Abstract
Neutrophils are an essential part of the innate immune system. To study their importance, experimental studies often aim to deplete these cells, generally by injecting anti-Ly6G or anti-Gr1 ...antibodies. However, these approaches are only partially effective, transient or lack specificity. Here we report that neutrophils remaining after anti-Ly6G treatment are newly derived from the bone marrow, instead of depletion escapees. Mechanistically, newly generated, circulating neutrophils have lower Ly6G membrane expression, and consequently reduced targets for anti-Ly6G-mediated depletion. To overcome this limitation, we develop a double antibody-based depletion strategy that enhances neutrophil elimination by anti-Ly6G treatment. This approach achieves specific, durable and controlled reduction of neutrophils in vivo, and may be suitable for studying neutrophil function in experimental models.
Understanding the immune compartment of tumors facilitates the development of revolutionary new therapies. We used a Kras(G12D)-driven mouse model of lung cancer to establish an immune signature and ...identified a contribution of Gr1+ neutrophils to disease progression. Depletion experiments showed that Gr1+ cells (1) favor tumor growth, (2) reduce T cell homing and prevent successful anti-PD1 immunotherapy, and (3) alter angiogenesis, leading to hypoxia and sustained Snail expression in lung cancer cells. In turn, Snail accelerated disease progression and increased intratumoral Cxcl2 secretion and neutrophil infiltration. Cxcl2 was produced mainly by neutrophils themselves in response to a factor secreted by Snail-expressing tumor cells. We therefore propose a vicious cycle encompassing neutrophils and Snail to maintain a deleterious tumor microenvironment.
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•An unsupervised immune signature allows stratification of poorly immunogenic tumors•Neutrophil depletion reverts immune exclusion allowing anti-PD1 treatment efficacy•Neutrophils alter angiogenesis, increasing hypoxia and stabilizing Snail•Snail favors neutrophil homing, induces partial EMT, and enhances tumor progression
Faget et al. extract an immune signature from lung tumors pointing to neutrophils as contributors to disease progression. They show that neutrophils inhibit immunotherapy efficacy and alter angiogenesis, increasing tumor hypoxia and Snail expression. Snail enhances tumor growth and neutrophil recruitment, establishing an amplification loop favoring cancer aggressiveness.
Abstract This study shows for the first time that normal brain tissue toxicities after WBI can be reduced with increased dose rate. Spatial memory is preserved after WBI with mean dose rates above ...100 Gy/s, whereas 10 Gy WBI at a conventional radiotherapy dose rate (0.1 Gy/s) totally impairs spatial memory.
To rapidly assess healthy tissue toxicities induced by new anti-cancer therapies (i.e., radiation alone or in combination with drugs), there is a critical need for relevant and easy-to-use models. ...Consistent with the ethical desire to reduce the use of animals in medical research, we propose to monitor lung toxicity using an ex vivo model. Briefly, freshly prepared organotypic lung slices from mice were irradiated, with or without being previously exposed to chemotherapy, and treatment toxicity was evaluated by analysis of cell division and viability of the slices. When exposed to different doses of radiation, this ex vivo model showed a dose-dependent decrease in cell division and viability. Interestingly, monitoring cell division was sensitive enough to detect a sparing effect induced by FLASH radiotherapy as well as the effect of combined treatment. Altogether, the organotypic lung slices can be used as a screening platform to rapidly determine in a quantitative manner the level of lung toxicity induced by different treatments alone or in combination with chemotherapy while drastically reducing the number of animals. Translated to human lung samples, this ex vivo assay could serve as an innovative method to investigate patients’ sensitivity to radiation and drugs.
The anti-Ly6G antibody is used to deplete Ly6G
pos
neutrophils and study their role in diverse pathologies. However, depletion is never absolute, as Ly6G
low
neutrophils resistant to depletion ...rapidly emerge. Studying the functionality of these residual neutrophils is necessary to interpret anti-Ly6G-based experimental designs. In vitro, we found anti-Ly6G binding induced Ly6G internalization, surface Ly6G paucity, and primed the oxidative burst of neutrophils upon TNF α co-stimulation. In vivo, we found neutrophils resistant to anti-Ly6G depletion exhibited anti-neutrophil-cytoplasmic-antibodies. In the pre-clinical Kras
Lox-STOP-Lox-G12D/WT
; Trp53
Flox/Flox
mouse lung tumor model, abnormal neutrophil accumulation and aging was accompanied with an N2-like SiglecF
pos
polarization and ly6g downregulation. Consequently, SiglecF
pos
neutrophils exposed to anti-Ly6G reverted to Ly6G
low
and were resistant to depletion. Noting that anti-Ly6G mediated neutrophil depletion alone had no anti-tumor effect, we found a long-lasting rate of tumor regression (50%) by combining anti-Ly6G with radiation-therapy, in this model reputed to be refractory to standard anticancer therapies. Mechanistically, anti-Ly6G regulated neutrophil aging while radiation-therapy enhanced the homing of anti-Ly6G-boundSiglecF
neg
neutrophils to tumors. This anti-tumor effect was recapitulated by G-CSF administration prior to RT and abrogated with an anti-TNFα antibody co-administration. In summary, we report that incomplete depletion of neutrophils using targeted antibodies can intrinsically promote their oxidative activity. This effect depends on antigen/antibody trafficking and can be harnessed locally using select delivery of radiation-therapy to impair tumor progression. This underutilized aspect of immune physiology may be adapted to expand the scope of neutrophil-related research.
