Biology subjects

Mitchell, A.

Publications and source records attributed to Mitchell, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Single-point mutation alters odorant receptor sensitivity associated with host plant specialization in Spodoptera moths

Host specialization in herbivorous insects is often associated with divergence in chemosensory abilities. Here, we investigated the possible contribution of odorant receptors (ORs) in host plant restriction in the lily moth Spodoptera picta, a species specialized on Amaryllidaceae. Manual annotation of S. picta ORs in its genome revealed a repertoire similar in size and composition to those of its polyphagous sister species, S. littoralis and S. litura, suggesting that specialization did not involve major gene loss or expansion in the lily moth. To assess functional divergence beyond gene number, we applied a large scaled structure-based virtual screening approach to the entire OR repertoires of these three Spodoptera species, generating ligand-binding profiles for 120,591 volatile compounds. Among 69 1:1:1 OR orthologs, 24 exhibited divergent predicted binding spectra. We pinpointed OR29 that we also found to be highly expressed in both male and female antennae of S. picta through a RNAseq approach. Functional assays demonstrated that S. picta OR29 acquired heightened sensitivity to limonene enantiomers, volatiles emitted by host Amaryllidaceae inflorescences. Site-directed mutagenesis revealed that a single amino acid substitution within the predicted binding region underlies this shift in sensitivity. These results show that host specialization in S. picta has not been accompanied by significant OR repertoire remodeling, but rather by subtle molecular changes that fine-tune receptor sensitivity to host-derived volatiles.

molecular biology↗

bioRxiv2026

Cell-state-dependent responses to PLK1 inhibition reveal a non-canonical microtubule-endolysosomal vulnerability in quiescent leukemia stem cells

Relapse in cancer is frequently driven by therapy-resistant quiescent cancer stem cells. Conventional chemotherapy has been designed to target proliferating tumor cells and is generally presumed to be ineffective against non-cycling cancer stem cells. Using acute myeloid leukemia (AML) as a model, we challenge this prevailing view by showing that inhibition of the mitotic master regulator Polo-like kinase 1 (PLK1), a kinase extensively pursued for antiproliferative cancer therapy, unexpectedly eradicates quiescent leukemia stem cells (LSC) through a mechanism distinct from its canonical mitotic function. In proliferating AML cells, PLK1 inhibition (PLK1i) induced G2/M arrest and mitotic catastrophe. In contrast, quiescent LSC underwent apoptosis independent of mitotic arrest, revealing a cell-state-dependent mode of drug action. Mechanistically, PLK1i initiated a multi-step process through disruption of a previously unrecognized, stem cell-specific interaction between PLK1 and MAP1A, resulting in perturbed vesicle trafficking and endolysosomal homeostasis characterized by altered receptor internalization, vesicle accumulation and lysosomal dysfunction, ultimately culminating in apoptotic cell death. Combinatorial pharmacologic perturbation studies established microtubule regulation as a critical determinant of quiescent LSC survival, while ex vivo and in vivo assays demonstrated depletion of functionally-defined LSC following PLK1i. These findings identify a previously unrecognized role for PLK1 in intracellular trafficking and establish MAP1A-dependent control of vesicle homeostasis as a mechanistic determinant of cancer stem cell survival. More broadly, this study demonstrates that classical antimitotic compounds, including microtubule-targeting agents and PLK1 inhibitors, can eradicate both cycling leukemic blasts and quiescent LSC through distinct, cell state-dependent mechanisms, challenging proliferation-centric models of chemotherapy action.

cancer biology↗

bioRxiv2026

Prefrontal brain-to-brain synchrony during human group hunting: Evidence from fNIRS hyperscanning

