Antitumor immune effects of preoperative sitravatinib and nivolumab in oral cavity cancer: SNOW window-of-opportunity study

Abstract

Background: Sitravatinib, a tyrosine kinase inhibitor that targets TYRO3, AXL, MERTK and the VEGF receptor family, is predicted to increase the M1 to M2-polarized tumor-associated macrophages ratio in the tumor microenvironment and have synergistic antitumor activity in combination with anti-programmed death-1/ligand-1 agents. SNOW is a window-of-opportunity study designed to evaluate the immune and molecular effects of preoperative sitravatinib and nivolumab in patients with oral cavity squamous cell carcinoma.

Methods: Patients with newly-diagnosed untreated T2-4a, N0-2 or T1 >1 cm-N2 oral cavity carcinomas were eligible. All patients received sitravatinib 120 mg daily from day 1 up to 48 hours pre-surgery and one dose of nivolumab 240 mg on day 15. Surgery was planned between day 23 and 30. Standard of care adjuvant radiotherapy was given based on clinical stage. Tumor photographs, fresh tumor biopsies and blood samples were collected at baseline, at day 15 after sitravatinib alone, and at surgery after sitravatinib-nivolumab combination. Tumor flow cytometry, multiplex immunofluorescence staining and single-cell RNA sequencing (scRNAseq) were performed on tumor biopsies to study changes in immune-cell populations. Tumor whole-exome sequencing and circulating tumor DNA and cell-free DNA were evaluated at each time point.

Results: Ten patients were included. Grade 3 toxicity occurred in one patient (hypertension); one patient required sitravatinib dose reduction, and one patient required discontinuation and surgery delay due to G2 thrombocytopenia. Nine patients had clinical-to-pathological downstaging, with one complete response. Independent pathological treatment response (PTR) assessment confirmed a complete PTR and two major PTRs. With a median follow-up of 21 months, all patients are alive with no recurrence. Circulating tumor DNA and cell-free DNA dynamics correlated with clinical and pathological response and distinguished two patient groups with different tumor biological behavior after sitravatinib alone (1A) versus sitravatinib-nivolumab (1B). Tumor immunophenotyping and scRNAseq analyses revealed differential changes in the expression of immune cell populations and sitravatinib-targeted and hypoxia-related genes in group 1A vs 1B patients.

Conclusions: The SNOW study shows sitravatinib plus nivolumab is safe and leads to deep clinical and pathological responses in oral cavity carcinomas. Multi-omic biomarker analyses dissect the differential molecular effects of sitravatinib versus the sitravatinib-nivolumab and revealed patients with distinct tumor biology behavior.

Trial registration number: NCT03575598.

Keywords: clinical trials as topic; head and neck neoplasms; immunotherapy; macrophages; tumor biomarkers.

Conflict of interest statement

Competing interests: DVA: honoraria from GSK, MSD and Pfizer. AP: research funding from BMS. AS: consultant for (Advisory Board), Merck (compensated), Bristol-Myers Squibb (compensated), Novartis (compensated), Oncorus (compensated), Janssen (compensated). Grant/Research support from (Clinical Trials): Novartis, Bristol Myers Squibb, Symphogen, AstraZeneca/Medimmune, Merck, Bayer, Surface Oncology, Northern Biologics, Janssen Oncology/Johnson & Johnson, Roche, Regeneron, Alkermes, Array Biopharma/Pfizer, GSK. ARH: consultant for (Advisory Board) Bristol Myers Squibb (compensated), Merck (compensated), GlaxoSmithKline (compensated), Eisai (compensated). Grant/Research support from (Clinical Trials): Karyopharm, Bristol Myers Squibb, GlaxoSmithKline, Merck, Boerhinger-Ingelheim, AstraZeneca/Medimmune, Roche/Genentech, Macrogenics, Astellas Pharma, Janssen. PO, RS, HD are employees and shareholders of Mirati Therapeutics. NA, QA are employees of Neogenomics Laboratories. LS: consulting/advisory arrangements with Merck, Pfizer, Celgene, AstraZeneca, Morphosys, Roche, Oncorus, Symphogen, Seattle Genetics, GlaxoSmithKline, Voronoi, Arvinas, Tessa, Navire, Relay, Rubius, Janpix, Daiichi Sanyko; stock ownership of Agios (spouse); leadership position in Treadwell Therapeutics (spouse); and institution receives clinical trials support from Novartis, Bristol Myers Squibb, Pfizer, Boerhinger-Ingelheim, GlaxoSmithKline, Roche/Genentech, Karyopharm, AstraZeneca, Merck, Celgene, Astellas, Bayer, AbbVie, Amgen, Symphogen, Intensity Therapeutics, Mirati Therapeutics, Shattucks, Avid.

© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY. Published by BMJ.

Figures

Figure 1

ctDNA (MTM/mL) (A) and cfDNA (ng/mL) (B) dynamics following study treatment. (A) Two-dimensional line charts showing MTM/mL at each of the three time points. (B) Two-dimensional line charts showing cfDNA in ng/mL at each of the three time points. The symbols represent each individual patient. The colors represent the groups according to the distinct tumor biological behavior following treatment. cfDNA, cell-free DNA; ctDNA, circulating tumor DNA; D15, day 15; MTM, mean tumor molecules; PRE, pretreatment; SRG, pre-surgery.

Figure 2

ctDNA dynamics correlated with tumor changes following sitravatinib and sitravatinib plus nivolumab. Charts showing log-scale changes in ctDNA and cfDNA (Y-axis) at each time point (X-axis) in each individual patient. Tumor photographs performed during study at each of the corresponding time points are shown above the line charts for each patient. Arrows indicate the location of the primary tumor. ctDNA, circulating tumor DNA; cfDNA, cell-free DNA; D15, day 15; MTM, mean tumor molecules; PRE, pretreatment; SRG, pre-surgery.

Figure 3

Changes in tumor-associated macrophage populations following sitravatinib alone (D15) and sitravatinib plus nivolumab (SRG). (A) Uniform Manifold Approximation and Projection (UMAP) plots showing the integration of 15 samples from five SNOW patients at three time points. Colors represent cell types. (B) Multiplexing immuno-fluorescence staining in tumor biopsies at pre-treatment (PRE), day 15 (D15) and pre-surgery (SRG) using NeoGenomics MultiOmyx panels showing changes in macrophages subpopulations: M1 type (CD68+CD163–) shown in red, M2 type (CD68+CD163+) shown in yellow and M1 intermediate type (CD68+HLA-DR+CD163–) shown in magenta. Upper images show H&E staining of tissue sample. (C) Ratio of M1/M2 macrophages detected using IHC and scRNAseq measurements. Orange: 1A=responders to sitravatinib; blue: 1B=responders to sitravatinib–nivolumab; black=unclassifiable. IHC, immunohistochemistry; NK, natural killer; MDSC, myeloid-derived suppressor cell; scRNAseq, single-cell RNA sequencing.

Figure 4

Single-cell RNA sequencing pathways enrichment analysis comparing Group 1A versus Group 1B patients at pre-surgery time point. Bar plot showing Gene Set Enrichment Analysis NES for the MSigDB (Molecular Signatures Database) hallmark pathways. Significant hits (adjusted p value