Smith, Katherine E R

Abstract Purpose: Rearranged during transfection (RET) alterations are oncogenic drivers across solid tumors. Selective RET inhibitors (SRI) selpercatinib and pralsetinib have transformed outcomes for patients with RET-altered malignancies. Limited knowledge exists on genomic mechanisms of resistance to SRI. Experimental Design: We established “RETgistry,” a global consortium of patients with advanced RET-altered solid tumors who received SRI and underwent postprogression tissue or plasma biopsies assessed by next-generation sequencing. Frequencies of secondary RET resistance mutations and acquired non-RET gene alterations were determined. Progression-free survival (PFS) and time to treatment discontinuation (TTD) on first SRI were estimated with the Kaplan–Meier method. Results: RETgistry included 109 patients with RET-altered advanced solid tumors (lung, n = 94; thyroid, n = 15) who underwent 143 post-SRI progression biopsies (tissue, 91; plasma, 52). The median PFS and TTD were 13.9 months [95% confidence interval (CI), 10.1–16.6] and 17.3 months (95% CI, 14–20.2), respectively. Secondary RET mutations were detected in 20 (14%) biopsies [lung cancer, 15 (12.4%) and thyroid carcinoma, 5 (22.7%)]. Common acquired off-target alterations involved MET (18.2%; amplification, 15%), TP53 (8.2%), APC (7.6%), KRAS (7.1%), KEAP1 (5.9%), and CDKN2A/B (5.3%). MET alterations were enriched in post-SRI versus pre-SRI specimens (full cohort, 17.6% vs. 2.0%, P = 0.022; lung cancer, 19.1% vs. 2.1%, P = 0.022). Conclusions: The prevalence of secondary RET mutations after SRI was low, underscoring a greater role for off-target resistance. Recurrent acquired alterations involving tumor suppressor genes or upstream regulators of MAPK and PI3K pathways were identified, most commonly MET amplification. Continued efforts to characterize SRI resistance biology are critical to guide the development of novel therapeutic strategies.

PURPOSE BRAF and MEK inhibitors are standard treatments in histiocytic disorders, such as Erdheim-Chester disease (ECD). Some patients lack MAPK-pathway alterations, making these treatments less effective. METHODS We describe three patients with histiocytic disorders who have novel non-MAPK pathway alterations. These alterations were studied through genomic and in silico analyses when applicable, then treated with off-label medications rationally selected on the basis of genomic alterations. RESULTS Patient 1 had rapidly progressive ECD involving the CNS. A CSF1R in-frame deletion (p.S560_P566del) was identified, and in silico modeling predicted a gain-of-function mutation. This alteration was targeted with pexidartinib, which led to a clinical complete response (CR) within 2 months, and a partial response (PR) on imaging after 3 months. After 15 months, the disease became resistant to pexidartinib and transformed to histiocytic sarcoma. Patient 2 has skin-only involvement of a xanthogranuloma disorder. A KIF5B-FGFR1 fusion was identified on RNA sequencing and targeted with pemigatinib. At 24 months of follow-up, she remains in a clinical PR. Patient 3 has ECD involving the bone marrow, gastrointestinal tract, and subcutaneous tissues. A MEF2C-FLT3 fusion was identified and targeted with sorafenib. He achieved a clinical CR and radiographic PR within 3 months, which has continued for 30 months. CONCLUSION We report three patients with histiocytic disorders harboring novel alterations who had sustained responses to off-label kinase inhibitors specific to their histiocytic disorder. Pathogenic variants outside of the MAPK pathway, including variants of unknown significant, may be targeted with readily available small molecules.

Abstract Immune related adverse events (irAEs) from immune checkpoint inhibitors (ICIs) are well documented for nearly every organ system, except for the reproductive system. Mouse models have demonstrated that ICIs can decrease ovarian follicular reserve and impair oocyte maturation, which may have implications for fertility (PMID: 36008687). There are no studies investigating the effect of ICIs on fertility in humans. We sought to evaluate ovarian function in women treated with ICIs compared to untreated women by evaluating non-invasive surrogate markers of ovarian reserve, anti-müllerian hormone (AMH) and antral follicle counts (AFC). We enrolled women 18 - 40 years old diagnosed with stage III or IV melanoma. Participants underwent phlebotomy for AMH levels and a pelvic ultrasound to assess AFC. The medical record was retrospectively reviewed to obtain data on patient characteristics, treatments, and outcomes. A total of 14 patients with stage IIIA - IV melanoma participated. Six patients received ICI treatment before AMH and AFC testing while 8 had no prior ICI exposure. The ICI group ages ranged from 22-40 years with a median age of 31 years. The untreated group ages ranged from 25-39 years with a median age of 32 years. The time between starting an ICI and enrolling in the study ranged from 28 - 351 days. ICI regimens included ipilimumab/nivolumab, nivolumab, pembrolizumab, and atezolizumab (on a clinical trial). Four of these six patients experienced an irAE during their course, including: dermatitis, hepatitis, colitis, thyroid dysfunction, and adrenal insufficiency. Median AMH levels were lower for patients treated with ICIs (2.9 vs 4.0 ng/mL), as was median AFC (22.5 vs 23.5 ng/mL). Only three patients’ clinicians discussed fertility preservation. None of the patients underwent fertility preservation. Our study found lower AMH and AFC in women with melanoma who were treated with an ICI compared to untreated women. Although AMH and AFC are surrogate markers for ovarian reserve and fertility, and our study is a small cohort, these early results suggest that ICIs may impact ovarian function. Age is less likely to be a confounding factor as the median age of the treated group was younger than the untreated group. While these results are preliminary, when considered in conjunction with recent pre-clinical data showing changes within the ovaries after ICI treatment, fertility consultation should be considered for young women prior to ICI initiation. We are investigating associations between AMH, AFC, age, ICI exposure, and future pregnancies in an ongoing prospective study. Citation Format: Ashley Hickman, Katherine Smith, Zaraq Khan, Elizabeth Ann Enninga, Yulian Zhao, Elizabeth Cathcart-Rake, Haidong Dong, Lisa Kottschade, Svetomir Markovic, Heather Montane, Anastasios Dimou, Yiyi Yan. Investigating the effect of immune checkpoint inhibitors (ICI) on ovarian function in young patients with melanoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6644.

