Inhibition of MYC translation through targeting of the newly identified PHB-eIF4F complex as a therapeutic strategy in CLL.

Dysregulation of messenger RNA (mRNA) translation, including preferential translation of mRNA with complex 5' untranslated regions such as the MYC oncogene, is recognized as an important mechanism in cancer. Here, we show that both human and murine chronic lymphocytic leukemia (CLL) cells display a high translation rate, which is inhibited by the synthetic flavagline FL3, a prohibitin (PHB)-binding drug. A multiomics analysis performed in samples from patients with CLL and cell lines treated with FL3 revealed the decreased translation of the MYC oncogene and of proteins involved in cell cycle and metabolism. Furthermore, inhibiting translation induced a proliferation arrest and a rewiring of MYC-driven metabolism. Interestingly, contrary to other models, the RAS-RAF-(PHBs)-MAPK pathway is neither impaired by FL3 nor implicated in translation regulation in CLL cells. Here, we rather show that PHBs are directly associated with the eukaryotic initiation factor (eIF)4F translation complex and are targeted by FL3. Knockdown of PHBs resembled FL3 treatment. Importantly, inhibition of translation controlled CLL development in vivo, either alone or combined with immunotherapy. Finally, high expression of translation initiation-related genes and PHBs genes correlated with poor survival and unfavorable clinical parameters in patients with CLL. Overall, we demonstrated that translation inhibition is a valuable strategy to control CLL development by blocking the translation of several oncogenic pathways including MYC. We also unraveled a new and direct role of PHBs in translation initiation, thus creating new therapeutic opportunities for patients with CLL.

CMTM6 and CMTM7: New leads for PD-L1 regulation in breast cancer cells undergoing EMT.

Programmed death-ligand 1 (PD-L1) expression has long been used as a biomarker to stratify patients with cancer who will benefit from anti-PD-1/PD-L1 immunotherapy. However, the use of PD-L1 as a biomarker to guide treatment decisions has recently been called into question due to its dynamic and heterogeneous expression within each tumor and among different tumors as well as during tumor cell plasticity. Therefore, understanding the molecular basis of PD-L1 expression would enable delineating its value as a reliable biomarker in the clinic. Here, we provide our perspective on the involvement of CMTM6 and CMTM7 as new lead candidates for the regulation of PD-L1 in breast tumors undergoing an epithelial to mesenchymal transition.

The genomic landscape of nonsmall cell lung carcinoma in never smokers.

Lung cancer is the number one cause of cancer-related death worldwide with cigarette smoking as its major risk factor. Although the incidence of lung cancer in never smokers is rising, this subgroup of patients is underrepresented in genomic studies of lung cancer. Here, we assembled a prospective cohort of 46 never-smoking, nonsmall cell lung cancer (NSCLC) patients and performed whole-exome and low-coverage whole-genome sequencing on tumors and matched germline DNA. We observed fewer somatic mutations, genomic breakpoints and a smaller fraction of the genome with chromosomal instability in lung tumors from never smokers compared to smokers. The lower number of mutations, enabled us to identify TSC22D1 as a potential driver gene in NSCLC. On the other hand, the frequency of mutations in actionable genes such as EGFR and ERBB2 and of amplifications in MET were higher, while the mutation rate of TP53, which is a negative prognostic factor, was lower in never smokers compared to smokers. Together, these observations suggest a more favorable prognosis for never smokers with NSCLC. Classification of somatic mutations into six-substitution type patterns or into 96-substitution type signatures revealed distinct clusters between smokers and never smokers. Particularly, we identified in never smokers signatures related to aging, homologous recombination damage and APOBEC/AID activity as the most important underlying processes of NSCLC. This further indicates that second-hand smoking is not driving NSCLC pathogenesis in never smokers.

Targeting autophagy inhibits melanoma growth by enhancing NK cells infiltration in a CCL5-dependent manner.

