Mismatch Repair Proficient Colorectal Cancer Research Articles

Abstract CT116: First-in-human study of AZD8853, an anti-growth and differentiation factor 15 (GDF15) antibody, in patients (pts) with advanced/metastatic solid tumors

Abstract Background: The cytokine GDF15 is overexpressed in solid malignant tumors such as colorectal, lung and urothelial cancer, where it modulates T cells, dendritic cells (DCs) and myeloid-derived cells, driving the tumor microenvironment toward an immunosuppressive, tumor-promoting state. AZD8853 is a humanized immunoglobulin G1 monoclonal antibody that binds to, and neutralizes, GDF15. Anti-GDF15 treatment increased T cell proliferation and DC activation, leading to an antitumor immune response in preclinical studies of anti-PD-L1 resistant models. In vitro and in vivo preclinical data support the potential antitumor activity of AZD8853 in pts with selected advanced/metastatic cancers. Methods: This Phase I/IIa, first-in-human, open-label study (NCT05397171) assesses the safety, pharmacokinetics (PK), pharmacodynamics (PD) and preliminary efficacy of AZD8853 in pts with histologically or cytologically confirmed locally advanced, unresectable or metastatic mismatch repair-proficient colorectal cancer (pMMR-CRC), non-small-cell lung cancer (NSCLC) and urothelial carcinoma (UC). Up to 165 pts will be enrolled in 3 parts: Part A, dose escalation; Part B, pharmacodynamics expansion; and Part C, efficacy expansion. All pts receive AZD8853 IV. Eligible pts are ≥18 years old with ≥1 measurable target lesion per RECIST v1.1, ECOG PS of 0/1, life expectancy ≥12 weeks and adequate organ and bone marrow function. Pts with NSCLC must have had ≥1 prior line of systemic treatment in the advanced/metastatic setting, and no sensitizing EGFR or ALK aberrations. Pts with pMMR-CRC must have had ≥2 prior treatments in the advanced/metastatic setting. Pts with UC must have had ≥1 prior treatment in the advanced/metastatic setting including platinum-containing therapy and/or a PD-(L)1-inhibitor. Pts with Grade ≥2 unresolved toxicities from prior therapy, symptomatic CNS metastases or leptomeningeal disease, or prespecified active/ongoing infections are excluded. The primary objective is safety, including dose-limiting toxicities, adverse events (AEs), serious AEs and AEs leading to AZD8853 discontinuation. The secondary objectives include assessment of efficacy (objective response rate, disease control rate, duration of response, percentage change from baseline in target lesion size, change from baseline in circulating tumor DNA, and progression-free and overall survival), PK and immunogenicity. Changes in GDF15 serum levels are measured in Parts A and B. Tumoral CD8+ T cell infiltration is measured in a subset of pts from Part B using PET/CT imaging and IHC of paired biopsies. The study is currently recruiting at centers in the USA and Canada with additional sites planned in the UK, France and Spain. Citation Format: Benedito A. Carneiro, Maria Diab, Brian A. Van Tine, Anthony F. Shields, Albiruni Abdul Razak, John F. Hilton, Rafael Santana-Davila, Elhan Sanai, Jorge Zeron-Medina, Veronique Bragulat, Kath Lowery, Arthur Lambert, John Hood, Rakesh Kumar, Duncan Jodrell, Patricia LoRusso. First-in-human study of AZD8853, an anti-growth and differentiation factor 15 (GDF15) antibody, in patients (pts) with advanced/metastatic solid tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 2 (Clinical Trials and Late-Breaking Research); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(8_Suppl):Abstract nr CT116.

Biomarker analysis to predict response in patients with metastatic mismatch repair proficient colorectal cancer treated with regorafenib and nivolumab.

