Atezolizumab in combination with bevacizumab enhances antigen-specific T-cell migration in metastatic renal cell carcinoma

Abstract

Anti-tumour immune activation by checkpoint inhibitors leads to durable responses in a variety of cancers, but combination approaches are required to extend this benefit beyond a subset of patients. In preclinical models tumour-derived VEGF limits immune cell activity while anti-VEGF augments intra-tumoral T-cell infiltration, potentially through vascular normalization and endothelial cell activation. This study investigates how VEGF blockade with bevacizumab could potentiate PD-L1 checkpoint inhibition with atezolizumab in mRCC. Tissue collections are before treatment, after bevacizumab and after the addition of atezolizumab. We discover that intra-tumoral CD8(+) T cells increase following combination treatment. A related increase is found in intra-tumoral MHC-I, Th1 and T-effector markers, and chemokines, most notably CX3CL1 (fractalkine). We also discover that the fractalkine receptor increases on peripheral CD8(+) T cells with treatment. Furthermore, trafficking lymphocyte increases are observed in tumors following bevacizumab and combination treatment. These data suggest that the anti-VEGF and anti-PD-L1 combination improves antigen-specific T-cell migration.

Conflict of interest statement

This study was sponsored by Genentech, Inc., a member of the Roche Group, which provided the study drug. Some of the authors of this manuscript are employees of Genentech/Roche. The remaining authors declare no competing financial interests.

Figures

Figure 1. Antitumour activity of atezolizumab and bevacizumab combination.

(a) Tumour burden over time in RCC patients. Plot of patients with RCC measuring the maximum reduction from baseline in the sum of the longest diameter for target lesions. CR, complete response; PD, progressive disease; PR, partial response; SD, stable disease. (b) Duration of study treatment for patients with RCC.

Figure 2. Gene expression tumour biomarkers following treatments.

Levels of on-treatment tumour samples are shown relative to the baseline levels (dotted line) for the patients where the pre-treatment, post-bevacizumab (bev; blue circles) and post-combination biopsies were collected. Vascular signature genes (ANGPT2, CD34, DLL4, EGFL7 and ESM1) are shown in black, CD8 T effector genes (CD8A, CD8B, EOMES, GZMA, GZMB, IFNG and PRF1) are shown in red, Th1 chemokines (CXCL10, CXCL11, CXCL13 and CXCL9) are shown in blue and natural killer (NK) cell genes (GZMB, KLRK1 and SLAMF7) are shown in green. atezo, atezolizumab.

Figure 3. Protein expression of immune and vasculature markers in baseline and on-treatment tumour samples.

(a) Representative images of CD8 (blue), CD31 (blue), PD-L1 (brown) and MHC-I (brown) by IHC from patient 3 tumours. (b) Quantitation of CD31, CD8 and MHC-I IHC. P values were determined by paired t-test. The line in the middle of the box is plotted at the median. Lines above and below the boxes represent variability outside the upper and lower quartiles. (c) IHC images for the indicated triple and double stains of serial sections from patient 3 tumours. A scale bar for each image representing 50 or 100 μm is shown. atezo, atezolizumab; bev, bevacizumab.

Figure 4. Analyses of antigen-specific T cells in the blood and T-cell repertoire.

(a) Flow cytometry analyses of antigen-specific T cells (dextramer positive). Representative data from two HLA-A2 positive patients with blood draws matched to tumour biopsy time points are shown. (b) Increased expression of chemokines, including CX3CL1 with treatment in tumours. The line in the middle of the box is plotted at the median. Lines above and below the boxes represent variability outside the upper and lower quartiles. (c) TCRβ sequencing from patient 6 TILs before and after treatment. The top clones (up to 25) for each group are shown. Prevalence of trending TCRβ clone populations are shown in red, blue, green, orange, and purple. (d) TCRβ sequencing of patient 2, 3, and 6 pre-treatment PBMCs, post-bevacizumab (bev) and –atezolizumab (atezo) PBMCs, and post-bevacizumab and -atezolizumab TILs. The top clones (up to 25) for each group are shown.