Orca, wolves, chimpanzees and humans share a similarly impressive capacity for group hunting, where individuals coordinate behaviour together to capture prey. Studying hunting behaviours has important implications for understanding how behaviour in group contexts may be indicative of cognitive decline. Despite growing interest in brain circuits for prey capture, the brain regions involved in tracking prey during a hunt and the behaviours in group hunt linked to success remain unclear. Here we combined functional near infrared spectroscopy (fNIRS) and a virtual minecraft world to examine behaviour, brain dynamics and brain synchrony involved in group hunting behaviour. We focused on the prefrontal cortex (PFC) due to its known role in planning and social coordination and recorded from pairs of individuals as they either cooperated to hunt another person (prey) or simply followed another person. Hunters were more successful if they managed to keep a smaller distance to the prey and moved at speeds that were more synchronised with their co-predator. At high-range frequencies for fNIRS (0.1-0.2Hz), we found greater brain-to-brain synchrony in lateral and medial (frontopolar) PFC regions during hunting compared with chance levels. Together, these findings provide insights into what behaviours and brain dynamics associated with successful group hunting.

neuroscience↗

bioRxiv2026

Establishment of Stable Immortalized Human Choroidal Melanocytes for Ocular Research

PurposeThe short lifespan of primary normal choroidal melanocytes (NCMs) in vitro represents a major barrier to mechanistic, functional, and translational studies of choroid biology and uveal melanoma (UM). This study aimed to establish and characterize immortalized human NCM lines that retain melanocytic function, maintain a non-cancerous profile, and are amenable to gene editing. MethodsNCMs from four donors were immortalized by lentiviral transduction of Cyclin-dependent kinase 4 (CDK4R24C), Cyclin D1, and human Telomerase reverse transcriptase (hTERT), establishing NCM-K4DT lines. Their morphology, melanocytic marker expression, proliferation and functional properties (melanin synthesis, tyrosinase activity) were evaluated. Genomic stability was assessed by targeted mutation profiling, karyotyping, and copy number variation analysis. The tumorigenicity was tested in immunodeficient mice. Plasmid-based CRISPR/Cas9 editing was performed to determine their suitability for gene editing. ResultsNCM-K4DT lines retained dendritic-shaped morphology, pigmentation, and expression of PMEL, TYRP1, Melan-A, and SOX10. Cells exhibited enhanced proliferative capacity with preserved cell cycle regulation. Melanin production and tyrosinase activity were comparable to primary NCMs. Genomic profiling confirmed the absence of UM-associated driver mutations and chromosomal abnormalities. In vivo growth assays demonstrated no tumorigenic potential. Notably, NCM-K4DT cells were efficiently edited by CRISPR/Cas9. ConclusionsNCM-K4DT lines represent stable, non-cancerous, and genetically tractable models for studying choroidal melanocyte biology, modeling UM-associated mechanisms, and advancing therapeutic development in ocular research.

cell biology↗

bioRxiv2025

Chain-length regulation by WzzE is necessary for, but genetically separable from, cyclic enterobacterial common antigen synthesis

Enterobacterial common antigen (ECA) is a conserved glycan that supports outer membrane impermeability and intrinsic antibiotic resistance in Enterobacterales. ECA exists in outer membrane diacylglycerol-phosphate- and lipopolysaccharide-linked forms, and a cyclic periplasmic form (ECACYC). Intriguingly, ECACYC both affects the outer-membrane permeability barrier and functions in regulation of diacylglycerol-phosphate-linked ECA levels. While the length of linear ECA polymers generated by WzyE is regulated by the co-polymerase WzzE, WzzE is also required for ECACYC biogenesis and no ECACYC is synthesized in its absence. To uncover WzzE functions necessary for ECACYC biosynthesis, we generated plasmid-borne and chromosomal wzzE mutants in Escherichia coli K-12 and quantified their effects on linear chain-length regulation and on ECACYC synthesis. Mutations affecting linear ECA chain-length regulation in either transmembrane helix 2 or the periplasmic domain abolished ECACYC synthesis. Notably, two variants at residue F104 exhibited identical linear ECA chain-length regulation which was similar to wild type, but resulted in sharply different ECACYC production: chromosomal WzzEF104Y produced near wild-type levels of ECACYC, whereas WzzEF104H produced approximately two-fold less ECACYC. This difference in ECACYC production vs. linear chain-length regulation demonstrates ECACYC synthesis is genetically separable from this regulation. Thus, WzzE-mediated chain-length regulation is necessary for ECACYC biogenesis, but not sufficient.

