Periodic Reporting for period 4 - residualCRC (MECHANISMS BEHIND RESIDUAL DISEASE IN COLORECTAL CANCER and MODELLING OF THERAPIES THAT PREVENT RELAPSE)
Período documentado: 2025-03-01 hasta 2025-08-31
Colorectal cancer (CRC) is one of the leading causes of cancer death worldwide. Even after surgery to remove the primary tumor, about one in three patients will experience metastatic relapse, due to a few cancer cells that linger undetected by current clinical tests. This minimal residual disease” (MRD) has been difficult to study, leaving a major gap in our understanding and treatment capacity. Our project set out to identify the cancer cells that survive surgery or chemotherapy, to understand how they evade the immune system, and to reveal new ways to eliminate MRD before it causes relapse. These goals are crucial for society, as preventing metastatic recurrence would reduce mortality, improve quality of life, and lower long-term healthcare costs.
By the end of the project, we have generated innovative pre-clinical models of metastatic relapse that, for the first time, allowed to identify a group of cells, marked by the protein EMP1, as the cell of origin of residual disease and metastatic recurrence. We revealed a window of immune vulnerability that can be exploited with neoadjuvant immunotherapy.
In parallel, we uncovered CRC persister cells that survive chemotherapy, marked by the protein Mex3a. Upon exposure to chemotherapy, CRC cells convert to a plastic, fetal intestine-like program that they share with EMP1⁺ cells.
Cellular plasticity also underlies resistance to targeted therapy with KRAS inhibitors: CRCs evade treatment by switching into a cancer stem-like state. Combining KRAS inhibitors with agents that target cancer stem cells enhanced KRAS-directed therapy, opening a new therapeutic avenue focused on blocking cell plasticity.
Our research on how tumour cells evade the action of the immune system defined a dual barrier imposed by TGF-β signaling on immune cells. First, TGF-β prevents the recruitment of fresh CD8+ T lymphocytes from peripheral blood. Second, it reprograms stromal macrophages to produce the protein osteopontin (encoded by SPP1), which in turn suppresses the proliferation of the T cells. These findings nominate SPP1 as a promising therapeutic target.
Together, these discoveries provide actionable biomarkers, inspire and/or support ongoing clinical trials, and establish cancer-cell plasticity and minimal-residual disease biology as new therapeutic frontiers. The project therefore delivers fundamental insights with immediate translational potential.
By the end of the project, we have generated innovative pre-clinical models of metastatic relapse that, for the first time, allowed to identify a group of cells, marked by the protein EMP1, as the cell of origin of residual disease and metastatic recurrence. We revealed a window of immune vulnerability that can be exploited with neoadjuvant immunotherapy.
In parallel, we uncovered CRC persister cells that survive chemotherapy, marked by the protein Mex3a. Upon exposure to chemotherapy, CRC cells convert to a plastic, fetal intestine-like program that they share with EMP1⁺ cells.
Cellular plasticity also underlies resistance to targeted therapy with KRAS inhibitors: CRCs evade treatment by switching into a cancer stem-like state. Combining KRAS inhibitors with agents that target cancer stem cells enhanced KRAS-directed therapy, opening a new therapeutic avenue focused on blocking cell plasticity.
Our research on how tumour cells evade the action of the immune system defined a dual barrier imposed by TGF-β signaling on immune cells. First, TGF-β prevents the recruitment of fresh CD8+ T lymphocytes from peripheral blood. Second, it reprograms stromal macrophages to produce the protein osteopontin (encoded by SPP1), which in turn suppresses the proliferation of the T cells. These findings nominate SPP1 as a promising therapeutic target.
Together, these discoveries provide actionable biomarkers, inspire and/or support ongoing clinical trials, and establish cancer-cell plasticity and minimal-residual disease biology as new therapeutic frontiers. The project therefore delivers fundamental insights with immediate translational potential.
Over the course of the project we developed new experimental models and protocols that, for the first time, allowed detailed study of minimal residual disease in CRC. In brief:
• We identified EMP1 as a biomarker for disseminated tumour cells (DTCs).
• We characterized the transcriptome of EMP1+ cells, revealing similarities to intestinal fetal progenitors.