9538 Background: Uveal melanoma has a poor prognosis following metastasis to the liver. Immune checkpoint inhibitors (ICIs) are often utilized in patients with metastatic uveal melanoma (MUM); however, ICIs in MUM have low response rates. We studied the association of immune infiltrates in MUM liver metastases with survival outcomes for all patients, in addition to evaluating if immune infiltrates improve survival for patients treated with ICIs. Methods: We included MUM patients with liver metastases who had pre-treatment liver biopsies available at Mayo Clinic Rochester. Demographics, disease characteristics, treatments, and clinical outcomes were obtained from the electronic medical record. Core biopsies of liver metastases were reviewed for the presence of lymphocytes at interface of the tumor and liver parenchyma, and for tumor-infiltrating lymphocytes (TILs). If TILs were present, then additional immunohistochemistry (IHC) staining was performed for CD3, CD4, CD8, PD1, CD25, and FoxP3. Overall survival (OS) was estimated with the Kaplan-Meier method and compared between groups with Cox proportional hazards regression. Median OS and hazard ratios (HR) were calculated and p-values (p) <0.05 were considered statistically significant. Results: 140 patients with MUM who had pre-treatment liver biopsies were identified. 122 biopsies were evaluable for the presence of lymphocytes, either at the tumor-liver interface, or for TILs. 80/122 (65.6%) biopsies showed presence of any lymphocyte and 27/80 had TILs. 27 biopsies had sufficient tissue for further IHC classification. CD8+ T cells were identified in 27/27 samples, T cell PD1 positivity ≥1% in 4/27 samples, and regulatory T cell markers (CD25 and/or FoxP3) in 18/27 samples. Median OS for the entire cohort was 16 months (mo). Patients still alive at the time of data analysis had at least one year of follow-up available (range 13-124 mo). Univariate analyses (UVA) showed no significant improvement in OS based on the presence of any lymphocytes (HR 1.01, p 0.98), lymphocytes at the tumor-liver interface (HR 1.10, p 0.64), TILs (HR 0.78, p 0.27), >50% CD8+ T cells (HR 0.88, p 0.77), or absence of FoxP3+ T cells (HR 0.68, p 0.40). In the 69 patients treated with ICIs (pembrolizumab, nivolumab, or ipilimumab plus nivolumab), neither the presence of any lymphocytes (HR 1.19, p 0.56) nor TILs (HR 0.68, p 0.24) were associated with an OS benefit. Conclusions: We present a large cohort of patients with MUM where we characterized immune infiltrates in liver metastases and evaluated clinical outcomes. OS did not significantly improve with the presence of any lymphocytes, TILs, CD8+ T cells, or with the absence of FoxP3+ T cells. These results were consistent across all patients whether they were treated with ICIs or not. Our findings suggest that lymphocytes in MUM are functionally dormant and highlights the need for novel immune-activating treatment strategies.