228 Background: We previously conducted a phase I/Ib study with regorafenib and nivolumab in patients with refractory metastatic mismatch repair proficient (pMMR) colorectal cancer (CRC). This study aimed to investigate the biomarkers that predict the treatment response. Methods: Out of the 51 patients who received regorafenib and nivolumab, 22 archival pretreatment tumor samples were subjected to the Xerna TME Panel, a machine learning-based RNA-sequencing biomarker assay and were classified into one of four TME biomarker subtypes: Angiogenesis (A), Immune Active (IA), Immune Desert (ID), or Immune Suppressed (IS). Potential predictive biomarkers including the TME subtypes, KRAS (wild type vs mutant), PD-L1 (negative vs. positive, samples with > 1% tumor cells for PD-L1 were considered positive), CD8 expression (low vs. high), and Treg cells (low vs. high) in tumor microenvironment were evaluated for correlation with overall survival (OS), progression free survival (PFS) and disease control rate (DCR, defined as complete response + partial response + stable disease). Results: Among the 22 patients, 16 (72.7%) had liver metastasis and 15 (68.2%) had lung metastasis. KRAS mutation was found in 16 (68.2%) patients. 11/21 (52.4%) were positive for PD-L1. 12 (54.5%) had high CD8 expression, whereas 9/21 (42.9%) had high Treg cells in tumor microenvironment. Ten (45.5%) patients were classified as biomarker-positive (IA + IS subtypes) and 12 (54.5%) were biomarker-negative (A + ID) based on Xerna TME panel. Two (9.1%) patients achieved partial response, 12 (54.5%) had stable disease, and five (22.7%) developed progressive disease. The median PFS was 5.6 months and median OS was 13.1 months. No significant correlation was observed between RAS mutation (p = 0.664, p = 0.609), PD-L1 expression (p = 0.287, p = 0.173), CD8 (p = 0.152, p = 0.456) and PFS or OS. Low Treg was found to be associated with prolonged PFS (median: 9.8 vs. 1.9 months, p = 0.011) but not OS (p = 0.280). Similarly, only low Treg level was related with DCR (83.3% vs. 33.3%, p = 0.032). While not reaching statistical significance, Xerna TME biomarker-positive patients showed trends for higher median PFS (7.9 months vs. 4.1 months, p = 0.254), median OS (15.75 months vs. 11.9 months, p = 0.378), and higher DCR (70% vs. 58%, p = 0.675) compared to biomarker-negative patients. Additionally, the two patients with partial responses were Xerna TME biomarker-positive. Conclusions: Our study demonstrated that low Treg in tumor microenvironment is correlated with better prognosis in patients with refractory metastatic pMMR CRC who were treated with regorafenib plus nivolumab. Xerna TME panel analysis of these patients also showed trends for predictive clinical benefit. Prospective and larger cohort studies are needed to better define predictive biomarkers for this combination in the future.

A phase I study of a mutant KRAS-targeted long peptide vaccine combined with ipilimumab/nivolumab in resected pancreatic cancer and MMR-proficient metastatic colorectal cancer.

TPS814 Background: Novel strategies are needed to improve immune responses in “cold” tumors such as pancreatic ductal adenocarcinoma (PDAC) and mismatch repair-proficient colorectal cancer (MMRp CRC). As a frequent oncogenic driver, mutant KRAS (mKRAS) neoantigens are attractive targets to augment anti-tumor immunity in both diseases. Recently, adoptive transfer of mKRAS G12D-specific T cells has shown durable tumor regressions in patients with metastatic CRC and PDAC (Tran et. al., 2020; Leidner et. al., 2022). Furthermore, our preclinical work has demonstrated that combining a mKRAS neoantigen vaccine with immune-modulating agents prevents progression of premalignant lesions to PDAC in mice (Keenan et. al., 2014). Based on this rationale, our study pairs a pooled synthetic long peptide (SLP) mKRAS vaccine with dual checkpoint blockade to assess safety and immunogenicity in patients with resected PDAC and chemorefractory MMRp CRC. Methods: This is a first-in-human, single-arm, open-label phase I trial evaluating a pooled SLP mKRAS vaccine combined with ipilimumab/nivolumab (ipi/nivo) in patients with resected PDAC (Cohort A, n = 12) and MMRp metastatic CRC (Cohort B, n = 12) The vaccine consists of poly-ICLC adjuvant admixed with SLPs corresponding to six common mKRAS subtypes: G12D, G12R, G12V, G12A, G12C, and G13D. In priming phase, the mKRAS vaccine is given on days 1, 8, 15, and 22 along with ipi/nivo. In boost phase, the mKRAS vaccine is given on weeks 13, 21, 29, 37, and 45 along with nivo alone. Cohort A patients who remain disease-free can continue to receive boost vaccines in a 12-month extended treatment phase. Eligible patients must have molecular tumor testing that demonstrates one of the six KRAS mutations listed above. Cohort A patients must be disease-free following completion of adjuvant chemotherapy within 6 months prior to study entry. Cohort B patients must have confirmed MMRp status, exposure to ≥ 2 prior lines of standard chemotherapy, and measurable disease amenable to biopsies at baseline and week 7. The co-primary endpoints of this study are safety and T cell response. Adverse events will be graded per NCI CTCAE v5.0. T cell response will be determined by the maximal percent change in IFNγ-producing mKRAS-specific T cell density within 16 weeks post-vaccination compared to baseline. Secondary endpoints include disease control and objective response rates at 16 weeks per RECIST v1.1/iRECIST (Cohort B only) as well as disease-free/progression-free and overall survival. Correlative studies will examine treatment-associated changes in T cell receptor (TCR) repertoire diversity by next-generation TCR sequencing of peripheral blood and tumor specimens. Patient accrual began in May 2020 and is completed for Cohort A. Enrollment is currently ongoing for Cohort B. Study drug support provided by Bristol Myers Squibb. Clinical trial information: NCT04117087 .

Genetic and pharmacological modulation of DNA mismatch repair heterogeneous tumors promotes immune surveillance.

