Background
Esophagogastric adenocarcinoma (EGA) presents a substantial global health challenge as the number of cases continues to rise. The current standard approach for treating localized EGA ...involves a combination of triplet chemotherapy, which consists of a platinum compound, a fluoropyrimidine, and a taxane (known as FLOT), followed by surgery. In cases of metastatic EGA with HER2-positive status or in certain studies with localized EGA, the use of HER2-targeted antibodies such as trastuzumab has shown improved responses. Recently, the addition of programmed cell death protein 1 (PD-1) inhibitors, such as pembrolizumab, when combined with 5-FU, platinum-based chemotherapy, and trastuzumab, has demonstrated significant enhancements in response rates for HER2-positive metastatic EGA. However, there is currently insufficient evidence regarding this treatment approach in localized HER2-positive disease.
Methods
The PHERFLOT study is an open-label, single-arm, multicenter, exploratory phase II trial designed to assess the efficacy, safety, and tolerability of perioperative pembrolizumab, FLOT, and trastuzumab in patients with previously untreated localized HER2-positive EGA. In total, 30 patients will be recruited. The co-primary end points are pathological complete response rate and disease-free survival rate after 2 years. Secondary objectives include safety and tolerability, efficacy in terms of progression-free survival and objective response rate and translational markers, such as blood-based signatures (e.g., immune repertoire changes or emergence of anti-HER2 resistance variants) or microbiota signatures that may correlate with immune activation and therapy response.
Discussion
Recent evidence from phase II clinical trials demonstrated improved efficacy through the addition of trastuzumab to perioperative FLOT. Furthermore, in advanced or metastatic EGA, the combination of trastuzumab, FLOT, and the PD1-inhibitor pembrolizumab significantly improved treatment response. The PHERFLOT study aims to assess the efficacy and safety of this treatment approach in HER2-positive–localized EGA, potentially identifying a promising new perioperative regimen for localized EGA, which then needs to be confirmed within a randomized trial. Furthermore, the accompanying translational program of the study might help to improve the stratification of suitable patients and to identify potential translational targets for future clinical trials.
Clinical trial registration
https://clinicaltrials.gov
, identifier NCT05504720.
BackgroundIn patients with microsatellite stable (MSS) metastatic colorectal cancer (mCRC), immune checkpoint blockade is ineffective, and combinatorial approaches enhancing immunogenicity need ...exploration.MethodsWe treated 43 patients with predominantly microsatellite stable RAS/BRAF wild-type mCRC on a phase II trial combining chemotherapy with the epidermal growth factor receptor antibody cetuximab and the programmed cell death ligand 1 (PD-L1) antibody avelumab. We performed next-generation gene panel sequencing for mutational typing of tumors and liquid biopsy monitoring as well as digital droplet PCR to confirm individual mutations. Translational analyses included tissue immunohistochemistry, multispectral imaging and repertoire sequencing of tumor-infiltrating T cells. Detected PD-L1 mutations were mechanistically validated in CRISPR/Cas9-generated cell models using qRT-PCR, immunoblotting, flow cytometry, complement-dependent cytotoxicity assay, antibody-dependent cytotoxicity by natural killer cell degranulation assay and LDH release assay as well as live cell imaging of T cell mediated tumor cell killing.ResultsCirculating tumor DNA showed rapid clearance in the majority of patients mirroring a high rate of early tumor shrinkage. In 3 of 13 patients expressing the high-affinity Fcγ receptor 3a (FcγR3a), tumor subclones with PD-L1 mutations were selected that led to loss of tumor PD-L1 by nonsense-mediated RNA decay in PD-L1 K162fs and protein degradation in PD-L1 L88S. As a consequence, avelumab binding and antibody-dependent cytotoxicity were impaired, while T cell killing of these variant clones was increased. Interestingly, PD-L1 mutant subclones showed slow selection dynamics reversing on avelumab withdrawal and patients with such subclones had above-average treatment benefit. This suggested that the PD-L1 mutations mediated resistance to direct antitumor effects of avelumab, while at the same time loss of PD-L1 reduced biological fitness by enhanced T cell killing limiting subclonal expansion.ConclusionThe addition of avelumab to standard treatment appeared feasible and safe. PD-L1 mutations mediate subclonal immune escape to avelumab in some patients with mCRC expressing high-affinity FcγR3a, which may be a subset experiencing most selective pressure. Future trials evaluating the addition of avelumab to standard treatment in MSS mCRC are warranted especially in this patient subpopulation.Trial registration numberNCT03174405.
