to study due to the vast numbers of genes affected. Here we established a CRISPR knockout- and activation-linked assay (CRISPR-KOALA), enabling high-throughput bidirectional genetic screens in immunocompetent mouse models of cancer. We developed a compendium of the ten most frequent human chromosome-arm-level alterations in basal-like breast cancer (BLBC), a disease type that is driven by large copy-number alterations (CNAs)^5,6,7,8. Using CRISPR-KOALA, we screened the mouse orthologues of 3,752 genes on these arms and identified 90 cancer driver genes, the function of the vast majority of which is unknown. These genes drive distinct signalling pathways including MAPK, HIPPO and WNT, reflecting the high degree of BLBC heterogeneity. Manipulating the identified cancer driver genes overcomes the need for CNAs in Trp53-mutant BLBC mouse models. Mechanistically, we identify that PLGRKT is a potent oncogene that lies on chromosome 9p and show that its tumour-promoting activity is associated with highly stress-resistant mitochondria and an increased ability to detoxify reactive oxygen species. Together, our findings reveal that arm-level CNAs can function to select specific driver genes to promote heterogeneous biological processes.

As solid tumors progress, the tumor microenvironment (TME) becomes increasingly immunosuppressive, impairing cytotoxic T-cell activity and limiting the efficacy of the immune checkpoint blockade. However, the mechanistic drivers of this immunosuppression remain poorly understood. Here, we identify a tumor-derived lipid–neutrophil–adenosine axis as a critical regulator of immune suppression in advanced colorectal cancer (CRC). We show that fatty acids enriched in tumor interstitial fluid reprogram neutrophils to generate adenosine via PPARα activation, leading to T-cell suppression. Using AB928, a dual A2aR/A2bR adenosine receptor antagonist currently in clinical trials, we restored T-cell proliferation, effector function, and tumor-killing capacity in vitro and in vivo. Importantly, AB928 synergized with anti-PD-1 therapy to enhance survival in an autochthonous model of metastatic CRC. Our findings define a metabolic immune evasion mechanism in the TME and provide a rationale for targeting neutrophil-derived adenosine signaling to improve immunotherapy responses in CRC and other solid tumors.

ZFTA–RELA⁺ ependymomas are malignant brain tumours defined by fusions formed between the putative chromatin remodeller ZFTA and the NF-κB mediator RELA¹. Here we show that ZFTA–RELA⁺ cells produce itaconate, a key macrophage-associated immunomodulatory metabolite². Itaconate is generated by cis-aconitate decarboxylase 1 (ACOD1; also known as IRG1). However, the production of itaconate by tumour cells and its tumour-intrinsic role are not well established. ACOD1 is upregulated in a ZFTA–RELA-dependent manner. Functionally, itaconate enables a feed-forward system that is crucial for the maintenance of pathogenic ZFTA–RELA levels. Itaconate epigenetically activates ZFTA–RELA transcription by enriching for activating H3K4me3 via inhibition of the H3K4 demethylase KDM5. ZFTA–RELA⁺ tumours enhance glutamine metabolism to supply carbons for itaconate synthesis. Antagonism of ACOD1 or glutamine metabolism reduces pathogenic ZFTA–RELA levels and is potently therapeutic in multiple in vivo models. Mechanistically, ZFTA–RELA epigenetically suppresses PTEN expression to upregulate PI3K–mTOR signalling, a known driver of glutaminolysis. Finally, suppression of ACOD1 or a combination of glutamine antagonism with PI3K–mTOR inhibition abrogates spinal metastasis. Our data demonstrate that ZFTA–RELA⁺ ependymomas subvert a macrophage-like itaconate metabolic pathway to maintain expression of the ZFTA–RELA driver, which implicates itaconate as a candidate oncometabolite. Taken together, our results position itaconate upregulation as a previously unappreciated driver of ZFTA–RELA⁺ ependymomas. Our work has implications for future drug development to reduce pathogenic ZFTA–RELA expression for this brain tumour, and will advance our understanding of oncometabolites as a new class of therapeutic dependencies in cancers.

Gliomas account for ~80% of primary malignant brain tumors. Many CNS WHO grade 2–3 and some grade 4 gliomas harbor mutant isocitrate dehydrogenase 1 (mIDH1), which causes a gain of function mutation (IDH1 R132H) leading to the production of 2-hydroxyglutarate (2HG). Mutant IDH1-induced 2HG, through epigenetic reprogramming elicits an immune-permissive tumor microenvironment (TME). An immunosuppressive mechanism in the glioma TME involves adenosine production via the ectoenzyme CD73. This study investigates mIDH1’s influence on CD73 expression and adenosine levels. We demonstrate that mIDH1 glioma cells exhibit reduced CD73 expression, driven by DNA hypermethylation, leading to reduced adenosine levels. Since wtIDH1 gliomas have high CD73 expression, we evaluated CD73 blockade as an immunotherapy target. We show that CD73 inhibition used as monotherapy, did not improve survival in wtIDH1 glioma-bearing mice. However, when combined with immune-stimulatory Ad-TK (adenoviral vectors encoding herpes simplex virus thymidine kinase) and Ad-Flt3L (adenoviral vectors encoding FMS-like tyrosine kinase 3 ligand) gene therapy, CD73 blockade significantly enhanced therapeutic efficacy and increased anti-glioma effector T cell activity. These findings reveal that CD73 inhibition used in combination with immune stimulatory Ad-TK/Ad-Flt3L gene therapy may be an effective treatment for wtIDH1 gliomas, which could be readily translated to the clinical arena.