Patients affected by colorectal cancer (CRC) with DNA mismatch repair deficiency (MMRd), often respond to immune checkpoint blockade therapies, while those with mismatch repair-proficient (MMRp) tumors generally do not. Interestingly, a subset of MMRp CRCs contains variable fractions of MMRd cells, but it is unknown how their presence impacts immune surveillance. We asked whether modulation of the MMRd fraction in MMR heterogeneous tumors acts as an endogenous cancer vaccine by promoting immune surveillance. To test this hypothesis, we use isogenic MMRp (Mlh1+/+) and MMRd (Mlh1-/-) mouse CRC cells. MMRp/MMRd cells mixed at different ratios are injected in immunocompetent mice and tumor rejection is observed when at least 50% of cells are MMRd. To enrich the MMRd fraction, MMRp/MMRd tumors are treated with 6-thioguanine, which leads to tumor rejection. These results suggest that genetic and pharmacological modulation of the DNA mismatch repair machinery potentiate the immunogenicity of MMR heterogeneous tumors.

Biomarker concordance between primary colorectal cancer and ovarian metastases: a Dutch cohort study.

The genetic characteristics and mismatch repair (MMR) status of the primary tumor and corresponding metastases in colorectal cancer (CRC) are generally considered to be highly concordant. This implies that either the primary or metastatic tumor can be used for testing gene mutation and MMR status. However, whether this is also true for CRC and their ovarian metastases is currently unknown. Ovarian metastases generally show a poorer response to systemic therapy compared to other metastatic sites. Differences in biomarker status between primary CRC and ovarian metastases could possibly explain this difference in therapy response. The study cohort was selected from CRC patients treated in two Dutch hospitals. Eligible patients with CRC and ovarian metastasis who were surgically treated between 2011 and 2018 were included. CRC and corresponding ovarian metastatic tissues were paired. Gene mutation status was established using next-generation sequencing, while the MMR status was established using either immunohistochemistry or microsatellite instability analysis. Matched samples of CRC and ovarian metastasis from 26 patients were available for analysis. A biomarker concordance of 100% was detected. Complete biomarker concordance was found between MMR proficient CRC and their matching ovarian metastasis. Biomarker testing of MMR proficient CRC tissue appears to be sufficient, and additional testing of metastatic ovarian tissue is not necessary. Differences in therapy response between ovarian metastases and other metastases from CRC are thus unlikely to be caused by differences in the genetic status.

Identification of genes and cellular response factors related to immunotherapy response in mismatch repair-proficient colorectal cancer: a bioinformatics analysis

Mismatch repair-proficient (pMMR) colorectal cancers (CRCs) are thought to be primarily resistant to immune checkpoint inhibitor (ICI) monotherapy. However, recent clinical trials have reported that early-to-mid stage (non-metastatic) CRC responds well to ICI monotherapy. We hypothesized that the efficacy of immunotherapy is linked to a series of gene expression profiles that can characterize the pMMR CRC disease stage. Using The Cancer Genome Atlas (TCGA) CRC data sets, we first investigated transcriptomic features that continuously changed (were continuously upregulated or downregulated) with pMMR CRC disease-stage progression. We defined these gene sets as stage-associated genes. The deconvolution algorithm then enriched these genes with the dynamic changes in the cell type populations of the CRC tumor microenvironment (TME). Finally, the stage-associated genes were cross-referenced to the current transcriptome profile data on ICI treatment of pMMR CRC, which revealed the gene set specifying an effective pMMR tumor response. In total, 774 genes were found to increase in expression and 845 genes to decrease in expression as the stage increased. Using deconvolution methods, we discovered 2 major disease stage-associated alterations in the cellular composition of pMMR CRCs, including changes in cell types involved in host immune responses and tumor cell metastasis. The central memory CD8+ T cell population decreased as the pMMR CRC disease stage increased, but the endothelial cell populations associated with proliferation and metastasis increased. Using a different cell type annotation set (LM22), we discovered that as the disease progressed, M1 macrophages and CD8+ T cells decreased in the TME. In mismatch repair-deficient patients with CRC, however, such a decrease was not observed. Finally, we identified 27 signature genes that can be used to assess ICI efficacy in treatment-naïve patients with pMMR CRC. The current study sought to identify the underlying molecular mechanisms, pathways, and cell landscapes that explain why early-to-mid stage pMMR CRC responds well to ICI treatment. This analysis might be valuable for the selection of patients who might benefit from immunotherapeutic strategies.

Efficacy of neoadjuvant immunotherapy in advanced colorectal cancer: a meta-analysis of cross-sectional studies.

Although neoadjuvant immunotherapy is being widely studied, there is no consensus on its efficacy in microsatellite-stable (MSS) or mismatch repair proficient (pMMR) colorectal cancer (CRC). This meta-analysis aimed to evaluate studies on neoadjuvant immunotherapy for advanced CRC to assess its efficacy and provide new clinical guidelines. We searched literature databases to identify studies that assessed the efficacy of neoadjuvant immunotherapy in advanced CRC. The outcomes evaluated were pathological complete response (pCR), major pathological response (MPR), R0 resection, and anal preservation rates. Heterogeneity among the included studies was assessed by sensitivity analysis, and publication bias was evaluated using Begg and Egger tests. Eleven articles were included in the analysis. The pCR, MPR, R0 resection, and anal preservation rates reported in these studies were 39 and 49, 97, and 76%, respectively. The MSI-H and MSS groups had pooled pCR rates of 70 and 24%, respectively. The pCR rates for the induction, consolidation, and concurrent immuno-chemoradiotherapy (CRT) subgroups were 43, 33, and 27%, respectively, and those for the single and double immunotherapy subgroups were 34 and 40%, respectively. Neoadjuvant immunotherapy combined with CRT is effective in treating MSI-H/dMMR advanced CRC. It could also be a new first-line therapeutic option for MSS/pMMR advanced CRC.

