Periodic Reporting for period 3 - BIOCELLPHE (Ultrasensitive BIOsensing platform for multiplex CELLular protein PHEnotyping at single-cell level)
Período documentado: 2023-10-01 hasta 2025-09-30
BIOCELLPHE tackles the difficulty of detecting and analyzing circulating tumor cells (CTCs), which are extremely rare cancer cells found in the bloodstream. Because current technologies often fail to capture and characterize these cells with enough sensitivity or precision, clinicians lack important information that could improve diagnosis, monitor disease progression, and guide personalized treatment—especially in breast cancer. Improving CTC detection matters for society because it supports earlier and more accurate clinical decisions, ultimately contributing to better outcomes for patients.
To address this challenge, BIOCELLPHE set out to develop an affordable, portable lab‑on‑a‑chip platform capable of identifying multiple protein markers on single CTCs with exceptional sensitivity. The project’s objectives included creating engineered bacterial biosensors able to recognize cancer‑associated proteins, developing optimized SERS‑based detection components, building nanostructured substrates for signal enhancement, and designing AI‑driven analytical tools for reliable data interpretation. These elements were combined into a prototype optofluidic device that demonstrated the feasibility of sensitive CTC detection in both laboratory and preclinical settings, laying the foundation for future personalized cancer diagnostics.
To address this challenge, BIOCELLPHE set out to develop an affordable, portable lab‑on‑a‑chip platform capable of identifying multiple protein markers on single CTCs with exceptional sensitivity. The project’s objectives included creating engineered bacterial biosensors able to recognize cancer‑associated proteins, developing optimized SERS‑based detection components, building nanostructured substrates for signal enhancement, and designing AI‑driven analytical tools for reliable data interpretation. These elements were combined into a prototype optofluidic device that demonstrated the feasibility of sensitive CTC detection in both laboratory and preclinical settings, laying the foundation for future personalized cancer diagnostics.
Throughout the project, BIOCELLPHE brought together experts in synthetic biology, microengineering, nanotechnology, artificial intelligence, and cancer diagnostics to develop a platform for identifying protein markers on CTCs. These efforts were organised into several work packages, each contributing essential components to the final technology.
Engineering bacteria to recognize cancer markers:
The project began by developing E. coli capable of attaching to proteins on breast cancer cells circulating in the bloodstream. Researchers produced small antibody‑like molecules (“nanobodies”) and displayed them on the bacterial surface to bind cancer‑related proteins. Fluorescent markers were added so attachment to cancer cells could be visually confirmed.
Creating new biological sensing systems:
A major part of the work focused on designing mechanisms allowing the bacteria not only to bind cancer cells but also generate a detectable signal. This led to a new class of outer‑membrane sensors, Bacterial Antigen Receptors (BARs), designed to recognise external markers and trigger signalling. Although a fully functional pathway was not completed, key internal components were developed for future optimisation. A patent was filed for this sensing technology.
Producing optical reporter molecules for ultrasensitive detection:
To create measurable signals, the team engineered bacteria to produce Raman reporters, molecules that generate unique optical fingerprints under SERS. Using AI, the most promising reporters were selected based on optical performance and ease of production. Four top candidates were identified, their expression optimised, and three were integrated into the chromosome. These strains reliably produced strong signals for detection.
Developing nanostructured materials and AI tools for SERS:
Researchers created gold nanoparticles and nanostructured surfaces to amplify Raman signals. Two reproducible sensing platforms were produced, one showing particularly high sensitivity. An AI model, SpectralConFormer, was developed to analyse SERS data, accurately identifying and quantifying single or mixed reporters and outperforming conventional methods.
Building and testing the lab‑on‑a‑chip for clinical use:
A modular lab‑on‑a‑chip system was built to isolate, sort, and analyse individual cells using microfluidics. It was tested with model cell lines and later with patient samples. A full clinical framework was established, including ethics approvals, data protection, and standardised protocols. Blood samples from early‑stage and metastatic breast cancer patients showed that engineered bacteria could recognise and label CTCs under clinically relevant conditions. Samples from healthy donors confirmed specificity.
Dissemination and exploitation:
The project produced 25 open‑access publications, participated in 60+ events, created communication materials, organised training, filed a patent, registered a clinical study, and collaborated with other EU projects via Horizon Results Booster. Partners also reviewed IP opportunities and explored future exploitation pathways.
Engineering bacteria to recognize cancer markers:
The project began by developing E. coli capable of attaching to proteins on breast cancer cells circulating in the bloodstream. Researchers produced small antibody‑like molecules (“nanobodies”) and displayed them on the bacterial surface to bind cancer‑related proteins. Fluorescent markers were added so attachment to cancer cells could be visually confirmed.
