Kymera Therapeutics

Cancer immunotherapy has achieved remarkable clinical efficacy across multiple cancers but remains ineffective for most patients, and the mechanisms underlying resistance remain incompletely understood. Here, we perform in vivo CRISPR-Cas9 screens in female mice and identify DNA methyltransferase 3 A (DNMT3A) as a key driver of tumor immune evasion through both methyltransferase-dependent and -independent mechanisms. DNMT3A represses TNFRSF1A and TNFRSF1B expression, enabling tumor cells to evade TNFα-induced killing. Independently of its methyltransferase activity, DNMT3A interacts with NFYA to activate cholesterol biosynthesis, leading to zymosterol accumulation in tumor interstitial fluid. Zymosterol impairs SREBP2 cleavage and promotes HMGCR degradation in CD8⁺ T cells, thereby suppressing cholesterol biosynthesis and compromising their antitumor effector functions. Targeting DNMT3A with antisense oligonucleotides overcomes resistance to immune checkpoint blockade. Collectively, these findings establish DNMT3A as an integrated epigenetic and metabolic checkpoint in tumor immune evasion and a potential target for immunotherapy.

Anti-CD19 chimeric antigen receptor T-cell therapy (CAR-T) has transformed the treatment of relapsed/refractory (R/R) non-Hodgkin lymphoma (NHL). However, toxicities including cytokine release syndrome (CRS) and immune effector cell-associated neurotoxicity syndrome (ICANS) remain significant. Prophylactic corticosteroid use increased after cohort 6 of ZUMA-1, which suggested reduced severe CRS and ICANS, although the lack of a comparator arm limits interpretation. To evaluate the impact of steroid prophylaxis on outcomes and toxicities in a real-world setting. We conducted a retrospective, single-institution analysis of adults (≥18 years) with R/R NHL treated with CD28 costimulated axicabtagene ciloleucel (axi-cel) or brexucabtagene autoleucel (brexu-cel) at The Ohio State University (2018-2023). All patients received steroid prophylaxis (dexamethasone 10 mg daily × 3 from infusion day). Outcomes including toxicities (CRS, ICANS) and response rates, survival outcomes, and hazard ratios were compared using logistic regression, log-rank test, and Cox model. 137 patients received CAR-T (axi-cel n=118, brexu-cel n=19); 27.0% received steroid prophylaxis (PPX). Baseline characteristics were similar between groups, although primary refractory disease differed. CRS and ICANS prevalence did not significantly differ between cohorts. Any-grade CRS occurred in 97.3% of patients with PPX vs 88.0% with no PPX (p=0.13), with grade 3-4 CRS in 2.7% vs 12.0% (p=0.13). ICANS occurred in 75.7% vs 68.0% (p=0.38), with grade 3-4 ICANS in 24.3% vs 32.0% (p=0.38). Median time to CRS onset was delayed in the PPX group (4 vs 3 days, p<0.01), with no difference in ICANS onset. Overall response rates (ORR) were 75.7% vs 69.0% (p=0.44); complete response rates (CRR) were 62.2% vs 54.0% (p=0.39). OS and PFS did not differ. Prophylactic corticosteroids did not impact CRS, ICANS, response rates, or survival. Although CRS onset may be modestly delayed, these findings do not support routine steroid prophylaxis in CAR-T-treated NHL patients.

Determining how chemotherapeutic drugs alter the MHC class I-associated immunopeptidome is essential for designing rational combinatorial therapies to enhance antitumor CD8+ T-cell activity, such as immune checkpoint inhibitor Abs. We investigated the impact of a first-in-class inhibitor of cellular NEDDylation, MLN4924, on self-peptide presentation using 2 human tumor cell lines: HeLa cells secreting the human MHC class I protein HLA-A*11:01 and the glioblastoma cell line U87-MG. MLN4924 indirectly inactivates the cullin-RING ligases (CRLs), a large family of E3 ubiquitin ligases responsible for targeting numerous substrates for degradation by the ubiquitin-proteasome system. MLN4924 treatment prevented cell division and altered the transcriptome of HeLa cells in a predictable manner. However, MLN4924 treatment did not alter the cell-surface levels of MHC class I proteins. An analysis of the immunopeptidome and the source proteins, which yielded peptides for presentation, revealed some unexpected findings. Greater than 50% of the peptides presented, and more than two-thirds of the source proteins from which peptides were derived, remained unaltered after MLN4924 treatment. Furthermore, alterations to the transcriptome could not predict the observed changes to the immunopeptidome. Of the approximately 100 CRL substrates that were presented by HLA-A*11:01, most were presented independent of MLN4924 treatment. Alternative degradation pathways and the presence of intron sequences in the specific mRNA transcripts may help explain how these proteins yield peptides for Ag presentation. In summary, MLN4924 treatment only partially affects the immunopeptidomic profile, suggesting it could be used in combinatorial cancer immunotherapies.