Early radiologic signal of responsiveness to immune checkpoint blockade in microsatellite-stable/mismatch repair-proficient metastatic colorectal cancer

BackgroundImmune checkpoint blockade (ICB) results in radiologic tumour response dynamics that differ from chemotherapy efficacy measures and require an early signal of clinical utility.MethodsPreviously untreated, unresectable microsatellite-stable (MSS)/mismatch repair-proficient (pMMR) colorectal cancer (CRC) patients were randomly assigned to the oxaliplatin-based Nordic FLOX regimen (control arm) or repeat sequential two FLOX cycles and two ICB cycles (experimental arm). The radiologic response was assessed every 8 weeks. In this post hoc analysis, we explored early target lesion (TL) dynamics as indicator of ICB responsiveness. Progression-free survival (PFS) was the primary endpoint.ResultsUsing a landmark analysis approach, we categorised experimental-arm patients into ≥10% (N = 19) or <10% (N = 16) TL reduction at the first post-baseline response assessment. Median PFS for the groups was 16.0 (95% confidence interval (CI), 12.3–19.7) and 3.9 months (95% CI, 2.3–5.5), respectively, superior and inferior (both P < 0.01) to the median PFS of 9.8 months (95% CI, 4.9–14.7) for control arm patients (N = 31).ConclusionsRadiologic TL reduction of ≥10% at the first post-baseline response assessment identified patients with ICB-responsive metastatic MSS/pMMR-CRC. This pragmatic measure may be used to monitor patients in investigational ICB schedules, enabling early treatment adaptation for unresponsive cases.Trial registrationClinicalTrials.gov number, NCT03388190 (02/01/2018).

Abstract 2030: A single-cell spatially resolved map of colorectal cancer identifies novel spatial relationships between cancer cells and the microenvironment

Abstract Understanding the tumor microenvironment (TME) requires more than just a catalog of cell types and gene programs. It is critical to see the spatial organization of the cells are and where they form multicellular interaction networks. Here we present a single-cell spatially resolved transcriptomic analysis of human mismatch repair deficient (MMRd) and proficient (MMRp) colorectal cancer (CRC) specimens. High tumor mutational burden MMRd tumors are known to have an immune response characterized by higher cytolytic T cell infiltrates compared to MMRp tumors, making them an ideal system for spatial single-cell profiling and understanding how the immune-driven programs differ between these tumors. MERFISH is a massively multiplexed single molecule imaging technology which can simultaneously capture and measure the quantity and distribution of hundreds to thousands of RNA species within single cells across a tissue1. We designed a MERFISH library of over 450 genes including genes important to proliferation, apoptosis, immune signaling, immune cell type pathways and other critical pathways in CRC. Patient samples, obtained commercially or through MGH, were hybridized with the designed MERFISH library and stained with a cell boundary marker to delineate cells across the tissue. We performed unsupervised clustering to identify cell types and we calculated spatial statistics to characterize how the cell type distribution varied between MMRd and MMRp tumors. We identified the cellular composition of each tumor, including immune and stromal cells, and the spatial distribution of these cell types. We were able to readily identify all cell types and states previously discovered by single-cell RNA sequencing2 in intact patient specimens, thus providing an accurate map of the cellular composition and spatial organization of these cells in the tumor microenvironment. We transformed these cell types into neighborhoods and discovered a highly organized spatial distribution of most cell types throughout the tumor. While spatial organization was observed in both MMRp and MMRd CRC we saw a significant shift in spatial organization between these tumor classifications notably in the immune population. Further, previously predicted multicellular interaction networks2 appeared as spatially organized structures in the tissue and were distinct in MMRd versus MMRp tumor specimens. Our data provide a richness of concrete hypotheses about which cells are working together, how these cells function cooperatively, and where these cells are located which will be critical in advancing therapy in these immunologically distinct types of colorectal cancer. These cancer maps are critical to truly understand the biology of CRC as well as identify avenues for the development of future therapies for CRC patients. Citation Format: Colles Price, Jonathan H. Chen, Karin Pelka, Sherry Chao, Michael Therrien, Timothy Wiggin, Nicolas Fernandez, Jiang He, George Emanuel, Genevieve Boland, Nir Hacohen. A single-cell spatially resolved map of colorectal cancer identifies novel spatial relationships between cancer cells and the microenvironment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2030.

