Faculty Bibliography

The gut microbiome (GMB) has been associated with outcomes of immune checkpoint blockade therapy in melanoma, but there is limited consensus on the specific taxa involved, particularly across different geographic regions. We analyzed pre-treatment stool samples from 674 melanoma patients participating in a phase-III trial of adjuvant nivolumab plus ipilimumab versus nivolumab, across three continents and five regions. Longitudinal analysis revealed that GMB was largely unchanged following treatment, offering promise for lasting GMB-based interventions. In region-specific and cross-region meta-analyses, we identified pre-treatment taxonomic markers associated with recurrence, including Eubacterium, Ruminococcus, Firmicutes, and Clostridium. Recurrence prediction by these markers was best achieved across regions by matching participants on GMB compositional similarity between the intra-regional discovery and external validation sets. AUCs for prediction ranged from 0.83-0.94 (depending on the initial discovery region) for patients closely matched on GMB composition (e.g., JSD ≤0.11). This evidence indicates that taxonomic markers for prediction of recurrence are generalizable across regions, for individuals of similar GMB composition.

Glycosylation is a hallmark of cancer biology, and altered glycosylation influences multiple facets of melanoma growth and progression. To identify glycosyltransferases, glycans, and glycoproteins essential for melanoma maintenance, we conducted an in vivo growth screen with a pooled shRNA library of glycosyltransferases, lectin microarray profiling of benign nevi and melanoma patient samples, and mass spectrometry-based glycoproteomics. We found that α-2,3 sialyltransferases ST3GAL1 and ST3GAL2 and corresponding α-2,3-linked sialosides are upregulated in melanoma compared to nevi and are essential for melanoma growth in vivo and in vitro. Glycoproteomics revealed that glycoprotein targets of ST3GAL1 and ST3GAL2 are enriched in transmembrane proteins involved in growth signaling, including the amino acid transporter Solute Carrier Family 3 Member 2 (SLC3A2/CD98hc). CD98hc suppression mimicked the effect of ST3GAL1 and ST3GAL2 silencing, inhibiting melanoma cell proliferation. We found that both CD98hc protein stability and its pro-survival effect in melanoma are dependent upon α-2,3 sialylation mediated by ST3GAL1 and ST3GAL2. In summary, our studies reveal that α-2,3-sialosides functionally contribute to melanoma maintenance, supporting ST3GAL1 and ST3GAL2 as novel therapeutic targets in these tumors.

KRAS^G12C inhibitors (adagrasib and sotorasib) have shown clinical promise in targeting KRAS^G12C-mutated lung cancers; however, most patients eventually develop resistance. In lung patients with adenocarcinoma with KRAS^G12C and STK11/LKB1 co-mutations, we find an enrichment of the squamous cell carcinoma gene signature in pre-treatment biopsies correlates with a poor response to adagrasib. Studies of Lkb1-deficient KRAS^G12C and Kras^G12D lung cancer mouse models and organoids treated with KRAS inhibitors reveal tumors invoke a lineage plasticity program, adeno-to-squamous transition (AST), that enables resistance to KRAS inhibition. Transcriptomic and epigenomic analyses reveal ΔNp63 drives AST and modulates response to KRAS inhibition. We identify an intermediate high-plastic cell state marked by expression of an AST plasticity signature and Krt6a. Notably, expression of the AST plasticity signature and KRT6A at baseline correlates with poor adagrasib responses. These data indicate the role of AST in KRAS inhibitor resistance and provide predictive biomarkers for KRAS-targeted therapies in lung cancer.

BACKGROUND:Dickkopf-related protein 1 (DKK1) is a Wingless-related integrate site (Wnt) signaling modulator that is upregulated in prostate cancers (PCa) with low androgen receptor expression. DKN-01, an IgG4 that neutralizes DKK1, delays PCa growth in pre-clinical DKK1-expressing models. These data provided the rationale for a clinical trial testing DKN-01 in patients with metastatic castration-resistant PCa (mCRPC). METHODS:(combination) for men with mCRPC who progressed on ≥1 AR signaling inhibitors. DKK1 status was determined by RNA in-situ expression. The primary endpoint of the phase 1 dose escalation cohorts was the determination of the recommended phase 2 dose (RP2D). The primary endpoint of the phase 2 expansion cohorts was objective response rate by iRECIST criteria in patients treated with the combination. RESULTS:18 pts were enrolled into the study-10 patients in the monotherapy cohorts and 8 patients in the combination cohorts. No DLTs were observed and DKN-01 600 mg was determined as the RP2D. A best overall response of stable disease occurred in two out of seven (29%) evaluable patients in the monotherapy cohort. In the combination cohort, five out of seven (71%) evaluable patients had a partial response (PR). A median rPFS of 5.7 months was observed in the combination cohort. In the combination cohort, the median tumoral DKK1 expression H-score was 0.75 and the rPFS observed was similar between patients with DKK1 H-score ≥1 versus H-score = 0. CONCLUSION/CONCLUSIONS:DKN-01 600 mg was well tolerated. DKK1 blockade has modest anti-tumor activity as a monotherapy for mCRPC. Anti-tumor activity was observed in the combination cohorts, but the response duration was limited. DKK1 expression in the majority of mCRPC is low and did not clearly correlate with anti-tumor activity of DKN-01 plus docetaxel.