• We linked Kras mutations to the presence of EMP1+ cells in CRCs.
• Micrometastases were composed of Emp1+ cells which, over time, regenerated the heterogeneity of CRC metastases.
• Ablation of EMP1+ cells prior to primary tumour surgery, prevented disease relapse and cured mice bearing aggressive CRCs, demonstrating that EMP1+ cells are the cell of origin of CRC relapse.
• We dissected the cellular composition of residual-disease niches and found abundant T cell immune infiltrates, which were progressively remodelled and reduced as metastases evolved.
• Thus, we tested neoadjuvant immune checkpoint blockade (ICB), which also prevented disease relapse and cured mice. These results support clinical trials testing neoadjuvant ICB for microsatellite-stable CRCs.
• We investigated EMP1+ cells dependency on KRAS signaling.
• Upon treatment with KRAS inhibitors, CRCs switched to a cancer stem-cell state characterized by high Wnt signaling and marked by LGR5.
• Ablation of Lg5+ cells increased KRAS-inhibitor efficacy.
• These studies revealed cell plasticity as a novel therapeutic target space in CRC.
• Plasticity also underlies chemotherapy resistance by CRC cells.
• Upon chemotherapy, CRCs adopt an intestinal fetal-like phenoptype akin to EMP1+ DTCs which eventually also regenerate full-blown metastases.
• We uncovered Mex3a⁺ as a biomarker for persister cells in triple mutant tumours (APC, KRAS P53; or APC, KRAS, TGF-β). These cells survived chemotherapy in a low proliferative state.
• Knock-out of the Mex3a gene prevented the plastic switch towards the oncofetal state.
• We dissected immune evasion orchestrated by TGF-β signaling revealing a dual immune barrier in evolving metastases.
• TGF-β signaling prevented the recruitment of peripheral CD8 T cells, needed for effective ICB.
• We identified stromal SPP1 as a potential target to enhance ICB downstream of TGF-β signaling.
These findings have been widely disseminated through high-impact publications, conference presentations, and collaborations with clinical researchers. Together, they provide actionable biomarkers and therapeutic strategies to explore in clinical trials.
• We identified EMP1 as a biomarker for disseminated tumour cells (DTCs).
• We characterized the transcriptome of EMP1+ cells, revealing similarities to intestinal fetal progenitors.
• We linked Kras mutations to the presence of EMP1+ cells in CRCs.
• Micrometastases were composed of Emp1+ cells which, over time, regenerated the heterogeneity of CRC metastases.
• Ablation of EMP1+ cells prior to primary tumour surgery, prevented disease relapse and cured mice bearing aggressive CRCs, demonstrating that EMP1+ cells are the cell of origin of CRC relapse.
• We dissected the cellular composition of residual-disease niches and found abundant T cell immune infiltrates, which were progressively remodelled and reduced as metastases evolved.
• Thus, we tested neoadjuvant immune checkpoint blockade (ICB), which also prevented disease relapse and cured mice. These results support clinical trials testing neoadjuvant ICB for microsatellite-stable CRCs.
• We investigated EMP1+ cells dependency on KRAS signaling.
• Upon treatment with KRAS inhibitors, CRCs switched to a cancer stem-cell state characterized by high Wnt signaling and marked by LGR5.
• Ablation of Lg5+ cells increased KRAS-inhibitor efficacy.
• These studies revealed cell plasticity as a novel therapeutic target space in CRC.
• Plasticity also underlies chemotherapy resistance by CRC cells.
• Upon chemotherapy, CRCs adopt an intestinal fetal-like phenoptype akin to EMP1+ DTCs which eventually also regenerate full-blown metastases.
• We uncovered Mex3a⁺ as a biomarker for persister cells in triple mutant tumours (APC, KRAS P53; or APC, KRAS, TGF-β). These cells survived chemotherapy in a low proliferative state.
• Knock-out of the Mex3a gene prevented the plastic switch towards the oncofetal state.
• We dissected immune evasion orchestrated by TGF-β signaling revealing a dual immune barrier in evolving metastases.
• TGF-β signaling prevented the recruitment of peripheral CD8 T cells, needed for effective ICB.
