Background and rationale - CRC and PDAC are the second and fourth most common cause of
cancer death, respectively. Patients affected by these cancers die of liver MTS.
Conventional therapies are active in either primary tumors but fail in metastatic
disease. The effectiveness of immunotherapy by immune checkpoint blockade and adoptive
cell therapy with tumor-specific T cells is only partial or temporary in MTS from solid
tumors, because of local immunosuppressive mechanisms. This is particularly relevant for
hepatic MTS, in which a natural tolerogenic milieu may further sustain immunosuppression
induced by cancer cells and infiltrating immune cells, such as tumor associated
macrophages (TAMs), myeloid-derived suppressor cells (MDSCs), neutrophils, and regulatory
T or B cells.
In this regard, evidence suggests that progression of primary or metastatic CRC
correlates with poor immune infiltration and inflammation. Moreover, our published
results in primary PDAC support a correlation between tumor progression and a strong type
2 anti-inflammatory/immunosuppressive signature. Efficacy of chemotherapy is known to
rely also on reactivation and/or repolarization of tumor targeting immune responses.
Pre-surgical chemotherapy may induce beneficial effects and potentially targetable
changes in the immune profile of CRC or PDAC MTS.
The investigators hypothesize that a combination of: i) direct cancer cell killing by
adoptively transferred tumor-specific T cells and ii) indirect control of tumor
progression via reprogramming of the tumor microenvironment would control hepatic MTS of
CRC and PDAC.
Thus the investigators propose a thorough characterization of the metastatic milieu in
these two cancer types to rationally shape, on the basis of similarities and differences,
innovative ATMPs.
Experimental results obtained from biological samples will be correlated: i) with
clinical data from patients at the time of enrollment and along their follow-up; and ii)
for a subgroup of patients, with biochemical and immunological data from peripheral blood
samples routinely collected as a part of their clinical follow-up, with the additional
aim to identify novel biologic parameters and/or signatures to help the stratification of
patients and the tailoring of treatments.
Objectives - The investigators want to study:
the tumor mutational burden and the composition of the metastatic tumor
microenvironment in the liver;
the inhibitory pathways that constrain immunity in liver MTS;
the antigenic and clonal breadth of anti-tumor T cell responses;
the spatial-temporal variations of effector and regulatory immune cells upon
therapy;
the characteristics of tumor cells and their microenvironment in liver MTS versus
(vs) primary tumor;
the longitudinal variations of clinical parameters as well as blood-derived
biological and immunological markers during the medical follow-up of patients.
Moreover, they want to collect and biobank patients'samples to support the
validation and selection of the newl ATMPs developed by the other partners of the
translational program.
The primary objectives will be:
to identify immune suppressive/exhaustion pathways
to develop a library of tumor-specific TCR (N≥5/tumor type) and identify new CAR
targets (N≥1/tumor type) in CRC and PDAC MTS
Study design - This clinical protocol aims at deeply characterizing MTS tumor cells and
their microenvironment, including possibly informative dynamic modifications from
pro-tumor to anti-tumor profiles that may follow chemotherapy, which is administered
pre-surgery (neo-adjuvant) to a substantial fraction of patients. PDAC and CRC MTS
patients will be therefore characterized by either cross-sectional or longitudinal
investigations on chemotherapy-naïve vs -treated patients.
In addition, the investigators aim at comparing hepatic MTS with primary tumor, in order
to highlight dynamic qualitative modifications in cancer cells, immune infiltrates and
not-immune microenvironment that could facilitate disease progression toward metastatic
spread. These modifications may offer a window of opportunity for the optimal application
of our ATMPs. To this purpose, primary non-metastatic PDAC patients undergoing surgical
resection will be enrolled in this study and followed-up for monitoring the development
of metachronous liver MTS, that, whenever possible, will be characterized and compared to
their matched/not matched, previously collected, primary tumor samples. Peripheral blood
samples at the time of liver MTS diagnosis will be also collected and analyzed. Also, in
those CRC MTS patients undergoing synchronous resection of primary CRC and hepatic MTS,
comparative analyses will be carried out on tumor from both sites, allowing intra- and
inter-patient profiling of both tumor niches.
Moreover, results from multi-level research analyses will be correlated with clinical
data collected from all patients at the time of enrollment in the study and during their
clinical follow-up, whenever possible for up to 2 years, considering PDAC patients, and 3
years, considering CRC.
In particular, for metastatic CRC patients, the clinical follow-up will be associated
with concurrent collection of blood samples at serial timepoints, that will be
prospectively biobanked for further analyses aimed at: i) studying biological and
immunological correlates of the patient's clinical status; ii) evaluating key
discriminants previously identified in blood samples analyzed at the time of liver MTS
surgery, to serve as early predictors of liver progression or recurrence and possibly as
indicators for therapeutic stratification.
The investigators expect to collect tissue/blood samples and clinical data from at least:
200 metastatic CRC patients (either synchronous or metachronous);
150 non-metastatic PDAC patients;
75 synchronous PDAC MTS patients and about 20 metachronous PDAC MTS patients, the
latter originally enrolled as primary non-metastatic patients for the longitudinal
study, for a total of 95 metastatic PDAC patients;
30 healthy volunteers, as negative controls.
Different biological samples (peripheral blood, primary and/or metastatic tumor tissue,
portal blood) will be collected, in the course of the standard diagnostic and therapeutic
care of those patients, and will be in part directly analyzed and in part biobanked for
further studies. In parallel, clinical data will be collected from patients at the time
of enrollment in the study and, whenever possible, for a follow-up period of up to 2
years, for PDAC patients, and 3 years, for CRC patients.
The statistical analysis of collected data will be performed throughout the clinical
study, starting 6 months after the enrollment of the first patient until 6 months after
the end of patients' follow-up, for a total of 7 years. The study will last 7 years.
Different biological samples (peripheral blood, primary and/or metastatic tumor tissue,
portal blood) will be collected, in the course of the standard diagnostic and therapeutic
care of those patients, and will be in part directly analyzed and in part biobanked for
further studies. In parallel, clinical data will be collected from patients since the
time of enrollment in the study for a follow-up period of 2 years, for PDAC patients, and
of 3 years, for CRC patients.
The statistical analysis of the collected clinical and experimental data will be carried
out all along the duration of the clinical study, starting 6 months after the enrollment
of the first patient until the end of patients' follow-up, for a total of 7 years. The
study will last 7 years.
The research activities will be organized and integrated in 6 work packages (WPs):
Collection and biobanking of the material from patients with CRC and PDAC MTS to the
liver and with primary non-metastatic resectable PDAC.
Definition of the tumor mutational burden, epigenetic and gene expression profile of
the CRC and PDAC metastatic liver at bulk and at single cell level.
Evaluation of the molecular and cellular composition of CRC and, if possible, PDAC
liver MTS by spatial transcriptomics technologies (NICHE-seq and Visium).
Characterization of the immune landscapes of CRC and, if possible, PDAC liver MTS by
high dimensional flow cytometry.
Validation of the molecular results obtained in 2) and 4).
Definition of the antigenic landscape and TCR repertoire of CRC and PDAC liver MTS.
The results obtained by the above mentioned WPs will be crossed with clinical follow-up
data and serve to calibrate the program activities focused on novel ATMPs development.