microbiology↗

bioRxiv2025

Impaired envelope integrity in the absence of SanA is linked to increased Lipid II availability and an imbalance of FtsI and FtsW activities

In Gram-negative bacteria, the outer membrane (OM) acts in conjunction with the peptidoglycan (PG) cell wall as a barrier against physical, osmotic, and chemical environmental stressors including antibiotics. SanA, an inner membrane protein in Escherichia coli K-12, is required for vancomycin resistance at high temperatures (>42 {degrees}C) and impacts sodium dodecyl sulfate (SDS) resistance during stationary phase reached from carbon limitation. However, its function remains unknown. Here, we show that {Delta}sanA has a synthetic genetic interaction with {Delta}wecA, a mutation that increases the availability of the isoprenoid carrier for PG synthesis. Specifically, the {Delta}sanA {Delta}wecA strain demonstrated heightened SDS-EDTA sensitivity, activation of the Rcs stress response, and increased cell length. Further investigation tied the SDS-EDTA sensitivity to increased lipid II available for PG synthesis. Spontaneous suppressor mutants of this phenotype harbored point mutations in prc, which encodes tail specific protease, or ftsI, which encodes the cell division DD-transpeptidase, a target of Prc. We focused on the ftsI mutations and demonstrated that the ftsI mutations had increased cell length but nevertheless enhanced PG incorporation at the septum compared to the {Delta}sanA mutant, returning PG incorporation to wild-type levels. Moreover, other mutations affecting septal PG synthesis, but not divisome assembly, also suppressed the SDS-EDTA sensitivity. These findings suggest that, in the absence of SanA, increased lipid II availability perturbs the balance between septal PG synthesis, lateral PG elongation, and other envelope biogenesis pathways, which leads to increased OM permeability. IMPORTANCEThe Gram-negative cell envelope is a barrier that protects the cell from environmental stress. Therefore, the synthesis of each layer of this envelope needs to be closely coordinated throughout growth and division. Here, we investigated SanA, a protein in Escherichia coli K-12 that affects envelope permeability under cellular stress, including nutrient limitation and high temperature. We found that SanA plays a key role in maintaining the permeability barrier when precursor levels for peptidoglycan (PG) synthesis are elevated, linking envelope integrity to balanced septal PG production during cell division. Our results suggest that SanA modulates substrate availability to preserve envelope function, and that in its absence, imbalanced substrate flux to septal PG synthesis disrupts septum formation and compromises barrier integrity.

microbiology↗

bioRxiv2025

Landscapes of missense variant impact for human superoxide dismutase 1

Amyotrophic lateral sclerosis (ALS) is a progressive motor neuron disease for which important subtypes are caused by variation in the Superoxide Dismutase 1 gene SOD1. Diagnosis based on SOD1 sequencing can not only be definitive but also indicate specific therapies available for SOD1-associated ALS (SOD1-ALS). Unfortunately, SOD1-ALS diagnosis is limited by the fact that a substantial fraction (currently 26%) of ClinVar SOD1 missense variants are classified as "variants of uncertain significance" (VUS). Although functional assays can provide strong evidence for clinical variant interpretation, SOD1 assay validation is challenging, given the current incomplete and controversial understanding of SOD1-ALS disease mechanism. Using saturation mutagenesis and multiplexed cell-based assays, we measured the functional impact of over two thousand SOD1 amino acid substitutions on both enzymatic function and protein abundance. The resulting missense variant effect maps not only reflect prior biochemical knowledge of SOD1 but also provide sequence-structure-function insights. Importantly, our variant abundance assay can discriminate pathogenic missense variation and provides new evidence for 41% of missense variants that had been previously reported as VUS, offering the potential to identify additional patients who would benefit from therapy approved for SOD1-ALS.