Abstract 2613: A modified Luria-Delbrück assay allows quantification of colorectal cancer persister cells’ mutation rate

Abstract When cancer cells are exposed to lethal doses of targeted therapies, the emergence of a subpopulation of drug-tolerant persister cells (DTPs) is often observed. We previously reported that colorectal cancer (CRC) cells exposed to targeted therapies activate an adaptive mutability stress response, involving DNA damage induction and a switch to low-fidelity DNA replication. Therefore, targeted treatment might lead to increased mutation rate in DTPs, but mutation rates of cancer cells during treatment have not been quantitatively assessed. Here, we combined biological experiments and mathematical modelling to characterize emergence and dynamics of DTPs. From this, we extrapolated parameters governing dynamics of cancer cells populations and designed a modified Luria-Delbrück assay on mammalian cells (MC-LD) to quantify mutations rates of CRC cells under standard growth conditions and during exposure to targeted therapy. We selected mismatch repair proficient CRC cell lines sensitive to different clinically used therapeutic agents, and derived clones to be used for experiments. By monitoring cell dynamics in drug-response growth assays, we found that CRC cells exposed to targeted therapy display a biphasic killing curve reaching a stable plateau, a pattern indicative of emergence of DTPs. By fitting model estimation to population growth assays, we predicted that, even if a subgroup of DTPs predated treatment, the majority of them emerged only upon exposure to targeted therapies. We also observed that DTPs slowly replicate under treatment, as shown by Carboxy fluorescein succinimidyl ester (CFSE) analysis and staining with 5-ethynyl-2’-deoxyuridine (EdU). We used these population dynamics parameters to design the MC-LD assay. CRC clones were plated in several 96-multiwell plates each, and after an expansion phase in standard culture conditions, treatment was added. After 3-4 weeks, a minority of wells showed growth of resistant colonies: based on the measured growth rates, we could predict that the resistant cells arose before treatment by spontaneous mutation. The remaining wells contained a homogenous population of DTPs. After several weeks of treatment, when pre-treatment resistant clones would have already emerged, late-emerging resistant colonies appeared in a subset of wells in which DTPs had previously been detected. Using the number of residual DTPs and resistant colonies to infer mutation rates, we found a 7- to 50-fold increase (depending on the cell line) in DTPs’ mutation rate compared to sensitive cells.In conclusion, we developed a new assay which allows quantitative comparisons of spontaneous and drug-induced mutation rates in cancer cells and showed that adaptive mutability in DTPs leads to increased mutation rates. This approach could be used to measure whether and how a wide range of environmental conditions affect DTP phenotype and mutation rates in mammalian cells. Citation Format: Alberto Sogari, Mariangela Russo, Simone Pompei, Mattia Corigliano, Giovanni Crisafulli, Andrea Bertotti, Marco Gherardi, Federica Di Nicolantonio, Marco Cosentino Lagomarsino, Alberto Bardelli. A modified Luria-Delbrück assay allows quantification of colorectal cancer persister cells’ mutation rate [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2613.

An integrative in-silico analysis discloses a novel molecular subset of colorectal cancer possibly eligible for immune checkpoint immunotherapy

BackgroundHistorically, the molecular classification of colorectal cancer (CRC) was based on the global genomic status, which identified microsatellite instability in mismatch repair (MMR) deficient CRC, and chromosomal instability in MMR proficient CRC. With the introduction of immune checkpoint inhibitors, the microsatellite and chromosomal instability classification regained momentum as the microsatellite instability condition predicted sensitivity to immune checkpoint inhibitors, possibly due to both high tumor mutation burden (TMB) and high levels of infiltrating lymphocytes. Conversely, proficient MMR CRC are mostly resistant to immunotherapy. To better understand the relationship between the microsatellite and chromosomal instability classification, and eventually discover additional CRC subgroups relevant for therapeutic decisions, we developed a computational pipeline that include molecular integrative analysis of genomic, epigenomic and transcriptomic data.ResultsThe first step of the pipeline was based on unsupervised hierarchical clustering analysis of copy number variations (CNVs) versus hypermutation status that identified a first CRC cluster with few CNVs enriched in Hypermutated and microsatellite instability samples, a second CRC cluster with a high number of CNVs mostly including non-HM and microsatellite stable samples, and a third cluster (7.8% of the entire dataset) with low CNVs and low TMB, which shared clinical-pathological features with Hypermutated CRCs and thus defined Hypermutated-like CRCs. The mutational features, DNA methylation profile and base substitution fingerprints of these tumors revealed that Hypermutated-like patients are molecularly distinct from Hypermutated and non-Hypermutated tumors and are likely to develop and progress through different genetic events. Transcriptomic analysis highlighted further differences amongst the three groups and revealed an inflamed tumor microenvironment and modulation Immune Checkpoint Genes in Hypermutated-like CRCs.ConclusionTherefore, our work highlights Hypermutated-like tumors as a distinct and previously unidentified CRC subgroup possibly responsive to immune checkpoint inhibitors. If further validated, these findings can lead to expanding the fraction of patients eligible to immunotherapy.