• We identified stromal SPP1 as a potential target to enhance ICB downstream of TGF-β signaling.
These findings have been widely disseminated through high-impact publications, conference presentations, and collaborations with clinical researchers. Together, they provide actionable biomarkers and therapeutic strategies to explore in clinical trials.
There were no effective therapies to prevent relapse after resection of the primary tumor in stage 2 and 3 CRC patients. The reason is that, before this project, the identity and features of residual cancer cells were largely unknown. We have gone beyond the state of the art by:
• Identifying the seeds of relapse. We discovered the CRC cells that seed metastases. Using advanced mouse models and patient samples, we described two key cell types. The first, marked by the protein EMP1, persists after removal of the primary tumor, and is the only cell type found in micro metastases. The second, marked by Mex3a, survives chemotherapy in a slow-dividing state that allows the tumor to regenerate once treatment stops. By defining the molecular “fingerprints” of these two populations, we have provided practical biomarkers—EMP1 and Mex3a—that pathologists can potentially use to detect high-risk patients, monitor minimal residual disease, and guide early, targeted treatments aimed at preventing relapse.
• Revealing a window of immune vulnerability. We discovered that micrometastases, while still small, can be eliminated by immune checkpoint therapy, supporting clinical trials of neoadjuvant immunotherapy for microsatellite-stable colorectal cancer, a group that currently has very limited options.
• Uncovering a novel oncofetal program. We showed that invasive and chemotherapy-exposed cells can adopt a fetal-like genetic state, explaining how they resist treatment and regenerate tumors. Research on this novel Oncofetal state has become a hot topic in the field.
• Establishing cancer-cell plasticity as a therapeutic target. We demonstrated that plasticity underlies resistance not only to chemotherapy but also to modern KRAS inhibitors, and that blocking this process—by combining KRAS inhibitors with agents that target stem-like cells—dramatically improves treatment efficacy.
These breakthroughs create entirely new treatment concepts: giving immunotherapy before surgery, and designing drug combinations that prevent or reverse plastic state changes. In the final months of the project we continue to analyze how the tumor microenvironment evolves over time and to map the full spectrum of plastic states that cancer cells adopt under therapy. These studies aim to translate our discoveries into clinical trials and, ultimately, into therapies that prevent relapse and improve survival for patients with colorectal and other epithelial cancers.
• Identifying the seeds of relapse. We discovered the CRC cells that seed metastases. Using advanced mouse models and patient samples, we described two key cell types. The first, marked by the protein EMP1, persists after removal of the primary tumor, and is the only cell type found in micro metastases. The second, marked by Mex3a, survives chemotherapy in a slow-dividing state that allows the tumor to regenerate once treatment stops. By defining the molecular “fingerprints” of these two populations, we have provided practical biomarkers—EMP1 and Mex3a—that pathologists can potentially use to detect high-risk patients, monitor minimal residual disease, and guide early, targeted treatments aimed at preventing relapse.
• Revealing a window of immune vulnerability. We discovered that micrometastases, while still small, can be eliminated by immune checkpoint therapy, supporting clinical trials of neoadjuvant immunotherapy for microsatellite-stable colorectal cancer, a group that currently has very limited options.
• Uncovering a novel oncofetal program. We showed that invasive and chemotherapy-exposed cells can adopt a fetal-like genetic state, explaining how they resist treatment and regenerate tumors. Research on this novel Oncofetal state has become a hot topic in the field.
• Establishing cancer-cell plasticity as a therapeutic target. We demonstrated that plasticity underlies resistance not only to chemotherapy but also to modern KRAS inhibitors, and that blocking this process—by combining KRAS inhibitors with agents that target stem-like cells—dramatically improves treatment efficacy.
These breakthroughs create entirely new treatment concepts: giving immunotherapy before surgery, and designing drug combinations that prevent or reverse plastic state changes. In the final months of the project we continue to analyze how the tumor microenvironment evolves over time and to map the full spectrum of plastic states that cancer cells adopt under therapy. These studies aim to translate our discoveries into clinical trials and, ultimately, into therapies that prevent relapse and improve survival for patients with colorectal and other epithelial cancers.