genetics↗

bioRxiv2025

Bacterial population dynamics during colonization of solid tumors

Bacterial colonization of tumors is widespread, yet the dynamics during colonization remain underexplored. Here we discover strong variability in the sizes of intratumor bacterial clones and use this variability to infer the mechanisms of colonization. We monitored bacterial population dynamics in murine tumors after introducing millions of genetically barcoded Escherichia coli cells. Results from intravenous injection revealed that roughly a hundred bacteria seeded a tumor and that colonizers underwent rapid, yet highly nonuniform growth. Within a day, bacteria reached a steady-state and then sustained load and clone diversity. Intratumor injections, circumventing infection bottlenecks, revealed that the nonuniformity persists and that the sizes of bacterial progenies followed a scale-free distribution. Theory suggested that our observations are compatible with a growth model constrained by a local niche load, global resource competition, and noise. Our work provides the first dynamical model of tumor colonization and may allow distinguishing genuine tumor microbiomes from contamination.

systems biology↗

bioRxiv2024

Distinct gene regulatory networks govern hematopoietic and leukemia stem cells

The underlying gene regulatory networks (GRN) that govern leukemia stem cells (LSC) in acute myeloid leukemia (AML) and hematopoietic stem cells (HSC) are not well understood. Here, we identified GRNs by integrating gene expression (GE) and chromatin accessibility data derived from functionally defined cell populations enriched for HSC and LSC. We analyzed n=32 LSC+ and n=32 LSC-cell fractions from n=22 AML patients, along with n=7 stem and n=10 progenitor enriched cell populations sorted from human umbilical cord blood (hUCB), producing a database of n{approx}17,000 transcription factor (TF) regulatory interactions for hUCB-HSPC and AML. We developed an iterative algorithm that associates the degree of chromatin openness with TF binding preferences, and the GE of candidate TF and target genes within 100kb upstream of transcription start sites. A putative regulatory structure was found to be enriched in HSC-enriched cell populations, comprising TF-target gene interactions between ETS1, EGR1, RUNX2, and ZNF683 oriented in a self-reinforcing configuration. A regulatory loop comprising FOXK1 and MEIS1, rather than the 4-factor HSC subnetwork, was detected in the LSC-specific GRN. The core HSC and LSC TF networks were extended using protein-protein interaction (PPI) data to determine connectivity with interacting genes whose expression strongly associated with LSC/HSC frequency estimates, producing a database of n=103,516 PPI target pathways. The effect of perturbing genes along the identified pathways on functional HSC and LSC frequency was predicted based on statistical regression analyses. To validate GRN predictions, we used pharmacologic and CRISPR targeting, in addition to re-examining published functional data associated with several network nodes that were predicted to impact stemness. Notably, we found that inhibition of CDK6 in AML samples markedly reduced LSC numbers as assessed in de novo serial xenotransplantation studies (fold change {approx} 10), as predicted by the LSC GRN model. Additionally, in-house CRISPR-based knockdown of ETS1 resulted in a significant decrease in HSC quiescence-associated microRNA-126 expression, and increased HSC frequency. Taken together, our models provide a comprehensive view of the underlying regulatory structures governing functional human HSC and LSC. This approach has translational potential as it can be used as a high-throughput in-silico screening tool for the systematic identification of gene targets for LSC elimination and HSC expansion.

bioinformatics↗

bioRxiv2024

Transcriptional Characterization of Resistance in Early Drug Response

Therapeutic resistance is a major cause of cancer treatment failure, with increasing evidence suggesting a non-genetic basis. This non-genetic resistance is often due to drug-resistant transcriptional cell states, either induced by treatment or pre-existing in some cells. However, the connection between early cellular drug response and long-term resistance is poorly understood. Moreover, it is unknown whether resistance-associated early transcriptional responses are evolutionarily conserved. Integrating long-term drug resistance and early drug response data across multiple cancer cell lines, bacteria, and yeast, our findings indicate that cancer states in drug-naive populations and shortly after treatment share transcriptional properties with fully resistant populations, some of which are evolutionarily conserved. CRISPR-Cas9 knockout of resistant states markers increased sensitivity to Prexasertib in ovarian cancer cells. Finally, early resistant state signatures discriminated therapy responders from non-responders across multiple human cancer trials, and distinguished premalignant breast lesions that progress to malignancy from those that do not.

cancer biology↗

bioRxiv2024

FLT3 is genetically essential for ITD-mutated leukemic stem cells but dispensable for human hematopoietic stem cells