INTRODUCTION/BACKGROUND:Pathologists are staging thymic epithelial tumors (TET) according to the 8th UICC/AJCC TNM system. Within the French RYTHMIC network, dedicated to TET, agreement on pathologic tumor stage (pT) among the pathology panelists was difficult. The aim of our study was to determine the interobserver reproducibility of pT at an international level, to explore the source of discrepancies and potential interventions to address these. METHODS:An international panel of pathologists was recruited through the International Thymic Malignancy Interest Group (ITMIG). The study focused on invasion of mediastinal pleura, pericardium, and lung. From a cohort of cases identified as challenging within the RYTHMIC network, we chose a series of test and validation cases (n = 5 and 10, respectively). RESULTS:Reproducibility of the pT stage was also challenging at an international level as none of the 15 cases was classified as the same pT stage by all ITMIG pathologists. The agreement rose from slight (κ = 0.13) to moderate (κ = 0.48) between test and validation series. Discussion among the expert pathologists pinpointed two major reasons underlying discrepancies: 1) Thymomas growing with their "capsule" and adhering to the pleurae, pericardium, or lung were often misinterpreted as invading these structures. 2) Recognition of the mediastinal pleura was identified as challenging. CONCLUSION/CONCLUSIONS:Our study underlines that the evaluation of the pT stage of TET is problematic and needs to be addressed in more detail in an upcoming TNM classification. The publication of histopathologic images of landmarks, including ancillary tests could improve reproducibility for future TNM classifications.

Advancements in the classification of lung adenocarcinoma have resulted in significant changes in pathological reporting. The eighth edition of the tumour-node-metastasis (TNM) staging guidelines calls for the use of invasive size in staging in place of total tumour size. This shift improves prognostic stratification and requires a more nuanced approach to tumour measurements in challenging situations. Similarly, the adoption of new grading criteria based on the predominant and highest-grade pattern proposed by the International Association for the Study of Lung Cancer (IASLC) shows improved prognostication, and therefore clinical utility, relative to previous grading systems. Spread through airspaces (STAS) is a form of tumour invasion involving tumour cells spreading through the airspaces, which has been highly researched in recent years. This review discusses updates in pathological T staging, adenocarcinoma grading and STAS and illustrates the utility and limitations of current concepts in lung adenocarcinoma.

INTRODUCTION/UNASSIGNED:Quantitative, multiplexed imaging is revealing complex spatial relationships between phenotypically diverse tumor infiltrating leukocyte populations and their prognostic implications. The underlying mechanisms and tissue structures that determine leukocyte distribution within and around tumor nests, however, remain poorly understood. While presumed players in metastatic dissemination, new preclinical data demonstrates that blood and lymphatic vessels (lymphovasculature) also dictate leukocyte trafficking within tumor microenvironments and thereby impact anti-tumor immunity. Here we interrogate these relationships in primary human cutaneous melanoma. METHODS/UNASSIGNED:We established a quantitative, multiplexed imaging platform to simultaneously detect immune infiltrates and tumor-associated vessels in formalin-fixed paraffin embedded patient samples. We performed a discovery, retrospective analysis of 28 treatment-naïve, primary cutaneous melanomas. RESULTS/UNASSIGNED:Here we find that the lymphvasculature and immune infiltrate is heterogenous across patients in treatment naïve, primary melanoma. We categorized five lymphovascular subtypes that differ by functionality and morphology and mapped their localization in and around primary tumors. Interestingly, the localization of specific vessel subtypes, but not overall vessel density, significantly associated with the presence of lymphoid aggregates, regional progression, and intratumoral T cell infiltrates. DISCUSSION/UNASSIGNED:We describe a quantitative platform to enable simultaneous lymphovascular and immune infiltrate analysis and map their spatial relationships in primary melanoma. Our data indicate that tumor-associated vessels exist in different states and that their localization may determine potential for metastasis or immune infiltration. This platform will support future efforts to map tumor-associated lymphovascular evolution across stage, assess its prognostic value, and stratify patients for adjuvant therapy.

BACKGROUND:We previously reported a higher incidence of a pathogenic germline variant in the kinase insert domain receptor (KDR) in melanoma patients compared to the general population. Here, we dissect the impact of this genotype on melanoma tumor growth kinetics, tumor phenotype, and response to treatment with immune checkpoint inhibitors (ICIs) or targeted therapy. METHODS:The KDR genotype was determined and the associations between the KDR Q472H variant (KDR-Var), angiogenesis, tumor immunophenotype, and response to MAPK inhibition or ICI treatment were examined. Melanoma B16 cell lines were transfected with KDR-Var or KDR wild type (KDR-WT), and the differences in tumor kinetics were evaluated. We also examined the impact of KDR-Var on the response of melanoma cells to a combination of VEGFR inhibition with MAPKi. RESULTS:= 0.0159), and KDR-Var cells showed synergistic cytotoxicity to the combination of dabrafenib and lenvatinib. CONCLUSIONS:Our data demonstrate a role of germline KDR-Var in modulating melanoma behavior, including response to treatment. Our data also suggest that anti-angiogenic therapy might be beneficial in patients harboring this genotype, which needs to be tested in clinical trials.