Pembrolizumab and maraviroc in refractory mismatch repair proficient/microsatellite-stable metastatic colorectal cancer – The PICCASSO phase I trial

BackgroundPD-1/PD-L1 inhibitors do not show activity in mismatch repair proficient (MMRp) colorectal cancer. Inhibition of C–C motif chemokine receptor 5 (CCR5) leads to an antitumoral activation of macrophages, affecting immune cell infiltrates. PICCASSO is a phase I trial exploring safety and efficacy of pembrolizumab and maraviroc in refractory MMRp CRC. MethodsTwenty patients received pembrolizumab and maraviroc (core period, eight cycles), followed by pembrolizumab monotherapy. Primary endpoint was the feasibility rate (patients without treatment-related grade ≥3 immune-related adverse events, treatment-related grade ≥4 adverse events, or any toxicity-related premature withdrawal of treatment). Secondary endpoints included safety/toxicity, overall response rate (ORR), progression-free survival (PFS), and overall survival (OS). Optional biopsies of liver metastases were performed for analyses of the micromilieu. ResultsThe feasibility rate was 94.7% [90% CI 77.4–99.7%], with one grade 4 hyperglycemia and no additional ≥ grade 3 treatment-related toxicities. ORR according to RECIST was 5.3%. Median PFS according to RECIST was 2.10 months [95%CI 1.68–2.30], median OS 9.83 months [95% CI, 5.59–20.02]. Disease control rate of poststudy salvage treatment was >70%. Translational analyses showed an increase of antitumoral chemokines during treatment; eotaxin, a chemokine involved in chemotaxis, was identified as a biomarker linked to OS. ConclusionsTherapy with pembrolizumab and maraviroc was feasible and showed a beneficial toxicity pattern. Clinical activity in MMRp CRC patients was limited with prolonged disease stabilizations observed in single patients. Efficacy of poststudy salvage treatment and OS was higher than expected in this heavily pretreated population. This trial is registered at clinicaltrials.govNCT03274804.

Neoadjuvant immunotherapy in microsatellite stability or mismatch repair proficient colorectal cancer

Immunotherapy has become an important treatment option for microsatellite instability-high (MSI-H) and mismatch repair deficient (dMMR) colorectal cancer. From late-line to first-line treatment, and even in neoadjuvant setting for early stage colorectal cancer, promising efficacy was observed with immunotherapy. In microsatellite stability (MSS) or mismatch repair proficient (pMMR) colorectal cancer, the researches of neoadjuvant immunotherapy have been conducted constantly. This paper focuses on the recent researches and progress of neoadjuvant immunotherapy for MSS or pMMR colorectal cancer. Neoadjuvant immunotherapy alone led to a good pathological response in a subset of patients. Studies of induction or consolidation immunotherapy before or after neoadjuvant chemoradiotherapy or concurrent immunotherapy during radiotherapy showed higher pathological complete remission (pCR) rates as compared to standard chemoradiotherapy. Studies on sequential dual immunotherapy after radiochemotherapy and targeted therapy combined with neoadjuvant immunotherapy are ongoing. At present, most of these are pilot studies with small sample size. More researches and long-term follow-up are needed to prove the efficacy of neoadjuvant immunotherapy in MSS or pMMR colorectal cancer.

Abstract P061: Dendritic cell paucity in mismatch repair-proficient colorectal cancer liver metastases limits the efficacy of immune checkpoint blockade

Abstract Liver metastasis is a major cause of mortality for patients with colorectal cancer (CRC). Mismatch repair-proficient (pMMR) CRCs make up about 95% of metastatic CRCs, and are unresponsive to immune checkpoint blockade (ICB) therapy. Here we show that mouse models of orthotopic pMMR CRC liver metastasis accurately recapitulate the inefficacy of ICB therapy in patients, whereas the same pMMR CRC tumors are sensitive to ICB therapy when grown subcutaneously. To reveal local, nonmalignant components that determine CRC sensitivity to treatment, we compared the microenvironments of pMMR CRC cells grown as liver metastases and subcutaneous tumors. We found a paucity of both activated T cells and dendritic cells in ICB-treated orthotopic liver metastases, when compared to their subcutaneous tumor counterparts. Furthermore, treatment with FMS-like tyrosine kinase 3 ligand (Flt3L) plus ICB therapy increased dendritic cell infiltration into pMMR CRC liver metastases and improved mouse survival. Lastly, we show that human CRC liver metastases and microsatellite stable (MSS) primary CRC have a similar paucity of T cells and dendritic cells. These studies indicate that orthotopic tumor models, but not subcutaneous models, should be used to guide human clinical trials. Our findings also posit dendritic cells as antitumor components that can increase the efficacy of immunotherapies against pMMR CRC. Citation Format: William W. Ho, Igor L. Gomes-Santos, Shuichi Aoki, Meenal Datta, Kosuke Kawaguchi, Nilesh P Talele, Sylvie Roberge, Jun Ren, Hao Liu, Ivy X Chen, Patrik Andersson, Sampurna Chatterjee, Ashwin S. Kumar, Zohreh Amoozgar, Qixian Zhang, Peigen Huang, Mei Rosa Ng, Vikash P Chauhan, Lei Xu, Dan G. Duda, Jeffrey W. Clark, Mikael J. Pittet, Dai Fukumura, Rakesh K Jain. Dendritic cell paucity in mismatch repair-proficient colorectal cancer liver metastases limits the efficacy of immune checkpoint blockade [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2021 Oct 5-6. Philadelphia (PA): AACR; Cancer Immunol Res 2022;10(1 Suppl):Abstract nr P061.