Leukemic stem cells (LSCs) fuel acute myeloid leukemia (AML) growth and relapse, but therapies tailored towards eradicating LSCs without harming healthy hematopoietic stem cells (HSCs) are lacking. FLT3 is considered an important therapeutic target due to frequent mutation in AML and association with relapse. However, there has been limited clinical success with FLT3 targeting, suggesting either that FLT3 is not a vulnerability in LSC, or that more potent inhibition is required, a scenario where HSC toxicity could become limiting. We tested these possibilities by ablating FLT3 using CRISPR/Cas9-mediated FLT3 knock-out (FLT3-KO) in human LSCs and HSCs followed by functional xenograft assays. FLT3-KO in LSCs from FLT3-ITD mutated, but not FLT3-WT AMLs, resulted in short-term leukemic grafts of FLT-3-KO edited cells that disappeared by 12 weeks. By contrast, FLT3-KO in HSCs from fetal liver, cord blood and adult bone marrow did not impair multilineage hematopoiesis in primary and secondary xenografts. Our study establishes FLT3 as an ideal therapeutic target where ITD+ LSC are eradicated upon FLT3 deletion, while HSCs are spared. These findings support the development of more potent FLT3-targeting drugs or gene-editing approaches for LSC eradication to improve clinical outcomes. KEY POINTSThe FLT3 gene is essential for ITD-mutated leukemic stem cells (LSCs) to establish and propagate leukemia. Normal human hematopoietic stem cells (HSCs) do not require FLT3 to engraft and sustain hematopoiesis. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=179 SRC="FIGDIR/small/601363v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@b77f56org.highwire.dtl.DTLVardef@109011dorg.highwire.dtl.DTLVardef@282e04org.highwire.dtl.DTLVardef@1e51e3b_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2024

Colibactin-induced damage in bacteria is cell contact independent

The bacterial toxin colibactin, produced primarily by the B2 phylogroup of Escherichia coli, underlies some cases of colorectal cancers. Colibactin crosslinks DNA and induces genotoxic damage in both mammalian and bacterial cells. While the mechanisms facilitating colibactin delivery remain unclear, results from multiple studies supported a delivery model that necessitates cell-cell contact. We directly tested this requirement in bacterial cultures by monitoring the spatiotemporal dynamics of the DNA damage response using a fluorescent transcriptional reporter. We found that in mixed-cell populations, DNA damage saturated within twelve hours and was detectable even in reporter cells separated from colibactin producers by hundreds of microns. Experiments with distinctly separated producer and reporter colonies revealed that the intensity of DNA damage decays similarly with distance regardless of colony contact. Our work reveals that cell contact is inconsequential for colibactin delivery in bacteria and suggests that contact-dependence needs to be reexamined in mammalian cells as well. ImportanceColibactin is a bacteria-produced toxin that binds and damages DNA. It has been widely studied in mammalian cells due to its potential role in tumorigenesis. However, fundamental questions about its impact in bacteria remain underexplored. We used E. coli as a model system to study colibactin toxicity in neighboring bacteria and directly tested if cell-cell contact is required for toxicity, as has previously been proposed. We found that colibactin can induce DNA damage in bacteria hundreds of microns away and that the intensity of DNA damage presents similarly regardless of cell-cell contact. Our work further suggests that the requirement for cell-cell contact for colibactin-induced toxicity also needs to be reevaluated in mammalian cells.

microbiology↗

bioRxiv2024

Morphometrics as a conservation tool for rapid in-situ distinction of native and introduced iguanas: comparing Iguana delicatissima and Iguana iguana