While blocking tumor growth by targeting autophagy is well established, its role on the infiltration of natural killer (NK) cells into tumors remains unknown. Here, we investigate the impact of targeting autophagy gene Beclin1 (BECN1) on the infiltration of NK cells into melanomas. We show that, in addition to inhibiting tumor growth, targeting BECN1 increased the infiltration of functional NK cells into melanoma tumors. We provide evidence that driving NK cells to the tumor bed relied on the ability of autophagy-defective tumors to transcriptionally overexpress the chemokine gene CCL5 Such infiltration and tumor regression were abrogated by silencing CCL5 in BECN1-defective tumors. Mechanistically, we show that the up-regulated expression of CCL5 occurred through the activation of its transcription factor c-Jun by a mechanism involving the impairment of phosphatase PP2A catalytic activity and the subsequent activation of JNK. Similar to BECN1, targeting other autophagy genes, such as ATG5, p62/SQSTM1, or inhibiting autophagy pharmacologically by chloroquine, also induced the expression of CCL5 in melanoma cells. Clinically, a positive correlation between CCL5 and NK cell marker NKp46 expression was found in melanoma patients, and a high expression level of CCL5 was correlated with a significant improvement of melanoma patients' survival. We believe that this study highlights the impact of targeting autophagy on the tumor infiltration by NK cells and its benefit as a novel therapeutic approach to improve NK-based immunotherapy.

Identification of beta-arrestin-1 as a diagnostic biomarker in lung cancer.

BACKGROUND: Distinguishing lung adenocarcinoma (ADC) from squamous cell carcinoma (SCC) has a tremendous therapeutic implication. Sometimes, the commonly used immunohistochemistry (IHC) markers fail to discriminate between them, urging for the identification of new diagnostic biomarkers. METHODS: We performed IHC on tissue microarrays from two cohorts of lung cancer patients to analyse the expression of beta-arrestin-1, beta-arrestin-2 and clinically used diagnostic markers in ADC and SCC samples. Logistic regression models were applied for tumour subtype prediction. Parallel reaction monitoring (PRM)-based mass spectrometry was used to quantify beta-arrestin-1 in plasma from cancer patients and healthy donors. RESULTS: Beta-arrestin-1 expression was significantly higher in ADC versus SCC samples. Beta-arrestin-1 displayed high sensitivity, specificity and negative predictive value. Its usefulness in an IHC panel was also shown. Plasma beta-arrestin-1 levels were considerably higher in lung cancer patients than in healthy donors and were higher in patients who later experienced a progressive disease than in patients showing complete/partial response following EGFR inhibitor therapy. CONCLUSIONS: Our data identify beta-arrestin-1 as a diagnostic marker to differentiate ADC from SCC and indicate its potential as a plasma biomarker for non-invasive diagnosis of lung cancer. Its utility to predict response to EGFR inhibitors is yet to be confirmed.

Inhibition of HIF1α-Dependent Upregulation of Phospho-l-Plastin Resensitizes Multiple Myeloma Cells to Frontline Therapy.

The introduction of novel frontline agents in multiple myeloma (MM), like immunomodulatory drugs and proteasome inhibitors, has improved the overall survival of patients. Yet, MM is still not curable, and drug resistance (DR) remains the main challenge. To improve the understanding of DR in MM, we established a resistant cell line (MOLP8/R). The exploration of DR mechanisms yielded an overexpression of HIF1α, due to impaired proteasome activity of MOLP8/R. We show that MOLP8/R, like other tumor cells, overexpressing HIF1α, have an increased resistance to the immune system. By exploring the main target genes regulated by HIF1α, we could not show an overexpression of these targets in MOLP8/R. We, however, show that MOLP8/R cells display a very high overexpression of LCP1 gene (l-Plastin) controlled by HIF1α, and that this overexpression also exists in MM patient samples. The l-Plastin activity is controlled by its phosphorylation in Ser5. We further show that the inhibition of l-Plastin phosphorylation restores the sensitivity of MOLP8/R to immunomodulatory drugs (IMiDs) and proteasome inhibitors (PIs). Our results reveal a new target gene of DR, controlled by HIF1α.

Exosomes released by chronic lymphocytic leukemia cells induce the transition of stromal cells into cancer-associated fibroblasts.

