Mavorixafor, an Orally Bioavailable CXCR4 Antagonist, Increases Immune Cell Infiltration and Inflammatory Status of Tumor Microenvironment in Patients with Melanoma

Abstract

Purpose: Mavorixafor is an oral, selective inhibitor of the CXCR4 chemokine receptor that modulates immune cell trafficking. A biomarker-driven phase Ib study (NCT02823405) was conducted in 16 patients with melanoma to investigate the hypothesis that mavorixafor favorably modulates immune cell profiles in the tumor microenvironment (TME) and to evaluate the safety of mavorixafor alone and in combination with pembrolizumab.

Experimental design: Serial biopsies of melanoma lesions were assessed after 3 weeks of mavorixafor monotherapy and after 6 weeks of combination treatment for immune cell markers by NanoString analysis for gene expression and by multiplexed immunofluorescent staining for in situ protein expression. Serum samples taken at biopsy timepoints were evaluated for key chemokine and cytokine alterations using the Myriad Rules Based Medicine multiplex immunoassays.

Results: Within the TME, mavorixafor alone increased CD8⁺ T-cell infiltration, granzyme B signal, antigen presentation machinery, and both tumor inflammatory signature (TIS) and IFNγ gene expression signature scores. Increases in the key serum cytokines CXCL9 and CXCL10 were further enhanced when mavorixafor was combined with pembrolizumab. Adverse events (AE), as assessed by the investigator according to NCI Common Terminology Criteria for Adverse Events (v4.03), related to either mavorixafor or pembrolizumab (≥15%) were diarrhea, fatigue, maculopapular rash, and dry eye. Reported AEs were all ≤ grade 3.

Conclusion/discussion: Treatment with single-agent mavorixafor resulted in enhanced immune cell infiltration and activation in the TME, leading to increases in TIS and IFNγ gene signatures. Mavorixafor as a single agent, and in combination with pembrolizumab, has an acceptable safety profile. These data support further investigation of the use of mavorixafor for patients unresponsive to checkpoint inhibitors.

Significance: Despite survival improvements in patients with melanoma treated with checkpoint inhibitor therapy, a significant unmet medical need exists for therapies that enhance effectiveness. We propose that mavorixafor sensitizes the melanoma tumor microenvironment and enhances the activity of checkpoint inhibitors, and thereby may translate to a promising treatment for broader patient populations.

FIGURE 1

Study schematic. Sixteen patients received 400 mg mavorixafor every day for 9 weeks. Following 3 weeks of mavorixafor monotherapy, 6-week combination therapy was initiated by the addition of pembrolizumab (intravenous infusion). Sampling occurred at baseline, week 4 (post-monotherapy), and week 9 (post-combination therapy).

FIGURE 2

T-cell infiltration of the melanoma TME after mavorixafor therapy. Increased infiltration of the melanoma TME by CD8⁺ cells labeled by IHC predose (A) or at the end of mavorixafor monotherapy (B). Eight of 11 patient samples had a demonstrable increase in the fold change of CD8⁺ cell within the melanoma sample when compared before and after monotherapy (C). CTL signature scores for available pairs of patient biopsies following mavorixafor monotherapy (D). CD8⁺ T cells at the melanoma tumor interface with normal tissue were labeled using multiplex IHC (E). CD8⁺ cells/mm² using HALO image analysis and plotted against distance from the tumor boundary in 25 μm bands (F). Labeled cells within 100 μm inside or outside of the tumor boundary with normal tissue were quantified (G).

FIGURE 3

Mavorixafor-activated immune cell activities in TME. IHC labeling for granzyme B at predose (A) and postdose timepoints (B). The fold change of granzyme B positivity posttreatment for all evaluable samples (C). Quantification was performed using HALO software and the entire tumor area was scored. RNA expression levels for GZMB for 5 patients with both pre- and post-mavorixafor monotherapy treatment-evaluable biopsies (D). Data shown (C and D) is from all evaluable patient samples available at the time of analysis. Biopsies of melanoma lesions from patient #5 stained by mIF for CD8⁺ cells, Ki-67 (proliferating cells), and a cocktail of melanoma-specific antibodies to label tumor (E--G). Images represent the graphical output from the nearest-neighbor analysis module, with unlabeled cells rendered as gray.

FIGURE 4

CXCR4 inhibitor affects the TME. Increased gene expression scores for IFNγ (A) and TIS (B) as determined by NanoString analysis of RNA extracted from patient biopsy samples at predose and post-mavorixafor timepoints. Gene expression scores for antigen processing and presentation genes as determined by NanoString analysis for 9 patients receiving Mavorixafor monotherapy (C). Data shown are from all evaluable patient samples available at the time of analyses.

FIGURE 5

Repertoire diversity of patients 3, 5, and 8 demonstrating cumulative clonal frequency. Repertoire diversity obtained by plotting the cumulative frequency of a selected number of the most frequent clones using the function topSeqsPlot. Each of the top sequences is represented by its own color, with less-frequent clones assigned a single color (violet).