To noninvasively assess vascular hemodynamics with photoacoustic imaging (PAI) and blood oxygenation level-dependent (BOLD) magnetic resonance (MR) imaging in phantoms and in an animal model. In vivo studies were performed with institutional animal care and use committee approval. In vitro experiments were performed by using a tissue-mimicking phantom in multiple oxygenation conditions (n = 6) to compare PAI measurements and BOLD MR imaging measurements. PAI and T2-weighted spin-echo-based BOLD MR imaging were performed to assess tumor response to carbogen (95% O2, 5% CO2) in mice with head and neck tumors before (n = 11) and after (n = 9) treatment with a vascular disrupting agent (VDA). Two-tailed Pearson correlation analysis was performed to determine the correlation between the parameters measured with PAI and BOLD MR imaging in vitro. Two-tailed paired t tests were used to compare change in tumor hemoglobin oxygen saturation (sO2) levels and BOLD signal in response to carbogen. Changes in PAI and BOLD signal intensity before and after VDA treatment were analyzed for significance by using analysis of variance with repeated measures. Phantom measurements yielded good correlation between photoacoustically derived sO2 levels and BOLD signal intensity (r = 0.937, P = .005) and partial pressure of oxygen (r = 0.981, P = .005). In vivo hemodynamic response to carbogen was characterized by a significant increase in tumor sO2 levels (P = .003) and BOLD signal (P = .001). When compared with pretreatment estimates, treatment with VDA resulted in a significant reduction in the tumor hemodynamic response to carbogen at PAI (P = .030). Carbogen-based functional imaging with PAI and BOLD MR imaging enables monitoring of early changes in tumor hemodynamics after vascular targeted therapy.

Hypoxia is a recognized characteristic of tumors that influences efficacy of radiotherapy (RT). Photoacoustic imaging (PAI) is a relatively new imaging technique that exploits the optical characteristics of hemoglobin to provide information on tissue oxygenation. In the present study, PAI based measures of tumor oxygen saturation (%sO₂) were compared to oxygen-enhanced magnetic resonance imaging (MRI) measurements of longitudinal relaxation rate (R1 = 1/T1) and ex-vivo histology in patient derived xenograft (PDX) models of head and neck cancer. PAI was utilized to assess early changes (24 h) in %sO₂ following RT and chemoRT (CRT) and to assess changes in salivary gland hemodynamics following radiation. A significant increase in tumor %sO₂ and R1 was observed following oxygen inhalation. Good spatial correlation was observed between PAI, MRI and histology. An early increase in %sO₂ after RT and CRT detected by PAI was associated with significant tumor growth inhibition. Twenty four hours after RT, PAI also detected loss of hemodynamic response to gustatory stimulation in murine salivary gland tissue suggestive of radiation-induced vascular damage. Our observations illustrate the utility of PAI in detecting tumor and normal tissue hemodynamic response to radiation in head and neck cancers.

The negative impact of tumor hypoxia on radiotherapeutic efficacy is well recognized. However, an easy to use, reliable imaging method for assessment of tumor oxygenation in routine clinical practice remains elusive. Photoacoustic imaging (PAI) is a relatively new imaging technique that utilizes a combination of light and ultrasound (US) to enable functional imaging of tumor hemodynamic characteristics in vivo. Several clinical trials are currently evaluating the utility of PAI in cancer detection for breast, thyroid, and prostate cancer. Here, we evaluated the potential of PAI for rapid, label-free, non-invasive quantification of tumor oxygenation as a biomarker of radiation response in head and neck cancer.

Purpose: This research investigates the association between benzodiazepines (BZD) and cancer patient survival outcomes, the pancreatic cancer tumor microenvironment, and cancer-associated fibroblast (CAF) signaling. Experimental Design: Multivariate Cox regression modeling was used to retrospectively measure associations between Roswell Park cancer patient survival outcomes and BZD prescription records. IHC, H&E, Masson's trichrome, RNAscope, and RNA sequencing were used to evaluate the impact of lorazepam (LOR) on the murine PDAC tumor microenvironment. ELISA and qPCR were used to determine the impact of BZDs on IL6 expression or secretion by human-immortalized pancreatic CAFs. PRESTO-Tango assays, reanalysis of PDAC single-cell sequencing/TCGA data sets, and GPR68 CRISPRi knockdown CAFs were used to determine the impact of BZDs on GPR68 signaling. Results: LOR is associated with worse progression-free survival (PFS), whereas alprazolam (ALP) is associated with improved PFS, in pancreatic cancer patients receiving chemotherapy. LOR promotes desmoplasia (fibrosis and extracellular matrix protein deposition), inflammatory signaling, and ischemic necrosis. GPR68 is preferentially expressed on human PDAC CAFs, and n-unsubstituted BZDs, such as LOR, significantly increase IL6 expression and secretion in CAFs in a pH and GPR68-dependent manner. Conversely, ALP and other GPR68 n-substituted BZDs decrease IL6 in human CAFs in a pH and GPR68-independent manner. Across many cancer types, LOR is associated with worse survival outcomes relative to ALP and patients not receiving BZDs. Conclusions: We demonstrate that LOR stimulates fibrosis and inflammatory signaling, promotes desmoplasia and ischemic necrosis, and is associated with decreased pancreatic cancer patient survival.

High macrophage infiltration in cancer is associated with reduced survival in animal models and in patients. This reflects a shift in the macrophage response from a tumor-suppressive to tumor-supportive program governed by transcriptional events regulated by the inflammatory milieu. Although several transcription factors are known to drive a prometastatic program, those that govern an antimetastatic program are less understood. IFN regulatory factor-8 (IRF8) is integral for macrophage responses against infections. Using a genetic loss-of-function approach, we tested the hypothesis that IRF8 expression in macrophages governs their capacity to inhibit metastasis. We found that: (a) metastasis was significantly increased in mice with IRF8-deficient macrophages; (b) IRF8-deficient macrophages displayed a program enriched for genes associated with metastasis; and (c) lower IRF8 expression correlated with reduced survival in human breast and lung cancer, as well as melanoma, with high or low macrophage infiltration. Thus, a macrophage ^hi IRF8 ^hi signature was more favorable than a macrophage ^hi IRF8 ^lo signature. The same held true for a macrophage ^lo IRF8 ^hi vs. a macrophage ^lo IRF8 ^lo signature. These data suggest that incorporating IRF8 expression levels within a broader macrophage signature or profile strengthens prognostic merit. Overall, to our knowledge, our findings reveal a previously unrecognized role for IRF8 in macrophage biology to control metastasis or predict outcome.