TRIPODD: a Novel Fluorescence Imaging Platform for In Situ Quantification of Drug Distribution and Therapeutic Response

McMahon, Nathan P.; Solanki, Allison; Wang, Lei G.; Montaño, Antonio R.; Jones, Jocelyn; Samkoe, Kimberley S.; Tichauer, Kenneth M.; Gibbs, Summer L.

doi:10.1007/s11307-021-01589-x

Cited by 5 publications

(6 citation statements)

References 66 publications

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“…1B-D). The targeted and untargeted behavior of the iPAI probes has been extensively validated in silico, in vitro, and in vivo as previously reported [58,75].…”

Section: Synthesis Of Fluorescently Labeled Erlotinibmentioning

confidence: 80%

“…After image acquisition, a custom MATLAB script (10.5281/zenodo.4004647) was used to calculate ratiometic DTA images on a per pixel basis for each tissue image. DTA, as previously reported [75], was calculated as Drug Target Availability (DTA) =…”

Section: Pharmacokinetic Study Of Ipai Probesmentioning

confidence: 99%

“…In summary, the ideal assay to measure therapeutic response requires concomitant measurement of: 1) the extent and heterogeneity of DTA across a tissue as well as 2) drug interactions with the drug target protein along with off-target effects that allow for correlations to be made between efficacy, toxicity and dosing 13 . To bridge this unmet analytical gap, we have previously reported our novel fluorescence imaging platform, T herapeutic R esponse I maging through P roteomic and O ptical D rug D istribution ( TRIPODD ) 75 . TRIPODD facilitates simultaneous single-cell quantification of DTA with iPAI and the associated tumor biology through accurate segmentation of spatially aligned tumor cells based on Ab-oligo cyCIF.…”

Section: Introductionmentioning

confidence: 99%

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In situ single-cell therapeutic response imaging facilitated by the TRIPODD fluorescence imaging platform

McMahon,

Solanki,

Wang

et al. 2024

Theranostics

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Purpose: Small molecule drugs such as tyrosine kinase inhibitors (TKIs) targeting tumoral molecular dependencies have become standard of care for numerous cancer types. Notably, epidermal growth factor receptor (EGFR) TKIs (e.g., erlotinib, afatinib, osimertinib) are the current first-line treatment for non-small cell lung cancer (NSCLC) due to their improved therapeutic outcomes for EGFR mutated and overexpressing disease over traditional platinum-based chemotherapy. However, many NSCLC tumors develop resistance to EGFR TKI therapy causing disease progression. Currently, the relationship between in situ drug target availability (DTA), local protein expression and therapeutic response cannot be accurately assessed using existing analytical tools despite being crucial to understanding the mechanism of therapeutic efficacy. Procedure: We have previously reported development of our fluorescence imaging platform termed TRIPODD (Therapeutic Response Imaging through Proteomic and Optical Drug Distribution) that is capable of simultaneous quantification of single-cell DTA and protein expression with preserved spatial context within a tumor. TRIPODD combines two complementary fluorescence imaging techniques: intracellular paired agent imaging (iPAI) to measure DTA and cyclic immunofluorescence (cyCIF), which utilizes oligonucleotide conjugated antibodies (Ab-oligos) for spatial proteomic expression profiling on tissue samples. Herein, TRIPODD was modified and optimized to provide a downstream analysis of therapeutic response through single-cell DTA and proteomic response imaging. Results: We successfully performed sequential imaging of iPAI and cyCIF resulting in high dimensional imaging and biomarker assessment to quantify single-cell DTA and local protein expression on erlotinib treated NSCLC models. Pharmacodynamic and pharmacokinetic studies of the erlotinib iPAI probes revealed that administration of 2.5 mg/kg each of the targeted and untargeted probe 4 h prior to tumor collection enabled calculation of DTA values with high Pearson correlation to EGFR, the erlotinib molecular target, expression in the tumors. Analysis of single-cell biomarker expression revealed that a single erlotinib dose was insufficient to enact a measurable decrease in the EGFR signaling cascade protein expression, where only the DTA metric detected the presence of bound erlotinib. Conclusion:We demonstrated the capability of TRIPODD to evaluate therapeutic response imaging to erlotinib treatment as it relates to signaling inhibition, DTA, proliferation, and apoptosis with preserved spatial context.