9065 Background: Rearranged during transfection ( RET) gene alterations are the oncogenic driver in diverse tumor types, including RET fusions identified in 1-2% of non-small cell lung cancers (NSCLC). While RET-selective tyrosine kinase inhibitors (TKIs) selpercatinib and pralsetinib are effective, acquired drug resistance remains a challenge. Here, we report the initial results from the RETgistry, an international consortium aimed at elucidating mechanisms of resistance to RET TKIs across RET-altered solid tumors. Methods: This was a retrospective analysis performed across 16 institutions. Patients (pts) were eligible if they had advanced solid tumor harboring an oncogenic RET alteration, received ≥1 RET TKI with disease progression, and underwent resistant tumor or liquid biopsy for next-generation sequencing (NGS). Results: A total of 103 time-distinct biopsies (62 tissue, 30 plasma, 11 paired tissue/plasma) were included in analysis, obtained from 88 pts with progression on a RET-selective TKI [selpercatinib (n = 70), pralsetinib (n = 14), selpercatinib and pralsetinib sequentially (n = 4)]. Pts had the following tumor types: 72 NSCLC (69% KIF5B-RET, 21% CCDC6-RET, 10% other RET fusion), 13 medullary thyroid cancer (TC) (54% RET M918T, 46% other RET mutation), and 2 papillary and 1 anaplastic TC (all, RET fusion). Median age was 58 (range, 21-86); 48% were male with 89% never/light smokers. Median duration of RET TKI preceding biopsies [first-line, n = 32 (31%); second-line, n = 42 (41%); third-/greater-line, n = 29 (28%)] was 16.5 months (mos) (95% CI, 14.0-19.6); median PFS was 14.1 mos (95% CI, 9.3-17.0). Resistant biopsies were obtained at median 15.0 mos from TKI initiation (range, 1.8-58.8). Acquired RET mutations were detected in 14 (14%), most common being G810 substitutions in 12 (12%). Potential off-target resistance gene alterations identified in 43 cases (42%) included MET amplification (14%), BRAF V600E or fusion (2%), KRAS gain or mutation (5%), ERBB2 amplification (2%), EGFR amplification (3%), ROS1 fusion (1%), and activating PIK3CA mutation or PTEN loss (4%). No resistant lung cancer biopsy demonstrated small cell transformation. The duration of TKI therapy (HR 0.64, p = 0.11) or PFS (HR 0.75, p = 0.33) did not differ according to the presence of on-target vs off-target resistance. NGS analyses of pre-RET-selective TKI tumors (n = 92) revealed frequent co-occurring alterations in TP53 (29%) and CDKN2A/B (12%). Conclusions: On-target resistance to RET inhibition due to acquired RET mutations was less common than off-target resistance, identified in 14%. The majority of RET TKI resistance is mediated by off-target mechanisms such as bypass receptor tyrosine kinase activation. Further studies are warranted to enable the development of therapeutic strategies to address resistance in pts with RET-altered tumors. The RETgistry accrual and data analyses continue.

Abstract Oncologic viruses (OV) can induce tumor death through both direct oncolysis and immune-mediated destruction. Our study investigates the use of a novel OV with a Vesicular Stomatitis Virus (VSV) vector modified to express interferon-beta (IFN-b) and Tyrosinase Related Protein 1 (TYRP1) (VSV-IFNb-TYRP1). VSV is an attractive vector since humans are not naturally infected with VSV; thus, pre-existing immunity is minimal. IFN-b leads to increased T-cell responses and tumor cell apoptosis, while TYRP1, a melanocyte differentiation antigen, increases immunogenicity of the therapy. We conducted a Phase 1 clinical trial with a 3+3 design in patients with metastatic uveal melanoma. VSV-IFNb-TYRP1 was injected into a liver metastasis under image guidance, then administered on the same day as a single intravenous (IV) infusion. IV doses started at dose level (DL) 1 1 × 1010 TCID50, then escalated to DL2 3 × 10 10, DL3 1 × 1011, and DL4 3 × 1011. The dose delivered intratumorally (IT) varied based on the IV dose. The primary endpoints were safety and maximum tolerated dose (MTD). Efficacy was a secondary endpoint. Correlative studies focused on understanding viral pharmacokinetics (PK) and immunological responses induced by VSV-IFNb-TYRP1 therapy. Twelve patients with previously treated metastatic uveal melanoma were enrolled. Median follow up was 19.1 months. Four DLs were evaluated with one patient at DL4 experiencing dose limiting toxicities (DLTs), including decreased platelet count (grade 3), increased aspartate aminotransferase (AST), and cytokine release syndrome (CRS). Non-DLTs at DL4 were fatigue, fever, CRS, hematological toxicities (decreased platelets and lymphocytes), and AST elevation. Four patients had stable disease (SD) and 8 had progressive disease (PD) as their best response. ELIspot data show that three of the four patients with response to TYRP1 also had a response to gp100, suggesting possible epitope spreading. These four patients received immune checkpoint inhibitor treatment as the next line of therapy, which led to clinical benefit for two patients. Our study evaluated VSV-IFNb-TYRP1 administered via IT and IV routes in a previously treated population of metastatic uveal melanoma patients and found no major safety events. Although there were no clear objective responses to VSV-IFNb-TYRP1, dose-dependent immunogenicity to melanoma antigens was seen. While OV response alone is not sufficient for clinical benefit, the evidence of epitope spreading is encouraging. Future studies of VSV-IFNb-TYRP1 will evaluate combinations with other therapies. Citation Format: Katherine E. Smith, Adam Weisbrod, Carrie Strand, Jacob Allerd, Jose S. Pulido, Alysha Newsom, Lianwen Zhang, Nandakumar Packiriswamy, Heather Montane, Lisa Kottschade, Anastasios Dimou, Yiyi Yan, Markovic Svetomir, Kah Whye Peng, Richard G. Vile, Matthew S. Block, Roxana S. Dronca. Phase I trial of vesicular stomatitis virus expressing human interferon beta and tyrosinase related protein 1 (VSV-IFNb-TYRP1) in metastatic ocular melanoma [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 CT207.