Biology subjects

Khan, A. G.

Publications and source records attributed to Khan, A. G..

6 recordsLinked to original sources

bioRxiv2025

Development of antibody-drug conjugates targeting L1CAM to treat metastatic cancer

Effective treatment for metastatic cancer has remained elusive due to the persistence of drug-resistant metastasis stem cells (MetSCs) that drive relapse. MetSCs are tumor cell subpopulations enriched for their ability to reinitiate and sustain metastatic growth, displaying phenotypic plasticity and resistance to chemotherapy. These cells express the L1 cell adhesion molecule (L1CAM), a transmembrane protein detected in numerous human solid tumor types and at multiple disseminated organ sites. As a selective surface marker of MetSCs, L1CAM is a promising candidate for molecularly targeted drugs aimed at eliminating metastases, yet strategies to date have not achieved clinical success. Here, we develop antibody-drug conjugates to deliver highly toxic PNU-159682 payloads to L1CAM-expressing cells. We report the generation of monoclonal antibodies (mAb) with high binding affinity, specificity and selectivity for the human L1CAM extracellular domain. Optimized L1CAM-targeting mAbs were conjugated to PNU-159682 to generate ADC variants with both cleavable and non-cleavable linkers, with an average drug-antibody-ratio (DAR) of four. ADCs derived from three antibodies targeting various epitopes of the L1CAM extracellular portion potently killed cells exhibiting varying levels of surface L1CAM expression. L1CAM ADCs given as monotherapy resulted in robust tumor control and extended survival in mice harboring subcutaneous L1CAM+ xenografts or L1CAM+ lung metastases from triple-negative basal breast cancer and lung adenocarcinoma. Safety analyses with mouse cross-reactive antibodies indicate a feasible therapeutic window. Our findings offer strong proof-of-concept to support the preclinical development of these novel L1CAM ADCs as therapeutic agents for advanced solid tumors.

cancer biology↗

bioRxiv2024

High Throughput Variant Libraries and Machine Learning Yield Design Rules for Retron Gene Editors

The bacterial retron reverse transcriptase system has served as an intracellular factory for single-stranded DNA in many biotechnological applications. In these technologies, a natural retron non-coding RNA (ncRNA) is modified to encode a template for the production of custom DNA sequences by reverse transcription. The efficiency of reverse transcription is a major limiting step for retron technologies, but we lack systematic knowledge of how to improve or maintain reverse transcription efficiency while changing the retron sequence for custom DNA production. Here, we test thousands of different modifications to the retron-Eco1 ncRNA and measure DNA production in pooled variant library experiments, identifying regions of the ncRNA that are tolerant and intolerant to modification. We apply this new information to a specific application: the use of the retron to produce a precise genome editing donor in combination with a CRISPR-Cas9 RNA-guided nuclease (an editron). We use high-throughput libraries in S. cerevisiae to additionally define design rules for editrons. We extend our new knowledge of retron DNA production and editron design rules to human genome editing to achieve the highest efficiency retron-Eco1 editrons to date.

bioengineering↗

bioRxiv2024

NMDA receptor activation drives early synapse formation in vivo

The rules governing neural circuit formation in mammalian central nervous systems are poorly understood. NMDA receptors are involved in synaptic plasticity mechanisms in mature neurons, but their contribution to circuit formation and dendritic maturation remains controversial. Using pharmacological and genetic interventions to disrupt NMDA receptor signaling in hippocampal CA1 pyramidal neurons in vitro and in vivo, we identify an early critical window for a synapse-specific function in wiring Schaffer collateral connections and dendritic arborization. Through in vivo imaging, we show that NMDA receptors are frequently activated during early development and elicit minute-long calcium transients, which immediately precede the emergence of filopodia. These results demonstrate that NMDA receptors drive synapto- and dendritogenesis during development, challenging the view that these processes are primarily mediated by molecular cues.

neuroscience↗

bioRxiv2024

An experimental census of retrons for DNA production and genome editing

Retrons are bacterial immune systems that use reverse transcribed DNA as a detector of phage infection. They are also increasingly deployed as a component of biotechnology. For genome editing, for instance, retrons are modified so that the reverse transcribed DNA (RT-DNA) encodes an editing donor. Retrons are commonly found in bacterial genomes; thousands of unique retrons have now been predicted bioinformatically. However, only a small number have been characterized experimentally. Here, we add substantially to the corpus of experimentally studied retrons. We synthesized >100 previously untested retrons to identify the natural sequence of RT-DNA they produce, quantify their RT-DNA production, and test the relative efficacy of editing using retron-derived donors to edit bacterial genomes, phage genomes, and human genomes. We add 62 new empirically determined, natural RT-DNAs, which are not predictable from the retron sequence alone. We report a large diversity in RT-DNA production and editing rates across retrons, finding that top performing editors outperform those used in previous studies, and are drawn from a subset of the retron phylogeny.

bioengineering↗

bioRxiv2022

Attentional modulation is orthogonal to disinhibition by VIP interneurons in primary visual cortex

Attentional modulation of sensory processing is a key feature of cognition, yet its neural circuit basis is poorly understood. A candidate mechanism is the disinhibition of pyramidal cells through vasoactive intestinal peptide (VIP) and somatostatin (SOM) positive interneurons. However, the interaction of attentional modulation and VIP-SOM disinhibition has never been directly tested. We used all-optical methods to bi-directionally manipulate VIP interneuron activity as mice performed an attention switching task. We measured the activity of VIP, SOM and parvalbumin (PV) positive interneurons and pyramidal neurons identified in the same tissue and found that although activity in all cell classes was modulated by both attention and VIP manipulation, their effects were orthogonal. Attention and VIP-SOM disinhibition relied on distinct patterns of changes in activity and reorganisation of interactions between inhibitory and excitatory cells. Circuit modelling revealed a precise network architecture consistent with multiplexing strong yet non-interacting modulations in the same neural population.

neuroscience↗

bioRxiv2021

Learning and attention increase visual response selectivity through distinct mechanisms

Selectivity of cortical neurons for sensory stimuli can increase across days as animals learn their behavioral relevance, and across seconds when animals switch attention. While both phenomena are expressed in the same cortical circuit, it is unknown whether they rely on similar mechanisms. We imaged activity of the same neuronal populations in primary visual cortex as mice learned a visual discrimination task and subsequently performed an attention switching task. Selectivity changes due to learning and attention were uncorrelated in individual neurons. Selectivity increases after learning mainly arose from selective suppression of responses to one of the task relevant stimuli but from selective enhancement and suppression during attention. Learning and attention differentially affected interactions between excitatory and PV, SOM and VIP inhibitory cell classes. Circuit modelling revealed that cell class-specific top-down inputs best explained attentional modulation, while the reorganization of local functional connectivity accounted for learning related changes. Thus, distinct mechanisms underlie increased discriminability of relevant sensory stimuli across longer and shorter time scales.

neuroscience↗