Abstract 4969
While multiparameter flow cytometry (MPF) is an integral part in the diagnosis, disease staging and response evaluation for hematologic malignancies such as acute leukemia or ...non-hodgkin-lymphoma, MPF for multiple myeloma (MM) is still mostly restricted to research studies or only performed by specialised laboratories experienced in the technique of immunophenotyping. Furthermore, the exact phenotype of malignant myeloma cells is still a matter of debate. Recently, we have identified CD229, a surface marker belonging to the family of signaling lymphocytic activation molecules (SLAM) involved in lymphocyte activation as a potential novel target for diagnosis and treatment of MM. CD229 is expressed on freshly isolated myeloma cells including their clonogenic precursors and several myeloma cell lines. In order to further validate our findings from a previous pilot study, we now analysed 151 samples from 81 patients with suspected or proven MM or monoclonal gammopathy of uncertain significance (MGUS) via MPF.
Between May 2010 and May 2012, specimens (bone marrow (n=142), peripheral blood (n=10), cells from isolated plasmocytoma (n=1)) from patients (pts) with MM (n=65), plasmocytoma (n=1), MGUS (n=6), lymphoplasmacytic lymphoma (n=1) and patients with suspected MM (n=8) were simultaneously analysed via cytology and 8-colour MPF. 19 pts. were analysed at least 3 times during the course of their disease so that CD229 expression could be followed under therapy. Plasma cells were specified using surface markers CD38, CD138, CD45 and cytoplasmatic light chain restriction. Antigens analysed on plasma cells were CD19, CD28, CD33, CD56, CD81, CD117, CD200, CD221 and CD229.
Although plasma cell numbers determined by MPF were constantly considerably lower compared to simultaneously determined cytology results, linear regression model showed a highly significant correlation between plasma cell percentages in bone marrow measured by MPF with cytology (p<0. 0001). Plasma cell enumeration in pB also showed a strong correlative trend between cytologic and MPF results, however, due to lower numbers (n=10), this was not statistically significant (p = 0. 057).
CD229 could be detected on all atypical plasma cells irrespective if they were found in MGUS or MM samples. The median of mean fluorescence intensity (MFI) of CD229 expression on plasma cells was 3, 63 (range −144. 1 – 34, 23). Median MFI on MM samples (3, 62; range −144 − 34, 23; n=131) did not differ from MFI on atypical plasma cells in pts with MGUS (3, 74, range 1. 07 – 8, 65; n=9). CD229 expression was highest on atypical plasma cells with expression of CD56 compared to CD56 negative plasma cells (p<0. 0001). This was confirmed when correlation of marker expression was evaluated. CD229 expression was clearly correlated with expression of CD56 (n=141, p = 0. 03), CD117 (n=139, p = 7E–08) and CD200 (n=140; p = 0. 03), while it was inversely correlated with expression of CD19 (n=140; p = 0. 03). Serial CD229 expression (>= twice) was determined in 39 patients. Except for three samples, where plasma cell counts became less than 1% of bone marrow cells, CD229 expression remained stable throughout the various analyses.
While the exact function of the immunoreceptor CD229 on myeloma cells is still unknown, CD229 allows identification of atypical plasma cells by MPF. Our results show that CD229 is constantly expressed on atypical plasma cells independent of therapy and can be used in addition to other surface markers for determination of malignant plasma cell phenotype and to monitor minimal residual disease (MRD) under treatment.
No relevant conflicts of interest to declare.
Abstract 4993
Extramedullary organ impairment in patients with multiple myeloma (MM) is a very rare event, mostly occurring during disease relapse after high-dose chemotherapy with autologous or ...allogeneic stem cell transplantation. Recent data reported by different authors suggest a very unfavorable outcome and rapid clinical course for patients with extramedullary relapses. Often the extramedullary manifestations are associated with special biological features, such as loss of CD56 expression or plasmablastic cell morphology.
In this context, only one FISH study of 9 MM patients reported that the incidence of TP53 deletions in the bone marrow of those MM patients with central nervous system (CNS) involvement was about 75% higher than in the MM patient without CNS involvement. However, there is a lack of genetic data for specific extramedullary manifestations. We herein report the first cytogenetic investigation of extramedullary organ infiltrations compared to bone lesions or consecutive soft tissue impairment of MM patients using cIg-FISH on paraffine embedded sections.
We investigated paraffin-embedded sections of different extramedullary organ manifestations of 13 MM patients and 11 MM patients with bone or soft tissue impairment originating from a bone lesion. Extramedullary organ involvements comprised biopsies from skin, pleura, pleural effusion, uterus, liver, CNS, subcutaneous soft tissue, lymph node, and thyroid gland, attained at different stages of the disease. The second group consisted of bone lesions or surrounding soft tissue, like muscle, which was infiltrated per continuitatem. For investigation of paraffine-embedded samples, we further developed the conditions of the well known cIg-FISH method to utilize all the advantages of this very sensitive method. We evaluated the most important prognostic chromosomal regions in MM using the following FISH-probes:
13q14 (D13S25), 17p13 (TP53), 8q24 (MYC), t(4;14) (FGFR3/MMSET;IGH).