HIV persistence in CD4+ T cells and brain microglia is maintained by host mechanisms that enforce viral latency. We identify ENL and the deubiquitinase USP7 as a host complex that sustains HIV latency. Pharmacologic inhibition or genetic loss of ENL reactivated latent HIV. Proteomic analyses revealed an ENL-USP7-BRD4 regulatory axis associated with latency maintenance, where USP7 maintained BRD4 protein stability. Targeted USP7 degradation using PROTAC induced latency reversal and destabilized BRD4. Chromatin immunoprecipitation studies showed that ENL was required for USP7 and BRD4 occupancy at the HIV nuc-1 region but not at the 5' long terminal repeat. Single-nucleus RNA sequencing further implicated this pathway in HIV persistence within the central nervous system. Importantly, degradation of either ENL or USP7 reactivated latent HIV in resting CD4+ T cells and brain microglia from individuals receiving antiretroviral therapy. These findings identify the ENL-USP7-BRD4 axis as a therapeutically actionable mechanism sustaining HIV latency and viral reservoirs.

The ubiquitin proteasome system (UPS) and autophagy cooperatively orchestrate plant immune homeostasis, while diverse phytopathogens deploy secreted effectors to disrupt their crosstalk for successful colonization. Proteaphagy, the autophagic turnover of intact 26S proteasome holoenzymes, is a conserved stress response co-opted by Pseudomonas syringae effector HopM1 to cripple host proteolysis. However, how the Huanglongbing (HLB) pathogen 'Candidatus Liberibacter asiaticus' (CLas) simultaneously manipulates UPS and autophagy to subvert citrus immunity remains largely uncharacterized. Here, we functionally characterize the secreted CLas effector SDE5640, which exacerbates HLB susceptibility in sweet orange (Citrus sinensis). SDE5640 directly binds the 19S proteasome chaperone CsPSMD10, whose homodimerization via Cys163/Cys190 disulfide bonds is required for immune suppression. CsPSMD10 overexpression enhances citrus susceptibility to CLas, whereas the dimerization-deficient CsPSMD10-C190S mutant confers robust HLB resistance. Mechanistically, CsPSMD10 targets CsATG8 isoforms for UPS-dependent degradation to restrain autophagic flux; SDE5640 competitively occupies the Ank2/Ank3 domains of CsPSMD10 to disrupt CsPSMD10-CsATG8c complexes, thereby stabilizing CsATG8c and triggering autophagy dependent on CsPSMD10. Moreover, SDE5640-induced autophagy drives selective autophagic degradation of the 20S proteasome catalytic subunit CsPBA1, which ultimately disrupts cellular proteasome activity and attenuates salicylic acid-mediated citrus immunity. Collectively, our findings identify a novel SDE5640-CsPSMD10-CsATG8c-CsPBA1 regulatory cascade, illustrating how a single CLas effector synchronously rewires the UPS-autophagy crosstalk to facilitate pathogen proliferation.

Approximately 96% of myelofibrosis (MF) patients express wild-type (WT) p53. MF hematopoietic stem/progenitor cells (HSPCs) overexpress MDM2 that degrades WTp53, thereby supporting MF HSPC survival. Although MDM2 small-molecule inhibitors (SMIs) are clinically effective, their benefit is compromised by an inability to sufficiently deplete MF HSPCs. KT-253 degrades MDM2 and overcomes the p53-dependent feedback loop that restores MDM2 levels observed with the use of MDM2 SMIs such as AMG-232. We evaluated the effects of KT-253 using MPN cell lines, primary MF CD34+ cells, and a mouse xenograft model. KT-253 activity was correlated with activation of the p53 pathway and upregulation of p53 downstream genes, including p21, NOXA, and PUMA in WTp53 UKE-1 cells but not p53 mutant cells. KT-253 treatment induced apoptosis of primary MF CD34+ cells at a tenfold lower concentration than AMG-232 and prevented the upregulation of MDM2 seen with AMG-232. Low doses of KT-253 reduced total MF colony and JAK2V617F+ colony numbers to a far greater extent than log-higher doses of AMG-232 (60% and 71% versus 20% and 10%, respectively). Mouse xenograft studies indicated that KT-253 more effectively depleted JAK2V617F+ MF marrow-repopulating cells. These findings support the further clinical evaluation of MDM2 degraders for the treatment of MF patients.