Creating new biological sensing systems:
A major part of the work focused on designing mechanisms allowing the bacteria not only to bind cancer cells but also generate a detectable signal. This led to a new class of outer‑membrane sensors, Bacterial Antigen Receptors (BARs), designed to recognise external markers and trigger signalling. Although a fully functional pathway was not completed, key internal components were developed for future optimisation. A patent was filed for this sensing technology.
Producing optical reporter molecules for ultrasensitive detection:
To create measurable signals, the team engineered bacteria to produce Raman reporters, molecules that generate unique optical fingerprints under SERS. Using AI, the most promising reporters were selected based on optical performance and ease of production. Four top candidates were identified, their expression optimised, and three were integrated into the chromosome. These strains reliably produced strong signals for detection.
Developing nanostructured materials and AI tools for SERS:
Researchers created gold nanoparticles and nanostructured surfaces to amplify Raman signals. Two reproducible sensing platforms were produced, one showing particularly high sensitivity. An AI model, SpectralConFormer, was developed to analyse SERS data, accurately identifying and quantifying single or mixed reporters and outperforming conventional methods.
Building and testing the lab‑on‑a‑chip for clinical use:
A modular lab‑on‑a‑chip system was built to isolate, sort, and analyse individual cells using microfluidics. It was tested with model cell lines and later with patient samples. A full clinical framework was established, including ethics approvals, data protection, and standardised protocols. Blood samples from early‑stage and metastatic breast cancer patients showed that engineered bacteria could recognise and label CTCs under clinically relevant conditions. Samples from healthy donors confirmed specificity.
Dissemination and exploitation:
The project produced 25 open‑access publications, participated in 60+ events, created communication materials, organised training, filed a patent, registered a clinical study, and collaborated with other EU projects via Horizon Results Booster. Partners also reviewed IP opportunities and explored future exploitation pathways.
Single-cell protein analysis offers critical insights into disease mechanisms and holds strong potential to advance personalized medicine. However, existing methods for characterizing surface biomarkers at the single-cell level remain limited in sensitivity, multiplexing capability, cost and portability, underscoring the need for fundamentally new approaches. BIOCELLPHE aimed to address these limitations by developing a highly sensitive, multiplexed and portable platform for the detection of surface proteins in CTCs, while reducing operational complexity and cost. Upon successful implementation, BIOCELLPHE was expected to deliver a robust, low-cost diagnostic device with ultrasensitive performance, significantly extending the current feasibility limits of single-cell protein phenotyping.
Beyond its technological objectives, BIOCELLPHE has established a new research and innovation framework to address unresolved diagnostic and healthcare challenges, particularly the profiling of circulating tumor cells for cancer diagnostics. By engineering synthetic trans-envelope signaling pathways in bacteria, the project has tackled a long-standing challenge in synthetic biology, sensing large extracellular targets such as proteins, viruses, or cells, thereby unlocking applications previously inaccessible in medical diagnostics and microbial therapeutics. BIOCELLPHE generated new knowledge across multiple disciplines, including programmable bacterial signaling, metabolic engineering, AI-driven design and data analysis, and SERS-based LoC optofluidic systems. These advances are expected to stimulate further research in artificial intelligence, nanotechnology, materials science, microfluidics, and plasmonics, while enabling new industrial opportunities. Socio-economically, BIOCELLPHE aligns with the global shift toward precision medicine, improving treatment stratification and potentially reducing healthcare costs. While its initial focus is cancer, once successfully developed, the technology could be broadly applicable to other diseases, infectious pathogens and sectors such as environmental monitoring, food safety, industrial processing, and security, highlighting its wide-ranging transformative potential.
Beyond its technological objectives, BIOCELLPHE has established a new research and innovation framework to address unresolved diagnostic and healthcare challenges, particularly the profiling of circulating tumor cells for cancer diagnostics. By engineering synthetic trans-envelope signaling pathways in bacteria, the project has tackled a long-standing challenge in synthetic biology, sensing large extracellular targets such as proteins, viruses, or cells, thereby unlocking applications previously inaccessible in medical diagnostics and microbial therapeutics. BIOCELLPHE generated new knowledge across multiple disciplines, including programmable bacterial signaling, metabolic engineering, AI-driven design and data analysis, and SERS-based LoC optofluidic systems. These advances are expected to stimulate further research in artificial intelligence, nanotechnology, materials science, microfluidics, and plasmonics, while enabling new industrial opportunities. Socio-economically, BIOCELLPHE aligns with the global shift toward precision medicine, improving treatment stratification and potentially reducing healthcare costs. While its initial focus is cancer, once successfully developed, the technology could be broadly applicable to other diseases, infectious pathogens and sectors such as environmental monitoring, food safety, industrial processing, and security, highlighting its wide-ranging transformative potential.