1296P Discovery of novel germline genetic biomarkers of melanoma recurrence impacting exonic and long non-coding RNA (lncRNA) transcripts T. Kirchhoff1 , D. Simpson2 , T. Hekal2 , R. Ferguson2 , E. Kazlow2 , U. Moran2 , Y. Lee3 , A. Izsak3 , M.A. Wilson4 , R. Shapiro4 , A. Pavlick4 , I. Osman3 1
Perlmutter Cancer Center, New York University, New York, NY, USA,2
Perlmutter Cancer Center, New York University Medical Center, New York, NY, USA,3
Dermatology, New York University Medical Center, New York, NY, USA,4
Department of Medicine, New York University - Perlmutter Cancer Center - Langone Medical Center, New York, NY, USA Background: One challenge in clinical management of cutaneous melanoma (CM) is the limited predictability of recurrence and hence the progression to advanced stages that associate with less favorable outcomes. Aggressive follow-up care and novel adju-vant therapy strategies in early-stage patients with high-risk of recurrence would likely improve CM-specific survival and reduce mortality. We developed a genome-wide approach to identify germline genetic determinants of melanoma prognosis as putative personalized biomarkers for patients at risk of CM recurrence.
Methods: Exploring both the coding and non-coding transcribed genome we per-formed germline whole genome sequencing (WGS) and tumor RNA-seq on 96 CM patients with tumor/blood matched specimens. All patients were of primary stages I-IIB and of Ashkenazi Jewish ancestry to reduce genetic heterogeneity. We compared 48 patients that recurred in < 4 years versus 48 patients with recurrence in > 6 years. Univariate and multivariate logistic regression, gene-burden analysis (SKAT), and dif-ferential expression analyses of both mRNA and lncRNAs were used to identify germ-line regions associated with melanoma recurrence.
Results: Several gene regions were associated with melanoma recurrence, with NEGR1 and MGST3 both passing SKAT levels of significance (p < 1e-05), and logistic regres-sion analysis on common WGS variants found over 100 variants with significance p < 1e-03. In addition, we found 200 differentially expressed putative lncRNAs (p < 0.05). The analysis of germline WGS found rs199818927, a 1bp insertion previ-ously associated with both NEGR1 and lncRNA LINC01360, among our top 5 most sig-nificant regression results (OR¼ 9.107 p ¼ 2.31e-05).
Conclusions: This initial phase of our large scale whole genome scan has uncovered germline variants in several coding loci and putative lncRNAs that associate with mela-noma recurrence. Most notably, we identified germline variation in a lncRNA near NEGR1, a putative tumor suppressor that has been shown to be under-expressed in advanced cancers, indicating its putative role in cancer progression to metastatic stages. We are currently expanding the patient cohorts and validating these results. Legal entity responsible for the study: Tomas Kirchhoff.
Funding: NIH.
Disclosure: All authors have declared no conflicts of interest.
1297P Pilot study: Localizing target lymph node using a magnetic marker allows reliable and representative judgement of pathological responses after neo-adjuvant ipilimumab (IPI) 1 nivolumab (NIVO) in macroscopic stage III melanoma
A.C.J. van Akkooi1
, B. Schermers2 , V. Franke1 , M. Wouters1 , C.L. Zuur3 , W.M.C. Klop3 , E.A. Rozeman4
, B.A. van de Wiel5
, T.J.M. Ruers1
, C.U. Blank6 1
Surgical Oncology, Netherlands Cancer Institute/Antoni van Leeuwenhoek hospital (NKI-AVL), Amsterdam, Netherlands,2Surgical Oncology, The Netherlands Cancer Institute Antoni van Leeuwenhoek Hospital, Amsterdam, Netherlands,3
Head and Neck Surgery and Oncology, Netherlands Cancer Institute/Antoni van Leeuwenhoek hospital (NKI-AVL), Amsterdam, Netherlands,4
Medical Oncology, The Netherlands Cancer Institute, Amsterdam, Netherlands,5
Pathology, Netherlands Cancer Institute/Antoni van Leeuwenhoek Hospital (NKI-AVL), Amsterdam, Netherlands,6
Medical Oncology, The Netherlands Cancer Institute Antoni van Leeuwenhoek Hospital, Amsterdam, Netherlands
Background:The outcome of high risk stage III melanoma patients (pts) is poor, with a 5-year overall survival (OS) rate of < 50%. Adjuvant (adj) high dose IPI , adj NIVO and pembrolizumab improved RFS. Neo-adjuvant (neoadj) treatment may be an even more favorable approach as immune checkpoint inhibition is of greatest value at the moment of TCR triggering and therefore dependent on the amount of antigen present. In our previous phase Ib OpACIN study the pathological RR (pRR) was 78% in the neoadj arm, and to date after 25 months of median follow-up none of the responders has relapsed. This raises the question whether such pts need to undergo complete lymph node dissection (CLND). A prerequisite for such an approach would be an anal-ysis method that reliably indicates pathological response within the whole lymph node bed, without the need for CLND.
Methods:To address this question an in-house developed magnetic marker was placed ultrasound-guided at baseline into the largest regional lymph node metastasis of pts participating in OpACIN-neo, (NCT02977052), a phase 2 trial aiming at identification of the optimal neoadj combination scheme of IPI and NIVO in stage IIIB/C melanoma pts followed by CLND.