Immune-based anti-cancer strategies combined with radiation therapy (RT) are actively being investigated but many questions remain, such as the ideal treatment scheme and whether a potent immune ...response can be generated both locally and systemically. In this context, tumor-associated tertiary lymphoid structures (TLS) have become a subject of research. While TLS are present in several types of cancer with strong similarities, they are especially relevant in medullary breast carcinoma (MBC). This suggests that MBC patients are ideally suited for investigating this question and may benefit from adapted therapeutic options. As RT is a corner-stone of MBC treatment, investigating interactions between RT and TLS composition is also clinically relevant. We thus first characterized the lymphoid structures associated with MBC in a patient case report and demonstrated that they closely resemble the TLS observed in a genetical mouse model. In this model, we quantitatively and qualitatively investigated the cellular composition of the tumor-associated TLS. Finally, we investigated TLS regulation after hypo-fractionated RT and showed that RT induced their acute and transient depletion, followed by a restoration phase. This study is the first work to bring a comprehensive and timely characterization of tumor-associated TLS in basal conditions and after RT. It highlights cellular targets (i.e., Tregs) that could be selectively modulated in subsequent studies to optimize anti-tumor immune response. The study of TLS modulation is worth further investigation in the context of RT and personalized medicine.
Neutrophils are the most abundant circulating leucocytes and are essential for innate immunity. In cancer, pro- or antitumor properties have been attributed to tumor-associated neutrophils (TAN). ...Here, focusing on TAN accumulation within lung tumors, we identify GLUT1 as an essential glucose transporter for their tumor supportive behavior. Compared with normal neutrophils, GLUT1 and glucose metabolism increased in TANs from a mouse model of lung adenocarcinoma. To elucidate the impact of glucose uptake on TANs, we used a strategy with two recombinases, dissociating tumor initiation from neutrophil-specific
deletion. Loss of GLUT1 accelerated neutrophil turnover in tumors and reduced a subset of TANs expressing SiglecF. In the absence of GLUT1 expression by TANs, tumor growth was diminished and the efficacy of radiotherapy was augmented. Our results demonstrate the importance of GLUT1 in TANs, which may affect their pro- versus antitumor behavior. These results also suggest targeting metabolic vulnerabilities to favor antitumor neutrophils. SIGNIFICANCE: Lung tumor support and radiotherapy resistance depend on GLUT1-mediated glucose uptake in tumor-associated neutrophils, indicating that metabolic vulnerabilities should be considered to target both tumor cells as well as innate immune cells. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/9/2345/F1.large.jpg.
Immune checkpoint blockade (ICB) with PD-1 or PD-L1 antibodies has been approved for the treatment of non-small cell lung cancer (NSCLC). However, only a minority of patients respond, and sustained ...remissions are rare. Both chemotherapy and antiangiogenic drugs may improve the efficacy of ICB in mouse tumor models and patients with cancer. Here, we used genetically engineered mouse models of
;
NSCLC, including a mismatch repair-deficient variant (
;
;
) with higher mutational burden, and longitudinal imaging to study tumor response and resistance to combinations of ICB, antiangiogenic therapy, and chemotherapy. Antiangiogenic blockade of vascular endothelial growth factor A and angiopoietin-2 markedly slowed progression of autochthonous lung tumors, but contrary to findings in other cancer types, addition of a PD-1 or PD-L1 antibody was not beneficial and even accelerated progression of a fraction of the tumors. We found that antiangiogenic treatment facilitated tumor infiltration by PD-1
regulatory T cells (T
), which were more efficiently targeted by the PD-1 antibody than CD8
T cells. Both tumor-associated macrophages (TAMs) of monocyte origin, which are colony-stimulating factor 1 receptor (CSF1R) dependent, and TAMs of alveolar origin, which are sensitive to cisplatin, contributed to establish a transforming growth factor-β-rich tumor microenvironment that supported PD-1
T
Dual TAM targeting with a combination of a CSF1R inhibitor and cisplatin abated T
, redirected the PD-1 antibody to CD8
T cells, and improved the efficacy of antiangiogenic immunotherapy, achieving regression of most tumors.
The physico-chemical and biological response to conventional and UHDR electron and proton beams was investigated, along with conventional photons. The temporal structure and nature of the beam ...affected both, with electron beam at ≥1400 Gy/s and proton beam at 0.1 and 1260 Gy/s found to be isoefficient at sparing zebrafish embryos.
The incidence of human papillomavirus-associated head and neck squamous cell carcinoma (HPV
-HNSCC) is rising worldwide and although current therapeutic modalities are efficient in the majority of ...patients, there is a high rate of treatment failures. Thus, novel combination approaches are urgently needed to achieve better disease control in patients with HPV
-HNSCC. We investigated the safety and therapeutic efficacy of a novel fibroblast activation protein (FAP)-targeted CD40 agonist (FAP-CD40) in combination with local hypofractionated radiation in a syngeneic HPV
-HNSCC model.
Using an established orthotopic model, we treated tumor-bearing mice with local hypofractionated radiotherapy (2 × 6 Gy) alone or in combination with a systemic administration of the FAP-CD40 antibody. Following up the mice, we evaluated the changes in the tumor microenvironment (TME) by immunofluorescence, FACS, and NanoString RNA analysis.
The suboptimal radiotherapy regimen chosen failed to control tumors in the treated mice. The FAP-CD40 administered in monotherapy transiently controlled tumor growth, whereas the combined therapy induced durable complete responses in more than 80% of the tumor-bearing mice. This notable efficacy relied on the radiotherapy-induced remodeling of the TME and activation of the CD8
T-cell-cDC1 axis and was devoid of the systemic toxicity frequently associated with CD40-targeted therapy. Moreover, the robust immunologic memory developed effectively prevented tumor relapses, a common feature in patients with HNSCC.
Our study provides proof of concept, as well as mechanistic insights of the therapeutic efficacy of a bispecific FAP-CD40 combined with local radiotherapy in a FAP
-HNSCC model increasing overall survival and inducing long-term antitumor immunity.
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