Exosomes derived from solid tumor cells are involved in immune suppression, angiogenesis, and metastasis, but the role of leukemia-derived exosomes has been less investigated. The pathogenesis of chronic lymphocytic leukemia (CLL) is stringently associated with a tumor-supportive microenvironment and a dysfunctional immune system. Here, we explore the role of CLL-derived exosomes in the cellular and molecular mechanisms by which malignant cells create this favorable surrounding. We show that CLL-derived exosomes are actively incorporated by endothelial and mesenchymal stem cells ex vivo and in vivo and that the transfer of exosomal protein and microRNA induces an inflammatory phenotype in the target cells, which resembles the phenotype of cancer-associated fibroblasts (CAFs). As a result, stromal cells show enhanced proliferation, migration, and secretion of inflammatory cytokines, contributing to a tumor-supportive microenvironment. Exosome uptake by endothelial cells increased angiogenesis ex vivo and in vivo, and coinjection of CLL-derived exosomes and CLL cells promoted tumor growth in immunodeficient mice. Finally, we detected α-smooth actin-positive stromal cells in lymph nodes of CLL patients. These findings demonstrate that CLL-derived exosomes actively promote disease progression by modulating several functions of surrounding stromal cells that acquire features of cancer-associated fibroblasts.

BAT3 modulates p300-dependent acetylation of p53 and autophagy-related protein 7 (ATG7) during autophagy.

Autophagy is regulated by posttranslational modifications, including acetylation. Here we show that HLA-B-associated transcript 3 (BAT3) is essential for basal and starvation-induced autophagy in embryonic day 18.5 BAT3(-/-) mouse embryos and in mouse embryonic fibroblasts (MEFs) through the modulation of p300-dependent acetylation of p53 and ATG7. Specifically, BAT3 increases p53 acetylation and proautophagic p53 target gene expression, while limiting p300-dependent acetylation of ATG7, a mechanism known to inhibit autophagy. In the absence of BAT3 or when BAT3 is located exclusively in the cytosol, autophagy is abrogated, ATG7 is hyperacetylated, p53 acetylation is abolished, and p300 accumulates in the cytosol, indicating that BAT3 regulates the nuclear localization of p300. In addition, the interaction between BAT3 and p300 is stronger in the cytosol than in the nucleus and, during starvation, the level of p300 decreases in the cytosol but increases in the nucleus only in the presence of BAT3. We conclude that BAT3 tightly controls autophagy by modulating p300 intracellular localization, affecting the accessibility of p300 to its substrates, p53 and ATG7.

A high rate of telomeric sister chromatid exchange occurs in chronic lymphocytic leukaemia B-cells.

Cancer cells protect their telomere ends from erosion through reactivation of telomerase or by using the Alternative Lengthening of Telomere (ALT) mechanism that depends on homologous recombination. Chronic lymphocytic leukaemia (CLL) B cells are characterized by almost no telomerase activity, shelterin deregulation and telomere fusions. To characterize telomeric maintenance mechanisms in B-CLL patients, we measured their telomere length, telomerase expression and the main hallmarks of the ALT activity i.e. C-circle concentration, an extra-chromosomal telomere repeat (ECTR), and the level of telomeric sister chromatid exchange (T-SCE) rate. Patients showed relative homogenous telomere length although almost no TERT transcript and nearly no C-circle were evidenced. Nevertheless, compared with normal B cells, B-CLL cells showed an increase in T-SCE rate that was correlated with a strong down-regulation of the topoisomerase III alpha (TOP3A) expression, involved in the dissolution of Holliday Junctions (HJ), together with an increased expression of SLX1A, SLX4, MUS81 and GEN1, involved in the resolution of HJ. Altogether, our results suggest that the telomere maintenance mechanism of B-CLL cells do not preferentially use telomerase or ALT. Rather, the rupture of the dissolvasome/resolvasome balance may increase telomere shuffling that could homogenize telomere length, slowing telomere erosion in this disease.

Randomized Phase II Study of Cabazitaxel Versus Methotrexate in Patients With Recurrent and/or Metastatic Squamous Cell Carcinoma of the Head and Neck Previously Treated With Platinum-Based Therapy.