Results:So far, 11 pts participated in this side trial of OpACIN-neo. No complications from marker placing were observed, and all magnetic markers were retrieved during the CLND after 6 weeks of neoadj IPIþNIVO. 10/11 marked lymph-nodes (LN were representative in their response for the whole CLND specimen, i.e. index node showed
a complete or partial response (PR), all others on CLND showed the same or better responses. Only 1 case was incongruent, as the index LN had 60% vital tumor (no response) compared to another LN (40% vital tumor, PR).
Conclusions:Our early exploratory data from this pilot study indicate that marked LN in stage III melanoma could serve as response indicators for the outcome of the whole CLND after neoadj IPIþNIVO. If confirmed, our data can open the path towards response-driven extent of lymph-node dissection in macroscopic stage III melanoma. Clinical trial identification:NCT02977052.
Legal entity responsible for the study:Alexander C.J. van Akkooi. Funding:Has not received any funding.
Disclosure:A.C.J. van Akkooi: Consulting or advisory: Amgen, Novartis, MSD Oncology, Merck; Travel, accommodations, expenses: Amgen, Roche, Novartis; Research funding: Amgen, Novartis. C.U. Blank: Personal fees, Advisory role: MSD, BMS, Roche, GSK, Novartis, Pfizer, Lilly; Grants: BMS, Novartis, outside the submitted work. All other authors have declared no conflicts of interest.
1298P Discovery of KIRREL as a biomarker for prognostic stratification of patients with thin melanoma
S. Lundgren, H. Fagerstro¨m-Vahman, L. Ben-Dror, B. Nodin, K. Jirstrom
Department of Clinical Sciences, Oncology and Pathology, Lund University, Faculty of Medicine, Lund, Sweden
Background: There is a great unmet clinical need to identify patients with thin primary cutaneous melanomas (T1, Breslow thickness 1 mm) who have a high risk for tumor recurrence and death from melanoma. Kin of IRRE-like protein 1 (KIRREL/NEPH1) is expressed in podocytes and involved in glomerular filtration, but its expression in human cancer has not yet been reported. Screening in the Human Protein Atlas portal revealed a particularly high expression of KIRREL in melanoma, both at the mRNA and protein levels. In this study, we followed up on these findings and examined the prognostic value of KIRREL in a population-based cohort of melanoma.
Methods: Immunohistochemical analysis of KIRREL was performed on tissue microar-rays with a subset of primary tumors and paired lymph node metastases from an origi-nal cohort of 268 incident cases of melanoma in the Malmo¨ Diet and Cancer study. Kaplan Meier analysis and Cox proportional hazards modelling were used to assess the relationship between KIRREL expression and time to recurrence (TTR) and mela-noma-specific survival (MSS). The prognostic value of KIRREL mRNA expression was examined in 102 melanoma cases in The Cancer Genome Atlas (TCGA).
Results: Membranous/cytoplasmic expression of KIRREL was detected in 158/185 (85.4%) primary tumours and 18/19 (94.7%) metastases, in various fractions and intensities. High expression of KIRREL was significantly associated with several unfav-ourable clinicopathological factors. KIRREL expression was not prognostic in tumours >1 mm thickness, but in T1 tumours (n¼ 106, median thickness 0.58, range 0.08-1.00), high expression of KIRREL was significantly associated with a reduced TTR, independent of and outperforming absolute thickness in mm and ulceration (HR¼ 4.54, 95% CI 1.01-20.45), and borderline significantly associated with MSS. High mRNA levels of KIRREL were associated with a significantly reduced overall sur-vival in the TCGA (p¼ 0.028).
Conclusions: KIRREL is not only a novel potential diagnostic marker for melanoma, but may also be a useful prognostic biomarker for improved stratification of patients with thin melanoma. These findings may be of high clinical relevance and therefore merit further validation.
Legal entity responsible for the study: Lund University.
Funding: Swedish Cancer Society, the Swedish Research Council; the Swedish Government Grant for Clinical Research, the Mrs Berta Kamprad Foundation, Lund University Faculty of Medicine and University Hospital Research Grants. Disclosure: All authors have declared no conflicts of interest.
1299P BRAF V600E mutation in melanoma sustains IFN-gamma inducible PD-L1 expression by coactivating STAT1 and increasing protein translation
M.M. Wasylecka-Juszczynska, P. Gorniak, M. Szydłowski, A. Polak, P. Juszczynski Department of Experimental Hematology, Institute of Hematology and Transfusion Medicine, Warsaw, Poland
Background: Approximately 40-60% of melanoma patients have activating BRAF mutations. Targeting BRAF inhibits proliferation of melanoma cells and increases their immunogenicity. PD-1 immune checkpoint blockade is another breakthrough in mela-noma therapy, shown to effectively restore T cell function. These observations indicate that combinatorial use of BRAF inhibitors and immunotherapy might be a rational therapeutic strategy. Herein, we studied the role of BRAF V600E in regulating the expression of PD-L1 in melanoma cells.
Methods: CD274 gene transcript abundance and PD-L1 protein level was measured with qPCR, Western-blot and FACS. Identification of signaling pathways responsible for regulation of PD-L1 expression was performed using western blot, FACS and