Invasive alien species severely impact native and endemic species, disproportionately affecting insular species, and this is especially true for Caribbean reptiles. The Lesser Antillean iguana (Iguana delicatissima) experienced a drastic range decline strongly driven by hybridization with non-native green iguanas (NNGI, Iguana iguana species complex). With numerous NNGI populations within the Lesser Antilles, it is a matter of time before these arrive on the last I. delicatisisma inhabited islands, whereupon rapid in-situ identification of non-native and hybrid animals is essential. However, only a few scale and coloration characters allow in-situ identification of NNGI, which are compromised by introgression. Here we assessed the differentiating power of an additional 20 scale and length-dependent characteristics between the I. delicatissima population on St. Eustatius and the established NNGI population on St. Maarten, the main source of arriving stowaway iguanas on St. Eustatius. We identified 14 length-dependent characteristics that significantly differ between I. delicatissima and NNGIs, with multi-variate analysis showing clear morphospace separation and high assignment accuracy of predictive models (>91%). Additionally, the number of femorals and supra digital scales significantly differs between these iguanas. Morphospace knowledge of St. Eustatius I. delicatissima now allows rapid identification of any divergent iguanas using easy to obtain measurements and meristics. Given intraspecific variation in I. delicatissima and green iguanas, we recommend that these characteristics are assessed and validated for other populations, including hybrid individuals. Our work demonstrates the urgent need to invest in baseline morphometric datasets to aid rapid in-situ conservation efforts once NNGI arrive and start to hybridize with I. delicatissima.

zoology↗

bioRxiv2023

Precise single-cell transcriptomic mapping of normal and leukemic cell states reveals unconventional lineage priming in acute myeloid leukemia

In acute myeloid leukemia (AML), genetic mutations distort hematopoietic differentiation, resulting in the accumulation of leukemic blasts. Yet, it remains unclear how these mutations intersect with cellular origins and whether they converge upon similar differentiation patterns. Single-cell RNA sequencing (scRNA-seq) has enabled high-resolution mapping of the relationship between leukemia and normal cell states, yet this application is hampered by imprecise reference maps of normal hematopoiesis and small sample sizes among patient cohorts. As a first step we constructed a reference atlas of human bone marrow hematopoiesis from 263,519 single-cell transcriptomes spanning 55 cellular states, that was benchmarked against independent datasets of immunophenotypically pure hematopoietic stem and progenitor cells. Using this reference atlas, we mapped over 1.2 million single-cell transcriptomes spanning 318 AML, mixed phenotype acute leukemia (MPAL), and acute erythroid leukemia (AEL) samples. This large-scale analysis, together with systematic mapping of genotype-to-phenotype associations between driver mutations and differentiation landscapes, revealed convergence of diverse genetic alterations on twelve recurrent patterns of aberrant differentiation in AML. This included unconventional lymphoid and erythroid priming linked to RUNX1 and TP53 mutations, respectively. We also identified non-genetic determinants of AML differentiation such as two subgroups of KMT2A-rearranged AML that differ in the identity of their leukemic stem cells (LSCs), likely reflecting distinct cellular origins. Furthermore, distinct LSC-driven hierarchies can co-exist within individual patients, providing insights into AML evolution. Together, precise mapping of normal and malignant cell states provides a framework for advancing the study and disease classification of hematologic malignancies thereby informing therapy development.

cancer biology↗

bioRxiv2023

Increased bleeding and thrombosis in myeloproliferative neoplasms mediated through altered expression of inherited platelet disorder genes

An altered thrombo-hemorrhagic profile has long been observed in patients with myeloproliferative neoplasms (MPNs). We hypothesized that this observed clinical phenotype may result from altered expression of genes known to harbor genetic variants in bleeding, thrombotic, or platelet disorders. Here, we identify 32 genes from a clinically validated gene panel that were also significantly differentially expressed in platelets from MPN patients as opposed to healthy donors. This work begins to unravel previously unclear mechanisms underlying an important clinical reality in MPNs. Knowledge of altered platelet gene expression in MPN thrombosis/bleeding diathesis opens opportunities to advance clinical care by: (1) enabling risk stratification, in particular, for patients undergoing invasive procedures, and (2) facilitating tailoring of treatment strategies for those at highest risk, for example, in the form of antifibrinolytics, desmopressin or platelet transfusions (not current routine practice). Marker genes identified in this work may also enable prioritization of candidates in future MPN mechanistic as well as outcome studies.

genomics↗

bioRxiv2023

Convergence of direction, location and theta in the rat anteroventral thalamic nucleus