LESSONS LEARNED: Cabazitaxel has activity in squamous cell carcinoma of the head and neck (SCCHN) and taxane-resistant cell lines. For the first time, cabazitaxel was investigated in incurable patients with recurrent SCCHN. Patients were randomly assigned to cabazitaxel every 3 weeks or weekly methotrexate.This phase II study did not meet its primary endpoint.Cabazitaxel has low activity in SCCHN.The toxicity profile in this population also was not favorable owing to the high rate of febrile neutropenia observed (17%). BACKGROUND: Cabazitaxel is a second-generation taxane that improves the survival of patients with metastatic castrate-resistant prostate cancer following docetaxel therapy. Cabazitaxel has activity in squamous cell carcinoma of the head and neck (SCCHN) and taxane-resistant cell lines. In this randomized phase II trial, we investigated cabazitaxel in patients with recurrent SCCHN. METHODS: Patients with incurable SCCHN with progression after platinum-based therapy were randomly assigned to cabazitaxel every 3 weeks (cycle 1, 20 mg/m2, increased to 25 mg/m2 for subsequent cycles in the absence of nonhematological adverse events [AEs] greater than grade 2 and hematological AEs greater than grade 3) or methotrexate (40 mg/m2/week). The patients were stratified according to their performance status and previous platinum-based chemotherapy for palliation versus curative intent. The primary endpoint was the progression-free survival rate (PFSR) at 18 weeks. RESULTS: Of the 101 patients, 53 and 48, with a median age of 58.0 years (range, 41-80), were randomly assigned to cabazitaxel or methotrexate, respectively. The PFSR at 18 weeks was 13.2% (95% confidence interval [CI], 5%-25%) for cabazitaxel and 8.3% (95% CI, 2%-20%) for methotrexate. The median progression-free survival was 1.9 months in both arms. The median overall survival was 5.0 and 3.6 months for cabazitaxel and methotrexate, respectively. More patients experienced serious adverse events with cabazitaxel than with methotrexate (54% vs. 36%). The most common drug-related grade 3-4 AE in the cabazitaxel arm was febrile neutropenia (17.3%). CONCLUSION: This study did not meet its primary endpoint. Cabazitaxel has low activity in recurrent SCCHN.

Granzyme B degradation by autophagy decreases tumor cell susceptibility to natural killer-mediated lysis under hypoxia.

Recent studies demonstrated that autophagy is an important regulator of innate immune response. However, the mechanism by which autophagy regulates natural killer (NK) cell-mediated antitumor immune responses remains elusive. Here, we demonstrate that hypoxia impairs breast cancer cell susceptibility to NK-mediated lysis in vitro via the activation of autophagy. This impairment was not related to a defect in target cell recognition by NK cells but to the degradation of NK-derived granzyme B in autophagosomes of hypoxic cells. Inhibition of autophagy by targeting beclin1 (BECN1) restored granzyme B levels in hypoxic cells in vitro and induced tumor regression in vivo by facilitating NK-mediated tumor cell killing. Together, our data highlight autophagy as a mechanism underlying the resistance of hypoxic tumor cells to NK-mediated lysis. The work presented here provides a cutting-edge advance in our understanding of the mechanism by which hypoxia-induced autophagy impairs NK-mediated lysis in vitro and paves the way for the formulation of more effective NK cell-based antitumor therapies.

Quantification of SAA1 and SAA2 in lung cancer plasma using the isotype-specific PRM assays.

The quantification of plasma proteins using the high resolution and accurate mass (HR/AM)-based parallel reaction monitoring (PRM) method provides an immediate benefit over the conventional SRM-based method in terms of selectivity. In this study, multiplexed PRM assays were developed to analyze isotypes of serum amyloid A (SAA) proteins in human plasma with a focus on SAA1 and SAA2. Elevated plasma levels of these proteins in patients diagnosed with lung cancer have been reported in previous studies. Since SAA1 and SAA2 are highly homologous, the available immunoassays tend to overestimate their concentrations due to cross-reactivity. On the other hand, when mass spectrometry (MS)-based assays are used, the presence of the several allelic variants may result in a problem of underestimation. In the present study, eight peptides that represent the target proteins at three different levels: isotype-specific (SAA1α,  SAA 1ß,  SAA1γ,  SAA2α,  SAA2ß), protein-specific (SAA1 or SAA2), and pan SAA (SAA1 and SAA2) were chosen to differentiate SAAs in lung cancer plasma samples using a panel of PRM assays. The measurement of specific isotypes, leveraging the analytical performance of PRM, allowed to quantify the allelic variants of both target proteins. The isotypes detected were corroborated with the genetic information obtained from the same samples. The combination of SAA2α and SAA2ß assays representing the total SAA2 concentration demonstrated a superior analytical outcome than the previously used assay on the common peptide when applied to the detection of lung cancer.

Verification of the biomarker candidates for non-small-cell lung cancer using a targeted proteomics approach.

