Cancer Res Treat.  2022 Apr;54(2):458-468. 10.4143/crt.2021.425.

Histone Deacetylase as a Valuable Predictive Biomarker and Therapeutic Target in Immunotherapy for Non-Small Cell Lung Cancer

• Shin HS
• Choi J
• Lee J
• Lee SY

• KMID: 2528215
• DOI: http://doi.org/10.4143/crt.2021.425

Abstract

Purpose
Histone deacetylase inhibitors (HDACis) are epigenetic regulators and used clinically for hematopoietic malignancies. Recently, HDACis have received attention as a factor that modulates the immune system. In this study, the role of histone deacetylase (HDAC) expression as a predictive marker in lung cancer patients who were treated with immune checkpoint inhibitors (ICIs) and the role of HDACi and ICI combination treatment in the mouse tumor model were analyzed.
Materials and Methods
The overall response rate (ORR) and progression-free survival (PFS) were analyzed by the expression of HDAC. In vitro assay, the mRNA and protein expression levels of cytokines and programmed death-ligand 1 (PD-L1) were analyzed after HDACi treatment. In vivo assay, TC-1 tumor-bearing mice were treated with HDACi and mouse programmed cell death 1 (PD-1) inhibitor.
Results
The HDAC6 low expression group showed high ORR and prolonged PFS. When the selective HDAC6 inhibitor was administered to the A549 cell line, the levels of interleukin-1β and interleukin-6 decreased and the expression of PD-L1 was reduced. Mice that received both the mouse PD-1 inhibitor and pan-HDACi had a smaller tumor size than that of the mice from the control group. Moreover, mice treated with the mouse PD-1 inhibitor and pan-HDACi generated greater numbers of E7-specific CD8+ T cells.
Conclusion
HDAC6 expression can predict the prognosis of non–small cell lung cancerpatients who were treated with ICIs. Furthermore, co-treatment with HDACi and PD-1 inhibitor was shown to decrease the tumor growth rate and create a favorable tumor microenvironment for cytotoxic T lymphocytes in the TC-1 mouse model.

Keyword

Non-small-cell lung carcinoma; Histone deacetylase; Histone deacetylase inhibitor; Tumor microenvironment; Immune checkpoint inhibitor; Interleukin; Programmed cell death 1

Figure

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  Fig. 1 Representative tumor microarray core stained with histone deacetylase 6 (HDAC6). The score was recorded as a percentage of tumor cells that showed any intensity of cytoplasmic immunoreactivity. (A) HDAC6-High group (HDAC expression 51%-100%). (B) HDAC6-Low group (HDAC expression 0%-50%).

• 

  Fig. 2 Patient flow chart. In total, 92 patients received immune checkpoint inhibitors; 46 patients were excluded based on the exclusion criteria. There were 11 and 35 patients in the histone deacetylase 6 (HDAC6)-Low and HDAC6-High groups, respectively. ECOG PS, Eastern Cooperative Oncology Group performance status.

• 

  Fig. 3 Waterfall plot for the overall objective response rate in patients with metastatic non–small cell lung cancer who received immune checkpoint inhibitors. The overall tumor size showed a greater decrease in the histone deacetylase 6 (HDAC6)–Low group. On average, the tumor size of the HDAC6-High group decreased by 13.4%±35.8%, and that of the HDAC6-Low group decreased by 38.6%±31.8%. PD, progressive disease; PR, partial response.

• 

  Fig. 4 Kaplan-Meier survival curves for progression-free survival (A) and overall survival (B). (A) Twenty-six patients showed progressive disease (PD) in the histone deacetylase 6 (HDAC6)–High group, whereas only four patients in the HDAC6-Low group showed PD (p=0.035). (B) Fifteen and three patients died in the HDAC6-High and HDAC6-Low groups, respectively (p=0.090).

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  Fig. 5 Reverse transcription polymerase chain reaction (RT-PCR) and Western blotting analysis of inflammatory cytokines. (A) A549 cells were treated in vitro with Nexturastat A dose-dependently, then mRNA levels of interleukin (IL)-1β and IL-6 were measured by RT-PCR. (B) Protein expression levels of IL-1β and IL-6 in A549 cells were measured by Western blot experiment after treatment with Nexturastat A. GAPDH, glyceraldehyde 3-phosphate dehydrogenase.

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  Fig. 6 Reverse transcription polymerase chain reaction (RT-PCR) and Western blotting analysis of programmed death-ligand 1 (PD-L1). (A) A549 cells were treated in vitro with Nexturastat A dose-dependently, then mRNA levels of PD-L1 were measured by RT-PCR. (B) Protein expression levels of PD-L1 in A549 cells measured by Western blot experiment after treatment with Nexturastat A or pan-HDACi (M166). GAPDH, glyceraldehyde 3-phosphate dehydrogenase; HDACi, histone deacetylase inhibitor.

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  Fig. 7 The TC-1 mouse model study. (A) Schematic representation of the anti-mouse programmed cell death 1 (PD-1) antibody (CD279) and new pan-HDACi (M166) treatment regimens. C57BL/6 mice (five per group) were subcutaneously injected with 2×105 TC-1 cells per mouse on day 0. Tumor-bearing mice were treated with CD279 and/or M166 beginning on day 10, as indicated in the timeline. CD279 (200 μg/mouse) was administered at intervals of 2–3 days. M166 was administered via intraperitoneal injection at a dose of 60 mg/kg body weight. (B) Tumor volumes of mice treated with PD-1 antibody and new pan-HDACi (M166). Mice that received both mouse PD-1 inhibitor and new pan-HDACi (M166) had a smaller tumor size compared to that of mice from the control group (p=0.03). (C) Characterization of E7-specific CD8+ T cells. Mice receiving both the mouse PD-1 inhibitor and pan-HDACi generated significantly more E7-specific interferon-γ–producing CD8+ T cells. HDACi, histone deacetylase inhibitors; ICI, immune check point inhibitor; IFN-γ, intracellular interferon γ.

Reference

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