The incidences in the group of extramedullary organ involvement vs. bone and soft tissue were as follows:
deletion of TP53: 23% vs. 18%
MYC-overrepresentation: 36% vs. 36%
deletion of 13q14: 18% vs. 36%
t(4;14) 42% vs. 30%
The incidence of TP53 deletions in organ infiltrations by malignant plasma cells was similar to bone or soft tissue involvement. Our result for MYC-overrepresentation is in line with the findings of recent analysis of bone marrow aspirates from patients with advanced multiple myeloma (32%) and showed again no difference between the two groups. Incidences of 13q14 deletion and t(4;14) varied slightly between the two groups, but are in range with other cIg-FISH-analyses on bone marrow plasma cells.
In this first cytogenetic investigation of extramedullary manifestations in MM patients, we showed similar frequencies of the chromosomal regions analyzed in the group of organ infiltrations by malignant plasma cells compared to bone or soft tissue involvement. All in all we could not detect differences in the incidence of the most relevant genetic aberrations of the investigated tissue probes compared to already well known bone marrow cytogenetics. Interestingly, this is in contrast to the findings of bone marrow probes in the study with patients with MM and CNS involvement, who showed a higher incidence of TP53 deletions. Concerning the group of soft tissues, originating from bone lesions this is not astonishing, maybe even expected. However, this work analyzed bone marrow of the MM patients and not the extramedullary manifestation, as we did. Further investigations or larger patient samples are needed to proof if these or other genetic aberrations are associated to the aggressive course of extramedullary manifestations of MM.
No relevant conflicts of interest to declare.
Esophagogastric adenocarcinoma (EGA) is one of the leading causes of cancer-related mortality worldwide. Therapeutic options are limited for patients with recurrent or metastatic disease. Targeted ...therapy may be a suitable treatment for selected patients, but its efficacy remains elusive.
Here, a 52-year-old male patient with advanced EGA Siewert Type II shows a significant response to combination therapy with olaparib and pembrolizumab. After progression following first- and second-line therapy, including a programmed cell death ligand 1 (PD-L1) inhibitor, next-generation sequencing of a tumor sample was performed to identify possible molecular targets. A mutation in RAD51C, a member of the homology-directed repair (HDR) system, was identified in addition to high PD-L1 expression. As a result, therapy with the poly-(ARD-Ribose) polymerase (PARP) inhibitor olaparib and the programmed cell death protein 1 (PD1)-inhibitor pembrolizumab was initiated. A durable partial response lasting for more than 17 months was observed. A second molecular profiling from a newly occurring subcutaneous metastasis showed a loss of FGF10 but no fluctuations in the gene alteration of RAD51C and SMARCA4. Interestingly, the new lesion showed HER2-positivity (immunohistochemistry 3+ and fluorescence in situ hybridization FISH-positivity) in 30% of tumor cells.
In this case, a long-lasting response to the combination of olaparib and pembrolizumab was observed despite previous treatment with a PD-L1 inhibitor. This case illustrates the need for further clinical trials to analyze the efficacy of PARP inhibitor combinations in EGA.
Ramucirumab and paclitaxel is the standard second-line therapy in patients with metastatic gastroesophageal adenocarcinoma. We report the efficacy and safety analyses of FOLFIRI and ramucirumab ...versus paclitaxel and ramucirumab after the failure of a platinum- and fluoropyrimidine-containing chemotherapy.
This multicenter, investigator initiated, phase II trial randomised patients with gastroesophageal adenocarcinoma to either FOLFIRI plus ramucirumab (RAM) (arm A) or paclitaxel plus RAM (arm B). The primary end-point was 6-month overall survival (OS) rate, with a proportion of ≥65% in arm A considered a positive signal for further investigation.
111 patients (65% of patients had prior docetaxel) were enrolled and 110 patients qualified for ITT population (arm A, 72; arm B, 38). The study did not meet the primary end-point for the comparison with historical control, as 6-month OS rate in the FOLFIRI plus RAM arm was 54% (95% CI 44–67). In between arm comparison, OS was similar (hazard ratio, HR 0.97 95% CI 0.62–1.52), while objective response rates (ORRs) and PFS were numerically better in arm A versus arm B (HR for PFS 0.73; ORR, 22% versus 11%). These differences were largely attributed to favourable efficacy results for arm A in docetaxel-pretreated patients (HR, 0.49; ORR, 25% versus 8%). In the safety population (n = 106), grade 3-5 adverse events were similar between arms (arm A, 75%; arm B, 68%).
The RAMIRIS trial demonstrated feasibility of FOLFIRI plus RAM. While the study was formally negative, it provided a signal to further investigate this combination for the group of patients with previous docetaxel therapy.
clinicaltrials.gov identifier: NCT03081143.
•FOLFIRI-ramucirumab is a safe and potent 2nd-line regimen.•FOLFIRI-ramucirumab is effective in docetaxel-pretreated patients.•FOLFIRI-ramucirumab is less-neurotoxic compared with paclitaxel-ramucirumab.
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