Lung cancer, with its high metastatic potential and high mortality rate, is the worldwide leading cause of cancer-related deaths. High-throughput "omics"-based platforms have accelerated the discovery of biomarkers for lung cancer, and the resulting candidates are to be evaluated for their diagnostic potential as noninvasive biomarkers. The evaluation of the biomarker candidates involves the quantitative measurement of large numbers of proteins in bodily fluids using advanced mass spectrometric techniques. In this study, a robust pipeline based on targeted proteomics was developed for biomarker verification in plasma samples and applied to verifying lung cancer biomarker candidates. Highly multiplexed liquid chromatrography-selected reaction monitoring (LC-SRM) assays for 95 potential tumor markers for non-small-cell lung cancer (NSCLC) were generated to screen plasma samples obtained from 72, early to late stage, patients. A total of 17 proteins were verified as potent tumor markers detectable in plasma and, where available, verified by enzyme-linked immunosorbent assays (ELISAs). A novel plasma-based biomarker, zyxin, fulfilled the criteria for a potential early diagnostic marker for NSCLC.

MicroRNA as biomarkers and regulators in B-cell chronic lymphocytic leukemia.

Early cancer detection and disease stratification or classification are critical to successful treatment. Accessible, reliable, and informative cancer biomarkers can be medically valuable and can provide some relevant insights into cancer biology. Recent studies have suggested improvements in detecting malignancies by the use of specific extracellular microRNAs (miRNAs) in plasma. In chronic lymphocytic leukemia (CLL), an incurable hematologic disorder, sensitive, early, and noninvasive diagnosis and better disease classification would be very useful for more effective therapies. We show here that circulating miRNAs can be sensitive biomarkers for CLL, because certain extracellular miRNAs are present in CLL patient plasma at levels significantly different from healthy controls and from patients affected by other hematologic malignancies. The levels of several of these circulating miRNAs also displayed significant differences between zeta-associated protein 70 (ZAP-70)(+) and ZAP-70(-) CLL. We also determined that the level of circulating miR-20a correlates reliably with diagnosis-to-treatment time. Network analysis of our data, suggests a regulatory network associated with BCL2 and ZAP-70 expression in CLL. This hypothesis suggests the possibility of using the levels of specific miRNAs in plasma to detect CLL and to determine the ZAP-70 status.

Glioblastoma-instructed microglia transition to heterogeneous phenotypic states with phagocytic and dendritic cell-like features in patient tumors and patient-derived orthotopic xenografts.

BACKGROUND: A major contributing factor to glioblastoma (GBM) development and progression is its ability to evade the immune system by creating an immune-suppressive environment, where GBM-associated myeloid cells, including resident microglia and peripheral monocyte-derived macrophages, play critical pro-tumoral roles. However, it is unclear whether recruited myeloid cells are phenotypically and functionally identical in GBM patients and whether this heterogeneity is recapitulated in patient-derived orthotopic xenografts (PDOXs). A thorough understanding of the GBM ecosystem and its recapitulation in preclinical models is currently missing, leading to inaccurate results and failures of clinical trials. METHODS: Here, we report systematic characterization of the tumor microenvironment (TME) in GBM PDOXs and patient tumors at the single-cell and spatial levels. We applied single-cell RNA sequencing, spatial transcriptomics, multicolor flow cytometry, immunohistochemistry, and functional studies to examine the heterogeneous TME instructed by GBM cells. GBM PDOXs representing different tumor phenotypes were compared to glioma mouse GL261 syngeneic model and patient tumors. RESULTS: We show that GBM tumor cells reciprocally interact with host cells to create a GBM patient-specific TME in PDOXs. We detected the most prominent transcriptomic adaptations in myeloid cells, with brain-resident microglia representing the main population in the cellular tumor, while peripheral-derived myeloid cells infiltrated the brain at sites of blood-brain barrier disruption. More specifically, we show that GBM-educated microglia undergo transition to diverse phenotypic states across distinct GBM landscapes and tumor niches. GBM-educated microglia subsets display phagocytic and dendritic cell-like gene expression programs. Additionally, we found novel microglial states expressing cell cycle programs, astrocytic or endothelial markers. Lastly, we show that temozolomide treatment leads to transcriptomic plasticity and altered crosstalk between GBM tumor cells and adjacent TME components. CONCLUSIONS: Our data provide novel insights into the phenotypic adaptation of the heterogeneous TME instructed by GBM tumors. We show the key role of microglial phenotypic states in supporting GBM tumor growth and response to treatment. Our data place PDOXs as relevant models to assess the functionality of the TME and changes in the GBM ecosystem upon treatment.