Retrosplenial cortex (RSC) is a cortical region that computes heading direction from landmark information, but how it does this remains unknown. Recently we found that its two major subregions receive differential projections from two anteroventral thalamic (AV) subfields; dorsomedial (AVDM) and ventrolateral (AVVL). To probe the respective contributions of these inputs we recorded single neurons and local field potentials from AV in rats during foraging. We observed and characterized neurons modulated by theta oscillations, heading direction, and a conjunction of these. Unexpectedly, we also discovered place cells (neurons modulated by location). Consistent with the notion that AV contains two parallel subcircuits, there was a prevalence of non-conjunctive cells in AVDM, and of conjunctive and place neurons in AVVL. This integration of spatial and movement signals in AV is consistent with a thalamic role in multimodal integration and may be important for supplying the spatial information that modulates RSC directional responding.

neuroscience↗

bioRxiv2022

Evolved bacterial resistance to the chemotherapy gemcitabine modulates its efficacy

Drug metabolism by the microbiome can influence anti-cancer treatment success. We previously suggested that chemotherapies with antimicrobial activity can select for adaptations in bacterial drug metabolism that can inadvertently influence the hosts chemoresistance. We demonstrated that evolved resistance against fluoropyrimidine chemotherapy lowered its efficacy in worms feeding on drug-evolved bacteria (Rosener et al., 2020). Here we examine a model system that captures local interactions that can occur in the tumor microenvironment. Gammaproteobacteria colonizing pancreatic tumors can degrade the nucleoside-analog chemotherapy gemcitabine and, in doing so, can increase the tumors chemoresistance. Using a genetic screen in Escherichia coli, we mapped all loss-of-function mutations conferring gemcitabine resistance. Surprisingly, we found that one third of resistance mutations increase or decrease bacterial drug breakdown and therefore can either lower or raise the gemcitabine load in the local environment. Experiments in three E. coli strains revealed that evolved adaptation converged to inactivation of the nucleoside permease NupC, an adaptation that increased the drug burden on co-cultured cancer cells. The two studies provide complementary insights on the potential impact of microbiome adaptation to chemotherapy by showing that bacteria-drug interactions transpire locally and systemically and can influence chemoresistance in the host.

systems biology↗

bioRxiv2022

KDM6A loss sensitizes human acute myeloid leukemia to PARP and BCL2 inhibition

Acute myeloid leukemia (AML) is a heterogeneous, aggressive malignancy with dismal prognosis and with limited availability of targeted therapies. AML exhibits epigenetic deregulation and transcriptional plasticity that contributes to pathogenesis. KDM6 proteins are histone-3 lysine-27 demethylases that play major context dependent roles in AML evolution and therapy resistance. Here, we demonstrate that KDM6 demethylase function critically regulates DNA damage repair (DDR) gene expression programs in AML. Mechanistically, KDM6 family protein expression is regulated by genotoxic stress, with deficiency of KDM6A (UTX) and KDM6B (JMJD3) impairing DDR transcriptional activation and compromising repair potential. Acquired KDM6A loss-of-function mutations have been implicated in chemoresistance, although a significant percentage of relapsed AML have upregulated KDM6A. Based on these mechanistic findings, olaparib treatment significantly reduced engraftment of patient-derived xenografts. Thus KDM6A-mutant human primary AML samples have increased susceptibility to Poly-(ADP-ribose)-polymerase (PARP) inhibition in vivo. Crucially, a higher KDM6A expression is correlated with venetoclax tolerance. Loss of KDM6A increased mitochondrial activity, BCL2 expression, and sensitized AML cells to venetoclax. Additionally, KDM6A loss was accompanied with a downregulated BCL2A1, which is commonly associated with venetoclax resistance. Corroborating these results, dual targeting of PARP and BCL2 was superior to PARP or BCL2 inhibitor monotherapy in inducing AML apoptosis, and primary AML cells carrying acquired KDM6A-domain mutations were even more sensitive to the combination. Together, our study illustrates a mechanistic rationale in support for a novel combination targeted therapy for human AML based on subtype heterogeneity, and establishes KDM6A as an important molecular regulator for determining therapeutic efficacy.

cancer biology↗