Peter Vermeulen

Colorectal liver metastases (CLM) exhibit distinct histopathological growth patterns (HGPs) at the metastasis-liver interface, clinically relevant for prognosis and potential predictive value. This study investigates the feasibility of using Fluorine-18 Fluorodeoxyglucose Positron Emission Tomography Computed Tomography ( ¹⁸ F-FDG PET/CT) to preoperatively classify HGPs in CLM and secondly investigate PET parameters as prognostic markers for overall survival (OS). In a prospective study at Umeå University Hospital (June 2013-December 2017), 78 patients with resectable CLM underwent contrast-enhanced liver MRI and ¹⁸ F-FDG PET/CT imaging. HGP classification was performed on tissue sections according to 2017 international guidelines. PET features (SUV _mean [standardised uptake value], SUV _max , metabolic tumour volume [MTV] and first-order texture metrics) were analysed to distinguish encapsulating (eHGP) from replacement (rHGP) patterns. In pretreated patients, multivariate logistic regression including both metabolic and texture parameters identified significant associations for parameters from the fixed level threshold algorithm (VuePoint Sharp IR) with small odds ratios. Overall, PET parameters showed limited ability to discriminate between eHGP and rHGP. OS was superior with eHGP versus rHGP (log-rank p = 0.045). In chemonaïve patients, SUV-based metrics showed unadjusted exploratory associations with OS after liver surgery, varying by reconstruction/delineation and were not significant after correction for multiple testing. FDG-PET-derived parameters were not reliably associated with HGP and did not support non-invasive HGP prediction in this cohort. eHGP was associated with superior survival, underscoring the value of HGP assessment for prognostication while alternative non-invasive biomarkers remain to be identified.

The antibody–drug conjugate trastuzumab deruxtecan has proven to be not only efficient in patients with HER2+ breast cancers (BC), but also in those patients with so-called HER2-low BC. HER2-low tumors are well described in the general BC population, but not in patients with invasive lobular carcinoma (ILC). Here, we aimed at analyzing the association of HER2-low with clinicopathological features and survival outcomes in patients with early-stage pure ILC. A multicentric retrospective cohort of patients diagnosed with stage I-III estrogen receptor positive (ER+) HER2 negative (HER2-) ILC between 01/01/2000 and 12/31/2020 was assembled. HER2- disease was categorized further by immunohistochemical (IHC) score into HER2 0, HER2 1+ and HER2 2+ following time appropriate ASCO/CAP guidelines from 2007 onward and by local guidelines prior to 2007. The association of HER2-low (HER2 1+ and 2+) with clinicopathological variables was assessed using multinomial logistic regression. Survival analyses were performed to evaluate the association of HER2-low with disease-free (DFS), distant recurrence-free (DRFS) and overall survival (OS). The data of 2098 patients with ER+ HER2- ILC was collected of which 1103 (52.6%) had a HER2-low tumor. Of these 716 (34.1%) had an IHC score of HER2 1+ and 387 (18.4%) of HER2 2+. In multivariable analysis, both tumor size of ≥ 2cm (OR: 1.37; 95%CI 1.01 – 1.87; p-value 0.042) and multifocality (OR: 1.55; 95%CI 1.11 – 2.15; p-value 0.009) were associated with HER2-low. HER2-low was associated with worse DFS (HR: 1.32; 95%CI 1.06 − 1.66; p-value 0.015) and OS (HR: 1.42; 95%CI 1.12 − 1.81; p-value 0.004) as compared to HER2 0. No association of HER2-low with DRFS was observed. HER2-low is present in more than half of the patients with early ER+ HER2- pure ILCs and is associated with larger tumor size and multifocality. HER2-low is associated with a worse DFS and OS as compared to HER2 0.

The behaviour of metastases in patients with liver-metastatic colorectal cancer (CRC) is still not adequately considered during treatment planning. However, studies in large cohorts have shown that the disease course in these patients depends on the histopathological growth pattern (HGP) of the liver metastases, with the desmoplastic (or encapsulated) pattern responsible for a favourable outcome and the replacement pattern for an unfavourable course. To increase our knowledge of cancer biology in general as well as to design clinical trials that take into account the diverse behaviour of liver metastases, it is necessary to know the cellular and molecular determinants of these growth patterns. For that purpose, we compared the transcriptome of tumour tissue (prospective cohort; n = 57) sampled very precisely at the transition of metastasis and adjacent liver, between the desmoplastic and replacement HGP. In addition, the mutational profiles for 46 genes related to CRC were extracted from the RNA sequencing reads. First, we show that the genetic constitution of a liver metastasis from colorectal cancer does not determine its HGP. Second, we show clear differences between HGPs regarding the expression of genes belonging to the Molecular Signatures Database hallmark gene sets. Biological themes of the replacement HGP reflect cancer cell proliferation and glucose metabolism, while the desmoplastic HGP is characterized by inflammation and immune response, and angiogenesis. This study supports the view that HGPs are a reflection of the biology of CRC liver metastases and suggests the HGPs are driven epigenetically rather than by specific gene mutations.

Supplementary Figure S3. Supplementary to Figure 3 in the main text. Average number of reads of the rim-enriched genes in the different clusters.

Supplementary Figure S2. Sample exclusion. Sample excluded from the study due to poor tissue quality.

Supplementary Figure S1. Gene distribution and bin radius analysis for sampled reads. Panel A shows the distribution of the number of genes (out of the 15 reads per bin) for each sample. Each bin contains eight unique genes on average. Panel B shows the radius of each bin, i.e., the distance between 15th nearest read and the bin center. The bin radius varied between 5 and 40 μm, with an average bin radius of 17 μm, corresponding to the approximate size of a cell.

Supplementary Table S1. Genes included in the panel together with their corresponding Entrez ID.

Supplementary Table S4. P-values for different neighborhood enrichment tests performed within the rim region in the 10 desmoplastic samples.

Supplementary Table S3. Detection oligo (DO) sequence for each fluorophore.

Supplementary Figure S4. Cluster concentration as a function of distance from liver and tumor. Clusters 0 to 2 and Cluster 8 show, respectively, a bias towards the tumor interface (inner rim) and liver interface (outer rim).