Cooperative effects of Janus and Aurora kinase inhibition by CEP701 in cells expressing Jak2V617F.

The Janus kinase 2 mutant V617F occurs with high frequency in myeloproliferative neoplasms. Further mutations affecting the Janus kinase family have been discovered mostly in leukaemias and in myeloproliferative neoplasms. Owing to their involvement in neoplasia, inflammatory diseases and in the immune response, Janus kinases are promising targets for kinase inhibitor therapy in these disease settings. Various quantitative assays including two newly developed screening assays were used to characterize the function of different small-molecule compounds in cells expressing Jak2V617F. A detailed comparative analysis of different Janus kinase inhibitors in our quantitative assays and the subsequent characterization of additional activities demonstrated for the first time that the most potent Jak2 inhibitor in our study, CEP701, also targets Aurora kinases. CEP701 shows a unique combination of both activities which is not found in other compounds also targeting Jak2. Furthermore, colony forming cell assays showed that Janus kinase 2 inhibitors preferentially suppressed the growth of erythroid colonies, whereas inhibitors of Aurora kinases preferentially blocked myeloid colony growth. CEP701 demonstrated a combined suppression of both colony types. Moreover, we show that combined application of a Janus and an Aurora kinase inhibitor recapitulated the effect observed for CEP701 but might allow for more flexibility in combining both activities in clinical settings, e.g. in the treatment of myeloproliferative neoplasms. The newly developed screening assays are high throughput compatible and allow an easy detection of new compounds with Janus kinase 2 inhibitory activity.

Real-world experience with isatuximab in the treatment of relapsed-refractory multiple myeloma: a case series from the Grand Duchy of Luxembourg.

BACKGROUND & OBJECTIVE: Anti-CD38 targeting has become an important pillar of the treatment for patients with multiple myeloma (MM). This evolution was spearheaded by daratumumab, but more recently isatuximab became the second CD38-directed monoclonal antibody to receive EMA approval for the treatment of patients with relapsed/refractory (RR) MM. In recent years, real-world studies have become increasingly important to confirm and solidify the clinical potential of novel anti-myeloma therapies. METHODS: This article describes the real-world experience with isatuximab-based therapy in a selection of four RRMM patients treated with an isatuximab-based treatment regimen in the Grand Duchy of Luxembourg. RESULTS: Three of the four cases described in this article consist of heavily pretreated patients who were previously exposed to daratumumab-based therapy. Interestingly, the isatuximab-based therapy provided clinical benefit to all three of these patients illustrating that prior exposure to an anti-CD38 mAb does not preclude a response to isatuximab. As such, these findings further support the design of larger prospective studies looking into the impact of prior daratumumab use on the efficacy of isatuximab-based therapy. In addition, two of the cases included in this report displayed renal insufficiency and the experience with isatuximab in these patients further supports the use of this agent in this setting. CONCLUSION: the clinical cases described illustrate the clinical potential of isatuximab-based treatment for RRMM patient in a real-world setting.

Lighting Up the Fire in the Microenvironment of Cold Tumors: A Major Challenge to Improve Cancer Immunotherapy.

Immunotherapy includes immune checkpoint inhibitors (ICI) such as antibodies targeting cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) or the programmed cell death protein/programmed death ligand 1 (PD-1/PD-L1) axis. Experimental and clinical evidence show that immunotherapy based on immune checkpoint inhibitors (ICI) provides long-term survival benefits to cancer patients in whom other conventional therapies have failed. However, only a minority of patients show high clinical benefits via the use of ICI alone. One of the major factors limiting the clinical benefits to ICI can be attributed to the lack of immune cell infiltration within the tumor microenvironment. Such tumors are classified as "cold/warm" or an immune "desert"; those displaying significant infiltration are considered "hot" or inflamed. This review will provide a brief summary of different tumor properties contributing to the establishment of cold tumors and describe major strategies that could reprogram non-inflamed cold tumors into inflamed hot tumors. More particularly, we will describe how targeting hypoxia can induce metabolic reprogramming that results in improving and extending the benefit of ICI.