A phase 1/2 clinical trial in patients with chemoresistant triple-negative breast cancer targets the inducible nitric oxide signaling pathway.

Immunosuppressive elements within the tumor microenvironment, such as tumor-associated macrophages (TAM), can present a barrier to successful anti-tumor responses by cytolytic T cells. Here we employed preclinical syngeneic p53 null mouse models of triple-negative breast cancer (TNBC) to develop a treatment regimen that harnessed the immunostimulatory effects of low-dose cyclophosphamide coupled with the pharmacologic inhibition of TAMs using either a small molecule CSF1R inhibitor or an anti-CSF1R antibody. This therapeutic combination was effective in treating several highly aggressive TNBC murine mammary tumor and lung metastasis models. Single cell RNA sequencing characterized tumor-infiltrating lymphocytes (TIL) including helper T cells and antigen-presenting B cells that were highly enriched in responders to combination therapy. In one model that exhibited long-term post-treatment tumor regression, high dimensional imaging techniques identified the close spatial localization of B220+/CD86+-activated B cells and CD4+ T cells in tertiary lymphoid structures that were present up to 6 weeks post-treatment. The transcriptional and metabolic heterogeneity of TAMs was also characterized in two closely related claudin-low/mesenchymal subtype tumor models with differential treatment responses. A murine TAM signature derived from the T12 model was highly conserved in human claudin-low breast cancers, and high expression of the TAM signature correlated with reduced overall survival in breast cancer patients. This TAM signature may help identify human claudin-low breast cancer patients that will benefit from the combination of cyclophosphamide and anti-CSF1R therapy. These studies illustrate the complexity of the tumor immune microenvironment and highlight different immune responses that result from rational immunotherapy combinations.

Purpose: A Phase 2 trial of stereotactic radiation therapy and in situ cytotoxic virus therapy in metastatic triple-negative breast cancer (mTNBC) patients followed by pembrolizumab (STOMP) was designed to evaluate dual approach of enhancing single-agent immune checkpoint blockade with ADV/HSV-tk plus valacyclovir gene therapy and SBRT in mTNBC patients. Methods: In this single-arm, open-label Phase 2 trial, mTNBC patients were treated with ADV/HSV-tk (5 x 1011 vp) intratumoral injection, followed by SBRT to the injected tumor site, then pembrolizumab (200mg, Q3w). The primary endpoint was clinical benefit rate (CBR, CR, PR or SD≥ 24 weeks per RECIST version1.1 at non-irradiated site). Secondary endpoints included duration on treatment (DoT), OS and safety. Exploratory endpoints included immune response to treatment assessed by correlative tissue and blood-based biomarkers. Results: 28 patients were enrolled and treated. CBR was seen in 6 patients (21.4%) including two CR (7.1%), one PR (3.6%) and three SD (10.7%). Patients with clinical benefit had durable responses, with median DoT of 9.6 months and OS of 14.7 months. The median OS was 6.6 months in the total population. The combination was well tolerated. Correlative studies with CyTOF and IMC revealed a significant increase of CD8 T cells in responders and of myeloid cells in non-responders. Conclusions: The median OS increased by more than 2-fold in patients with clinical benefit. The therapy is a well-tolerated treatment in heavily pretreated mTNBC patients. Early detection of increased effector and effector memory CD8 T cells and myeloids correlate with response and non-response, respectively.

Tumors accumulated with infiltrated immune cells (hot-tumors) have a higher response rate to immune checkpoint blockade, when compared to those with minimal T cell infiltration (cold-tumors). We report here that lung cancer patients with different racial backgrounds harbored distinct immune cell profiles in the tumor microenvironment. Compared to African Americans (AAs), Caucasian Americans (CAs) exhibited increased immune cell infiltration and vasculature, and increased survival. Changes of survival and immune profile were most pronounced among active smokers and non-smokers, compared to former smokers and total patients. Neighborhood analysis showed that immune cells accumulated around cancer cells in CAs but not AAs. Our findings reveal intrinsic biological differences between AA and CA lung cancer patients, suggesting that treatment plans should be tailored for patients with different racial backgrounds.

The sympathetic nervous system (SNS) is an important regulator of immune cell function during homeostasis and states of inflammation. Recently, the SNS has been found to bolster tumor growth and impair the development of antitumor immunity. However, it is unclear whether the SNS can modulate APC function. Here, we investigated the effects of SNS signaling in murine monocyte-derived macrophages (moMФ) and dendritic cells (DCs) and further combined the nonspecific β-blocker propranolol with a peptide cancer vaccine for the treatment of melanoma in mice. We report that norepinephrine treatment dramatically altered moMФ cytokine production, whereas DCs were unresponsive to norepinephrine and critically lack β2-adrenergic receptor expression. In addition, we show that propranolol plus cancer vaccine enhanced peripheral DC maturation, increased the intratumor proportion of effector CD8+ T cells, and decreased the presence of intratumor PD-L1+ myeloid-derived suppressor cells. Furthermore, this combination dramatically reduced tumor growth compared with vaccination alone. Taken together, these results offer insights into the cell-specific manner by which the SNS regulates the APC immune compartment and provide strong support for the use of propranolol in combination with cancer vaccines to improve patient response rates and survival.