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“…1B-D). The targeted and untargeted behavior of the iPAI probes has been extensively validated in silico, in vitro, and in vivo as previously reported [58,75].…”

Section: Synthesis Of Fluorescently Labeled Erlotinibmentioning

confidence: 80%

Section: Pharmacokinetic Study Of Ipai Probesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

In situ single-cell therapeutic response imaging facilitated by the TRIPODD fluorescence imaging platform

McMahon,

Solanki,

Wang

et al. 2024

Theranostics

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show abstract

“…This study depicts how protein therapeutics acting at the microscopic scale can further inform its tissue distribution and ultimate response. Finally, our collaborators developed TRIPODD (Therapeutic Response Imaging through Proteomic and Optical Drug Distribution) to evaluate in situ drug target availability with combination of paired-agent imaging and cyclic immunofluorescence [33]. The difference between TRIPODD with PAI presented in this study is that the imaging agents used are cell membrane permeable to achieve intracellular receptor imaging.…”

Section: Discussionmentioning

confidence: 99%

Improved Discrimination of Tumors with Low and Heterogeneous EGFR Expression in Fluorescence-Guided Surgery Through Paired-Agent Protocols

Wang

Folaron

et al. 2021

Mol Imaging Biol

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Purpose The goal of fluorescence-guided surgery (FGS) in oncology is to improve the surgical therapeutic index by enhancing contrast between cancerous and healthy tissues. However, optimal discrimination between these tissues is complicated by the nonspecific uptake and retention of molecular targeted agents and the variance of fluorescence signal. Paired-agent imaging (PAI) employs co-administration of an untargeted imaging agent with a molecular targeted agent, providing a normalization factor to minimize nonspecific and varied signals. The resulting measured binding potential is quantitative and equivalent to in vivo immunohistochemistry of the target protein. This study demonstrates that PAI improves the accuracy of tumor-to-healthy tissue discrimination compared to single-agent imaging for in vivo FGS. Procedures PAI using a fluorescent anti-epidermal growth factor receptor (EGFR) affibody molecule (ABY-029, eIND 122,681) with untargeted IRDye 700DX carboxylate was compared to ABY-029 alone in an oral squamous cell carcinoma xenograft mouse model at 3 h after dye administration (n = 30). Results PAI significantly enhanced tumor discrimination, as compared to ABY-029 alone in low EGFR-expressing tumors and highly heterogeneous populations including multiple cell lines with varying expression (diagnostic accuracy: 0.908 vs. 0.854 and 0.908 vs. 0.822; and ROC curve AUC: 0.963 vs. 0.909 and 0.957 vs. 0.909, respectively) indicating a potential for universal FGS image thresholds to determine surgical margins. In addition, PAI achieved significantly higher diagnostic ability than ABY-029 alone 0.25–5-h post injection and exhibited a stronger correlation to EGFR expression heterogeneity. Conclusion The quantitative receptor delineation of PAI promises to improve the surgical therapeutic index of cancer resection in a clinically relevant timeline.

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“…[2] While advances in fluorescence-based tissue histological techniques have enabled the visualization and quantification of target engagement in processed tissue samples, these assays are limited in their ability to simultaneously identify and characterize the phenotypes associated with the cell types present. [3] Furthermore, these approaches require either the introduction of a fluorophore or bioorthogonal handle which may affect the properties of the target molecule. Due to the inherent challenges with multiplexing fluorescence-based microscopy, combining biodistribution with cellular characterization is best suited to Imaging Mass Cytometry™ (IMC™).…”

Section: Introductionmentioning

confidence: 99%

Tellurophene‐Tagging of Teniposide Facilitates Monitoring by Mass Cytometry

et al. 2022

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Target engagement and the biodistribution of exogenously administered small molecules is rarely homogenous. Methods to determine the biodistribution at the cellular level are limited by the ability to detect the small molecule and simultaneously identify the cell types or tissue structures with which it is associated. The highly multiplexed nature of mass cytometry could facilitate these studies provided a heavy isotope label was available in the molecule of interest. Here we show it is possible to append a tellurophene to a known chemotherapeutic, teniposide, to follow this molecule in vivo. A semi-synthetic approach offers an efficient route to the teniposide analogue which is found to have similar characteristics when compared with the parent teniposide in vitro. Using mass cytometry we find the teniposide analogue has significant nonspecific binding to cells. In vivo the tellurium bearing teniposide produces the expected DNA damage in a PANC-1 xenograft model. The distribution of Te in the tissue is near the limits of detection and further work will be required to characterize the localization of this analogue with respect to cell type distributions.

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TRIPODD: a Novel Fluorescence Imaging Platform for In Situ Quantification of Drug Distribution and Therapeutic Response

Cited by 5 publications

References 66 publications

In situ single-cell therapeutic response imaging facilitated by the TRIPODD fluorescence imaging platform

In situ single-cell therapeutic response imaging facilitated by the TRIPODD fluorescence imaging platform

Improved Discrimination of Tumors with Low and Heterogeneous EGFR Expression in Fluorescence-Guided Surgery Through Paired-Agent Protocols

Tellurophene‐Tagging of Teniposide Facilitates Monitoring by Mass Cytometry

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