RNF43 pathogenic Germline variant in a family with colorectal cancer.

The role of RNF43 as a cause of an inherited predisposition to colorectal cancer (CRC) is yet to be fully explored. This report presents our findings of two individuals with CRC from a single family carrying a likely-pathogenic inherited germline variant in RNF43. The proband (III:1) and the proband's mother (II:2) were diagnosed with mismatch repair proficient CRCs at the age of 50 years and 65 years, respectively. Both patients had BRAFV600E mutated colon tumours, indicating that the CRCs arose in sessile serrated lesions. The germline variant RNF43:c.375+1G>A was identified in both patients. RNA studies showed that this variant resulted in an aberrantly spliced transcript, which was predicted to encode RNF43:p.Ala126Ilefs*50 resulting in premature termination of protein synthesis and was classified as a likely-pathogenic variant. Our report adds further evidence to the hereditary role of RNF43 as a tumour suppressor gene in colorectal tumorigenesis and supports the inclusion of RNF43 as a gene of interest in the investigation of CRC predispositions outside the setting of serrated polyposis.

BRAF Mutations as Actionable Targets: A Paradigm Shift in the Management of Colorectal Cancer and Novel Avenues.

BRAF mutations in colorectal cancer have been studied over the past several decades. BRAF V600E mutation, a class I mutation, is the most common oncogenic BRAF alteration in colorectal cancer. Until recently, the BRAF V600E mutation was not among actionable genes for colorectal cancer. However, recent discoveries have revealed therapeutic opportunities. The BRAF with or without MEK inhibition combined with epidermal growth factor receptor-directed therapy was recently found to be an effective therapy choice for patients with advanced-stage BRAF V600-mutant colorectal cancer. However, it is essential to distinguish patients with BRAF V600E-mutant mismatch repair-deficient colorectal cancer from those with mismatch repair-proficient colorectal cancer, as immune checkpoint inhibitor therapy is more appealing in this subset of patients with colorectal cancer. This review article discusses the molecular characteristics of class I, II, and III BRAF mutants and their impact on the clinical behavior of colorectal cancer. We also review the recent progress in the targetability of BRAF mutations in colorectal cancer, which has led to changes in clinical practice and elaborates on innovative therapeutic approaches to enhance the efficacy of BRAF-targeting therapies, to achieve more durable responses.

Phase II study of preoperative (PREOP) chemoradiotherapy (CTRT) plus avelumab (AVE) in patients (PTS) with locally advanced rectal cancer (LARC): The AVANA study.

3511 Background: Preop CTRT is considered the standard of care in the management of LARC. RT can induce antigen release from a low neoantigen-burden tumor (such as a mismatch repair proficient colorectal cancer) and activate dendritic cells leading to a CD8+ T lymphocyte-mediated anticancer immune response. In LARC patients, neoadjuvant CTRT increases PD-L1 expression in tumor cells, strongly suggesting a neoadjuvant combinatory strategy with RT and PD-1/PD-L1 pathway blockade. Based on such considerations, we have designed the AVANA study to investigate the role of Ave in combination with preop CTRT in LARC. Methods: This is an Italian multi-center, phase II study. Pts with resectable LARC, defined by the presence of at least one of the following features: cN+, cT4, high risk cT3, received standard preop CTRT (capecitabine 825 mg/sqm/bid 5 days/week+ 50.4 Gy in 28 fractions over 5.5 weeks) plus 6 cycles of Ave 10 mg/Kg every 2 weeks. Surgery with total mesorectal excision was performed at 8-10 weeks after the end of CTRT. The primary end-point was the pCR rate, defined as complete histological regression with no available tumor cells ypT0N0. Secondary end-points were R0 resection rate, tumor downstaging, local recurrence, sphincter preservation rate, progression-free survival, overall survival, safety profile, and the evaluation of exploratory predictive and/or prognostic biomarkers. Assuming as null hypothesis p0 a pCR rate of 15%, a significance level of 5% (one-side), and a power of 80%, a sample size of 101 pts was needed to detect an absolute increment of 10% in pCR rate (from 15% to 25%). The experimental regimen is considered for further studies if, in at least 22 pts, we observe a pCR. Results: From April 2019 to November 2020, a total of 101 resectable LARC pts were enrolled in 10 Italian Centers. The median age was 63 years (23-82), 62 (61.4%) pts were male, 93 (92%) had ECOG PS 0. At baseline, 94 (93%) and 16 (16%) pts had cN+ and cT4 LARC, respectively. All pts completed the induction phase. Out of 96 pts evaluable for pathological response, 22 (23%) pts achieved a pCR and 59 (61.5%) pts a major pathological response (a central review is ongoing). At this time, microsatellite status is available only in 39 pts, of which only one was instable. The rate of grade 3-4 non-immune and immune-related adverse events was 8% and 4%, respectively. Avelumab was early interrupted in 9 pts out 101, mainly due to toxicity. Conclusions: The combination of preop CTRT plus Ave showed a promising activity and a feasible safety profile. According to our statistical considerations, the experimental regimen will be considered for further studies. Updated results will be presented during the Congress. Sponsored by GONO and partially supported by Merck. EUDRACT 2017-003582-10. Clinical trial information: NCT03854799.

