MiR-34a, an important tumor-suppressing microRNA, is downregulated in several types of cancer; loss of its expression has been linked with unfavorable clinical outcomes in non-small-cell lung cancer (NSCLC), among others. MiR-34a represses several key oncogenic proteins, and a synthetic mimic of miR-34a is currently being tested in a cancer trial. However, little is known about the potential role of miR-34a in regulating DNA damage response and repair. Here, we demonstrate that miR-34a directly binds to the 3' untranslated region of RAD51 and regulates homologous recombination, inhibiting double-strand-break repair in NSCLC cells. We further demonstrate the therapeutic potential of miR-34a delivery in combination with radiotherapy in mouse models of lung cancer. Collectively, our results suggest that administration of miR-34a in combination with radiotherapy may represent a novel strategy for treating NSCLC.

Despite the potential to cure metastatic disease, immunotherapy on its own often fails outright or early on due to tumor immune evasion. To address this obstacle, we investigated combinations of anti-GITR, anti-PD1 and radiation therapy (XRT) in our previously developed anti-PD1 resistant 344SQ non-small cell lung adenocarcinoma preclinical tumor model. We hypothesized that targeting multiple mechanisms of immune evasion with this triple therapy would lead to an enhanced tumor-specific immune response and improve survival more so than any mono- or dual therapy. In a two tumor 344SQR murine model, treatment with anti-GITR, anti-PD1, and XRT led to significantly improved survival and an abscopal response, with half of the mice becoming tumor free. These mice showed durable response and increased CD4+ and CD8+ effector memory on tumor rechallenge. Regulatory T cells (Tregs) expressed the highest level of GITR at the tumor site and anti-GITR therapy drastically diminished Tregs at the tumor site. Anti-tumor effects were largely dependent on CD4+ T cells and partially dependent on CD8+ T cells. Anti-GITR IgG2a demonstrated superior efficacy to anti-GITR IgG1 in driving antitumor effects. Collectively, these results suggest that combinatorial strategies targeting multiple points of tumor immune evasion may lead to a robust and lasting antitumor response.

Radiation is used extensively to treat localized cancer, but improved understanding of its effects on the immune system has increased interest in its potential systemic (abscopal) effects, particularly in combination with checkpoint inhibitors such as anti-PD1. The majority of patients either do not respond or develop resistance to monotherapy over time. Here, we investigated the efficacy of OX40 (CD134) stimulation as an alternative immunotherapeutic approach in combination with radiotherapy (XRT) in a murine model of anti-PD1-resistant lung tumors. We established a bilateral tumor model in 129Sv/Ev mice using an anti-PD1-resistant lung tumor cell line. Primary tumors were treated with intratumoral injection of an OX40 agonist antibody, given as adjuvant therapy after XRT (36 Gy in three 12-Gy fractions), whereas secondary tumors were left untreated to investigate abscopal outcomes. The combination of XRT followed by OX40 stimulation effectively inhibited local and systemic antitumor growth, limited lung metastases, and improved survival rates. This treatment regimen augmented CD4 and CD8 T-cell expansion. XRT induced the expression of OX40 on T cells in tumors and spleens and increased the percentages of splenic CD103 dendritic cells. Our data extend the benefits of radiation to systemic disease control, especially when combined with anti-OX40 agonist to promote immunologically mediated abscopal effects. Moreover, this study provides a rational treatment approach and sequence to overcome anti-PD1-resistant poorly immunogenic tumors. .

Radiation therapy controls local disease but also prompts the release of tumor-associated antigens and stress-related danger signals that primes T cells to promote tumor regression at unirradiated sites known as the abscopal effect. This may be enhanced by blocking inhibitory immune signals that modulate immune activity through a variety of mechanisms. Indeed, abscopal responses have occurred in patients with lung cancer or melanoma when given anti-CTLA4 antibody and radiation. Other approaches involve expanding and reinfusing T or NK cells or engineered T cells to express receptors that target specific tumor peptides. These approaches may be useful for immunocompromised patients receiving radiation. Preclinical and clinical studies are testing both immune checkpoint–based strategies and adoptive immunotherapies with radiation.

Indoleamine 2,3-dioxygenase 1 (IDO1), involved in the catabolism of tryptophan (Trp) to kynurenine (Kyn) is an important regulator of tumor-mediated immunosuppression implicated in resistance to anti-PD1 immunotherapy. We investigated the role of IDO1 in an anti-PD1-resistant lung cancer model (344SQ_R) compared to the parental 344SQ tumors (344SQ_P). IDO1 was overexpressed in tumor-infiltrating leukocytes, and plasma Kyn levels were increased, in 344SQ_R vs. 344SQ_P. The IDO1 inhibitor INCB023843 retarded tumor growth and reduced lung metastases in 344SQ_R. IDO1 was expressed at higher levels in F4/80Gr1CD11b myeloid-derived suppressor cells (MDSCs) that were prominent in 344SQ_R. The INCB023843 reduced IDO1 expression and percentages of these MDSCs while increasing CD8 T cells infiltration, hence reactivating antitumor T-cell responses in 344SQ_R. Therefore, IDO1 inhibition holds promise for treating lung cancer that does not respond to anti-PD1 therapy.