Multicellular immune hubs and their organization in MMRd and MMRp colorectal cancer

Abstract Immune responses to cancer are highly variable, with mismatch repair-deficient (MMRd) tumors exhibiting more anti-tumor immunity than mismatch repair-proficient (MMRp) tumors. To understand the rules governing these varied responses, we transcriptionally profiled 371,223 cells from colorectal tumors and adjacent normal tissues of 28 MMRp and 34 MMRd patients. Analysis of 88 cell subsets and their 204 associated gene expression programs revealed extensive transcriptional and spatial remodeling across tumors. To discover hubs of interacting malignant and immune cells, we identified expression programs in different cell types that co-varied across patient tumors and used spatial profiling to localize coordinated programs. We discovered a myeloid cell-attracting hub at the tumor-luminal interface associated with tissue damage, and an MMRd-enriched immune hub within the tumor, with activated T cells together with malignant and myeloid cells expressing T-cell-attracting chemokines. By identifying interacting cellular programs, we thus reveal the logic underlying spatially organized immune-malignant cell networks.

Olaparib-mediated enhancement of 5-fluorouracil cytotoxicity in mismatch repair deficient colorectal cancer cells

BackgroundThe advances in colorectal cancer (CRC) treatment include the identification of deficiencies in Mismatch Repair (MMR) pathway to predict the benefit of adjuvant 5-fluorouracil (5-FU) and oxaliplatin for stage II CRC and immunotherapy. Defective MMR contributes to chemoresistance in CRC. A growing body of evidence supports the role of Poly-(ADP-ribose) polymerase (PARP) inhibitors, such as Olaparib, in the treatment of different subsets of cancer beyond the tumors with homologous recombination deficiencies. In this work we evaluated the effect of Olaparib on 5-FU cytotoxicity in MMR-deficient and proficient CRC cells and the mechanisms involved.MethodsHuman colon cancer cell lines, proficient (HT29) and deficient (HCT116) in MMR, were treated with 5-FU and Olaparib. Cytotoxicity was assessed by MTT and clonogenic assays, apoptosis induction and cell cycle progression by flow cytometry, DNA damage by comet assay. Adhesion and transwell migration assays were also performed.ResultsOur results showed enhancement of the 5-FU citotoxicity by Olaparib in MMR-deficient HCT116 colon cancer cells. Moreover, the combined treatment with Olaparib and 5-FU induced G2/M arrest, apoptosis and polyploidy in these cells. In MMR proficient HT29 cells, the Olaparib alone reduced clonogenic survival, induced DNA damage accumulation and decreased the adhesion and migration capacities.ConclusionOur results suggest benefits of Olaparib inclusion in CRC treatment, as combination with 5-FU for MMR deficient CRC and as monotherapy for MMR proficient CRC. Thus, combined therapy with Olaparib could be a strategy to overcome 5-FU chemotherapeutic resistance in MMR-deficient CRC.

CMTM6 expression in M2 macrophages is a potential predictor of PD-1/PD-L1 inhibitor response in colorectal cancer

BackgroundCMTM6 is a novel key regulator of PD-L1. High expression of both CMTM6 and PD-L1 may predict the benefit of PD-1 axis blockade in lung cancer. We aimed to investigate the expression pattern of CMTM6 between mismatch repair-defective (dMMR) and mismatch repair-proficient (pMMR) colorectal cancer (CRC) tissues and assess its correlation with the response to PD-1/PD-L1 pathway blockade.MethodsImmunohistochemistry (IHC) was used to analyze CMTM6 and PD-L1 expression and immune cell density in dMMR/pMMR CRC. Quantitative multiplex immunofluorescence (IF) was performed to detect CMTM6, PD-L1, CD4, CD8, CD68 and CD163 expression in CRC patients treated with PD-1/PD-L1 inhibitors.ResultIHC analysis showed that CMTM6 and PD-L1 were both expressed in tumor cells (TCs) and invasion front immune cells (ICs). CMTM6 and PD-L1 expression and CD4+, CD8+, CD68+ or CD163+ cell density were significantly higher in dMMR CRC patients than in pMMR CRC patients. CMTM6 expression was positively correlated with PD-L1 expression and CD163+ M2 macrophage density in dMMR CRC. IF analysis showed that the coexpression rate of CMTM6/PD-L1 and the expression rate of CMTM6 in CD8+ T cells and CD163+ M2 macrophages were significantly increased in the group that exhibited clinical benefit. CMTM6 expression in M2 macrophages was identified as the best biomarker for predicting the responsiveness to PD-1/PD-L1 inhibitors.ConclusionsCMTM6 expression in M2 macrophages may predict the PD-1/PD-L1 inhibitor response rate in CRC patients more accurately than dMMR/microsatellite instability-high (MSI-H) status. It can also identify pMMR CRC patients who could benefit from PD-1/PD-L1 inhibitors.